DMF File of Vildagliptin

DMF FILE OF VILDAGLIPTIN
Catalogue
3.2.S.1 Basic Information...............................................................................................................4
3.2.S.1.1 Drug Name................................................................................................................4
3.2.S.1.2 Structure.................................................................................................................... 4
3.2.S.1.3 Physicochemical Property........................................................................................ 4
3.2.S.2 Manufacture Information................................................................................................. 5
3.2.S.2.1 Manufacturer.............................................................................................................5
3.2.S.2.2 Manufacturing Technique & Process Control.......................................................... 5
3.2.S.2.3 Control of materials................................................................................................ 12
3.2.S.2.4 Controls of critical steps & Intermediates.............................................................. 23
3.2.S.2.5 Verification of process & Evaluation......................................................................35
3.2.S.2.6 Development of Production Technology................................................................ 38
3.2.S.3 Characterization.............................................................................................................. 50
3.2.S.3.1 Structure Confirmation & Physicochemical Property............................................50
3.2.S.3.2 Impurity...................................................................................................................63
3.2.S.4 Quality Control of API....................................................................................................... 72
3.2.S.4.1 Quality Standard..................................................................................................... 72
3.2.S.4.2 Analytical Procedure...............................................................................................75
3.2.S.4.2.1 Appearance.......................................................................................................75
3.2.S.4.2.2 Identification....................................................................................................79
3.2.S.4.2.3 Inspection.........................................................................................................80
3.2.S.4.2.4 Determination of Content................................................................................ 95
3.2.S.4.3 Verification of Analytical procedures ................................................................. 105
3.2.S.4.3.1 Methodological Verification of related substances inspection.........................105
3.2.S.4.3.2 Methodological Verification of enantiomer checking...................................... 161
3.2.S.4.3.3 Methodology validation of residual solvents & 3- amino -1- amantadine...... 174
3.2.S.4.3.4 Methodology validation of content.................................................................. 203
3.2.S.4.3.5 Verification of content determination by potentiometric titration....................220
3.2.S.4.3.6 Validation of ethyl chloroacetate method......................................................... 228
3.2.S.4.3.7 Impurities I........................................................................................................230
3.2.S.4.3.8Verification of trifluoroacetic acid method........................................................231
3.2.S.4.3.9Hygroscopicity...................................................................................................233
3.2.S.4.5 The basis of quality standard formulation..........................................................234
3.2.S.4.5.1 Appearance........................................................................................................234
3.2.S.4.5.2 Solubility...........................................................................................................234
3.2.S.4.5.3 Melting point.....................................................................................................234
3.2.S.4.5.4 Specific rotation................................................................................................235
3.2.S.4.5.5 Identification..................................................................................................... 236
3.2.S.4.5.6 Moisture............................................................................................................ 236
3.2.S.4.5.7 Sulfate............................................................................................................... 236
3.2.S.4.5.8 Chloride.............................................................................................................237
3.2.S.4.5.9 Related Substances........................................................................................... 237
3.2.S.4.5.10 Residual solvent &3- amino -1- amantadine..................................................243
2
3.2.S.4.5.11 Trifluoroacetic acid ....................................................................................... 245

3.2.S.4.5.12 Impurities I......................................................................................................245
3.2.S.4.5.13 Ethyl chloroacetate......................................................................................... 246
3.2.S.4.5.14 Enantiomer......................................................................................................247
3.2.S.4.5.15 Ignition residue............................................................................................... 248
3.2.S.4.5.16 Heavy metal.................................................................................................. 2518
3.2.S.4.5.17 Content（Potentiometric titration）............................................................ 2518
3.2.S.4.5.18 Content（HPLC）....................................................................................... 2529
3.2.S.4.5.19 Microbial limit.............................................................................................. 2552
3.2.S.5 Packaging materials& containers................................................................................. 2552
3.2.S.6STability.............................................................................................................................. 254
3.2.S.6.1 Stability summary................................................................................................ 254
3.2.S.6.2 Stability commitment & stability scheme after listing........................................257
3.2.S.6.3 Stability data........................................................................................................ 259
3.2.S.6.3.1 Influence factor test.......................................................................................... 260
3.2.S.6.3.2 Accelerated test................................................................................................. 263
3.2.S.6.3.3 Long term test................................................................................................... 270
3
3.2.S.1 Basic Information
3.2.S.1.1 Drug Name
Drug Name:Vildagliptin
Chemical name：(2S)-1-[[(3-Hydroxytricyclo[3.3.1.1[3,7]]dec-1-yl)amino]acetyl]-2-
pyrrolidinecarbonitrile
CAS Number ：274901-16-5
3.2.S.1.2 Structure
structural formula ：
Molecular formula：C17H25N3O2
Molecular weight：303.40
Spatial structure ： The structure of vildagliptin contains 1 chiral carbon atoms, the product is S
configuration
Crystal form ： This product is a crystalline solid which didn’t contain crystal water or solvent.
Crystal type is A form.
3.2.S.1.3 Physicochemical Properties
Table 3.2.S.1.3-1 Physicochemical properties of vildagliptin
Physical& chemical project Property description
Character White to yellowish or slightly gray crystalline powder
Melting point 146℃～151℃
Soluble in methanol, ethanol, DMSO, Water or 0.01mol/L hydrochloric acid solution,

Solubility
slightly soluble in acetonitrile or isopropanol
Specific rotation -84° ～ -90°
Under the condition of 25℃ and relative humidity 80% + 2%, the product can absorb
Hygroscopicity moisture and increase weight less than 0.2% in 24 hours. Therefore, the product has
almost no Humidifying.
4
3.2.S.2 Manufacture Information

3.2.S.2.1 Manufacturer
Manufacturer Name：Anhui Haikang Pharmaceutical Co. Ltd.
Address：No.21, Huancheng West Road, Daguan District,Anqing, Anhui, China.
Tel：+86-0556-5800026 Fax ：+86-0556-5800080
Factory Address：Anhui Haikang Pharmaceutical Co. Ltd. Workshop
3.2.S.2.2 Manufacturing Technique & Process Control
1. Route of Synthesis
Our company synthetic route as below：
质控点
HO HO
K2CO3 N
N
N
Cl NH2 DMF CH3CH2 OH H C
C O
O N
N C 10 H17NO C17H25 N3 O2
C 7H9ClN2O
Mol. Wt.: 172.61
Mol. Wt.: 167.25 步骤1 Mol. Wt.: 303.40
起始原料1 起始物料2 粗品：维达列汀
CH3 CH2OH
维达列汀成品
步骤2
Note 1：
Step 1：Step 1:DMF, potassium carbonate / heat preservation reaction temperature 55~60, 4.5~5h; concentration
temperature 90~95 C; anhydrous ethanol / dissolution temperature 60~70, 1~1.5h, crystallization -5~0 C, 8~9h;
isopropanol, purified water / 60~70 C, 1~1.5h, crystallization -5~0 C, 8~9h;
Step2：Absolute ethanol / solution temperature 60~70 C, 1~1.5h, crystallization temperature -5~0.,8~10h;
Note 2：SM-1：Starting material 1，(2S) -N- chloracetyl -2- cyanopyrrolidine
SM-2：Starting material 2，3- amino -1- amantadol
5
2. Process Flow Diagram

Step1：Initial synthesis of vildagliptin
(2S) -N- chloracetyl

-2- cyanopyrrolidine
Mixing and
dissolving
DMF
DMF
Heat preservation
3- amino -1- amantadol Heating up to
reaction at 55~60 C
55～60℃
4.5 ~ 5h
potassium carbonate
Send to Environmental
filter cake protection center
Colling to
filter
15～25℃
Vacuum concentration at the
filtrate end,vacuum degree is -0.09～
-0.10MPa ，and the temperature
inside the kettle is 90~95℃
Heating up to 60～ Send to

Colling to -5～
70℃ Environmental
0℃, filtrat
Heat preservation protection center
Stirring filter
and stirring 1 ~ 1.5h crystallization 8～
9h filtrat
Filter cake
Filter cake
Anhydrous
ethanol
6
Send to
Environmental
filtrat protection center
filtrat Vacuum drying 10 ~

12h, hot water Initial
Filter cake filter
temperature 60~70 product of
degrees, vacuum degree vildagliptin
Filter cake
-0.080 ~ -0.100MPa
Isopropanol
Step2：Vildagliptin Refining Medicinal

activated carbon
Anhydrous
ethanol Heat
Heating up Mixing and
filter
preservation
decolorizing 30 ~
to 60～70℃ and stirring
40min
Initial product of 1 ~ 1.5h
vildagliptin
Send to Environmental
filtrat protection center
Mixing Heat preservation
and and stirred
filtrat cooling to crystallization filter
-5 ~ 0 ℃ from 8 to 10h Vacuum drying 10 ~ 12h,
Filter hot water temperature
cake 60~70 degrees, vacuum
degree -0.080 ~
-0.100MPa
7
mixture、inner
Shatter
packing
packing cleaning cleaning
Test qualified
Outer packing Outer packing

material material
product storage
D Zone
8
3. Process description
The synthesis of Vildagliptin, use (2S) -N- chloracetyl -2- cyanopyrrolidine and
3- amino -1- amantadol as started material ， get target product by nucleophilic
substitution reaction ， get finished product after refined. Take representative as
following vildagliptin batches，Describe the process according to the process flow，
The amount of intermediates in the process description is represented by the verifying
process.The inventory rating of other batches shall be subject to the actual output of
the previous step.
Step 1：Condensation process，Synthesis of Initial vildagliptin
HO
HO
K2CO3
N
NH2 N
Cl H
O
C O C
N
N
Table 3.2.S.2.2-1 The material input ratio of Initial vildagliptin
Inventory
Working Molecular material Input
Material Name Rating Mole Ratio
Procedure Weight Ratio
(2S) -N- chloracetyl -2-

172.61 100.0kg 1.00 1.00
cyanopyrrolidine
Condensation
3- amino -1- amantadol 167.25 107.0 kg 1.07 1.10
process
potassium carbonate 138.19 128.0kg 1.28 1.60
N,N-Dimethylformamide / 400kg+200kg 4+2 /
Anhydrous ethanol / 158kg 1.58(2V) /
Post-processing Isopropanol / 157kg 1.57(2V) /
Purified water / 20kg 0.2(0.2V) /
9
1)Put (2S) -N- chloracetyl -2- cyanopyrrolidine into the 1000L Enamel glass reaction
kettle，adding 200kgDMF，Stir and dissolve to be used.
2)Open vacuum ，400kg DMF was pumped into the 1000 L Enamel glass reaction
kettle ， Close vacuum ， Open the agitator ， Add 3- amino -1- adamantanes and
potassium carbonate to the reactor，Open steam，Stirring and heating to 55~60 ℃；
3)When The temperature of the liquid in the kettle rise to 55~60 C, slowly adding (2S)
-N- chloracetyl -2- cyanopyrrolidine and DMF mixed solution ，control the liquid
temperature as 55~60℃ in Flow addition process ，after finished，continue to control
as 45~50℃, stir and react about 4.5~5 h；
4)When the reaction of heat preservation and agitation has reached 4.5~5h, Open the
circulating water，Colling to15~25℃，centrifugal filtration，collect filtrate into1000L
Enamel glass reaction kettle，open steam，Decompression and concentration；
5)There is no liquid outflow ， the temperature in the reactor is 90~95 ℃ ， and the
vacuum distillation ends; 158kg ethanol was added to the reactor and 1~1.5h was
stirred at 60~70℃ , liquid material into 500L Enamel glass reaction kettle and slowly
cool down to -5~0℃ ,stirred crystallization 8~9h .
6)The crystallization time has reached 8~9h and centrifuged. The Filter cake was
washed with isopropanol and the solid was collected;
7)adding 157kg isopropanol and 20.0kg purified water into the 200L Enamel glass
reaction kettle，adding the collected filter cake，open steam and hearting to 60~70℃
and stirring 1~1.5h，slowly colling to -5~0℃ and stirring the crystallization 8~9h；
The crystallization time has reached 8~9h ,centrifuged.，The Filter cake was washed
with isopropanol and the cake was collected;
8) The steam was controlled at 60~70 C, the vacuum degree was -0.080 ~ -0.100MPa,
and the vacuum drying 10~12h, and the 135.0kg, Initial of vildagliptin was obtained.
（The molar yield range of 70~80% ）.
Step3：Refining process，vildagliptin refined made
Table 3.2.S.2.2-3 The material input ratio of refining vildagliptin
10
Material name Molecular weight Inventory rating Material input ratio
Initial of vildagliptin 303.40 135.0kg 1.00
Anhydrous ethanol / 213.3kg 1.58(2V)
Medicinal activated carbon / 6.75 kg 0.05
1)Adding anhydrous ethanol into the 500 L Enamel glass reaction kettle，open stir,
add the inventory rating of Initial vildagliptin into the reactor ，finished ，open steam
and hearting up to 60~70℃,stirring and keep warming 1~1.5h；
2)Put the medcinal activated carbon into the reactor ， finished ， continue to keep
warming, stir and decolorize 30~40min,filter；
3)The filtrate was transferred to 500L reactor and then cooled to -5~0 ℃, stirring and
crystallization 8~10h.
4)Centrifuge filtration, collection of filter solid, control steam 60~70 ℃ , vacuum
degree in -0.080 to -0.100MPa, vacuum drying 10~12h, get wildagliptin finished
products, after comminution, mixing and packing, finished product 120.0kg. (The
molar yield range of 80~90%)
4. list of major equipment Manufacture

Table 3.2.S.2.2-4 Vildagliptin mian production equipment
Specification /
Device Name Material Purpose Use steps
model
1000L Enamel glass reaction Substituent Initial vildagliptin
1000L Glass lined
kettle reaction Synthesis
1000LEnamel glass reaction Decompression Initial vildagliptin
1000L Glass lined
kettle distillation Synthesis
500LEnamel glass reaction Initial vildagliptin
500L Glass lined Crystallization
kettle Synthesis
Initial vildagliptin
Centrifuge LB1000 Stainless steel filter
Synthesis
11
Centrifuge LB1000 Stainless steel filter
Low temperature vacuum Initial of vildagliptin

FZG-15 Stainless steel Dry
oven Synthesis
500LEnamel glass reaction
500L Stainless steel Dissolution Refined
kettle
Bag filter SUS304 Stainless steel Pressure filtration Refined
500LEnamel glass reaction

500L Glass lined Crystallization Crystallization
kettle
Centrifuge LBF1000 Stainless steel filter filter
Low temperature vacuum

YZG-1300 Stainless steel Dry Dry
oven
Water Crusher YF3-1 Stainless steel Smash Smash
HD-50 Multi-motion mixer HD-50 Stainless steel Mixture Mixture
5. Large production scale range

The batch quantity of commercial large production area is 115~125kg.
3.2.S.2.3 Control of materials
This chapter provides all materials information used in vildagliptin production,
including starting materials, other raw materials reagents, solvents etc. There are
maybe more than one supplier for the same material, but the quality standard will not
change, and only can confirm our company quality standard then put into production.
12
Table 3.2.S.2.3-1 Summary of production materials of vildagliptin
Chemical
Serial structure or Use
Classification Name Standard
number molecular steps
formula
Quality standard
(2S) -N- chloracetyl N
Cl 1 of enterprise
-2- cyanopyrrolidine O C
1 N internal control
Sichuan Tong Sheng Biological Medicine Co.,
Supplier (producer)
Starting Ltd.
material Quality standard

3- amino -1-
1 of enterprise
amantadol
2 internal control
Jiangsu MAN TAI dazzling Biological

Supplier (producer)
Technology Co., Ltd.
GB/T 1587-2000
and Quality
potassium carbonate K2CO3 1 standard of
Other reagent 3
enterprise
internal control
Supplier (producer) Ji'nan Ren Yuan Chemical Co., Ltd.
10 Isopropanol 1,2 GB/T 7814-2008
Solvent Supplier (producer) Wuhu Sanyou Chemical Trade Co., Ltd

Anhydrous ethanol CH3CH2OH 1,2 GB678-2002
12 Jiangsu flower hall Biological Technology Co.,
Supplier (producer)
Ltd.
13
HG 2028-91and
Quality standard
DMF 1
13 of enterprise
internal control
Supplier (producer) Shandong Jin Hao Chemical Co., Ltd.
Quality standard
Medicinal activated
/ 2 of enterprise
Auxiliary carbon
14 internal control
reagent
Jiangsu Hong Jia Environmental Protection
Supplier (producer)
Technology Co., Ltd
1. Starting Material
1) Selection basis of starting materials
Vildagliptin ， chemical name is (2S) -1-[[(3- hydroxy tricyclic [3.3.1.1[3,7]]
decane -1- base) amino] acetyl]-2- cyano pyrrolidine，There is a C chiral center，S
configuration. In the present mature preparation process, the complete synthesis route
can be summarized as the route shown below:
14
O
HN SOCl2 HN NH3 HN Et3N
Cl
Cl
CONH2
OH O OMe
O
A-2 A-3
A-1
TFAA
Cl N N
Cl
O CONH2 O C
A-5 N
A-4
H O2N HO
HNO3/H2SO4 KOH/H2O
NH2 NH2 NH2
B-1 B-2 B-3
K2CO3
HO
N
N
H
O C
N
Vildagliptin
In order to ensure the safety and controllability of pharmaceutical production,

and considering factors such as production cost and waste disposal, etc. A-5（(2S) -N-
chloracetyl -2- cyanopyrrolidine）and B-3（3- amino -1- amantadol）are designated as
the starting material for synthesis, It is a commercially available chemical compound.
Both of them are significant chemical structural fragments of vildagliptin, and they
can be identified and formulated by means of commonly used analytical methods.Two
starting materials A-5 and B-3 are also included in the study of impurities in API. The
removal process is clear and will not affect the quality of API.
2）Starting Material Control Strategy
A 、(2S) -N- chloracetyl -2- cyanopyrrolidine
According to the manufacturer's (2S) -N- chloracetyl -2- cyanopyrrolidine
preparation route, it is found that (2S) -N- chloracetyl -2- cyanopyrrolidine is obtained
through the acylation of L- prolyl through the acylation reaction of L- prolyl, and then
trifluoroacetic anhydride dehydration reaction is obtained.
15
O Et3N N TFAA Cl N
HN Cl
Cl
Cl O Na2CO3 O
CONH2 CONH2 C
N
VGT-1
According to its synthetic route, the acylation reaction is fast and complete, so
the possibility of the residue of L- prolyamide in the starting material of (2S) -N-
chloracetyl -2- cyanropyrrolidone (VGT-2) is very small; the dehydration reaction is
slower, the VGT-1 has a certain amount of residue at the end point of the reaction, and
VGT-1 has the possibility of conversion of H from hydrolysis to impurity H. as well
as the possibility of enantiomers of (2S) -N- chloroacetyl -2- cyano pyrrolidine
(VGT-2).
Table 3.2.S.2.3-2 Impurity analysis, test results and control of (2S) -N-
chloroacetyl -2- cyano pyrrolidine
Test
Component Name Chemical Structure Source Control Strategy
results
(2S) -N- chloracetyl

Target product 99.123% ≥98.0%
-2- cyanopyrrolidine
l-prolinamide Starting material 0.125% ≤0.5%
Intermediate
VGT-1 0.136% ≤0.5%
（not taken out)
VGT-1
Impurity H decomposition 0.080% ≤0.1%
product
VGT-1 Isomer By-product 0.030% ≤0.1%
16
VGT-2 Isomer By-product 0.069% ≤0.1%
Other unknown Single impurity ≤0.5%

/ yes
impurities Total impurities ≤2.0%
Table 3.2.S.2.3-3 (2S) -N- chloroacetyl -2- cyano pyrrolidine: quality standard and
purity / related substances method
starting material 1
Chemical name：(2S) -N- chloracetyl -2- cyanopyrrolidine
Abbreviations or abbreviations：VGT-2
Molecular formula：C7H9ClN2O
molecular weight：172.61
Detection project Acceptability limit Method index reference
Appearance White to yellow powder Visual measurement
Related substances
Impurity H： ≤0.1%
Internal control standard
Single impurity： ≤0.5%
Total impurities： ≤2.0%
Enantiomer: ≤0.5% Internal control standard
Related substances
High performance liquid chromatography，Area normalization with correction factor
▪High performance liquid chromatograph equipped with ultraviolet detector
▪chromatographic column：Cyanic bond silica gel as a filling agent（such as Agilent ZORBAX
SB-CN，4.6×250mm，5µm）
▪mobile phase：With phosphate buffer solution [taking anhydrous potassium dihydrogen phosphate
17
1.3g, adding water 1000ml to dissolve, using two potassium hydrogen phosphate solution (taking
anhydrous hydrogen phosphate, two potassiumng water 10ml to dissolve) to adjust pH to 6.0]-
water acetonitrile (400:600:15).
▪Detection wavelength：210nm Flow rate：1.0ml/min
▪Injection volume：10μl Column temperature：35℃
Sample solution：Appropriate amount of the product is added to the mobile phase to dissolve and
dilute the solution containing about 1.0mg in each 1ml.
Determination method：Area normalization with correction factor
Correction factor for impurity H (relative to peak retention time 0.30) ： 0.8 ， Other impurity
correction factors：1.0
Enantiomer
High performance liquid chromatography, principal component self-control.
▪chromatographic column：The surface of silica gel is covalently bonded with amylose - three - (3-
chloro -4- methyl phenyl carbamate) as filler (e.g. CHIRALPAK IF 4.6 x 250mm, 5 m m).
▪Mobile phase ：Hexane - anhydrous ethanol -DEA（600:400:1）
▪Detection wavelength：220nm flow rate ：1.0ml/min
▪Injection volume：10μl column temperature：35℃
Sample solution: take appropriate amount of the product and dilute solution to dilute it to make the
solution containing about 1.0mg per 1ml.
Determination method ：（ The retention time of the enantiomer peak relative to the principal
component peak is about 0.65.）Principal component self control method
B、3- amino -1- amantadol control strategy

According to the preparation route of 3- amino -1- amantadol provided by the
manufacturer, the 3- amino -1- adamantol was obtained by nitrification and
alkalization in the nitric acid sulphuric acid system with amantadine as raw material.
18
Table 3.2.S.2.3-4 Impurity analysis, detection results and control of 3- amino -1-
amantadine (SM-2)
Component Name Chemical Structure Source Test Results Control Strategy
Target product
3- amino -1- amantadine 99.1% ≥98.0%
（SM-2）
Starting
Amantadine N.D. ≤0.1%
material
Other unknown impurity ≤1.0%
Table 3.2.S.2.3-5 Quality standard and purity / related substances method of 3-

amino -1- amantadine
starting material 2
Chemical name： 3- amino -1- amantadine
Abbreviations or abbreviations：SM-2
Molecular formula：C10H17NO
Molecular weight：167.25
Appearance White or off-white powder Visual measurement
Identify Chemical reaction method Internal control standard
Related substances
Amantadine： ≤0.1%
Internal control standard
Single impurity： ≤1.0%
Total impurities： ≤2.0%
Residual solvent Internal control standard
19
DCM ≤0.06%
Ignition residue ≤0.5% Internal control standard
Related Substances：
Gas chromatography, amantadine is calculated by external standard method, and other unknown
impurities are normalized by area normalization method.
▪The gas chromatograph is equipped with a hydrogen flame ionization detector (FID).
▪Chromatographic column: 6% cyanopropyl phenyl -94% Dimethyl Polysiloxane as stationary
phase capillary column (KB-624 30m x 0.53mm, 3 m)
▪Chromatographic conditions
Column temperature: maintain 3 minutes at 100 ℃ centigrade, then rise to 210 minutes at
10 ℃per minute for 30 minutes.
injector temperature：250℃。
Detector temperature：280℃
carrier gas： nitrogen
Shunt ratio：10:1
column flow rate：3.0ml/min
Injection volume：1μl
Reference solution: take the appropriate amount of amantadine hydrochloride and add DMSO to
dissolve and dilute the solution containing amantadine 0.1mg in each 1ml.
Sample solution: take appropriate amount of the product and add DMSO to dissolve and dilute
the solution containing about 100mg in each 1ml.
Determination method: amantadine was calculated by external standard method, and other
unknown impurities were calculated by area normalization method (peak area percentage%).
Residual solvent：
Gas chromatography，Calculation of external standard method
▪The gas chromatograph is equipped with a hydrogen flame ionization detector (FID).
▪Chromatographic column: 6% cyanopropyl phenyl -94%Dimethyl Polysiloxane as stationary
phase capillary column (KB-624 30m x 0.53mm, 3 m)
▪Chromatographic conditions
20
Column temperature: maintain 3 minutes at 100 degrees centigrade, then rise to 210 minutes
at 10 degrees per minute for 30 minutes.
injector temperature：250℃
Detector temperature：280℃
carrier gas： nitrogen
Shunt ratio：10:1
Column flow rate：3.0ml/min
Sample solution: take appropriate amount of the product and add DMSO to dissolve and dilute the
solution containing about 100mg in each 1ml.
Reference solution: take appropriate amount of dichloromethane and add DMSO to dissolve and
dilute the solution containing 0.06mg in each 1ml.
Determination method: external standard method
2. Other Reagents
Table 3.2.S.2.3-6 reagents for vildagliptin production
Quality standard
Reference
Name Project Name (enterprise internal Use Steps
Standard
control)
White powderor White powderor Granular
Character
Granular
①
Chloride ≤0.10% ≤0.10%
potassium
inspect (with KCl) 1
carbonate
②loss on
≤1.00% ≤1.00%
ignition
Potassium
≥98.5% ≥98.5%
carbonate Assay
21
3. Solvent
Table 3.2.S.2.3-7 The solvent used for vildagliptin production
Quality standard
Name Project Name Reference Standard (enterprise internal Use steps
control)
Transparency liquid Transparency liquid
Character
The infrared
The infrared absorption
absorption spectrum
spectrum of this
of this product should
Identify Infrared Atlas product should be
be consistent with
consistent with that of
Isopropanol that of reference 1
reference substance.
substance.
①water ≤0.20% ≤0.15%
②water
Inspect Should be clarified Should be clarified
solubility test
③benzene ≤2ppm ≤2ppm
Assaying ≥99.7% ≥99.7%
Colorless and
Colorless and
Character transparent oil
transparent oil liquid
liquid
The retention time of
The retention time of
the main peak should
DMF the main peak should 1
Identify be consistent with
be consistent with that
that of the control
of the control solution.
solution.
inspect(water) ≤0.10% ≤0.10%
Assaying ≥99.0％ ≥99.0％
Colorless, transparent Colorless, transparent
character
and volatile liquid and volatile liquid
The odour of
The odour of iodoform
iodoform should
should occur and the
Identify occur and the Yellow
Yellow precipitation
precipitation should
should be produced.
Anhydrous be produced.
1，2
Ethanol ①water ≤0.2% ≤0.2%
②Evaporative
≤0.002% ≤0.002%
residue
Inspect
③methanol ≤0.02% ≤0.02%
④Isopropanol ≤0.05% ≤0.05%
⑤benzene ≤10ppm ≤10ppm
purity ≥99.5% ≥99.5%
22
4. Auxiliary Reagents
Table 3.2.S.2.3-8 Auxiliary reagents used in vildagliptin production
Quality standard
Name Project Name Reference Standard (enterprise internal Use Steps
control)
Black powder ， Odorless Black powder ， Odorless
Character
and tasteless；no sand and tasteless；no sand
Loss on drying ≤10.0% ≤10.0%
Medicinal No turbidity No turbidity
2
activated carbon The difference between The difference between
the consumption of iodine the consumption of iodine
adsorbability
solution and blank solution and blank
solution should not be solution should not be less
less than 1.2ml than 1.2ml
5. Waste Disposal Scheme

The "three wastes" produced in the process of production mainly are waste liquid
and waste residue. The organic solvent ethanol, isopropanol and dichloromethane in
the waste liquid can be recycled. The wastewater is discharged by proper acid and
alkali neutralization, and the waste residue, such as sodium sulfate and activated
carbon, is sent to the waste disposal station.
Table 3.2.S.2.3-9 Vildagliptin produces three waste Disposal scheme
Production Process Waste Gas Waste Liquid Waste Residue

Ethanol, isopropanol and other
Potassium carbonate
recycling processes; organic waste and potassium
liquid biochemical treatment after chloride are sent to
Initial synthesis / the environmental
discharge; centralized treatment of protection center for
wastewater and centralized centralized
treatment.
discharge.
Concentrated
treatment of active
Purification / Ethanol recovery carbon to
environmental
protection center
23
3.2.S.2.4 Controls of critical steps & Intermediates

1. Key steps and key process parameters
Process route of wildagliptin：
HO HO
Cl N K2CO3
NH2 N
O N
C H
N O C
N
CH3CH2OH
维达列汀成品
The synthesis of vildagliptin，produced the target product by nucleophilic

substitution of (2S) -N- chloroacetyl -2- cyano pyrrolidine with 3- amino -1-
adamantane. The condensation process is the most close to the finished product, and it
is easy to produce the main process impurity disubstituted compound impurity F,
It has great influence on product quality, so it is a key step.
impurity F
Refining process directly affects the quality characteristics of finished products,

so it is also identified as a key step.
During the study of the synthesis process of vergentry, our company completed
the risk assessment of the process and quality of the variety, studied the key quality
attributes such as the related substances, enantiomers, specific rotation and so on,
determined the main process parameters of each step, and carried out a small test and
verified the verification of batch size.
It is proved that all the control parameters are within the controllable range of
industrial production, and it supports the rationality of the determination of various
process parameters and lays a solid foundation for the subsequent production of the
24
varieties.
25
Table 3.2.S.2.4-1 Study on the technology of vildagliptin and risk assessment of its main parameters
Whether or not
Main steps Set value of Developing the
Main process The influence of beyond the it is a key
and process scope of Seriousness Risk assessment
parameters scope on the process process
reactions parameters confirmation
parameter
When the adamantanol
dosage is too low, it will

It is confirmed that the
(2S) -N- increase the possibility of
feeding reaction can be
chloroacetyl -2- two substitution of
Condensati carried out normally
cyano 1.1:1 1.1~1.15:1 impurities. The excessive Middle NO
on process according to the ratio of
pyrrolidine:adamanta feed volume has little effect
1.1:1, and the parameter is
nol on the product quality, but
easy to control.
increases the pressure of
post treatment removal.
26
Too high reaction
temperature has the

This parameter is easy to
Reaction temperature 55~60℃ 55~60℃ possibility of increasing Middle NO
control
impurities, too low and
prolonged reaction time.
The reaction time is short
and the reaction is This parameter is easy to
Reaction time 4.5~5h 4~5h incomplete. The longer the Middle control and can be NO
reaction time has little effect monitored by TLC
on the result.
The low solvent content

This parameter is
Post treatment of affects the purity of the
2V 2V Middle confirmed by a small test Yes
ethanol addition product, and the product
and easy to control
yield is too high.
27
When the crystallization
temperature is too high, the

The crystallization time is
Crystallization product is not easy to
-5~0℃ -5~0℃ obtained through a small
conditions of post precipitate, and the Middle No
8~9h 8~10h test, which is easy to
treatment crystallization time is too
control.
short, which will affect the
yield of the product.
The excessive yield of
isopropanol affects the yield
of products, and the yield of

Solvent content of
water is too large, which This parameter is
wet products
2V+0.2V 2V+0.2V also affects the yield. The Middle confirmed by a small test Yes
(isopropanol +
solvent quantity is too and easy to control
purified water)
small, and the dissolution is
incomplete, and the quality
of products is affected.
28
temperature is too high, the

The crystallization time is
Crystallization product is not easy to
-5~0℃ -5~0℃ obtained through a small
conditions of post precipitate, and the Middle No
8~9h 8~10h test, which is easy to
treatment crystallization time is too
control.
short, which will affect the
60~70℃ 60~70℃
Drying temperature has The parameters are easy to
Decompression Decompression
Drying condition little effect on the quality of Low control by drying to No
drying drying
products. constant weight.
The low dosage of ethanol

The ratio of crystallization
affects the solubility and
Refining The amount of refined solvent is obtained through
2V 2V purity of the product, and High yes
process ethanol small test, which is easy to
the product yield is too
control.
high.
29
High temperature is not
conducive to safe
Dissolution This parameter is easy to
60~70℃ 60~70℃ production, and the low Middle No
temperature control
temperature affects the
dissolution effect.
temperature is too high, the The crystallization solvent
product is not easy to ratio and crystallization
Crystallization mode -5~0℃. 8~10h -5~0℃. 8~10h precipitate, and the Middle time are obtained through a No
crystallization time is too small test, which is easy to
short, which will affect the control.
60~70℃ 60~70℃
Drying temperature has The parameters are easy to
Decompression Decompression
Drying condition little effect on the quality of Low control by drying to No
drying drying
products. constant weight.
30
2. Intermediate control
The Initial product of vildagliptin
Step 1 is the condensation reaction process，the Initial product of vildagliptin was
obtained by (2S) -N- chloroacetyl -2- cyanopyrrolidine and 3- amino -1- adamantane
condensation reaction
HO
N
N
O O C
N
C N
N 杂质 F
HO HO
Cl N K2CO3
O N
C NH2 N
N H
O C
起始物料：起始原料：粗品： VGT N
1 VGT-2 2 SM-2
HO
HO
HO N
N NH
Cl N H
O C
N N
O CONH2 OH 杂质 K
N N
H
O
HO 杂质 I O 杂质 C
N HO
N
H C
O O
O NH2
杂质 D
HO N
N
N O 杂质 E
N 杂质 G
H C
O
O OH
It is known from the synthetic route that if there is a residue of VGT-1 in the
starting material (2S) -N- chloracetyl -2- cyanopyrrolidine, the impurity D may be
produced and then the impurity G is produced. If the chloroacetic acid remains, the
impurity I may be produced during the reaction process.
(2S) -N- chloracetyl -2- cyanopyrrolidine and 3- aminoalkanol nucleophilic
substitution reaction, the reaction process may occur in the amino site two
substitutional and produce impurity F; the target product may be degraded to impurity
C, and then degradation into the impurity E.
After several experimental results, the main impurity of the reaction liquid is the
31
impurity D produced by the residual VGT-1 in VGT-2, the unreacted VGT-2 and the
two substituted impurity F. After reacting, the above impurities can be controlled in
the standard range, and the other impurities can be effectively removed and the purity
of the liquid phase is more than 98%.The other 3- amino -1- amantadine is over fed,
and after treatment, the residue can also be controlled within the standard range.
The isomers of the target products are also one of the potential impurities, but the
chiral carbon in the structure is not a reaction site in the reaction, and it is not easy to
raceme or turn over. After many tests, the isomers are not detected in the Initial
product of VGT.
Table 3.2.S.2.4-5 The Initial product of vildagliptin test results
Second batches of
First batch of small test Third batches of small tests
Related substances（HPLC） small tests
150401 150403
150402
Impurity A（%） N.D N.D N.D
ImpurityB（%） N.D N.D N.D
ImpurityC（%） N.D 0.008 0.008
ImpurityD（%） 0.086 0.084 0.080
ImpurityE（%） N.D N.D N.D
ImpurityF（%） 0.095 0.111 0.136
ImpurityG（%） N.D N.D N.D
Other max single Impurity （%） 0.014 0.019 0.019
Number of impurities 3 4 4
purity（%） 99.805 99.778 99.757
Enantiomer（HPLC） N.D N.D N.D
Table 3.2.S.2.4-6 Analysis of organic impurities in Initial product of vildagliptin
32
Whether or not
Compound Chemical Structure Source to subscribe Control Limit
quality standard
Target
VGT / ≥98.0%
product
Single
VGT-2 Starting
No Impurity≤
（ImpurityB） material
0.5%
Single
VGT-1
VGT-1 No Impurity≤
（ImpurityA）
0.5%
By-products Single
ImpurityC /Degradatio No Impurity≤
n product 0.5%
By-products Single
ImpurityD /Degradatio No Impurity≤
n product 0.5%
33
By-products Single
ImpurityE /Degradatio No Impurity≤
n product 0.5%
Single
ImpurityF By-products No Impurity≤
1.0%
By-products Single
ImpurityG /Degradatio No Impurity≤
n product 0.5%
Single Impurity / / Yes ≤1.0%

Total impurity / / Yes ≤2.0%
Note: no known impurity can be controlled by a single impurity of over 1%.
Table 3.2.S.2.4-7Internal control standard and purity / impurity method of Initial

vildagliptin
The Initial Product of vildagliptin
34
Chemical name ：(2S) -1-[[(3- hydroxy tricyclic [3.3.1.1[3,7]] decane -1- base) amino] acetyl]-2-
cyano pyrrolidine
Abbreviations or abbreviations: Initial VGT
Molecular formula：C17H25N3O2
molecular weight：303.40
White to yellowish or grayish

Appearance Visual measurement
crystalline powder
Related substances
Single impurity： ≤1.0% Internal control standard
Total impurity： ≤2.0%
Related substances：
High performance liquid chromatography (HPLC), area normalization
▪Chromatographic column: Eighteen alkyl silane bonded silica gel as filler (e.g. Agilent Extend
C18, 4.6 x 250mm, 5 m)
▪Mobile phase: with phosphate buffer (two sodium dihydrogen phosphate 3.12g, adding water
1000ml to dissolve, adding three ethylamine 1ml, mixing, using phosphoric acid to regulate the
pH value to 2.80) - acetonitrile (96:4) as the mobile phase A, with phosphate buffer liquid
acetonitrile (75:25) as the mobile phase B. The gradient elution conditions are as follows:
Time (minutes) Mobile phase A (%) Mobile phase B（%）

0 100 0
25.0 100 0
50.0 0 100
50.01 100 0
65 100 0
▪Detection wavelength：210nm flow rate：1.0ml/min
▪Injection volume：20μl column temperature：35℃
Sample solution: the product is dissolved in mobile phase and diluted to make about 1ml solution
containing 2.5mg in each solution.
35
Determination method: area normalization method (peak area percentage%)
3.2.S.2.5 Verification of process & Evaluation

Vildagliptin，The chemical name is 2S (-1-[[) (3- hydroxy tricyclic [3.3.1.1[3,7]]
decane -1- base) amino] acetyl]-2- cyano pyrrolidine.
It is an oral antidiabetic drug researched by Novartis. In September 2007, it was
listed in Europe with the name of Galvus.
In November 2014 to May 2015, our company completed the small test of the
synthetic technology of API. On the basis of literature, the small test technology was
determined, and the laboratory scale was verified. The stable and reliable production
process was determined, which laid the foundation for further amplification of
production.
Our company carried out three batch process verification production in June ~ July
2014. It further proved the stability of the synthesis process and the reasonableness of
various parameters, so that the quality of the intermediate and finished product of the
production is in accordance with the standard. The whole process is carried out under
the verification plan and GMP's strict control. The raw materials involved in the
validation conform to the requirements described in 3.2.S.2.3, and the equipment,
facilities and tools meet the requirements of GMP. There is no deviation and anomaly
in the whole verification process. The specific verification results are shown in the
following tables.
Table 3.2.S.2.5-1 Production batch involved in process verification process
Steps Product name Verification involving batch

150601A
Step 1 VGT：Initial vildagliptin 150602A
150603A
150601
Final product / finished
Step 2 150602
product：vildagliptin
150603
36
Table 3.2.S.2.5-2 Verifying results of verris crude (initial product of VGT)

production process validation
Acceptability limit / Batch

Project
standard 150601A 150602A 150603A
Output — 134.0kg 135.9kg 139.2kg
Yield 70～80% 76.230% 77.31% 79.19%
White to yellowish or White White
White crystalline
Apperance grayish crystalline crystalline crystalline
powder
powder powder powder
Max single Max single
Single impurity ≤ Max single
impurity impurity
1.0%， impurity0.691%，
Related substances 0.751%，Total 0.715%，Total
Total impurities ≤ Total
impurities1.72 impurities1.66
2.0% impurities1.228%
7% 2%
Table 3.2.S.2.5-4 Summary of product (initial product of VGT) production process

validation results
Acceptability limit / Batch

project
standard 150601 150602 150603
Output — 115.0kg 115.9kg 120.0kg
Yield 60～70% 65.42% 65.94% 68.27%
White to yellowish or White White
White crystalline
Apperance grayish crystalline crystalline crystalline
powder
powder powder powder
Melting point 146℃~151℃ 149~151℃ 149~151℃ 149~151℃
Specific rotation -84°to -90° -88° -89° -88°
37
The retention time of the

main peak of the sample
solution should be
consistent with the
retention time of the main
peak of the reference
Identify conforms conforms conforms
solution.
The infrared absorption
spectrum of this product
should be consistent with
that of reference
substance.
Impurity A ： Impurity A：
ND ND
Impurity B： Impurity B：
ND ND Impurity A：ND
Impurity C： Impurity C： Impurity B：ND
Impurity A ≤0.15%， 0.00096% 0.00037% Impurity C：
Impurity B ≤0.15%， Impurity D： Impurity D： 0.0014%
Impurity C ≤0.15%， 0.020% 0.021% Impurity D：
Impurity D ≤0.15%， Impurity E： Impurity E： 0.030%
Impurity E ≤0.15%， ND ND Impurity E：ND
Related substances
Impurity F ≤0.15%， Impurity F： Impurity F： Impurity F：
Impurity G ≤0.15%， 0.00090% 0.0012% 0.0025%
Other single impurity Impurity G： Impurity G： Impurity G：ND
≤0.1%， ND ND Other single
Total impurities ≤0.5% Other single Other single impurity：0.040%
impurity ： impurity： Total impurities：
0.024% 0.025% 0.11%
Total Total
impurities： impurities：
0.091% 0.094%
Enantiomer ≤0.15% 0.0025% N.D 0.0040%
ethanol：0.2% ethanol：0.2% ethanol：0.2%
Methanol≤0.3%，
Isopropanol： Isopropanol： Isopropanol：
ethanol≤0.5%，
0.005% 0.004% 0.004%
Isopropanol≤0.5 %，
3- amino -1- 3- amino -1- 3- amino -1-
Residual solvents and dichloromethane≤
amantadol： amantadol： amantadol：0.05%
0.06%，
L- proline amide, 3- 0.05% 0.05%
triethylamine≤0.032%，
alkanol：0.03%
amino -1- Adamantane DMF≤0.088% methanol 、 methanol、
l-prolinamide≤0.1%
methanol、 dichlorometha dichloromethane、
3- amino -1- amantadol
dichlorometha
≤0.1% ne 、 triethylamine、
ne、
38
triethylamine、 triethylamine、 DMF、

DMF、
l-prolinamide DMF 、 l-prolinamide are
are no l-prolinamide no detectable
detectable
are no
detectable
Sulfate ≤0.02% conforms conforms conforms
Chloride ≤0.02% conforms conforms conforms
Ignition residue Residue≤ 0.1% 0.07% 0.08% 0.1%
Heavy metal ≤0.002% ＜0.002% ＜0.002% ＜0.002%
Water ≤ 0.5% 0.02% 0.02% 0.03%
bacterial count ： ≤
1000cfu/g
Microbial limit Mildew and yeast count： conforms conforms conforms
≤ 100cfu/g
Escherichia coli：N.D/g
Assay 98-102% 99.5% 99.6% 99.6%
The results show that the quality and yield of the product meet the requirements of the
specified requirements. At the same time, it shows that the production process of
verstin is reasonable and effective, with good stability and reproducibility, and it can
produce products that meet the quality standard.
3.2.S.2.6 Development of production process
1. Selection and optimization of route
Vildagliptin，chemical name is 2S -1-[[(3- hydroxytricyclic [3.3.1.1[3,7]] decane
-1- based) amino] acetyl]-2- cyanopyrrolidine, an oral antidiabetic drug researched by
Novartis. In September 2007, it was listed in Europe with the name of Galvus.
This product is a IV two peptidyl peptidase (DPP-IV) inhibitor which inhibits the
activity of the enzyme by combining with DPP-IV to inhibit the activity of the
enzyme. It reduces the concentration of glucagon in GLP-1 concentration and induces
insulin production in islet beta cells, thus reducing blood sugar, and has no significant
influence on body weight.
There are so many vildagliptin's synthetic reports[1,2,3,4,5,6,7,8,9,10] ， The main
39
methods of the literature are as follows:

1.1 The (S) -N- chloracetyl -2- aminoacyl pyrrolidine was obtained by
chloroacetylation of L- prolyl, and then (2S) -N- chloracetyl -2- cyanopyrrolidine was
obtained by dehydration, and the target product was obtained by nucleophilic
substitution reaction with 3- amino -1- adamantanol.[1,2,3,4,5].
1.2 The aminocarbonylation of L- proline through two tert butyl carbonate was
protected under the action of 1- (3- two methyaminopropyl) -3- ethyl two imide
(EDC), and then L- prolyamide was obtained by hydrochloric acid removal and BOC
protection, and then the target product [9] was prepared by 1.1 methods. The route is
cumbersome and the route is long, which is not conducive to production.
1.3 The carbonylation of L- proline after chloroacetylation of N, N- dicyclohexyl
40
carbon two imide (DCC) and ammonium bicarbonate was used. After dehydration, the
target product [10] was prepared by nucleophilic substitution reaction with 3- amino
-1- amantadol. The route is used in DCC, and the DCU generated will be more
difficult to remove in production.
On the basis of route 1.1, we selected (2S) -N- chloracetyl -2- cyanopyrrolidine and 3-
aminoalkanol to produce the target product by nucleophilic substitution reaction.
After refining, the finished product was obtained. The specific route is as follows:
HO HO
Cl N K2CO3
O N
C NH2 N
N H
O C
起始原料：起始原料：粗品： VGT N
1 SM-1 2 SM-2
精制
维格列汀成品
2. Determination of key process parameters

For the synthesis of vildagliptin, our company determined the final process through
literature retrieval and small trial research, and selected the domestic market, the
quality controlled (2S) -N- chloracetyl -2- cyanopyrrolidine and 3- aminoalkanol as
the starting material, and the final products were condensed and refined.1) Preparation
of initial vildagliptin ：
The condensation process is (2S) -N- chloracetyl -2- cyanpyrrolidone (VGT-2)
and 3- amino -1- amantanol nucleophilic substitution reaction to get vildagliptin. We
select DMF as a reaction solvent. After the reaction is finished, the dedrying solvent is
processed to obtain the coarse vecline.The Material input ratio and post treatment
conditions were investigated. The results are as follows:
41
Table 3.2.S.2.6-1 The influence of the ratio of condensation reaction on the yield
and quality of products
Typical batch Test conditions test result detection result

TLC showed that the
98.62% max single
reaction was
The molar ratio of impurity is 1.02%，
complete, the
150311-1 intermediates to adamantanol second max impurity
reaction result was
MVGT-2：M SM-2=1:1.15 is 0.17%
normal, and the yield
Isomer is not detected
is 75.4%
TLC showed that the
reaction was 97.58% max single
The molar ratio of complete, the impurity is 1.36%，
140511-2 intermediates to adamantanol reaction result was second max impurity
MVGT-2：M SM-2=1:1.05 normal,but the is 0.91%
impurity a bit large， Isomer is not detected
the yield is73.8%
TLC showed that the
98.90% max single
reaction was
complete, the
reaction result was
MVGT-2：M SM-2=1:1.1 is 0.17%
is 78.1%
TLC showed that the
99.11% max single
reaction was
complete, the
reaction result was
MVGT-2：M SM-2=1:1.3 is 0.17%
is 78.7%
150313-2 The molar ratio of TLC showed that the 98.86% max single
42
intermediates to adamantanol reaction was impurity is 0.77%，

MVGT-2：M SM-2=1:1.5 complete, the second max impurity
reaction result was is 0.21%
normal, and the yield Isomer is not detected
is 66.2%
TLC showed that the
98.88% max single
reaction was
complete, the
reaction result was
MVGT-2：M SM-2=1:2 is 0.21%
is 67.3%
In order to suppress the appearance of two substitutional impurities, the dosage

of 3- amino -1- amantadol is generally overdosed. The ratio of literature to material is
1:1.17, and the reaction results are normal.By comparing the different feeding
conditions, it is found that when the amount of 3- amino -1- adamantol is less than 1.1,
the VGT-2 and two substitutions of the product are increased, while the increase of 3-
amino -1- mamantol is not obvious. Considering the cost and post treatment pressure,
the proportion of the feeding material is 1.1 ~1.15.
Table 3.2.S.2.6-7 Effect of post treatment on quality and yield of products
Typical
Test conditions test result detection result
batch
The quality of the 98.84% max single
The residue was dissolved in
product is normal, impurity is 0.71%，
2VVGT-2 ethanol and
150320-1 and the yield is second max impurity is
crystallized overnight at
78.4% 0.18%
about 0 ℃.
The residue was dissolved in Normal product 98.11% max single
150318-1
1.5VVGT-2 ethanol and character，and the impurity is 1.68%，
43
crystallized overnight at yield is 73.4% second max impurity is

about 0 ℃. 0.16%
99.15% max single
Normal product impurity is 0.55%，
2.5VVGT-2 ethanol and
150318-2 character，and the second max impurity is
yield is 59.6% 0.17%
about 0 ℃ .
99.16% max single

3VVGT-2 ethanol and
yield is 54.5% 0.17%
about 0 ℃ .
98.89 % max single

2VVGT-2 ethanol and
yield is 71.3% 0.21%
about 10℃

2VVGT-2 ethanol and
99.44% max single
about 0 ℃ . after filtration,
the wet product was
yield is 53.6% 0.08%
dissolved directly by
2VVGT-2 ethanol and 8h
above 0 ℃ .
44

2VVGT-2 ethanol and
99.75%max single
about 0 C. after filtration, the
wet product was dissolved
yield is 57.5% 0.04%
directly with 2VVGT-2
isopropanol and 0.2V water
and 8h above 0 ℃.
After the reaction was completed, we used the treatment method of direct drying
solvent after the reaction liquid filtration and then dissolving the crystal with ethanol.
The experiment proved that the residual residue after dissolving was treated with
2VVGT-2 ethanol crystallization, and the yield of the crude product met the process
requirements. Through the comparison of experiments, we optimized the dissolution
of 2VVGT-2 ethanol and spent the night at about 0 ℃ . Too much solvent or high
temperature will affect the yield of the product.
In order to verify the stability and repeat-ability of the batch amplification process in
the future, we refer to the refining method to remove the crude product from the
ethanol without drying, and then directly purify it once again, reducing the residual
possibility of the impurities such as the adamantanol and reducing the pressure of the
refining process. The experimental results showed that the yield of the products
treated with isopropanol / water dissolution and re crystallization decreased by
10~15%, but the quality characters of the products improved. The risk of
disqualification after refining was reduced.
Table 3.2.S.2.6-8 Detection of initial products after two crystallization treatment

after reaction
Related Substances 150301 Initial 150302 Initial 150303 Initial product

（HPLC） product of VGT product of VGT of VGT
45
batch（65.1g） batch（67.3g） batch（68.4g）

yield（74.07%） yield（76.57%） yield（77.83%）
impurityA（%） N.D N.D N.D

impurityB（%） N.D N.D N.D
impurityC（%） N.D 0.008 0.008
impurityD（%） 0.086 0.084 0.080
impurityE（%） N.D N.D N.D
impurityF（%） 0.619 0.600 0.644
impurityG（%） N.D N.D N.D
Other max single
0.014 0.019 0.019
ipurity（%）
Number of
3 4 4
impurities
Purity（%） 98.674 98.555 98.437
Enantiomer（HPLC） N.D N.D N.D
3) Vildagliptin Refining：
The refinement of the crude products of vildagliptin is directly related to the
quality of the final products. After continuous experiments, we have investigated the
different refining conditions. In the premise of taking into account the quality and
yield, the isopropanol / water system was finally refined. Moreover, for the sake of
short operation, the isopropanol / water system refining operation was incorporated
into the crude preparation. It is also considered that isopropanol drying is difficult to
remove, easy to cause residue, and to ensure the best level of all the related substances,
followed by a recrystallization of ethanol, after this treatment, the finished products
can meet the requirements of API. The following results are focused on the
crystallization conditions. The results are as follows:
Table 3.2.S.2.6-9 Effect of refined feed ratio and crystallization conditions on
Yield and quality of products
46
Typical batch Test conditions test result detection result

he refining effect is 99.84% max single
2V ethanol dissolves and
obvious, the yield is impurity is 0.12%，
150321-1 crystallized overnight at
acceptable and the second max impurity
about 0℃
yield is 88.6% is 0.02%
The refining effect is
99.87% max single
2V isopropanol +0.25V was obvious, the yield is
impurity is 0.07%，
150321-2 purified and dissolved at slightly lower, and
second max impurity
about 0 ℃ the yield is
is 0.03%
low.76.8%
99.87% max single
2V isopropanol +0.15V was obvious, the
impurity is 0.10%，
150321-3 purified and dissolved at impurity is slightly
second max impurity
about 0 ℃ large and the yield is
is 0.02%
82.3%
The refining effect is 99.89% max single
2V isopropanol +0.2V was
150321-4 purified and dissolved at
about 0℃
yield is 74.8% is 0.02%
slightly lower, and second max impurity
about 0℃
the yield is 49.1% is 0.02%
-5℃ , leaving for overnight.
yield is 78.5% is 0.05%
47
99.92 % max single

2V isopropanol +0.2V was impurity is 0.06%，
obvious, the yield is
150325-3 purified and dissolved at second max impurity
acceptable and the
about 5℃ is 0.01%
yield is 82.7%

slightly lower, and second max impurity
-5℃ , leaving for overnight.
the yield is 69.8% is 0.04%
Batch 150320：98.87% max single impurity 0.75%，second max single
Before refining
impurity 0.21%
From the above experiments, it is known that isopropanol / purified water system
is purified and crystallized, and the refining effect is obvious. The increase of purified
water will lead to the reduction of the yield, and the less dosage is more difficult to
dissolve and the quality is a little worse.
In all aspects, for the preparation of the finished product of vildagliptin, we
finally selected the reaction after the reaction after the ethanol crystallization, and the
wet products were refined without drying, and then treated with isopropanol / purified
water. At this time, the crude product was generally close to or less than 0.1%. At last,
after a refining treatment of ethanol, the possible impurities were removed, the purity
of the product was improved and the pressure of isopropanol refining drying was
avoided.
Table 3.2.S.2.6-10 Final product quality of three batches of small tests
Batch 140301 Batch 140302 Batch 140303

Related substances
57.6g）（59.0g）（59.5g）
（HPLC）
Refined yield Refined yield Refined yield（86.99%）
48
（88.48%）（87.67%）
impurityA（%） N.D N.D N.D
impurityB（%） N.D N.D N.D
impurityC（%） N.D N.D N.D
impurityD（%） 0.004 0.008 0.006
impurityE（%） N.D N.D N.D
impurityF（%） 0.113 0.132 0.095
impurityG（%） N.D N.D N.D
Other max single
0.014 0.035 0.036
impurity（%）
Number of impurities 3 4 4
Purity（%） 99.649 99.741 99.686
Enantiomer（HPLC） 0.02 0.078 N.D
The laboratory scale test shows that the process is reproducible and the product meets
API requirements.
3. Major changes in production process during the process of process
development
For the research and development of vildagliptin, our company has carried out a
detailed experiment on the basis of reference materials and determined the best
process parameters through many experiments. After the synthetic process was
confirmed in March ~ April 2015, we carried out three consecutive batch of
laboratory process validation, with a batch size of about 50g. In June 2015, we carried
out three consecutive batch of process validation, with a batch about 100kg.
After determined the synthesis process of vildagliptin, the technological route
basically hasn’t changed, and the technological parameters haven’t greatly adjusted.
From small test to verification batch enlargement, the main changes are feeding scale,
site (lab to workshop), production equipment and so on.
Table 3.2.S.2.6-11 Summary of major changes in production process
49
Evaluation of the influence

Project Trial stage Magnification stage
of quality
The yield and purity did

batch About 50g About 100kg
not change
Main
500ml reaction 500L~1000L The yield and purity did
reaction
bottle reaction kettle not change
equipment
heating The yield and purity did
Heating sleeve Hot bath
equipment not change
major Cooling The yield and purity did
Ice bath Ice brine
equipment equipment not change
Magnetic stirring
Stirring Autoclave mixing The yield and purity did
and mechanical
equipment with paddle not change
agitation
Filter Brucell funnel + The yield and purity did
Centrifuge
equipment flask not change
After the process was basically determined, there was no obviously differences on
the samples testing results, such as characters, related substances, isomers, residual
solvents and content, and the quality of vildagliptin was not significantly changed
because of the changing of the batch, equipment and sites.
50
3.2.S.3 Characterization
3.2.S.3.1 Structural and physicochemical properties
1. Structural Confirmation
According to the technical guidance of the structural confirmation of the chemical
drug material, we company has researched on the structural confirmation of the
verifying batch of vildagliptin samples, we confirm the structural confirmation
methods using IR, UV, NMR, HRMS, TG, DSC, and powder X- ray diffraction
(XRPD) etc, and the stereostructure was confirmed by specific rotation and chiral
chromatography. The main results are as follows:
vildagliptin
Chemical Formula: C17H25N3O2
Exact Mass: 303.1947
Table 3.2.S.3.1-1 Vildagliptin structure confirms sample information
Corresponding batch
Sample Number Source
number
Anhui Hai Kang
VGT-S1 150601
Pharmaceutical Co, Ltd.,
Vildagliptin VGT-S2 150602
according to the
VGT-S3 150602
production process.
1.1 High Resolution Mass Spectrometer ( HRMS)

Instrument type：solariX
Results of determination：
Table 3.2.S.3.1-2 HRMS data and analysis of vildagliptin
Theoretical molecular Theoretical molecular
Sample
weight(M) formula(M)
303.1947 C17H25N3O2
VGT-S1 Conjectured molecular
Test value(M+1)
formula(M+1)
51
304.2020 C17H26N3O2
Conjectured molecular
Test value(M+Na)
formula(M+Na)
326.1840 C17H25N3NaO2
Discussion and explanation：

The molecular weight (M+1) of the sample is 304.2020, and the molecular formula
(M+1) is C17H26N3O2, the other molecular weight is 326.1840 (M+Na), and the
molecular formula (M+Na) is C17H25N3NaO2. The theoretical value of the exact
molecular weight is 303.1947 (M) and the molecular formula is C17H25N3O2. The
test results agree with the theoretical value.
Conclusion: the molecular formula (M) of this product is C17H25N3O2, which is
consistent with the structure of the target compound.
1.2 Infrared absorption spectroscopy (IR)
Instrument type: Nicolet Nexus 670 FTIR spectrometer
Calibration of the instrument:In accordance with the infrared spectrophotometry
correction and calibration method in the appendix IV C of the two appendix of the
Chinese Pharmacopoeia (2010 Edition), the instrument is calibrated with the infrared
absorption spectrum of polystyrene film as the standard.
Sample preparation methods:According to the sample preparation method in
the two appendix IV C of the "infrared preparation spectrum of drugs" (2010 Edition),
the KBr spectrum was used to record the spectrogram.
Determination of data list:

Table 3.2.S.3.1-3IR atlas data of theTrial product of vildagliptin
Perssad vibration
VGT -S1,cm-1 VGT -S2,cm-1 VGT-S3,cm-1 Strength
form
The telescopic
3293.42 3293.52 3293.16 s
vibration of N-H
2992.22， 2992.11， 2992.21，
The telescopic
2915.02， 2915.15,2849.2 2914.99， s
vibration of C-H
2848.95 2 2848.78
2237.90 2237.99 2237.86 w The telescopic
52
vibration of -CN
The telescopic
1656.20 1657.61 1656.97 s
vibration of C=O
1405.63， 1405.76， 1405.53， In-plane bending
s
1354.21 1354.25 1354.14 vibration of C-H
The telescopic
1253.45 1253.74 1253.22 m
vibration of C-N
The telescopic
1119.99 1120.09 1119.91 m vibration of C-O
(H)
Asymmetric
1054.04， 1054.17， 1054.07，
m telescopic vibration
1034.42 1034.65 1034.11
of C-O (H)
Note: s- is strong, m- is medium, w- is weak

Discussion and explanation:
The data from the above tables and drawings show that the main functional groups,
such as nitrile, carbonyl, hydroxyl and subamino groups, exist in the chemical
structure of the sample. Taking VGT-S1 as an example, 3293.42 cm-1 is the telescopic
vibration of N-H, 2992.22 cm-12915.02 cm-12848.95 cm-1 as the expansion vibration
of the fat C-H, the 2237.90 cm-1 is the telescopic vibration of CN, the 1656.20 cm-1
is the C=O telescopic vibration, and the 1405.63 cm-11354.21 is the inner bending
vibration of the fat family. 1253.45 1119.99 cm-1 is the stretching vibration of C-O
(H), and the 1054.04 cm-11034.42 cm-1 is the asymmetric stretching vibration of C-O
(H). These characteristics indicate that the functional groups in the chemical structure
of the product do exist and are consistent with the actual situation.
1.3 Ultraviolet absorption spectroscopy (UV)
Instrument type: TU-1810 type UV-Vis spectrophotometer.
Test conditions:
Acidic solution: take this product and mix it with 0.1mol/l HCl solution.
Alkaline solution: take this product and prepare the appropriate concentration
with 0.1mol/l NaOH solution.
Neutral solution: take this product and use water to prepare suitable concentration
solution.
53
Determination methods and results:
Spectrophotometric method (two Appendix IV A of Chinese Pharmacopoeia
2010 Edition) was scanned in the wavelength range of 190 to 400nm.
The test data are shown in table 3.2.S.3.1-4 and 3.2.S.3.1-8, 3.2.S.3.1-9
and3.2.S.3.1-10.
Table 3.2.S.3.1-4 UV absorption spectrum data of vildagliptin
Maximum
Solvent absorption Absorbance，A Absorption band
wavelength，nm
transition of alcohols
water 199.00 0.684
and amines n→σ*
0.1N
hydrochloric 216.00 0.310 -
acid
0.1N NAOH 219.00 0.545 -
By testing the maximum absorption wavelength of the sample in neutral, acidic
and alkaline solutions, it is observed that the samples have red shift in both acidic and
alkaline solutions, indicating that there are ionizable groups in the samples. The
results are in accordance with the structural characteristics of vildagliptin.
1.4 Nuclear magnetic resonance hydrogen spectrum（1H-NMR、1H-1H COSY）
Instrument model:ADVANCE III 500 BRUKER A&T Center BNU
Conditions for determination: the solvent was D2O.
Determination results：
Table 3.2.S.3.1-5 Data and analysis of vildagliptin's 1H-NMR Atlas

Proton Chemical shift Related
Proton Corresponding
sequence （ppm） Multiplicity protons(1H-1H
number protons
number VGT-S1 COSY)
54
H-2 4.651~4.703 m 1 H-3 CH
H-5 3.399~3.567 m 2 H-4 CH2
H-7 3.324~3.368 m 2 - CH2
H-3 2.163~2.209 m 2 H-2,H-4 CH2
H-9,H-13,
H-10,H-12 2.163~2.209 m 2 H-11,H-14,H- 2×CH
15
H-4 2.042~2.070 m 2 H-5,H-3 CH2
H-14,H-15 1.544~1.585 m 4 H-10,H-12 2×CH2
H-16 1.460~1.520 m 2 - CH2
H-9,H-13 1.460~1.520 m 4 H-10,H-12 2×CH2
H-11 1.419 m 2 H-10,H-12 CH2
S = single peak, d = double peaks, M = multiple peaks

There are 20 methylene hydrogen, 3 methylene hydrogen, 1 hydroxyl hydrogen
and 1 imino hydrogen in the structure. 1H-NMR shows 23 proton signals, of which
the hydroxyl hydrogen and imino hydrogen are not shown by solvent D2O, which is
consistent with the structure of target compounds.
The submethylene hydrogen of H-2 is connected with cyanyl group and is located at
the lowest field, and 1H-1H COSY is associated with H-3; H-5 is methylene hydrogen
on pyrrolidine, which is linked to N and shows a lower field, and the splitting points,
1H-1H COSY is associated with H-4, followed by H-7, associated with subino and
carbonyl groups, without related hydrogen; 2.163~2.209 should be H-3 The display is
associated with H-2 and H-4; there are also two sub methylhydrogen H-10 and H-12
in this position, 1H-1H COSY shows H-9/11/13/14/15 and so on; the next slightly
high field hydrogen should be H-4, 1H-1H COSY shows H-5 and H-3; The slightly
higher field is H-16, which has no hydrogen; the secondary high field should be
H-9/13, 1H-1H COSY shows that it is associated with H-10/12; the highest field
should be H-11, and 1H-1H COSY is associated with H-10/12.
Conclusion: the 1H-NMR and 1H-1H COSY maps and data are in good agreement
with the structure.
1.5 Nuclear magnetic resonance carbon spectrum（13C-NMR、DEPT、HSQC、
HMBC）
55
Instrument model：ADVANCE III 500 BRUKER A&T Center BNU
Determination conditions：The solvent is D2O。
Table 3.2.S.3.1-6 13C-NMR map data and analysis of vildagliptin

Chemical shift
Related Related hydrogen
Carbon number (ppm) DEPT
hydrogen HSQC HMBC
VGT-S1
C-6 172.119 - H-7,H-2 C
C-1 119.104 - H-2 C
C-17 69.948 - H-10/12/14/15/16 C
H-7/10/12,H-9/13
C-8 53.634 - C
/16
C-16 47.837 H-16 H-9/13/15 CH2
C-2 46.841 H-2 H-3/4/5 CH
C-5 46.492 H-5 H-2/3/4 CH2
C-14,C-15 42.798 H-14,H-15 H-16/13/12/11/9 2×CH2
C-7 42.055-42.170 H-7 H-5/9/13/16 CH2
H-10/12/11/14/15/
C-9,C-13 39.654 H-9,H-13 2×CH2
16
C-11 34.252 H-11 H-9/13/14/15/16 CH2
C-10,C-12 30.264 H-10,H-12 H-9/11/13/14/15 2×CH
C-3 29.408 H-3 H-2/5/4 CH2
C-4 24.564 H-4 H-2/5 CH2
Discussion and explanation：

The structure of the test sample was analyzed: there were 4 Quaternary carbons,
3 tertiary carbons, 10 secondary carbons, and a total of 17 carbons. The 13C-NMR
spectra showed that the carbon atom signal was consistent with the actual situation.
56
DEPT shows that there are 4 Quaternary carbons, the carbonyl carbon that
should be C-6 at the lowest field, followed by the C-1 cyanocarbon, which should be
C-17 again, connected to the hydroxyl group, and the highest field should be C-8 ,
linked to imino groups.
Analysis from DEPT 90: There are two groups of tertiary carbons, C-2 is
attached to the cyano group and is located in the lower field, followed by C10/1 on
adamantane.2。Analysis from the DEPT 135 map: There are 8 groups of secondary
carbons, which in turn should be C-16, C-5, C-14/15, C-7, C-9/13, C-11, C-3, C-4 .
The result is consistent with the structure of the test article.
Analysis from HSQC: C16-H16, C2-H2, C5-H5, C6-H6, C14/15-H14/15, C7-H7,
C9/13-H9/13, C11-H11, C-10/12 Directly related to H-10/12, C-3 and H-3, C-4 and
H-4.
Analysis from the HMBC spectrum: C6 and H7/H2, C1 and H2, C17 and
H10/12/14/15/16, C8 and H7/10/12/9/13/16, C16 and H9/13/15, C2 and H3/4/5, C5
and H2/3/4, C14/15 and H16/13/12/11/9, C7 and H5/9/13/16, C9/13 and H10/12/11/
14/15/16, C11 and H9/16/13/14/15, C10/12 and H9/11/13/14/15, C-3 and H2/5/4, C4
and H2/5 Remote Correlation
The two-dimensional spectrum results exactly match the structure of this
product.
Conclusion：The 13C-NMR, DEPT, HSQC, and HMBC spectra of this product agree
with the actual structure.
1.6 Differential scanning calorimetry (DSC)
Instrument model ：Netzsch STA449F3
Testing sample：Trial product
Determination conditions ： The starting temperature is 35 ° C and the ending
temperature is 350°C. Heating rate: 10.0°C/min.
Conclusion ： The test article began to absorb heat at 150.1 ° C with a peak top
temperature of 156.2 °C, which is consistent with the melting point reported in the
literature (US20080167479 reported melting point 150°C).
1.7 Thermogravimetric analysis（TG）
Instrument model：Netzsch STA449F3
Testing sample ：Trial product。
57
Determination conditions ： The starting temperature is 35 ° C and the ending
temperature is 350°C. Heating rate: 10.0°C/min
Conclusion ： The test sample had no apparent weightlessness before 200 ° C,
indicating no free or crystal water，losing of weight after 250°C，up to 300 °C a
total weight loss of 35.01%
1.8 Powder X-ray diffraction（XRPD）
Instrument ：Brooke B8 advance
Sample：Trial product
Determination conditions ： Cuk α radiation; graphite monochromator, tube
pressure 40 kV, tube flow 40 mA, 2 θ scanning range 4.5-50 ° , scanning speed
19.6850°/s.
Table 3.2.S.3.1-7 X-Ray Diffraction Pattern Data and Analysis of Vildagliptin
VGT-S1 VGT-S2 VGT-S3

2() Int.（%） 2() Int.（%） 2() Int.（%）
11.9284 17.34 11.8791 16.88 11.8778 12.76
13.3846 8.59 13.3309 15.12 13.3480 10.43
16.6082 32.64 16.5029 33.44 16.4833 24.23
17.0573 100.00 16.9865 100.00 17.0400 100.00
17.3368 40.47 17.1735 86.10 17.2430 44.16
20.0598 20.08 19.9961 38.20 20.0138 21.52
22.2968 22.88 22.2852 28.49 22.2965 16.87
25.8181 12.88 25.7167 8.78 25.7660 7.31
27.2195 19.25 27.1686 26.52 27.1884 18.75
27.9919 15.04 27.9525 15.22 27.9510 12.85
31.1962 6.62 31.1386 11.43 31.1443 6.61
Conclusion :All three batches of the test sample are crystalline materials，it’s the same
of powder’s X ray diffraction pattern ， it’s the same crystalline material 。 A
crystalline X-ray diffraction pattern reported in the literature has 16.6 ° , 17.1 ° ,
17.2°±0.3 or preferably 12.0°, 13.5°, 16.6°, 17.1°, 17.2°, 20.1°, 22.5°,
58
27.4° 28.1°± The 2-theta peak of 0.3 is consistent with the reported crystalline of
the sample (WO2006078593A2).
1.9 Specific Rotation
Instrument：Shanghai Precision Scientific Instrument Co., Ltd. WZZ-2B
Test conditions：Methanol was used as the solvent and the solution concentration was
97 mg/ml.
Results of determination：See Table 3.2.S.3.1-8 for details.
Table 3.2.S.3.1-8 Measurement value of specific rotation ratio and literature
value
Determination value Literature value

-86.70° -85º
Conclusion ： The measured curl values of samples are in agreement with those
reported in literatures (US20080167479, -85 degrees (C9.73, MeOH)).
1.10 Comprehensive Analysis：
The accurate inference of molecular formula and molecular weight：
According to high resolution mass spectrometry, the molecular weight (M+1) of the
test sample is 304.2020, and the molecular formula (M+1) is assumed to be
C17H26N3O2. ， In addition, the molecular weight of 326.1840 is (M+Na), and the
formula (M+Na) is assumed to be C17H25N3NaO2. ， The theoretical value of the
molecular weight of the test sample is 303.1947 (M) and the molecular formula is
C17H25N3O2. The test results are in accordance with the theoretical values.
Structural Inference：
1) Unsaturation calculation ()and inference
Due to the formula : =1 + n4+ (n3－n1) / 2 to calculating ，The unsaturation in the
structure of the compound should be 7 ， The adamantane ring contributes 3
unsaturation, the carbonyl group contributes 1 unsaturation, the pyrrole ring
contributes 1 unsaturation, and the nitrile group contributes 2 unsaturation, which is
consistent with the actual structure of the target compound.
2) Carbon classification and determination of the number of atoms such as
hydrocarbons：
59
a. The carbon spectrum confirms that this product has 17 carbon atoms; the hydrogen
spectrum confirms that there are 25 protons ； Combined with high resolution mass
spectrometry analysis, the number of elements in the molecule is consistent with
reality.
b. DETT 90 atlas shows that there are 2 tertiary carbon 、DEPT 135 atlas shows 10
secondary carbons，The DEPT spectrum results agree with the actual。
c. The proton and carbon atoms of the nuclear magnetic spectrum of the test sample
have been reasonably assigned ， The correlation between HSQC and HMBC
two-dimensional spectrum is in accordance with the actual situation。
3) In the thermal analysis, the thermal difference between the test sample's differential
heat and the thermogravimetric analysis is consistent。
4) The powder X-ray diffraction data of the three batches of verification batch
samples are basically the same，Description This product is a crystalline compound.
Thecrystalline of the three batches of validated batch samples is the same.
Thecrystalline is consistent with the type A reported in the literature.
5) The measurement of specific rotation shows that the product is an optical isomer
and the measured value is consistent with the literature value。
The above analysis and discussion show that the final structural formula of this
product is consistent with the structural characteristics of the target compound
2. Physical and chemical properties
The vildagliptin structure contains 1 carbon chiral neutral and the product is S
configuration。Its physicochemical properties include traits, specific rotation, melting
point, solubility, hygroscopicity, and crystal shape examination as follows.
2.1 Character
This product is white to slightly yellow or slightly gray crystalline powder.
2.2 Specific Rotation
According to Appendix VI E of the 2010 Pharmacopoeia of the Chinese
Pharmacopoeia, the test was conducted at 97 mg/ml, and the solvent was methanol.
The results are shown in the following table.
Table 3.2.S.3.1-9 The sample ‘s specific rotation testing result
60
Batches 140601 140701 140702
Specific Rotation -86.70º -87.31º -87.22º
2.3 Melting Point

The test product uses a capillary assay to determine the melting point，The specific
results are shown in the table below.
Table 3.2.S.3.1-10 Sample melting point measurement results
Batch number 140601 140701 140702

Melting point 147.5℃~148.5℃ 146.5℃~147.5℃ 146.5℃~147.5℃
2.4 Solubility
In accordance with the Chinese Pharmacopoeia 2010 Edition, Section VIII Test，This
product is soluble in methanol, ethanol, dimethyl sulfoxide, water or 0.01mol/L
hydrochloric acid solution, slightly soluble in acetonitrile or isopropanol；The results
are shown in the table below.
Table 3.2.S.3.1-11 Sample solubility determination

Sample Add solvent volume phenomeno
Solvent Conclusion
quantity （ml） n
Incomplete
0.9912 1
dissolution
Methyl alcohol freely soluble
All
1.0896 10
dissolution
Incomplete
0.9975 1
dissolution
Ethanol freely soluble
All
1.0543 10
dissolution
Incomplete
0.9984 1
dissolution
Dimethyl
All freely soluble
sulfoxide(DMSO)
1.0781 10 dissoulutio
n
61
Incomplete
0.9967 1
dissolution
Water All freely soluble
n
Incomplete
0.9974 1
dissolution
0.01mol/L
Depart of freely soluble
Hydrochloric Acid
n
Incomplete
0.0961 3
dissolution
Slightly
Acetonitrile All
dissolving
n
Incomplete
0.0997 3
dissolution
Slightly
Iso-Propyl alcohol All
dissolving
n
2.5 Humidifying
Tests were conducted in accordance with Appendix XIX J of the 2010 edition of the
Chinese Pharmacopoeia. The results are shown in the table below。
Chart 3.2.S.3.1-12 Determination of sample humidifying
Sample
Weighing After 24h
Batch +weighing Weight Humidifyi
bottle sample+weighing
number bottle weight gain（g） ng
weight（g） bottle weight （g）
（g）
150601 30.2278 31.1878 31.1887 0.0009 0.094%
150602 30.6868 31.6328 31.6336 0.0008 0.085%
150603 29.6916 30.6620 30.6628 0.0008 0.082%
After 24 hours of open placement, the weight gain of Vildagliptin was less
62
than 0.2% ， It can be seen from this ， According to the Chinese Pharmacopoeia's
definition of wetting characteristics and the definition of wetting and gaining
weight,Vildagliptin has no hygroscopicity.
2.6 Crystalline
Through，Novartis AG point a medicinal crystalline of vildagliptin，Named Type
A，It is characterized by an X-ray diffraction pattern at 16.6°, 17.1°, 17.2°±0.3
or preferably 12.0°, 13.5°, 16.6°, 17.1°, 17.2°, 20.1°, 22.5°, 27.4° 28.1°
± 0.3. -Theta peak ， Infrared spectra show significant absorption bands at about
3293cm, 2925-2853cm, 2238cm, 1658cm, 1455/1354cm, 1254cm, 1054-1035cm 。
Through comparison with literature X-ray diffraction and infrared spectrum data，The
Vildagliptin prepared by our company is crystalline form A, and thecrystalline of the
serial batch production sample is consistent with thecrystalline described in the
literature 。 The accelerated test and long-term test results show that the company's
production samples have good stability。
Chart 3.2.S.3.1-13 Data stability of Vildagliptin 's X- powder diffraction
pattern（Accelerated June and Long-term June Stability）result
Sample Feature2 peak()

11. 1 1 1 1 2 2 2 2
150601
928 3.385 6.608 7.057 7.337 0.060 2.297 7.219 7.992
11. 1 1 1 1 1 2 2 2
150602
879 3.331 6.503 6.987 7.173 9.996 2.285 7.169 7.952
11. 1 1 1 1 2 2 2 2
150503
878 3.348 6.483 7.040 7.243 0.014 2.296 7.188 7.951
150601 11. 1 1 1 1 2 2 2 2
(add 6) 909 3.359 6.529 7.010 7.255 0.024 2.303 7.194 7.993
150602 11. 1 1 1 1 2 2 2 2
(add 6) 989 3.412 6.589 7.072 7.304 0.074 2.414 7.281 8.066
150603 11. 1 1 1 1 2 2 2 2
(add 6) 967 3.401 6.574 7.093 7.303 0.071 2.356 7.239 8.013
150601 11. 1 1 1 1 2 2 2 2
(long 940 3.392 6.540 7.045 7.220 0.075 2.347 7.249 8.007
63
6)
150602
11. 1 1 1 1 2 2 2 2
(long
915 3.412 6.601 7.027 7.246 0.057 2.354 7.283 8.032
6)
150603
11. 1 1 1 1 2 2 2 2
(long
937 3.425 6.576 7.084 7.288 0.062 2.349 7.272 7.996
6)
2.7 Stereoscopic Structure

Vildagliptin has 1 chiral carbon atom in the structure, S-configuration，We identified
it by specific rotation,The specific rotation is consistent with the literature value。Its
R-isomer is classified as an impurity and has been set in the quality standard. It is
controlled below 0.15% in the finished product.
3.2.S.3.2 Impurity
Table 3.2.S.3.2-1 Vildagliptin Impurities analysis table
Impurities
Classification Name Chemical structure Control limit
source
Target
Vildagliptin /
product
Introduction
L-Prolinamid
0.1% of starting
e
materials
3-AMINO-1-
Starting
Organic impurities ADAMANT 0.1%
material
ANOL
VGT-1 Introduction
（Impurities 0.15% of starting
A） materials
64
VGT-2
Original
（Impurities 0.15%
product
B）
By-products/
Impurities C 0.15% degradation
products
By-products
Impurities D 0.15% /degradation
products
By-products
Impurities E 0.15% /degradation
products
Impurities F 0.15% By-products
65
By-products
Impurities G 0.15% /degradation
products
Impurities I 0.10% By-products
VGT
By-products
Isomer
0.15% /degradation
（Impurities
products
K）
Name Source Type Control limit
dichlorometh Introduction of starting The second cl

0.06%
ane materials ass of solvents
The Third
Iso-Propyl
Step 1 class of 0.5%
alcohol
solvents
Residual solvent N,N-dimethyl The second cl
Step 1 0.088%
-Formamide ass of solvents
The Third
Ethyl alcohol
Step 1.2 class of 0.5%
absolute
solvents
Introduction of starting
Triethylamine 0.032%
materials
Inorganic materials mainly use anhydrous potassium carbonate, inorganic
Inorganic impurities impurities,The part of thing maily through the sulfate(ion)、CI、Residue on ignition and
Heavy metal to control。
66
Analysis of Impurity Spectrum in Synthesis Process of Vildagliptin：
HO
HO
Cl N
K2CO3
O NH2
C N
N N
起始原料：
1 SM-1 起始原料： H
2 SM-2 O C
粗品： VGT N
CH3CH2OH
维达列汀成品
Synthesis of vildagliptin: The target product is obtained by the nucleophilic

substitution reaction of (2S)-N-chloroacetyl-2-cyanopyrrolidine and
3-amino-1-adamantanol, which is refined to obtain the finished product.
Analysis of impurities introduced by starting materials：The key starting material for
the synthesis of vildagliptin is (2S)-N-chloroacetyl-2-cyanopyrrolidine and
3-amino-1-adamantanol. The structure is as follows：
(2S)-N-chloroacetyl-2-cyanopyrrolidine
3-amino-1-adamantanol
After several batches of tests, the two starting materials can be effectively
removed during the process, and the residuals in the finished product can be
effectively controlled（≤0.1%）
2) The starting material is (2S)-N-chloroacetyl-2-cyanopyrrolidine synthesis

introduced impurity analysis：
67
Cl N
O COOH
杂质 H
O Et3N N TFAA Cl N
HN Cl
Cl
Cl O Na2CO3 O
CONH2 CONH2 C
N
VGT-1 起始原料
Cl N
Cl N
O C
O CONH2 N
杂质 J
In fact ,the step is two-step reaction，The firstL-prolinamide is first acylated to

give VGT-1 ， then through the Dehydration of trifluoroacetic anhydride to give
(2S)-N-chloroacetyl-2-cyanopyrrolidine (VGT-2).
And the Acylation reaction is fast and complete ， So the possibility of

L-prolinamide residues in the synthesis of (2S)-N-chloroacetyl-2-cyanopyrrolidine is
very low ； Dehydration reaction due to its slow conversion rate, there is a certain
amount of residual VGT-1 at the end of the reaction ，and the VGT-1 Possibility of
Converting to Impurity H by Hydrolysis；On the other hand，If there is an enantiomer
in L-prolinamide ， it will be possible to form VGT-1 enantiomers and (2S) -N-
chloroacetyl -2- cyano pyrrolidine enantiomers 。 Through the control of the raw
material's corporate standards, the impurities in the finished product could be
controlled below 0.15%.，Meeting the quality standards and the effectively control.
3) Condensation process introduced impurity analysis：
68
HO
N
N
O O C
N
C N
N 杂质 F
HO HO
Cl N K2CO3
O N
C NH2 N
N H
O C
起始原料：粗品： VGT N
2 SM-2
HO
HO
HO N
N NH
Cl N H
O C
N N
O CONH2 OH 杂质 K
N N
H
O
HO 杂质 I O 杂质 C
N HO
N
H C
O O
O NH2
杂质 D
HO N
N
N
N O
H C
O
O OH
Due to the synthetic route :If the (2S)-N-chloroacetyl-2-cyanopyrrolidine

contains residues of VGT-1，It will be possible to produce impurities D through a
condensation reaction ， The Impurities G are produced by hydrolysis ； If there is
residual chloroacetic acid, impurities may be generated during the reaction through a
condensation reaction.
(2S)-N-chloroacetyl-2-cyanopyrrolidine undergoes nucleophilic substitution
reaction with 3-amino-1-adamantanol，during the reaction，May be disubstituted at the
amino position to produce impurity F ； The target product may be degraded to
impurity C and then oxidatively degraded to impurity E. Based on the results of
several experiments, the main impurity of the reaction solution was impurity D and
unreacted (2S)-N resulting from the residual VGT-1 in
(2S)-N-chloroacetyl-2-cyanopyrrolidine. -chloroacetyl-2-cyanopyrrolidine and
disubstituted impurities F，After the reaction, the above impurities could be controlled
within the standard range, and other impurities can also be effectively removed. The
69
purity of the crude liquid is >98%.Another 3-amino-1-adamantanol is over-dosed,
after the post-treatment, the residual amount can also be controlled within the standard
rangeThe isomer of the target product is also one of the potential impurities，However,
the chiral carbon in the structure is not a reaction site during the reaction, and it is not
easy to race or flip ， the (2S)-N-chloroacetyl-2-cyanopyrrolidine was detected
repeatedly and isomers were not detected ， so the isomers of crude VGT are also
basically not detected.，it’s only to do test ,not control
4) Degrading impurity：
From the chemical structure analysis, and through the influence factors,
long-term, accelerated test and so on.，We found that impurity C, impurity D, impurity
E and impurity G are potential degradation impurities of vildagliptin, so they are all
set into quality standard ， As a known impurity (less than 0.15%).The specific
degradation pathways are as follows：
5) Genetic genotoxic impurities：

This product is not involved in the preparation of genetic genetic toxic impurities
alert unit.
Table of the 3.2.S.3.2-2 :Summary of detection results of various potential

impurities of vildagliptin in finished products
70
Name of the Source of
Testing project 150601 150602 150603
impurities impurity
Impurities A VGT-1 ND. ND. ND.
Impurities B VGT-2 ND. ND. ND.
By-product or
the potential
Impurities C ND. ND. ND.
degradation
products
By-product or
the potential
Impurities D 0.077 0.017 0.020
degradation
products
Related
By-product or
impurities
Impurities the potential
ND. ND. ND.
E degradation
products
Impurities
By-product ND. 0.022 0.033
F
By-product or
the potential
Impurities G ND. ND. ND.
degradation
products
Impurities I By-product ND. ND. ND.
others 0.046 0.076 0.050
Enantiomer
Enantiomers s’ impurities By-product ND. ND. ND.
K
(ND.=Not Detected)
Contrastive comparison between homemade raw materials and original
preparation
Comparing the impurities of the company's self-made raw material with the
original preparation agent. The result is shown in the figure below:
71
Table 3.2.S.3.2-3 quality comparison between domestic and imported raw materials
Batch
number 150601 150602 150603 Listed products
Project
Impurity A： ImpurityA：
ND. ND.
Impurity B： ImpurityB：
ND. Impurity A：ND. ND. ImpurityA：N.D
Impurity C： ImpurityC：
Impurity B：ND. ImpurityB：N.D
ND. ND.
Impurity C：ND. ImpurityC：
ImpurityD： ImpurityD：
Impurity D： 0.145%
0.077% 0.020%
0.017% ImpurityD：
ImpurityE： ImpurityE：
Impurity E：ND. 0.144%
ND. ND.
Impurity F ： ImpurityE：
Related ImpurityF： ImpurityF：
0.022% 0.019%
substances ND. 0.033%
Impurities G ： ImpurityF：ND.
ImpurityG： ImpurityG：
ND. ImpurityG：ND.
ND. ND.
Single maximum Impurity：
Single Single
unknown 0.015%
maximum maximum
impurity：0.076% Total
unknown unknown
Total Impurities： Impurities：
impurity： impurity ：
0.114% 0.322%
0.046% 0.050%
Total Total
Impurities： Impurities ：
0.155% 0.103%
Number of
3 3 3 4
impurity
Content 99.9% 99.7% 99.7% 99.8%
72
Our company's self-made sample related substances are all less than 0.15%, total
miscellaneous 0.1~0.3% ， The largest single 0.145% of the listed products, total
miscellaneous 0.322%，Self-made sample quality is not lower than the listed product。
The quality of vildagliptin produced by our company's self-made raw material
medicine is not lower than that of the original preparation agent. ，It shows that the
quality of raw materials produced by our company is good and can be used for the
production of pharmaceutical preparations.
3.2.S.4 Quality Control of API

3.2.S.4.1 Quality Standard
The Vildagliptin’s quality standard show in table 3.2.S.4.1-1。
Table 3.2.S.4.1-1: Vildagliptin quality standard

Check item Method Release standard limits Shelf life standard limit
The product is The product is
Visualizatio white to yellowish or white to slightly yellow or
n Micro-gray crystalline slightly gray crystalline
powder。 powder。
It’s easily soluble It’s easily soluble
in the Methanol、 in the Methanol、Ethanol、
Ethanol、Dimethyl Dimethyl
Determinati sulfoxide(DMSO)、 sulfoxide(DMSO)、water
on of water or in the or in the
Character
solubility 0.01mol/hydrochloric 0.01mol/hydrochloric acid
acid solution，Slightly solution，Slightly soluble
soluble in acetonitrile or in acetonitrile or
isopropanol. isopropanol.
Melting
point 146℃~151℃ 146℃~151℃
method
Specific Specific rotation Specific rotation
rotation method is the-84°and method is the-84°and
73
method -90° -90°
In the chromatogram In the chromatogram
recorded under the recorded under the
determination, the determination, the
retention time of the retention time of the main
⑴ HPLC
main peak of the test peak of the test solution
ways
solution should be the should be the same as the
same as the retention retention time of the main
time of the main peak of peak of the reference
Identification the reference solution.。 solution.。

The product's infrared
absorption spectrum The product's infrared
should be consistent absorption spectrum
⑵Infrared
with the reference should be consistent with
spectrophot （Chinese the reference（Chinese
ometry
Pharmacopoeia 2010 Pharmacopoeia 2010
Edition 2 Appendix Ⅳ Edition 2 Appendix Ⅳ C）
。
C）。
Impurities A：Not to Impurities A：Not to
exceed 0.15% exceed 0.15%
Impurities B：Not to Impurities B：Not to
Impurities C：Not to Impurities C：Not to
Relative HPLCmeth Impurities D：Not to Impurities D：Not to
substance od exceed 0.15% exceed 0.15%
Impurities E：Not to Impurities E：Not to
Impurities F：Not to Impurities F：Not to
Impurities G：Not to Impurities G：Not to
74
The another The another impurities ：
impurities ：Not to Not to exceed 0.1%
exceed 0.1% Total impurities：Not to
Total impurities：Not to exceed 0.5%
exceed 0.5%
Enantiomers not to Enantiomers not to

Enantiomers HPLC ways
Methanol not to Methanol not to exceed
exceed 0.3% 0.3%
Ethanol not to exceed Ethanol not to exceed
0.5% 0.5%
dichloromethane not to dichloromethane not to
Residual solvent
N,N-Dimethylformamid N,N-Dimethylformamide
and
e not to exceed 0.088% not to exceed 0.088%
L-prolinamide, GC method
Triethylaminenot to Triethylaminenot to
3-amino-1-adam
antanol
Iso-Propyl alcohol not Iso-Propyl alcohol not
to exceed 0.5% to exceed 0.5%
L-Prolinamide not to L-Prolinamide not to
3-AMINO-1-ADAMAN 3-AMINO-1-ADAMANT
TANOL not to exceed ANOL not to exceed
0.1% 0.1%
Fisher Moisture must not
Moisture Not exceed 0.5%
Method exceed 0.5%
Sulfate
Sulfate Not exceed 0.02% Not exceed 0.02%
test
Chloride CI test Not exceed 0.02% Not exceed 0.02%
Residue on Residue on Residual residue must Residual residue must not
ignition ignition not exceed 0.1% exceed 0.1%
75
Heavy Heavy metals cannot
Heavy metals cannot
Heavy metal Metal exceed twenty
exceed twenty millionths
Inspection millionths
Number of bacteria：not Number of bacteria：not
exceed 1000cfu/g exceed 1000cfu/g
Microbial Mold and yeast count： Mold and yeast count：not
Microbial Limit
limit test not exceed 100cfu/g exceed 100cfu/g
Escherichia coli：not Escherichia coli：not
exceed /g exceed /g
In terms of anhydrous, In terms of anhydrous,
Determination HPLC
C17H25N3O2 should be C17H25N3O2 should be
of content method
98.0% ~102.0%. 98.0% ~102.0%.
3.2.S.4.2 Analytical Procedure

The registration of raw materials for vildagliptin is 3 categories, and it has not
obtained reference quality standards for raw materials at home and abroad. Please
refer to the registration standard for vildagliptin tablets (JX20100243)，The relevant
guidelines and the Chinese Pharmacopoeia have established this quality standard.
The specific content of the analysis method as follows：
3.2.S.4.2.1 Descriptions
3.2.S.4.2.1.1Appearance
Operation：
Take appropriate amount of this product, set it on a white background, visual
observation under natural light.
Standard regulation：
Should be white to slightly yellow or slightly gray crystalline powder.
3.2.S.4.2.1.2 Solubility
Reagent：
Water （Purified Water）
Methyl alcohol（AR）
Dimethyl sulfoxide(DMSO)（AR）
76
Ethanol（AR）
Acetonitrile（AR）
Isopropanol（AR）
0.01mol/L HCI
Operation：
Take appropriate amount of this product, research into fine powder, as a test
product.
i ） Methanol alcohol(AR) 、 Ethanol 、 Dimethyl sulfoxide(DMSO) 、 Water or
0.01mol/L’s solubility in hydrochloric acid solution
Take the test sample 1.0g (slightly less than 1.0g) accurately weighed, set at
25 ° C ± 2 °C, add the solvent 1ml, shake vigorously every 5 minutes for 30
seconds, observe the dissolution within 30 minutes It should not be completely
dissolved.
Take the test sample 1.0g (slightly larger than 1.0g) accurately weighed, set at
25 °C ± 2 °C, conical flask, add solvent 10ml, strong shaking every 5 minutes
for 30 seconds, observed within 30 minutes Dissolution should be completely
dissolved.ii）, Solubility in acetonitrile or isopropanol
Take the test sample 0.1g (slightly less than 0.1g) accurately weighed, set in
a test tube at 25 ° C ± 2 ° C, add solvent 3ml, shake vigorously every 5
minutes for 30 seconds, observe the dissolution within 30 minutes It should not
be completely dissolved.
Take the test sample 0.1g (slightly larger than 0.1g) accurately weighed, set
at 25 °C ± 2 °C, conical flask, add solvent 10ml, strong shaking every 5
minutes for 30 seconds, observe within 30 minutes Dissolution should be
completely dissolved.
This product is soluble in methanol, ethanol, dimethyl sulfoxide, water or
0.01mol/L hydrochloric acid solution, slightly soluble in acetonitrile or isopropanol.
3.2.S.4.2.1.3 Melting point

Instrument:
Melting point apparatus
77
Operation：
Take a suitable amount of the test product into a fine powder and dry overnight
in a phosphorus pentoxide dryer. Insert one end of the capillary opening into the
pretreated test sample, re-invert the capillary tube, and seal the end of the capillary
tube. Let the test sample fall into the bottom of the tube, and then use a clean glass
tube of appropriate length (about 60cm) and place it vertically on a watch glass or
other suitable hard object, and place the above-mentioned capillary tube with the
sample into the glass tube. It’s allowed to fall freely and repeated several times to
tightly gather the test sample on the bottom of the capillary ；The height of the test
sample should be 3mm.When the temperature of the warming liquid rises to 136°C,
immerse the capillary tube containing the test sample in the temperature-adjusting
liquid so that it adheres to the thermometer, and the contents of the capillary tube are
required to fit in the middle of the mercury ball of the thermometer.；Continue heating
and stirring at a heating rate of 1.5°C/min. Observe the change of the test sample in
the capillary；Record the temperature at which the sample begins to liquefy within the
capillaries and the appearance of significant droplets is used as the initial melting
temperature, and all liquefaction temperatures are taken as the total melting
temperature.
standard regulation：
Should be 146℃~151℃。
3.2.S.4.2.1.4 Specific Rotation
Reagent:
methyl alcohol
Preparation of the test solution：
Take this product 2.42g, accurately weighed, set 25mL volumetric flask, add the
amount of methanol, shaking to dissolve, diluted with methanol to the mark, shakeing
Operating：
It is used the methyl alcohol to blank correction；let the Measuring tube to use
the Trial product solution to washing several times，then Injecting the test solution
slowly（Not to be produced bubble），put it in the polarimeter to detect readings. The
optical rotation of the obtained sample solution ； Reading the optical rotation three
times with the same method and take the average of three measurements.
78
Notice：
1 、 When the solution is formulated and measured, the temperature should be
adjusted to 20 C + 0.5 C.
2、Before measuring the test solution, blank solution is used for blank calibration.
After the test solution is determined, blank is added and recalibrated once to
determine whether there is any change in the zero point during the measurement.；If
there is a change in the zero point found during the second calibration, it should be
recalibrated and the optical rotation of the test solution should be determined again.
3、The measuring tube can not be heated and dried in the drying oven，due to the
coefficient of linear expansion of the glass tube and the metal nuts at both ends is
different, and heating may cause damage，After it can be dried or treated with ethanol
and other organic solvents and dried
4 、 After using an acidic or organic solvent, it must be washed and dried
immediately，In order to avoid causing metal corrosion or aging the rubber gasket in
the nut, it becomes sticky.When the instrument is not in use, silica gel can be placed in
the sample chamber to keep it dry.
Calculation formula：
 Dt  100    1
l 4  w  (1  S )
In the formulation：[α] as the specific rotation

α as the measured optical rotation
D is the Sodium spectrumof D line
t is the temperature at the time of measurement (°C)
l is the measuring tube length (dm)
w is the sample weight (g)
S is the moisture of the sample (%)
Standard Regulations:
79
Specific rotation should be -84°to -90°
3.2.S.4.2.1.5 Hygroscopicity
Weighing bottle pretreatment：
Take a dry stoppered glass weighing bottle (outer diameter 50mm, height
15mm)，Place artificial climate box on the day before the test (set temperature 25°C
±1°C，Within 80%±2% relative humidity, accurately weighed (W1)。
The operation of the test product:
Take the test product and tile it in the above weighing bottle，The thickness of the
test specimen is about 1mm，Precision weighed (W2)。The weighing bottle is opened
and placed in the same temperature and humidity conditions as the bottle cap for 24
hours.。Cover the weighing bottle cover, precision weighing (W3)。
Calculation：
W3 - W2
Weight gain percentage   100%
W2 - W1
Characterization of Hydration Characterization and Definition of Wet Weight Gain：

Deliquescene：Absorbing enough moisture to form liquid。
Extremely hygroscopic：Wet weight gain not less than 15%。
Having Hygroscopicity：Wet weight gain less than 15% but not less than 2%。
Having a little Hygroscopicity：Wet weight gain less than 2% but not less than 0.2%。
No or almost no moisture: Less than 0.2% wet weight gain。
Standard：
This product has no or almost no moisture。
3.2.S.4.2.2 Identification
3.2.S.4.2.2.1 High performance liquid chromatography
In the chromatogram recorded under the determination, the retention time of the main
peak of the test solution should be the same as the retention time of the main peak of
the reference solution.
3.2.S.4.2.2.2 Infrared spectrophotometry
Reagent：
KBr（Spectrography）
80
Operation：
Take the amount of this product, set in the agate mortar，Add dry KBr powder
as dispersant，Grinding evenly，Placed in the tablet mold to spread evenly，Pressure，
After removal of the pressure, the prepared test piece is taken out and placed in an
infrared spectrophotometer, and the infrared absorption spectrum of the scanning
infrared light is measured.
The other way is taking the standing Vildagliptin ，using the same ways to do
it .
This product's infrared absorption spectrum should be consistent with the
reference map.
3.2.S.4.2.3 Inspection
3.2.S.4.2.3.1 Related Substances
Reagent：
Sodium dihydrogen phosphate（AR）
Triethylamine（AR）
Phosphoric acid（AR）
Hydrochloric Acid（AR）
Water（Ultra pure water）
Acetonitrile（Chromatography level）
Chromatographic conditions：
Use eighteen alkyl silane bonded silica gel as filling agent
Chromatographic （Agilent Extend C18，4.6×250mm，5µm）
column
mobile phase Take phosphate buffer solution (take 3.12 g of sodium
dihydrogen phosphate dihydrate, add water 1000 ml to dissolve，Add
1ml of triethylamine, mix, adjust pH to 2.8 with phosphoric
acid)-acetonitrile (96:4) as mobile phase A, and phosphate
buffer-acetonitrile (75:25) as mobile phase B；
Gradient elution mobile phase A mobile phase B
Time （minute）
condition （%）（%）
81
0 100 0
25.0 100 0
50.0 0 100
50.01 100 0
65 100 0
Detection wavelength 210nm
Flow rate 1.0ml/minute
Column temperature 35℃
Injection volume 20μl
Diluter preparation：
0.2% (v/v) hydrochloric acid solution - acetonitrile (90:10)。
Preparation of Impurity Stock Solution：
Take 10mg each of impurity A, impurity B, impurity C, impurity D, impurity E,
impurity F, and impurity G ， Place 100ml volumetric flasks respectively and add
appropriate amount of diluent to shake.，Dilute to volume with diluent and shake. (100
μg/ml)
System suitability test solution preparation：

Take about 50mg of Vildagliptin Control and place it in a 20ml volumetric flask，
Add appropriate amount of diluent, shake to dissolve, add 0.5ml each impurity stock
solution, dilute to the mark with diluent, shake ， and then you will get 。（ main
component 2.5mg/ml，impurity2.5μg/ml）
Preparation of reference solution：
Take vildagliptin reference substance 25mg, accurately weighed, set in 100ml
volumetric flask，Add appropriate amount of diluent, shake to dissolve, dilute to the
mark with diluent，shaking well ；Precise amount to take 1ml，Setting in 100ml
volumetric flask, dilute to volume with diluent, shake.
Take this product 25mg, accurately weighed, set 10ml volumetric flask，Add the
appropriate amount of diluent to dissolve, dilute to the mark with diluent, shake。
82
System suitability test：
Accurately measure the system suitability test solution 20 μ l into the liquid
chromatograph, record the chromatogram 。 The order of peaks is impurity A
(VGT-1)）、Impurity D (synthetic amide), Impurity C (cyclic guanidine), Impurity G
(carboxylic acid metabolite), Vildagliptin, Impurity B (VGT-2), Impurity F
(disubstituted), Impurity E (diketopiperazines) ), the degree of separation of impurity
G and main peak should be greater than 0.5，Separation degree of other impurities,
separation degree of impurity and main peak should be greater than 1.5 (Remarks:
main peak tailing factor is larger, impurity G and main peak reach baseline separation,
resolution is less than 1.5, verified when impurity G and main peak separation is
greater than 0.5, Baseline separation can be achieved, so the degree of separation of
impurity G and main peak is set to be greater than 0.5) ； Repeat injection with the
reference solution, the relative standard deviation of the continuous 6-pin main peak
area must not exceed 5%. The signal to noise ratio of the main peak in the reference
solution chromatogram is not less than 40.
Operation：
Precisely take 20 μ l of test solution into the liquid chromatograph, record the
chromatogram chart.
Counting：
Single impurity AT WS DT
    P  f  100 %
（% w / w ） AS DS WT
AT is the peak area of the single impurity in the solution chromatogram of the
test sample (except for the blank solvent peak)
AS is the mean of the main peak area in the chromatogram of the reference
solution
WS is the amount of sample of Vildagliptin control (mg)
DS is the dilution factor of the control solution
WT is the sample weight (mg)
DT is the dilution factor of the test solution
P is the content of the Vildagliptin reference substance (%)
f is the correction factor for each impurity (impurity A: 0.5, impurity B: 0.3,
impurity C: 1.0, impurity D: 0.8, impurity E: 0.6, impurity F: 0.6, impurity G: 1.0)
83
Impurity A shall not exceed 0.15%, Impurity B shall not exceed 0.15%, Impurity C
shall not exceed 0.15%, Impurity D shall not exceed 0.15%, Impurity E shall not
exceed 0.15%, Impurity F shall not exceed 0.15%, Impurity G shall not exceed 0.15%,
Others Individual impurities must not exceed 0.1%, and the sum of impurities must
not exceed 0.5%.
3.2.S.4.2.3.2 Enantiomers
Reagent：
Ethyl alcohol absolute（Chromatography level）
AKOS BBS-00004389（Chromatography level）
Chromatographic column CHIRALPAK ®IF （4.6×250mm,5μm）
Mobile phase ethyl alcohol absolute AKOS BBS-00004389;（100:0.1）
Flow rate 0.5ml/minute
Preparation of enantiomer stock solutions：

Approximately 15 mg of the enantiomer control of vildagliptin is taken and
placed in a 100 ml volumetric flask ， Add proper amount of absolute ethanol to
dissolve, dilute to volume with absolute ethanol and shake well. (150μg/ml)
Formulation of system suitability solution：
Take about 100mg of Vildagliptin Control and place it in a 10ml volumetric
flask ， Add 1ml of enantiomer stock solution, add absolute amount of ethanol to
dissolve, dilute to volume with absolute ethanol, and shake well. 。（ The main
component 10mg/ml, enantiomers 15μg/ml）
Take this product 100mg, accurately weighed, set 10ml volumetric flask, add
absolute amount of ethanol to dissolve, dilute to volume with absolute ethanol and
84
shake well. (10mg/ml）
Take 15 mg of Vildagliptin reference substance, accurately weigh it, set it in a
100 ml volumetric flask, add enough ethanol to shake it to make it dissolve, dilute to
volume with absolute ethanol, and shake it. ； Precisely take 5ml, place in a 50ml
volumetric flask, dilute to volume with absolute ethanol and shake well. (15μg/ml)
Accurately measure the system suitability solution 10 μ l, inject the liquid
chromatograph, record the chromatogram，The degree of separation of the enantiomer
peak from the main component peak must not be less than 0.9（Remarks: The system
suitability solution peak order is the enantiomeric peak, the main component peak, the
main peak tail, the enantiomeric peak and the main component peak can be separated
but the degree of separation is less than 1.5, the verified enantiomer When the
peak-to-peak resolution is greater than 0.9, baseline separation can be achieved.
Therefore, the resolution of the enantiomeric peaks and the main component peaks of
the applicability solution of the defined system should not be less than 0.9.）；Precisely
take 10 μ l of the reference solution and repeat the injection. Record the
chromatogram. The RSD of the 6-pin main component peak area must not exceed
5.0%.
Operation：
Precisely take 10 μ l of the test solution and inject it into the liquid
chromatograph to record the chromatogram.
AT WS DT
Enantiomer s     P 100 %
AS DS WT
（% w / w ）
In the calculation ：
AT is the peak area of the enantiomer in the chromatogram of the test solution
AS is the mean of the main peak area in the chromatogram of the reference
solution
WS is the amount of sample of Vildagliptin control (mg)
85
DS is the dilution factor of the control solution
DT is the dilution factor of the test solution
P is the content of the Vigliptin reference substance (%)
standard regulation：
Enantiomers must not exceed 0.15%.
3.2.S.4.2.3.3 Remaining solvent and 3-AMINO-1-ADAMANTANOL
Chromatography condition：
Capillary column with 6% cyanopropylphenyl-94%
Chromatographic column dimethylpolysiloxane as stationary phase (KB-624
30m0.53mm, 3.00μm)
The temperature was increased by the program, the
initial temperature was 40°C, maintained for 5 minutes,
Column temperature
and the temperature was raised to 220°C at a rate of
25°C per minute for 10 minutes.
Inlet temperature 250℃
Detection port
280℃
temperature
Control mode Pressure control mode（30KPa）
Split ratio 10:1
carrier gas Nitrogen
Detector flame ionization detector（FID）
Preparation of residual solvent stock solution：

Take methanol 300 mg, ethanol 500 mg, methylene chloride 60 mg,
N,N-dimethylformamide (DMF) 88 mg, triethylamine 32 mg and isopropyl alcohol
500 mg，precisely weighed, set in the same 100ml volumetric flask, dilute to the mark
with dimethyl sulfoxide, shaked. (Methanol 3 mg/ml, Ethanol 5 mg/ml,
Dichloromethane 0.6 mg/ml, DMF 0.88 mg/ml, Triethylamine 0.32 mg/ml,
Isopropanol 5 mg/ml）
Preparation of aminoamide and 3-amino-1-adamantanol stock solution：
Take 100 mg of 3-amino-1-adamantanol, accurately weigh it, place it in a 100-ml
volumetric flask, add dimethyl sulfoxide and shake it to dissolve it, dilute to the mark
86
with dimethyl sulfoxide, and shaked.（1mg/ml）
Precisely weigh 5ml each of the above stock solutions, set in the same 50ml
volumetric flask, dilute to the mark with dimethylsulfoxide, and shake.（Methanol 0.3
mg/ml, Ethanol 0.5 mg/ml, Dichloromethane 0.06 mg/ml, DMF 0.088 mg/ml,
Triethylamine 0.032 mg/ml, Isopropanol 0.5 mg/ml, 3-Amino- 1-adamantane Alcohol
0.1mg/ml）
Take this product 1g, accurately weighed, set in 10ml volumetric flask, add dimethyl
sulfoxide to the appropriate amount of shaking to dissolve, dilute to the mark with
dimethyl sulfoxide, shake, that is, too. (100mg/ml)
Precisely weigh 1 μ of the reference solution, repeat to inject, and record the
chromatogram. The RSD % of the peak area of each component of 6 consecutive
needles should not exceed 10%. ； The order of peaks was methanol, ethanol,
isopropanol, dichloromethane, triethylamine, N,N-dimethylformamide, dimethyl
sulfoxide, 3-amino-1-adamantanol. ， The degree of separation between the peaks of
each component should meet the requirements.
operation：
Precisely weigh 1μ of the test solution and inject it into the gas chromatograph
to record the chromatogram.。
remaining solvent ATi WSi DT
    100%
（%，w / w） ASi DS WT
In the calculation ：
ATi is the peak area of each residual solvent in the chromatogram of the test solution
ASi is the average of the corresponding residual solvent peak area in the
chromatogram of the reference solution
DS is the control solution dilution factor

87
DT is the test solution dilution factor
WSi is the reference sample weight (mg)

Methanol must not exceed 0.3%, ethanol must not exceed 0.5%, isopropanol
must not exceed 0.5%, dichloromethane must not exceed 0.06%, triethylamine must
not exceed 0.032%, and N,N-dimethylformamide must not exceed 0.088%.
3-Amino-1-adamantanol must not exceed 0.1%.
3.2.S.4.2.3.4 Moisture
Reagent：
Methyl alcohol（AR）
Fisher's test solution（AR）
Pretreatment test：
Add the appropriate amount of methanol to the reaction flask and set it to the end
point.
Precisely weigh about 10mg of purified water and inject it into the reaction flask
until one titration is complete and record the data. Repeat 2 more times.
W1
Calculate F value according to the following formula： F 
V1
In the calculation ： F is the weight of water per 1 ml of Fisher's test solution

（mg）
W1 is the weight of purified water（mg）
V1 is the volume of Xius test solution consumed to titrate purified water.
（ml）
The RSD of the F values measured three times should not exceed 1.0%. The
average of the F values measured three times is the F value of the Fergus test solution
that was calibrated this time.
Sample determination：
Take this product 1.0g, accurately weighed, set the reaction flask, to be
completed by one titration, record data. Repeat the measurement twice and calculate
the water content separately. Use the average value as the report value.
88
Water content in the test article V2  F
 100%
（%，w/w） W2
In the Calculation：F is the weight of water (mg) per 1 ml of Fisher's test solution
W2 is the weight of the test sample (mg)
V2 is the volume of sample solution (ml) consumed for titration of the test sample
standard：
The moisture content must not exceed 0.5%.
3.2.S.4.2.3.5 Sulfate
Reagent：
Water（Purified Water）
Potassium sulphate（AR）
Barium chloride（AR）
Preparation of the test product solution：
Take this product 1.0g, accurately weighed, set in 50ml Nessler colorimetric tube,
add water to make it into 40ml, add hydrochloric acid to make it neutral, add dilute
hydrochloric acid 2ml, shake.
Preparation of standard potassium sulfate solution：
Weigh 0.181g of potassium sulfate and place it in a 1000ml volumetric flask.
Add enough water to dissolve and dilute to the mark. Shake well. （ Each 1ml is
equivalent to 100μg SO4）.
Precisely take 2.0ml of standard potassium sulfate solution and place it in a 50ml
Nessler colorimetric tube. Add about 40ml of water and dilute hydrochloric acid 2ml.
Shake well.
Operating：
For the test solution and the reference solution, add 5ml of 25% cesium chloride
solution, dilute to 50ml with water, shake well. ， Place for 10 minutes on the same
black background. Observe and compare from above the colorimetric tube. The
turbidity of the test tube should be lighter than the turbidity of the control tube.
Precautions：
89
（1） After adding 25% cesium chloride solution, shake it well to avoid
affecting turbidity.
（2） 25% cesium chloride solution stored for too long, if precipitated, can not
be used, should be reconfigured。
（3） The test tube and the control tube should be placed on a black
background and the turbidity should be observed from the top to the
bottom. When necessary, the position of the test tube and control tube can
be changed and observed。
(4)After the Nessler colorimetric tube is used, it should be washed with water
immediately and no brushing should be applied to avoid scratching the colorimetric
tube.
Standard:
Not more than 0.02%.
3.2.S.4.2.3.6 Chloride
Reagent：
Water（Purified Water）
Nitric acid（AR）
Sodium chlorideAR）
Silver nitrate test solution
Take this product 0.1g, accurately weighed, set in the beaker, add water to
dissolve into 25ml, add nitric acid to make it neutral, add dilute nitric acid 10ml, if the
solution is not clear, should be filtered；Set 50ml Nessler colorimetric tube, add water
to make it about 40ml, shake well.
Preparation of standard sodium chloride solution：
Weigh 0.165 g of sodium chloride into a 1000-ml volumetric flask, add water to
dissolve and dilute to volume, shake ； Before use, accurately take 10ml of stock
solution, place it in a 100ml volumetric flask, dilute to the mark with water, shake
well, and obtain (Each 1ml corresponds to 10μg of Cl).
Precisely take 2ml of standard sodium chloride solution, place in 50ml Nessler
colorimetric tube, add dilute nitric acid 10ml, add water to make about 40ml, shaked.
90
Operation：
For the test solution and the control solution, add 1.0 ml of silver nitrate solution,
dilute to 50 ml with water and shake well. ，Place in darkness for 5 minutes on the
same black background. Observe and compare from above the cuvette. The turbidity
of the test tube should be shallower than the turbidity of the control tube.
Precautions：
（1） The test solution and the control solution should be operated at the same
time, the order of adding reagents should be the same。
（2） It should be noted that in accordance with the order of operations, first
made of 40ml of aqueous solution, then add 1.0ml of silver nitrate test
solution, in order to avoid localized turbidity under a large concentration
of chloride, affecting the turbidity.
（3） The test tube and the control tube should be placed on the same black
surface. The turbidity should be observed from the top to the bottom. It is
easier to judge. When necessary, the position of the test tube and the
control tube can be changed and observed.
（4） After adding the silver nitrate test solution to the test solution and control
solution, immediately shake it thoroughly to prevent over-concentration
locally and affect the resulting turbidity; and place it in a dark place for 5
minutes to avoid direct light irradiation.
（5） After the Nessler colorimetric tube is used, it should be rinsed with water
immediately and no brushing is applied to avoid scratching the
colorimetric tube.
Standard:
Not more than 0.02%.
3.2.S.4.2.3.7 Residue on ignition
Reagent：
Sulfuric acid（AR）
Empty Crucible ‘s constant weight：
Take a clean oven and place it in a high-temperature furnace. Place the lid on the
lid and weigh it at 500°C~600°C to constant weight (W0).
Operation ：
91
Take this product 1.0g，put it in the burning Crucible，Precision weighing (W1），
Slow burning on the electric furnace (it should avoid the test article from escaping by
sudden expansion or burning of heat) ， Igniting until the test article is completely
charred black, and no longer smoke, let cool to room temperature. Sulfuric acid 1ml
was added dropwise to make the charcoal all wet and continue to heat in the electric
furnace until the sulfuric acid vapor was removed and the white smoke disappeared
completely (the above operation should be carried out in a fume hood). The crucible is
placed in a high-temperature furnace, and the crucible lid is placed on the crucible and
is ignited at 500-600°C. for complete ashing. The crucible is placed in a desiccator,
cooled to room temperature, accurately weighed, and then ignited at 500-600°C. To
constant weight (W2).
Calculation：
Burn residue
W - W0
 2  100%
（%，w / w） W1 - W0
In the calculation : W0 is the weight of a crucible that has been ignited to a
constant weight (g)
W1 is the total weight of the samples and crucibles
before ignition (g).
W2 is the total weight of the samples and crucibles after
burning and constant weight (g).
Precautions：
（1） The Crucible should be coded. The lid and code should be the same. The
temperature, the sequence, the cooling time in the dryer, and the weighing
sequence when removing from the high-temperature furnace should all be
the same; the same dehydrator placed in the same dryer should not
exceed 4 at most, otherwise it is difficult to reach the constant weight.。
（2） After the Crucible cools, the negative pressure is easily formed in the
dryer，Care should be taken to open the dryer so as not to disperse the
light residue in the bowl.
（3） When opening and closing the furnace door, care should be taken not to
damage the high-quality fire-resistant insulation layer.
92
Standard：
Residual residue must not exceed 0.1%.
3.2.S.4.2.3.8 Heavy Metal
Reagent：
Acetate buffer（pH3.5）
Standard lead solution（10μg/ml）
Thioacetamide test solution
Phenolphthalein indicator solution
Nitric acid（AR）
Ammonia test solution（AR）
Purified Water
Preparation of acetate buffer solution (pH3.5)：
After extracting ammonium acetate 25g and adding water 25ml to dissolve ，
Adding 7mol/L hydrochloric acid 38ml ， Use 2mol/L hydrochloric acid solution or
5mol/L ammonia solution to adjust pH value to 3.5 accurately and dilute water to
100ml.
Thioacetamide’s Test solution preparation：

Take thioacetamide 4g, add water to dissolve into 100ml，put it in the refrigerator
storage. take the mixed liquor before using （Consists of 1mol/L sodium hydroxide
solution 15ml, water 5.0ml and glycerin 20ml ） 5.0ml ， Add 1.0ml of the above
thioacetamide solution, heat on a water bath for 20 seconds, cool, and use
immediately.
Preparation of standard lead solution：
Accurately weigh 0.1599 g of lead nitrate dried to constant weight at 105 °C,
place in a 1000 ml volumetric flask, add nitric acid 5 ml and water 50 ml, dilute to the
mark with water, and shake as a stock solution.。Pre use ，Precisely take 10ml of stock
solution, place it in a 100ml volumetric flask, dilute to the mark with water, shake
well，Use on the same day (every 1 mg corresponds to 10 μg of Pb).
93
Preparation of sample solution：
i）A tube：Take 0.5ml~1ml of sulfuric acid, set the evaporating dish, heat to low
temperature to dilute the sulfuric acid vapor, add 0.5ml of nitric acid, and evaporate，
removal of nitrogen oxide vapor，Add 2ml hydrochloric acid, add water 15ml after
drying on water bath，Add ammonia test solution to phenolphthalein indicator solution
pink, add acetate buffer (pH3.5) 2ml，After solubilization, the solution was placed in a
Nessler colorimetric tube and 2.0 ml of a standard lead solution was added and diluted
with water to 25 ml.
ii）B tube：Take residue left under the residue of ignition residue, add nitric acid
0.5ml, and evaporate ， After diluting to nitrogen oxide vapor, let cool, add
hydrochloric acid 2ml，Set water bath to dry and add water 15ml，Add ammonia test
solution to phenolphthalein indicator，then add Acetate buffer（pH3.5）2ml，After
microtherm dissolved, it is placed in a Nessler colorimetric tube and diluted with
water to 25ml。
Precautions：
（1） The standard lead solution shall be prepared freshly diluted with standard
lead stock solution before use. The lead solution shall be used on the
same day. The glass containers used for preparing and storing the
standard lead solution shall not contain lead.
(2) When preparing the acetate buffer (pH 3.5), use a pH meter to adjust the pH
of the solution. Care should be taken to control the addition of the
thioacetamide test solution and the development of the thioacetamide test
solution reagent. Color time.
（3） During the inspection, the standard pipe (A pipe) and the test product pipe
(B pipe) shall be operated in parallel, and the reagents shall be added in sequence. The
amount of reagents to be added and the operating conditions shall be the same.
Operation ：
Separately add 2 ml of thioacetamide test solution to both A and B tubes. Shake
them well and place them for 2 minutes on the same white paper. From the top down
perspective, the color of the B tube is compared with that of the tube. Not deeper.
Standard：
94
Heavy metals should not exceed 20 parts per million.
3.2.S.4.2.3.9 Microbial limit
Inspection Method：
Plate method
Specific test operation：
1、Preparation of test solution: take the test sample 10g, dissolve the sample with
100mlph 7.0 sterile sodium chloride-peptone buffer, as 1:10 test solution, take 1:10
test solution 1ml plus 9mlph 7.0 Sterile sodium chloride - peptone buffer mix, as a
1:100 test solution.
2、Bacterial inspection process: take 1:10 and 1:100 test solution for each two,
each 1ml, were placed in a sterile petri dish, and then inject 15-20ml nutrient agar
medium with a temperature not exceeding 45 degrees, mix, After incubating, invert
30-35°C for 3 days, observe the culture results on a daily basis, and finally count.
3、Mold and Yeast Inspection Procedure: Take 1∶10 and 1:100 test solution for
each two, each 1ml, place in a sterile petri dish, and then inject 15-20ml rose sodium
agar cultured at a temperature not exceeding 45 ° C. Base, mix, solidify and invert
23-28°C for 5 days. Observe culture results daily and count.
4、The process of Escherichia coli examination: Take 3 bottles of 100ml bile salt
lactose medium, add 2 bottles of 10ml of test solution of 1:10, take 1 bottle into the
positive control room and add 10-100cfu of Escherichia coli Bacterial suspension;
third bottle added 10ml dilution as negative control ， The above three bottles were
used as enrichment cultures, cultured at 30--35°C for 18-24 hours, observing culture
results, positive for turbidity, and clarified as negative.；Each of the above three bottle
cultures was taken in 0.2 ml each and added to 5 ml of MUG medium, respectively,
and observed under UV light at 366 nm for 5 hours and 24 hours. At the same time,
the uninoculated MUG was used as the background control, and the tube was cultured
under ultraviolet light. Blue-white fluorescence, MUG-positive, no fluorescence,
MUG-negative.After observation, a few drops of enamel matrix test solution was
added along the wall of the culture tube, and the liquid surface was rose-red, which
was positive for the sputum matrix. It was the original color of the reagent and was
negative for sputum matrix. If the MUG is positive and the cesium matrix is positive,
the test product will be tested for Escherichia coli; if the MUG is negative and the
95
sputum matrix is negative, the test sample does not detect E. coli ； If there is
MUG-positive, Indigo-matrix negative or MUG-negative, Indigo-matrix positive, take
the corresponding enrichment cultures were streaked on McConkey agar plates or
beard Methylene blue agar plates, culture 18-24 hours, observe the colony
morphology , If the colony morphology is consistent or suspected, the suspect
colonies should be picked for isolation, purification, staining microscopy and IMViiC
test to confirm whether it is E. coli.
5 、 Check the operation process of live quail: direct inspection method, first
observe with the naked eye, with or without white spots or spots of other colors, and
then examine with 5-10 times magnifier.
3.2.S.4.2.4 Determination of content
Reagent：
Potassium Phosphate Monobasic（AR）
Potassium hydrogen phosphate anhydrous（AR）
acetonitrile（Chromatography level）
methyl alcohol（Chromatography level）
Water（Ultra-pure water）
Diligent：
0.2% (v/v) hydrochloric acid solution - acetonitrile (90:10).
Buffer salt preparation：
Weigh out potassium dihydrogen phosphate 1.3g, add water 1000ml to dissolve, mix,
use dipotassium hydrogen phosphate solution (take dipotassium hydrogen phosphate
trihydrate 2.0g, add water 10ml to dissolve) adjust pH to 6.5, use 0.45 μ m filter
Membrane filtration.
Preparation of mobile phase：
Phase A: A mixed solution of buffer-water-acetonitrile-methanol
(400:600:15:15).
Phase B: Buffer-acetonitrile-methanol (400:450:150) mixed solution。
Take this product 25mg, accurately weighed, set the 50ml volumetric flask, add
the amount of diluent, shake to dissolve, dilute to the mark with a diluent, shake.
96
(0.5mg/ml)
Taking 25mg Vildagliptin, then precision weighed，put it in the 50ml bottle ，Add
appropriate amount of diluent, shake to dissolve, dilute to the mark with diluent,
shake well（0.5mg/ml）
Preparation of system suitability solution：
Take about 50mg of Vildagliptin reference substance, place it into a 10ml
volumetric flask, dissolve it with thinner and dilute to the mark, shake (5mg/ml）；
Precisely take 1ml, place it in a 10ml volumetric flask, add 1ml of 3% H2O2 solution,
place it in a water bath at 80 ° C for about 3 hours, take it out, let cool to room
temperature, dilute to the mark with diluent, and shake well .
Chromatographic column：Waters XTerra ®MS C18（50×4.6mm，3.5μm）
Detection wavelength：210nm
Velocity：1.8ml/minute
Column temperature：35℃
Gradient elution program：
Time（minute） mobile phase A(%) mobile phase B (%)
0 100 0
1.0 100 0
3.0 90 10
8.0 30 70
8.1 100 0
11.0 100 0
System suitability：
With system suitability solution injection, the degree of separation of each
impurity and main component peak should be in accordance with the provisions ；
Repeat the injection with the reference solution and record the chromatogram at the
wavelength of 210 nm. The relative standard deviation of the continuous 5-pin main
peak area must not exceed 1.0%. The tailing factor of the Wigliptin peak should not
exceed 1.8.
97
Operation：
Precisely take 10 μ l of the above solution into the liquid chromatograph and
record the chromatogram.。
Calculation：
AT WS 1
Content（%）   P  100%
AS WT 1- S
In the calculation：
AT is the main peak area in the chromatogram of the test solution。
AS is the average of the main peak area in the chromatogram of the reference
solution。
WT is the test sample weight (mg).
WS is the reference amount (mg) of Wigliptin control.
P is Wigliptin reference substance content (%)。

S is the moisture content of the test sample (%)
Standard：
Calculated as anhydrous, the content of C17H25N3O2 should be 98.0% ~ 102.0%.
98
Weigelieting
Vildagliptin
C17H25N3O2 303.40
This product is
(2S)-1-[[(3-hydroxytricyclo[3.3.1.1[3,7]]decane-1-yl)amino]acetyl]-2-cyanopyrrolidin
e. Calculated on the anhydrous basis, containing C17H25N3O2 should be 98.0% ~
102.0%.
【Character】This product is white to slightly yellow or slightly gray crystalline
powder.
This product is soluble in methanol, ethanol, dimethyl sulfoxide, water or
0.01mol/L hydrochloric acid solution, slightly soluble in acetonitrile or isopropanol.
Melting point The melting point of this product (Appendix VIC of the second
edition of the Chinese Pharmacopoeia 2010) is 146°C to 151°C.
Specific rotation Take appropriate amount of this product, accurately weighed,
add methanol to dissolve and quantitatively dilute to make about 97mg solution per
1ml，Determined according to law (Appendix VIE of the 2010 edition of the Chinese
Pharmacopoeia), specific rotation from -84° to -90°.
（1）In the chromatogram recorded under the determination, the
【Identification】
retention time of the main peak of the test solution should be the same as the retention
time of the main peak of the reference solution.
（2） The infrared absorption pattern of this product should be consistent with
that of the reference substance (Appendix IV C of the Chinese Pharmacopoeia 2010
Edition).
【 Checking 】 Relative substance Take appropriate amount of this product,
accurately weighed ， Dissolve and dilute with thinner to make up about 2.5mg of
solution per 1ml.，as a test solution。Take appropriate amount of vildagliptin reference
99
substance, accurately weighed ，Dissolve with diluent and dilute to make a solution
containing about 2.5 μg per 1 ml as a reference solution。Take appropriate amounts
of impurities A, B, C, D, E, F, G and vildagliptin，Diluent was dissolved and diluted to
make a solution containing about 2.5 μg (single impurity) and 2.5 mg (vildagliptin),
respectively, as a system suitability test solution per 1 ml. 。 According to high
performance liquid chromatography (Chinese Pharmacopoeia 2010 Edition, Part II
Appendix VD), using octadecylsilane bonded silica as a filler (Agilent Extend C18
4.6 × 250mm, 5μm or equivalent performance of the column)；0.02mol/L sodium
dihydrogen phosphate solution (take 3.12g sodium dihydrogen phosphate dihydrate,
add water 1000ml to dissolve, add triethylamine 1ml, mix, adjust pH to 2.8 with
phosphoric acid)-acetonitrile (96:4) Mobile phase A, 0.02mol/L monobasic sodium
phosphate solution - acetonitrile (75:25) as mobile phase B；Gradient elution in the
following table, flow rate 1.0 ml/min, detection wavelength 210 nm, column
temperature 35 ° C 。 Take the system suitability solution 20 μ l, into the liquid
chromatograph, record the chromatogram，The order of peaks is impurity A, D, C, G,
vildagliptin, B, F, and E. The degree of separation of impurity G and main peak
should be greater than 0.5. The degree of separation between other impurities and
main peaks and impurities should meet the requirements.。Take the control solution 20
μl into the liquid chromatograph, record the chromatogram, the signal to noise ratio
of the main peak should not be less than 40, adjust the detection sensitivity, so that the
main component peak height is about 10% of the full scale。Precisely take 20 μl of
the test solution and the reference solution, and inject them into the liquid
chromatograph. Record the chromatograms.。If there are chromatographic peaks in the
chromatogram of the test solution consistent with the retention times of impurities A,
B, C, D, E, F, and G, calculate the corrected peak area according to the principal
component external standard method. Impurity A (correction factor) 0.5) Not to
exceed 0.15%, Impurity B (correction factor 0.3) shall not exceed 0.15%, Impurity C
shall not exceed 0.15%, Impurity D (correction factor 0.8) shall not exceed 0.15%,
Impurity E (correction factor 0.6) shall not exceed 0.15%, Impurity F (correction
factor 0.6) must not exceed 0.15%; Impurity G must not exceed 0.15%; Other
impurities shall not exceed 0.1% as calculated by the principal component external
standard method; total impurities shall not exceed 0.5%.
100
Time（minute） mobile phase A(%) mobile phase B (%)

0 100 0
25 100 0
50 0 100
50.01 100 0
65 100 0
Thinner: 0.2% (v/v) hydrochloric acid solution - acetonitrile (90:10)
Enantiomers Take this product about 100mg, accurately weighed, set 10ml
volumetric flask, add ethanol to dissolve and dilute to the mark, shake, as the test
solution 。 Another 15 mg of Vildagliptin reference substance was accurately
weighed and placed in a 100-ml volumetric flask. An appropriate amount of absolute
ethanol was added and shaken to dissolve. Diluted to the mark with absolute ethanol
and shake it. Precisely take 5 ml and set 50 ml. In the bottle, dilute to volume with
absolute ethanol and shake it as a reference solution。Take about 15mg of enantiomers
of vildagliptin, place it in a 100ml volumetric flask, add absolute amount of absolute
ethanol, shake to dissolve, dilute to volume with absolute ethanol, shake well, and use
it as an impurity stock solution. Take about 100mg of Vildagliptin reference substance,
place it in a 10ml volumetric flask, add absolute amount of absolute ethanol to
dissolve, add 1ml of impurity stock solution, dilute to volume with absolute ethanol,
and shake well as a system suitability solution。According to high performance liquid
chromatography (Chinese Pharmacopoeia 2010 Edition, Part II Appendix VD)
Determination, the surface of silica gel covalently bonded with
amylose-tris-(3-chloro-4-methylphenyl carbamate) as filling (CHIRALPAK® IF 4.6
×250mm, 5μm or equivalent column)；Ethanol-diethylamine (100:0.1) was used
as the mobile phase; flow rate was 0.5 ml/min, detection wavelength was 220 nm,
and column temperature was 35°C.Take the system suitability solution 10μl, into
the liquid chromatograph, record the chromatogram, the resolution of the peak of
enzymatic peak and the peak of vitiligo should not be less than 0.9。Precisely take 10
μ l of the test solution and the reference solution, and inject them into the liquid
chromatograph. Record the chromatograms.。If there is a chromatographic peak in the
101
chromatogram of the test solution that corresponds to the retention time of the
enantiomer peak of vidextigate, the peak area shall not exceed 0.15% by the principle
component external standard method.
3-Amino-1-adamantanol and residual solvents
3-AMINO-1-ADAMANTANOL、
methyl alcohol、Ethanol、Dichloromethane、N,N-Dimethylformamide、Triethylamine、
iso-Propyl alcohol Take appropriate amount of this product, accurately weighed,
add dimethyl sulfoxide solution and dilute to make a solution containing 100mg per
1ml as the test solution ； Separately take methanol, ethanol, methylene chloride,
N,N-dimethylformamide, triethylamine, isopropanol, and 3-amino-1-adamantanol,
respectively. ， Precisely weighed, dissolved and diluted with dimethyl sulfoxide to
produce approximately 300 μg of methanol per 1 ml, 500 μg of ethanol, 60 μg of
methylene chloride, 88 μg of N,N-dimethylformamide, 32 μg of triethylamine, and
isopropyl alcohol 500 μg, 3-amino-1-adamantanol 100 μg solution ， As a control
solution. According to the residual solvent determination method (Chinese
Pharmacopoeia 2010 Edition II Appendix III P third method), the capillary tube with
6% cyanopropylphenyl-94% dimethylpolysiloxane (or similar polarity) as a fixed
solution Columns (eg KB-624 30m0.53mm, 3.00μm) ； The starting temperature is
40°C, maintained for 5 minutes, and raised to 220°C at a rate of 25°C per minute for
10 minutes; the inlet temperature is 250°C ； The detector is a hydrogen flame
ionization detector (FID) with a detector temperature of 280 °C and a carrier gas of
nitrogen。Take 1μl of the reference solution and inject it into the gas chromatograph.
Record the chromatogram. The order of peaks is methanol, ethanol, isopropanol,
dichloromethane, triethylamine, N,N-dimethylformamide, dimethyl Asia. Sulfone,
3-amino-1-adamantanol,The degree of separation between chromatographic peaks
should meet the requirements。Precisely take 1μl of the test solution and the reference
solution, inject them into the gas chromatograph, and record the chromatograms. ，
According to the calculation of peak area by external standard method, methanol,
ethanol, isopropyl alcohol, dichloromethane, triethylamine, N,N-dimethylformamide
and 3-amino-1-adamantanol should not exceed 0.3. %, 0.5%, 0.5%, 0.06%, 0.032%,
0.088%, and 0.1% 。
102
Sulfate Take this product 1.0g, check according to law (Chinese
Pharmacopoeia 2010 version of the second Appendix VIII B), and the standard
potassium sulfate solution 2.0ml made of the reference solution can not be more
concentrated (0.02%).
Chloride Take this product 0.1g, check according to law (Chinese
Pharmacopoeia 2010 version of the second Appendix VIII A), and the standard
solution of sodium chloride 2.0ml made of the reference solution can not be more
concentrated (0.02%).
Moisture Take this product, according to the determination of moisture
(Chinese Pharmacopoeia 2010 version II Appendix VIII M first method A)
determination, the moisture content must not exceed 0.5%.
Residue on ignition Take this product 1.0g, check according to law (Chinese
Pharmacopoeia 2010 version II Appendix VIII N), leaving no residue left over 0.1%.
Heavy metal Take the residues left over from the residue on ignition, check
according to law (Chinese Pharmacopoeia 2010 Second Edition Appendix II H second
method), containing heavy metals may not exceed twenty millionths.
【 Determination of content 】 According to high performance liquid
chromatography (Chinese Pharmacopoeia 2010 Edition, Part II Appendix V D)
determination.
Chromatographic conditions and System suitability test Use
octadecylsilane-bonded silica gel as a filler (Waters XTerra MS C18, 50 x 4.6mm, 3.5
μ m or equivalent column) ； Phosphate buffer solution [take potassium dihydrogen
phosphate 1.3g, add water 1000ml to dissolve, use dipotassium hydrogenphosphate
solution (take dipotassium hydrogen phosphate trihydrate 2.0g, add water 10ml to
dissolve) adjust pH to 6.5]-water- Acetonitrile-methanol (400:600:15:15) for mobile
phase A，Phosphate Buffer-Acetonitrile-Methanol (400:450:150) as Mobile Phase B；
according to the below chart to gradient elution，velocity of flow is 1.8ml /minute，Column
temperature is 35 ° C, detection wavelength is 210nm 。 Take about 50mg of
Vildagliptin reference substance, place it into a 10ml volumetric flask, dissolve it
with 0.2% (v/v) hydrochloric acid solution - acetonitrile (90:10) and dilute it to
the mark.；Take 1ml, place in a 10ml volumetric flask, add 1ml of 3% H2O2 solution,
place in a water bath at 80°C for about 3 hours, remove, cool to room temperature,
and dilute with 0.2% (v/v) hydrochloric acid solution - acetonitrile (90:10) Scale,
103
shake, as system suitability solution。Take the system suitability solution 10μl, into
the liquid chromatograph, record the chromatogram，The separation of the peaks of
vitilagliptin and adjacent impurity peaks should not be less than 1.5；Precisely take 10
μ l of the reference solution, inject it into the liquid chromatograph, record the
chromatogram, and inject 5 times continuously.，The relative standard deviation of the
peak area of vildagliptin should not be greater than 1.0%, and the tailing factor of the
vildagliptin peak should not be greater than 1.8.
Time（minute） Mobile phase A（%） Mobile phase B（%）

0 100 0
1.0 100 0
3.0 90 10
8.0 30 70
8.1 100 0
11.0 100 0
Determination method Take this product 25mg, accurately weighed, set in a
50ml volumetric flask, add 0.2% (v/v) hydrochloric acid solution - acetonitrile (90:10)
dissolved and diluted to the mark, shake ， Precisely take 10 μ l into the liquid
chromatograph and record the chromatogram. Also take the Wigliptin reference
substance and measure it with the same method. Calculated by peak area according to
external standard method。
【Category】Hypoglycemic drugs
【Preparations】Vildagliptin tablet
【Storage】Sealed, stored at room temperature。
【Available period】Tentative 24 months
Impurity related information：
Impurity
Impurity structure
name
104
105
3.2.S.4.3 Verification of Analytical procedures
The traits, melting point, solubility, specific rotation, moisture, sulfate, chloride,
ignition residue, heavy metal, and identification (2) of the quality standard of
vildagliptin are routine check items ， The company validated the method for the
relevant substances, residual solvents, enantiomers, content, and microbial limits in
the quality standards.，The method of identification (1) is consistent with the content
determination method. The method verification results are in accordance with the
standards specified in the verification protocol.
Sample information for method validation is shown in the table below Table
3.2.S.4.3-1：Method validation sample information table
3.2.S.4.3.1 Methodological Verification of related substances inspection

Batch
Sample information batch Production Date Preparation site
number
Vildagliptin 2015.6.2~2015.6.
150601 115.0kg Anhui Haikang Pharmaceutical CO.,LTD
（Verification batch） 10
Vildagliptin 2015.6.11~2015.
（Verification batch） 6.19
Vildagliptin 2015.6.20~2015.
（Verification batch） 6.28
3.2.S.4.3.1 The results of the method verification of the material inspection are
summarized in Table 3.2.S.4.3.1-1.
project Validation results

The separation of impurity G from the main peak is 0.604, which is greater than 0.5; the separation of other impurities
from the main peak and the degree of separation between impurities are all greater than 1.5, which meets the system
System suitability requirements.
suitability The average peak area of the main peak of continuous 6 solution for the solution was 34947, RSD was 2.11%, less than
5.0%, and the signal to noise ratio (pin 1) of the main peak of the reference solution was 236.6, which was greater than 40,
which met the system suitability requirements.。
106
Blank solvent does not interfere with the determination of related substances in this product；The degree of separation of
impurity G and main peak is greater than 0.5, and the separation of other impurities and main peaks and impurities is
Specific greater than 1.5；The PDA spectrum of the degraded sample shows that the separation degree of impurity G and main peak
properties is greater than 0.5, and the separation degree of other impurities and main peaks is greater than 1.5. The increase of
impurities and the degradation of main peak basically reach the material balance, and the peak purity meets the
requirements.
LOQ LOD
Equivalent to Equivalent to
Components concentration working concentrati working
（μg/ml） concentration on（μg/ml） concentration
percentage percentage
vildagliptin 0.589 0.024% 0.177 0.007%
Sensitivity ImpurityA 0.632 0.025% 0.190 0.008%
ImpurityB 0.675 0.027% 0.202 0.008%
ImpurityC 0.669 0.027% 0.201 0.008%
ImpurityD 0.627 0.025% 0.188 0.008%
ImpurityE 0.594 0.024% 0.178 0.007%
ImpurityF 0.646 0.026% 0.194 0.008%
ImpurityG 0.659 0.026% 0.198 0.008%
107
Correlation
Components Linear range Linear equation
coefficient R2
0.589μg/ml~4.711μg/ y=15494.5368x-1752.
vildagliptin 0.9991
ml 3013
0.632μg/ml~5.059μg/ y=31276.9091x-766.3
Impurity A 1.0000
ml 035
0.675μg/ml~5.399μg/ y=49500.0709x-1409.
Impurity B 0.9999
ml 0141
0.669μg/ml~5.355μg/ y=16827.0595x-320.0
Impurity C 1.0000
ml 525
0.627μg/ml~5.012μg/ y=18572.2216x-2828.
Impurity D 0.9996
ml 1492
0.594μg/ml~4.756μg/ y=26151.5552x-436.0
Impurity E 0.9994
ml 853
Linear
0.646μg/ml~5.170μg/ y=25578.3979x+2144
range Impurity F 0.9995
ml .2483
0.659μg/ml~5.269μg/ y=17055.2108x-1537.
Impurity G 0.9999
ml 0353
The response and concentration of each component showed a linear relationship. The correction factors for each
impurity are:
The ascription of the peak Correction factor
vildagliptin 1.00
Impurity A 0.5
Impurity B 0.3
Impurity C 0.9
Impurity D 0.8
Impurity E 0.6
Impurity F 0.6
Impurity G 0.9
108
Recovery rate average
RSD(%)
Components value （%）
Method 1 Method 2 Method 1 Method 2
Impurity A 98.50 94.15 3.19 3.19
Impurity B 101.39 97.72 2.95 2.95
Recovery
Impurity C 106.39 101.83 4.34 4.34
rate
Impurity D 99.12 98.06 1.95 1.97
Impurity E 97.49 93.97 1.75 1.75
ImpurityF 93.28 87.31 4.68 4.68
ImpurityG 94.13 92.03 3.60 3.60
The average recovery rate of each impurity was between 80% and 120%, which met the verification standard
Unknown Total
Impurity Impurity D(%) Impurity 2(%)
1(%) impurity (%)
Sample 1 0.031 0.075 0.047 0.153
Sample 2 0.030 0.076 0.045 0.152
Sample 3 0.031 0.078 0.045 0.155
Repeatabilit Sample 4 0.032 0.080 0.047 0.159
y Sample 5 0.032 0.082 0.046 0.160
Sample 6 0.034 0.073 0.045 0.152
average value 0.032 0.077 0.046 0.155
RSD% 3.70 4.19 2.15 2.42
In the same solution containing 6 parts of Vildagliptin, impurity D (synthetic amide) and 2 unknown impurities were
detected. The impurity spectrum was the same. The RSD of each impurity detected was less than 10%. Claim.
Unknown Unknown Total l
Impurity D
Sample impurity 1 impurity 2 impurity
（%）
（%）（%）（%）
sample1 0.031 0.075 0.047 0.153
sample2 0.030 0.076 0.045 0.152
sample3 0.031 0.078 0.045 0.155
Repeatability
Intermediat sample4 0.032 0.080 0.047 0.159
e precision sample5 0.032 0.082 0.046 0.160
sample6 0.034 0.073 0.045 0.152
sample1 0.028 0.066 0.036 0.130
sample2 0.033 0.065 0.039 0.137
Intermediate
sample3 0.034 0.064 0.037 0.135
precision
sample4 0.033 0.066 0.037 0.136
sample5 0.032 0.068 0.042 0.141
109
sample6 0.032 0.066 0.041 0.139
average value / 0.032 0.072 0.042 0.146
RSD(%) / 5.09 9.01 9.80 7.04
Another experimenter reconfigures 6 samples of the test solution with different instruments on different dates. Impurity D
(synthetic amide) and 2 unknown impurities are detected. The impurity spectrum is the same. The RSD of the 12 data is
less than 10%. Meet the requirements.
Solution The test solution was allowed to stand at room temperature for 25 hours. The number of impurities detected was the same,
stability the amount of impurities detected was not changed significantly, and the solution was stable.。
1) When buffer pH is adjusted + 0.1, flow rate is adjusted - 0.1 ml/min, buffer salt concentration is adjusted + 5%,
column temperature is ± 5 °C, wavelength is adjusted ± 2 nm，After replacing the same brand of the same type of
column and replacing different brands of the same type of column, the peak sequence of each component did not change,
and the degree of separation met the requirements. The recovery of each impurity was between 80% and 120%, which was
consistent with the durability. Claim;
(2) When the pH value is -0.1, the degree of separation of the impurity G and the main peak is less than 0.5, which does
not meet the requirements of durability. Therefore, the pH of the mobile phase must be strictly controlled.；
（3）When the flow rate is +0.1, the unknown impurity is contained in the impurity E. The recovery rate of the impurity E
Durability is not between 80% and 120%, and does not meet the durability requirement. Therefore, the flow rate must be strictly
controlled.
（4） When the ratio is -1%, the separation degree of the impurity G and the main peak is less than 0.5, the degree of
separation of the unknown impurity and the impurity E is less than 1.5, and the recovery of the impurity B, G is not
between 80% and 120%；When the proportion is +1%, the degree of separation of impurity A and impurity D is less than
1.5, and the unknown impurity is contained in impurity E. The recoveries of impurities A, G, and E are not between 80%
and 120% and do not meet the durability requirements. Strictly control the ratio.
（5）When the buffer salt concentration is -5%, the unknown impurity is contained in the impurity E, so the buffer salt
concentration must be strictly controlled.。
Summary of method verification results of material inspection
3.2.S.4.3.1.1 Instrument and test medicine

Instrument and test medicine see table 3.2.S.4.3.1-2
Table ：3.2.S.4.3.1-1 Instrument and test medicine
Model Manufactures
LC-16 Shimadzu
Instrument Waters
waters
2996-2695
110
FA1004N Shanghai Jinghai Instrument Co., Ltd.

PHS-3C Shanghai Yidian Science Instrument Co., Ltd.
150301 Anhui Haikang Pharmaceutical Co., Ltd.
20131219 Shenzhen Zhenqiang Biotechnology Co., Ltd.
TH11937-
Shenzhen Zhenqiang Biotechnology Co., Ltd.
Test medicine 018
1424-093
A3
1497-003
A7
1424-069
A3
3.2.S.4.3.1.2 Related material analysis method establishment

（1）Impurity research strategy
Refer to 3.2.S.3 impurity analysis section，The list of organic impurity control
strategies for the raw material of vildagliptin is as follows:
Whether to
Impurity Control Limited
Impurity name Structural formula source set quality
information Strategy （%）
standards
Starting Starting
Residual
materials-Introdu L-Prolinamide materials-Intr 0.1 Yes
solvent
ce oduce
Under
Starting materials Starting
3-AMINO-1-ADA Residual 0.1 Yes
二 materials
MANTANOL solvent
Under the Starting

VGT-1 Impurities A relevant materials-Intr 0.15 Yes
substances oduce
111
Under the
Starting materials Process
Impurities B relevant 0.15 Yes
一 impurities
substances
Process
Under the
impurities/De
V225 Impurities C relevant 0.15 Yes
gradation
substances
impurities
Process
Under the
impurities/De
Vildagliptinacylami Impurities D relevant 0.15 Yes
gradation
de substances
impurities
Process
Under the
impurities/De
V226 Impurities E relevant 0.15 Yes
gradation
substances
impurities
Under the
Process
V227 Impurities F relevant 0.15 Yes
impurities
substances
Process
Under the
impurities/De
V221 Impurities G relevant 0.15 Yes
gradation
substances
impurities
112
VGT-2Und
Starting
er the
— Impurities H materials-Intr 0.1 No
relevant
oduce
substances
VGT-2Und
Starting
VGT-2 er
Impurities J materials-Intr 0.5 NO
Enantiomer enantiomer
oduce
conditions
Process
Under
Vildagliptin impurities/De
Impurities K enantiomer 0.15 Yes
enantiomer gradation
conditions
impurities
Process
— Impurities I HPLC-MS 0.1 NO
impurity
（2）Method selection
Reference to the Vildagliptin Tablets import registration standard (standard
number: JX20100243) related material determination method, select high
performance liquid chromatography (HPLC) for the determination of the related
substances.
The HPLC condition is：
Waters XTerra MS C18 （ 50×4.6mm ， 3.5μm ）； With phosphate buffer solution
[potassium dihydrogen phosphate 1.3g, adding water 1000ml to dissolve, with two
potassium hydrogen phosphate (three hydrous hydrogen phosphate, two potassium
2.0g, and water 10ml to dissolve) adjustment pH value to 6.50 + 0.05] as the mobile
phase A, acetonitrile 1000ml as the mobile phase B, methanol 1000ml as the liquid
phase C, water 1000ml as the mobile phase, gradient elution mode such as At the
same time, the flow rate is 1.8ml per minute, and the detection wavelength is 210nm.
Mobile phase A Mobile phase B Mobile phase C Mobile phase D

Time （min）
(%) (%) (%) (%)
0 38.8 1.5 1.5 58.2
113
1.0 38.8 1.5 1.5 58.2
3.0 39.0 6.0 3.0 52.0
8.0 40.0 32.0 11.0 17.0
8.1 38.8 1.5 1.5 58.2
11.0 38.8 1.5 1.5 58.2
Blank solvent：
0.2% hydrochloric acid solution - acetonitrile (90:10);
Preparation of single impurity location solution：
Take impurity C, impurity E, impurity F and impurity G each 1mg, respectively, set
10ml volumetric flask, add appropriate amount of diluent to shake to dissolve, dilute
to the mark with diluent, shake, that is, too. (0.1mg/ml)

Take vildagliptin 12.5mg, accurately weighed, set 25ml volumetric flask, add
appropriate amount of diluent to dissolve, dilute to the mark with diluent, shake, that
is.
Operating ：
Take 10 μ l of each of the above solutions into a liquid chromatograph (PDA
detector) and record the chromatograms.
Experimental results：
The test results are shown in Table 3.2.S.4.3.1-3, and the chromatograms are
shown in table 3.2.S.4.3.1-1~7.。
Table 3.2.S.4.31-1 Related Material Methods - Test Results
Maximum absorption
Peak ownership keep time Trailing factor
wavelength（nm）
Impurity C 3.348 1.24 195.4
Impurity E 5.878 1.01 196.5
Impurity F 7.382 1.01 198.9
Impurity G 3.042 0.58 201.2
Vildagliptin 5.063 0.94 197.7
In conclusion ：It can be seen from the above results：The maximum absorption of
each impurity peak and the main component peak at the end wavelength is at the same
114
time as the wavelength of the determination of the substance for this product, which is
in accordance with the detection wavelength of the related substances in the
Vildagliptin Tablets import registration standard; the impurity G trailing factor is 0.58,
the peak shape is poor, the expected sensitivity is inconsistent, and the
chromatographic conditions need to be further optimized.
(1) Method optimization
The result of the method optimization is shown in table 3.2.S.4.3.1-4.
Table 3.2.S.4.31-1The result of the optimization of material methods
Chromatographic Result ：
conditions
optimiz chromatographic column： Peak Trailing
Rt(min)
ation1 Agilent Extend attribution factor
C18(4.6×250mm,5μm) ND Impurity E ND
Detection wavelength：210nm 3.171 Impurity C 1.30
Preparation of buffer salt： 17.640 Impurity F 0.79
10mmol/LSodium 3.765 Impurity G 0.77
dihydrogen phosphate 4.385 Vildagliptin 1.83
solution phosphoric Conclusion: Impurity G is better than

acid（1000:0.5） condition 1 peak shape, impurity E does
mobile phase：Buffer salt - not peak under isocratic conditions, and

gradient conditions are used instead。
acetonitrile（90:10）
Impurities C, E, F, G
concentrations：
0.1mg/ml
Concentration of Vildagliptin：
0.5mg/ml
optimiz Column：Agilent Extend Mixed solution test results：
ation2 C18(4.6×250mm,5μm) Absorp
Peak
Detection wavelength：210nm Rt(mi Resolut tion
ownershi
Mobile phaseA ： n) ion wavele
p
10mmol/L-sodium ngth
115
dihydrogen phosphate （nm）
solution - phosphoric Impuritie
6.635 -- 210.0
acid（1000:0.5） sA
Mobile phase B：Acetonitrile 7.230 Unknown 0.93 /
Gradient elution method see the Impuritie
8.625 2.49 195.4
following table sC
Time 11.22 Impuritie
A(%) B(%) 4.14 198.9
（min） 3 sG
0 95 5 12.01 Vildaglipt
0.82 197.7
5 95 5 1 in
15 90 10 16.86 Impuritie
6.79 202.4
25 75 25 7 sB
30.01 95 5 23.13 Impuritie
18.97 200.0
40 95 5 8 sF
Injection volume: 10μl of a 25.29 Impuritie
10.14 195.4
single positioning 8 sE
solution; 20μl of Positioning solution test results：
mixed solution： Absorption
Peak
The main component 0.5mg/ml, Rt(min) wavelength
ownership
each impurity 0.005 （nm）
mg/ml 6.655 Impurities A 210.0
Test solution concentration: 16.914 Impurities B 202.4
0.5mg/ml 8.264 Impurities C 195.4
Impurity C positioning solution
6.019 Impurities D 190.7
concentration: 0.1mg/ml
25.285 Impurities E 196.5
Impurity E positioning solution
22.982 Impurities F 198.9
11.072 Impurities G 201.2
Impurity F positioning solution
Conclusion: From the above results, it
can be seen that impurities A, B, D, C,
Impurity G positioning solution
E, F, G, and main component peaks all
have maximum absorption at the end
Impurity A positioning solution
wavelength, and provisional 210 nm is
116
concentration: 0.1mg/ml used as the wavelength for
Impurity B positioning solution determination of related substances in
concentration: 0.1mg/ml this product; impurity A and unknown
Impurity D positioning solution The separation of the peak, impurity G,
concentration: 0.1mg/ml and the Vildagliptin peak did not meet
the requirements, and the conditions
were optimized.
Optimiz Buffer salt preparation: 10 mmol/L Rt(min) Peak ownership Resolution
ation 3 sodium dihydrogen phosphate 6.497 Impurities A --
solution + 0.2% triethylamine (pH 6.931 Impurities D 0.888
adjusted to 2.57 with phosphoric 7.694 Impurities C 1.958
acid) 10.705 Impurities G 4.543
Mobile Phase A: Buffered Salt - 11.509 Vildagliptin 0.691
Acetonitrile (75:25)
20.117 Impurities B 8.580
Mobile Phase B: Buffered Salt -
Acetonitrile (95:5)
Gradient elution method see the
Conclusion: From the above results, it can be
following table
seen that the degree of separation of impurity
Time（min） A(%) B(%)
A and impurity D, impurity G, and
0 0 100
vildagliptin peak does not conform to
15 0 100 regulations, and the conditions are optimized.
50 100 0
50.01 0 100
60 0 100
Mixed solution concentration：
main ingredient:1.0mg/ml ， Each
impurity 0.005 mg/ml
117
Optimiz Buffer salt preparation: 20 mmol/L Degree of
Rt(min) Peak attribution
ation4 sodium dihydrogen phosphate separation
solution (pH adjusted to 2.8 with 7.779 Impurities A --
phosphoric acid) 11.362 Impurities D 5.517
Mobile Phase A: Buffered Salt - 13.995 Impurities C 4.136
Acetonitrile (75:25) 17.708 Impurities G 2.868
Mobile Phase B: Buffered Salt - 20.086 Vildagliptin 0.910
Acetonitrile (96:4)
21.833 Impurities B 0.134
Gradient elution method see the
following table
Time （min） A(%) B(%)
Conclusion: From the above results, it can be
0 0 100
seen that the degree of separation of impurity
20 0 100 G from vildagliptin, vildagliptin, and impurity
50 100 0 B does not conform to regulations, and the
50.01 0 100 conditions are optimized.
65 0 100
Injection volume: 10μl
Mixed solution concentration:
The main ingredient 1.0mg/ml, each
impurity 0.005 mg/ml
Optimiz Buffer salt preparation: 20 mmol/L Peak Resoluti Tailing

Rt(min) s/n
ation5 sodium dihydrogen phosphate ownership on factor
solution + 0.1% triethylamine Impurities
7.409 -- 0.958 577
(phosphoric acid adjusted to pH 2. A
Mobile Phase A: Buffered Salt - Impurities
8.323 1.501 0.876 434
Acetonitrile (75:25) D
Mobile Phase B: Buffered Salt - Impurities
10.259 3.422 1.141 804
Acetonitrile (96:4) C
Gradient elution method see the Impurities
following table 13.227 2.203 0.930 169
G
Time （min） A(%) B(%) 15.982 Vildagliptin 1.608 2.103 35689
0 0 100 Impurities
21.933 6.145 1.187 749
25 0 100 B
50 100 0 Impurities
44.013 36.908 1.009 806
50.01 0 100 F
65 0 100 47.054 Impurities 9.087 1.219 1634
118
Injection volume: 10μl E
Mixed solution concentration: Conclusion: The separation degree of
The main ingredient 1.0mg/ml, each each impurity peak and the main peak and the
impurity 0.005 mg/ml degree of separation between impurities are in
compliance with the regulations.
From the above table, the component
with the lowest response factor is impurity G,
and the signal-to-noise ratio is approximately
160 at a concentration of 0.005 mg/ml.
The maximum daily dose of vildagliptin
tablets is 100 mg. According to the technical
guideline ≤ for chemical impurities
research, the reporting limit for the detection
of impurities in the raw material for
vildagliptin is 0.05%. The limit of
quantification should be approximately 50%
of the limit of the report, ie the limit of
quantitation is approximately 0.025%.
Impurity G peaks are similar to those of
Shantou, and in order to balance the
difference in baseline noise of different
detectors, the signal-to-noise ratio of the
quantitative limit concentration is estimated to
be about 40.
Calculated according to the lowest response
impurity G, the signal-to-noise ratio of 40
corresponds to a concentration of 1.25 μ
g/ml ), according to this concentration is
calculated for the 0.025% of the working
concentration of the test sample, the
concentration of the test sample is about
5mg/ml , In this condition, the injection
volume is 10μ
l. According to the preparation of the relevant
substances for the test solution concentration
of about 2.5mg/ml, injection volume adjusted
to 20μl.
Optimiz Buffer salt preparation: 20 mmol/L Peak Resoluti Tailing
Rt(min) s/n
ation 6 sodium dihydrogen phosphate ownership on factor
solution + 0.1% triethylamine Impurities
7.663 -- 0.968 1273
(phosphoric acid adjusted to pH 2.8 A
Mobile Phase A: Buffered Salt - 9.061 Impurities 2.324 0.846 560
119
Acetonitrile (75:25) D
Mobile Phase B: Buffered Salt - Impurities
11.138 3.797 1.122 1056
Acetonitrile (96:4) C
Gradient elution method see the Impurities
14.408 2.921 0.814 200
following table G
Time （min） A(%) B(%) 15.562 Vildagliptin 0.516 6.904 127915
0 0 100 Impurities
22.064 3.972 1.099 1675
25 0 100 B
50 100 0 Impurities
44.159 44.433 1.125 1841
50.01 0 100 F
65 0 100 Impurities
47.362 11.322 1.125 2410
Injection volume: 20μl E
Mixed solution concentration: Conclusion: When the concentration of the
The main component 2.5mg/ml, test sample is changed to 2.5mg/ml, the tailing
impurities 2.5μg/ml factor of the main peak is 6.904, and the
impurity G and the main peak can be
separated from the baseline. The separation of
other impurities and main component peaks
and the degree of separation between
impurities are all greater than 1.5. Determine
the chromatographic conditions for this
product related to the determination of the
conditions. (Chromatograms are as follows)
Note: ND is not detected.。
杂质 F 杂质 E
维格列汀
杂质 G
杂质 B
杂质 A
杂质 D
杂质 C
Chart 3.2.S.4.3.1-1: optimization of 6 mixed solution test results

3.2.S.4.3.1.3 Verification of material analysis methods
The relevant methodology was carried out with a single known impurity limit of 0.1%,
and the specific impurity limits were revised to 0.15% according to the guidelines for
120
the study of chemical drug impurities and the results of the stability test.
3.2.S.4.3.1.3.1 System Applicability
（1）operation
The separation of the impurity G and the principal component peak should be
greater than 0.5 with the application of the system application solution. The separation
degree between the other impurities and the main peaks and impurities should be
more than 1.5; the relative standard deviation of the main peak area of the 6
continuous 6 needles should not exceed 5%, and the signal to noise ratio of the main
peak is not less than 40.
（2）Result
The results of the test are shown in table 3.2.S.4.3.1-5~6. The chromatograms are
shown in table 3.2.S.4.3.1-43~50.
Table 3.2.S.4.3.1-5: results of system suitability test for solution
Peak Relative
Peak # Retention time Resolution
attribution retention time
2 ImpurityA 7.465 0.50 5.800
3 ImpurityD 8.628 0.58 2.061
4 ImpurityC 10.524 0.71 3.723
5 ImpurityG 13.461 0.91 2.582
6 Vildagliptin 14.791 1.00 0.604
7 ImpurityB 21.422 1.45 4.215
8 ImpurityF 44.354 3.00 44.942
9 ImpurityE 47.617 3.22 10.793
Table 3.2.S.4.3.1-5: repeat injection results of reference solution 6 needles.

The signal to noise
ratio of the main peak
Retention time of
Sampling Main peak area of the control solution
main peak
(the first pin)
1 36420 16.993
2 34881 17.031 236.670554
3 34733 16.419
121
4 34444 17.009
5 34585 17.131
6 34618 17.132
Mean Value 34947 16.953 /
RSD（%） 2.11 1.58 /
（3）conclusion
The test results show that the system meets the requirements of system
applicability.
3.2.S.4.3.1.3.2 Specificity
（1）Blank and selective investigation
（a）Preparation of sample solution：
Blank solvent:
0.2% (v/v) hydrochloric acid solution -acetonitrile (90:10).
The preparation of the impurity positioning solution:
Impurities A, impurity B, impurity C, impurity D, impurity E, impurity F and
impurities G 10.0mg, respectively, are placed in the 100ml bottle, and a proper
amount of diluent is added to dissolve, dilute the diluent to the scale, shake well.
(0.1mg/ml)
System applicability test solution:
The vildagliptin control product is about 50mg, in the 20ml bottle, the above
impurities are accurately measured to locate the solution each 0.5ml, dilute the diluent
to the scale, shake well, that is. (impurity 2.5 mu g/ml, 0.1%)
Preparation of the test product solution:
Take this product 25mg, precision weighing, put 10ml measuring bottle, add diluent to
shake and dissolve, dilute dilute to scale, shake well. (2.5mg/ml)
（b）operation：
The above blank solvent, the location solution of each impurity, the system
applicability test solution and the solution of the sample were 20 l each, and the liquid
chromatograph was injected into the liquid chromatograph and the chromatogram was
recorded.
（c）Require：
The blank solvent does not interfere with the determination of each component.
122
The separation degree between impurity G and the principal component peak should
be greater than 0.5, and the separation degree between other impurities and the main
peak and impurity should be greater than 1.5.
（d）test result：
The results of a single impurity solution are shown in table3.2.S.4.3.1-7.
Table 3.2.S.4.3.1-5: the result of the impurity location solution test
Peak Attribution Retention Time（min）
ImpurityA 7.464
ImpurityB 21.423
ImpurityC 10.549
ImpurityD 8.538
ImpurityE 44.198
ImpurityF 47.540
ImpurityG 13.829
Vildagliptin 14.813
The test results of the sample solution are shown in table 3.2.S.4.3.1-8.
Table 3.2.S.4.3.1-5: test results for test products
Peak Impurity Total Impurity

Peak# Rt(min) RRt
Attribution （%）（%）
1 5.334 unknow 0.36 0.032
2 8.561 impurityD 0.58 0.078
0.158%
3 14.813 vildagliptin 1.00 /
4 48.051 unknow 3.24 0.048
The test results of system suitability solutions are shown in chart 3.2.S.4.3.1-9, and the
chromatogram is shown in table 3.2.S.4.3.1-60.
peak# Rt(min) Peak Attribution RRt Resolution

1 7.465 ImpurityA 0.50 5.800
2 8.628 ImpurityD 0.58 2.061
3 10.524 ImpurityC 0.71 3.723
123
4 13.461 ImpurityG 0.91 2.582
5 14.791 Vildagliptin 1.00 0.604
6 21.422 ImpurityB 1.45 4.215
7 44.354 ImpurityF 3.00 44.942
8 47.617 ImpurityE 3.22 10.793
（e）conclusion：
In the chromatogram of blank solvent, there is no obvious interference peak at the
peak of principal component and impurity A, impurity B, impurity C, impurity D,
impurity E, impurity F and impurity G. In the sample solution, the impurity G and the
main peak can be separated from the baseline (the separation degree is greater than
0.5), and the separation degree between the other impurities and the main peaks and
impurities conforms to the requirements.
（2）Forced degradation
（a）Preparation of sample solution：
Blank solvent：
0.2% (v/v) hydrochloric acid solution- acetonitrile (90:10).
The preparation of the control solution:
Vildagliptin control product 25mg, precision set, 100ml bottle, add diluent
moderate shake to dissolve, dilute the diluent to the scale, shake well; precision take
1ml, set 100ml bottle, dilute the diluent to the scale, shake well, that is. (2.5 mu g/ml)
The preparation of vildagliptin storage liquid:
Take vildagliptin 500mg for precise determination, put 20ml in the measuring
bottle, add diluent to shake and dissolve, dilute dilute to scale, shake well. (25mg/ml)
Preparation of acid and alkali blank solution:
1mol/L NaOH solution 0.9ml and 1mol/L HCl solution 1ml were taken
respectively, and placed in the same 10ml volumetric flask, diluted with diluent to
scale, shake well. (remarks: 1mol/L NaOH solution is added to 0.9ml because it
maintains an acidic environment according to the import registration standard).
Preparation of oxidation blank solution：
Take 3% H2O2 solution 1ml, put 10ml in the measuring bottle, dilute dilute to
scale, shake well.
The preparation of compulsory degradation experimental solution is shown in
124
table 3.2.S.4.3.1-10.
Table 3.2.S.4.3.1-5: preparation of forced degradation test solution
Solution name Method of sample processing and configuration
The mother liquor 1ml is accurately measured and placed in the 10ml
Undegraded volumetric flask. Diluted with diluent to scale, shake well.
The precision measurement of mother liquid 1ml, set 10ml bottle, add 1mol/L
hydrochloric acid solution 1ml, 80 C water bath for 2 hours, remove cold to
Acid degradation
room temperature, add 1mol/L sodium hydroxide solution 0.9ml to neutralize,
dilute the diluent to the scale, shake well, that is to be obtained.
The precision amount of the mother liquid 1ml, 10ml bottle, add 1mol/L
Alkali sodium hydroxide solution 0.9ml, immediately add 1mol/L hydrochloric acid
degradation solution 1.0ml to neutralize, dilute the diluent to the scale, shake well, that is,
obtained.
The mother liquor 1ml was accurately measured. In the 10ml measuring bottle,
Oxidative
3% H2O2 solution 1ml was added, and placed in the room temperature for
degradation
about 3 minutes. Diluted with diluent to scale, shake it up.
Take this product 25mg, precision and set, set in 10ml bottle, at 100 centigrade
Solid heat oven to place 4 hours, remove cold to room temperature, add the diluent to
shake to dissolve, dilute the diluent to the scale, shake well, that is.
Take the mother liquor 1ml accurately and put it in a 10ml measuring bottle.
Liquid high
Place it in a water bath at 80 degrees for 2 hours. Remove it and cool it to room
temperature
temperature, dilute it to scale, shake it well.
Take this product about 0.5g, put in the colorless weighing bottle, spread in the
bottom of the bottle, irradiate under the ultraviolet lamp for 24 hours, after
Solid light taking out, the precision is called 25mg to set the 10ml bottle, dilute the diluent
to the scale, shake well.
The precision of the mother liquid 5ml, set in the colorless weighing bottle,
placed under the ultraviolet lamp under 2 hours, after taking out and put cold to
Liquid light
room temperature, completely transferred to the 50ml bottle, diluent to the
scale, shake well, that is.
（b）operation：
125
PDA detector was used to determine the sample solution and the purity of vildagliptin
（c）Result：
The results of the forced degradation test are shown in table 3.2.S.4.3.1-11.
126
Table 3.2.S.4.3.1-5: results of the forced degradation test
Impurity Total Impurity
Degradation Peak Degradation of Peak Single point Degree of
Rt（min） RRt quantity impurity increase
conditions attribution main peak（%） purity threshold separation
（%）（%）（%）
5.525 0.35 unknown 0.028 --
9.114 0.57 impurityD 0.074 1.0000 7.115
Undegraded 0.141 / / 0.999866
15.876 1.00 vildagliptin / 00 3.922
47.924 3.02 unknown 0.039 21.482
4.875 0.30 unknown 0.020 --
5.597 0.34 unknown 0.027 2.921
9.275 0.56 impurityD 6.941 7.221
Alkali 1.00000
11.642 0.71 impurityC 0.051 7.052 6.911 6.536 0.999850 4.049
degradation 0
16.424 1.00 vildagliptin / 3.034
47.552 2.90 impurityE 0.003 20.500
48.181 2.93 unknown 0.010 2.205
5.573 0.34 unknown 0.005 --
6.140 0.38 unknown 0.004 2.366
Acid 9.248 0.57 impurityD 0.526 1.00000 6.079
0.700 0.559 0.436 0.999860
degradation 11.444 0.71 impurityC 0.058 0 3.660
14.408 0.89 impurityG 0.083 2.194
16.158 1.00 vildagliptin / 0.690
127
48.043 2.97 unknown 0.023 20.696
4.874 0.30 unknown 0.033 --

5.608 0.34 unknown 0.020 2.558
High 6.250 0.38 unknown 0.004 2.224
temperature 9.406 0.57 impurityD 0.471 1.00000 5.650
1.285 1.145 1.040 0.999853
degradation 11.696 0.71 impurityC 0.604 0 3.724
of liquid 16.497 1.00 vildagliptin / 2.978
47.551 2.88 impurityE 0.133 20.490
48.213 2.92 unknown 0.019 2.678
5.618 0.35 unknown 0.078 --
7.498 0.47 unknown 0.004 6.626
9.119 0.57 impurityD 1.325 3.089
Oxidative 1.00000
11.264 0.71 impurityC 0.565 3.415 3.274 3.054 0.999861 3.757
degradation 0
15.943 1.00 vildagliptin / 3.061
39.819 2.50 unknown 0.496 14.568
47.960 3.01 unknown 0.948 18.397
5.512 0.35 unknown 0.025 --
Degradation
9.007 0.57 impurityD 0.082 1.00000 7.014
of liquid 0.164 0.023 -1.144 0.999869
11.140 0.71 impurityC 0.013 0 3.897
light
15.707 1.00 vildagliptin / 2.993
128
47.275 3.01 unknown 0.005 20.786
47.940 3.05 unknown 0.038 2.196
5.665 0.34 unknown 0.023 --
9.452 0.57 impurityD 0.059 7.254
Degradation
11.797 0.71 impurityC 0.006 1.00000 3.717
of solid 0.145 0.004 -0.056 0.999850
16.660 1.00 vildagliptin / 0 2.909
light
46.236 2.78 unknown 0.023 17.364
48.031 2.88 unknown 0.033 4.395
4.767 0.30 unknown 0.053 --
5.499 0.35 unknown 0.033 2.753
8.983 0.57 impurityD 0.667 6.853
High 9.901 0.63 unknown 0.009 1.602
temperature 11.096 0.71 impurityC 0.246 1.00000 3.014
1.171 1.031 0.954 0.999870
degradation 13.403 0.85 impurityG 0.016 0 5.621
of solid 15.681 1.00 vildagliptin / 1.460
43.901 2.80 unknown 0.027 18.731
47.252 3.01 impurityE 0.095 12.342
47.949 3.06 unknown 0.026 2.930
129
（d）conclusion
The PDA spectrum of the degraded samples showed that the separation degree of the
impurity G and the peak of vildagliptin was more than 0.5, and the separation degree
of the other impurities and the peaks of Vildagliptin was greater than 1.5. The amount
of impurities and the degradation of the main peak reached the material balance, and
the purity of the peak peak was in line with the requirements.
3.2.S.4.3.1.3.3 Sensitivity (quantitative limit and detection limit)
（1）operation
According to the test results of the concentration of the sample in the process of
testing, 0.025% of the concentration of the sample was set as the quantitative limit
concentration. According to this, the sample solution containing Vildagliptin and
0.025% of the impurities were used as the quantitative limiting solution, and 3.3 times
the dilution of the quantitative limit solution was used as the detection limit solution.
LOQ solution was repeatedly injected into 6 needles, and LOD solution was
repeatedly injected into 3 injections.
（2）Results
The test results are shown in table 3.2.S.4.3.1-12~14.
130
Table 3.2.S.4.3.1-5: quantitative limit test results -1

average
Component 1 2 3 4 5 6 RSD（%）
value
peak area 19009 18886 18882 19145 19052 19064 19006 0.55
The signal to noise ratio of
ImpurityA
the first needle in the 360.4 / /
quantitative limit solution
peak area 32042 31841 31801 31981 32198 32038 31984 0.46
ImpurityB
peak area 10897 10926 10869 10909 10743 10823 10861 0.63
ImpurityC
peak area 9230 8246 8130 8759 9152 9014 8755 5.36
ImpurityD
131
peak area 14568 14590 14538 14570 14492 14532 14548 0.24
ImpurityE
peak area 15445 13825 15574 13682 13370 14408 14384 6.5
ImpurityF
peak area 9760 9317 9343 9743 9415 9983 9594 2.85
ImpurityG
peak area 8744 8913 8588 8760 8365 8506 8646 2.29
Vildagliptin
Table 3.2.S.4.3.1-5: quantitative limit test results -2

Component Concentration（μg/ml） The percentage equivalent to the concentration of the
132
sample
（%）
ImpurityA 0.632 0.025
ImpurityB 0.675 0.027
ImpurityC 0.669 0.027
ImpurityD 0.627 0.025
ImpurityE 0.594 0.024
ImpurityF 0.646 0.026
ImpurityG 0.659 0.026
Vildagliptin 0.589 0.024
Table 3.2.S.4.3.1-5: test limit test results

The percentage
Concentration equivalent to the
Component 1 2 3
（μg/ml） concentration of the
sample（%）
Signal-to-noise ratio 82.46 80.36 102.77
impurityA 0.190 0.008
Peak area 5407 5420 5354
impurityB 0.202 0.008
Peak area 8882 8970 8911
133

impurityC 0.201 0.008
Peak area 2847 2834 2865
impurityD 0.188 0.008
Peak area 3580 3396 3374
impurityE 0.178 0.007
Peak area 3872 3848 3946
impurityF 0.194 0.008
Peak area 3585 4145 4288
impurityG 0.198 0.008
Peak area 2208 2267 2294
vildagliptin 0.177 0.007
Peak area 2440 2504 2297
（3）conclusion：
The maximum daily dose of Vildagliptin Tablets is 100mg. According to ICH, the report limit of the raw materials of vildagliptin is 0.05%.
Impurities A, impurity B, impurity C, impurity D, impurity E, impurities F, impurity G, and vildagliptin quantitative limit solution concentration
are lower than the reported limit concentration, which is about 50% of the limited concentration of the report, which meets the sensitivity
requirements.
134
3.2.S.4.3.1.3.4 Solution stability
After being placed at normal temperature, the sample solution was measured in
accordance with the determination method of related substances, and the number of
impurities and the RSD (%) of the amount of impurities were investigated.
The test results are shown in table 3.2.S.4.3.1-15.
Table 3.2.S.4.3.1-5: test results for solution stability
Avera
RSD
Point of time 0h 8h 16h 25h ge
（%）
Value
Number of impurities 3 3 3 3 -- --
ImpurityA（%） ND ND ND ND ND --
ImpurityB（%） ND ND ND ND ND --
ImpurityC（%） ND ND ND ND ND --
ImpurityD（%） 0.075 0.075 0.074 0.077 0.075 1.71
ImpurityE（%） ND ND ND ND ND --
ImpurityF（%） ND ND ND ND ND --
ImpurityG（%） ND ND ND ND ND --
Unknow impurity1（%） 0.031 0.029 0.033 0.033 0.031 5.93
Unknow impurity2（%） 0.047 0.042 0.041 0.042 0.043 5.68
Total impurity（%） 0.153 0.146 0.148 0.152 0.150 2.17
note：ND is not detected
conclusion：The test solution is stable at room temperature for 25 hours.
3.2.S.4.3.1.3.5 linear
（1）Linear range
The limits of all known and unknown impurities are 0.1%, the concentration is equal
to 2.5 g/ml, and the range of linear investigation is 0.025% (quantitative limit) to
0.2%, which is equivalent to the range from 0.625 Mu to 5 g/ml.
（2）Solution preparation：
Preparation of impurity storage liquid:
Take impurities A, impurity B, impurity C, impurity D, impurity E, impurity F
and impurity G control products each 10mg, and set up the 100ml bottle, respectively,
135
add the diluent to the scale, shake well with the diluent to the scale, and use the
diluent as the storage liquid (100 mu g/ml); the precision quantity of the storage liquid
is 25ml, and the same 500ml measuring bottle is placed in the same 500ml measuring
bottle. Dilute the diluent to the scale and shake it as a storage solution. (5 mu g/ml)
The preparation of vildagliptin storage liquid:
Vildagliptin reference substance 10mg, precisely weigh, set in 100ml bottle, add
diluent and shake to dissolve, dilute the diluent to the scale, shake well, as a storage
liquid (100 mu g/ml); precision 25ml, 100ml bottle, diluent diluent to the scale, shake
well, as a storage liquid. (25 mu g/ml)
The preparation of linear solution:
According to table 3.2.S.4.3.1-16, take the volume of the impurity storage liquid
and the vildagliptin storage liquid. In the corresponding measuring bottle, dilute the
diluent to the scale and shake well.
Table 3.2.S.4.3.1-5: collocation method for linear sample solution
vildagliptin Impurity vildaglipti Impurity The percentage
Linear Volume of a
storage storage nstorage storage relative to the
investigation measuring
liquid ① liquid① liquid② liquid② concentration of the
solution bottle（ml）
（ml）（ml）（ml）（ml） sample（%）
L1 / / 5 25 200 0.025%
L2 / / 2 10 50 0.04%
L3 / / 5 25 100 0.05%
L4 / / 5 25 50 0.1%
L5 1 1 / / 20 0.2%
（3）operation：
The sample solutions were determined according to the related substances
determination method of vildagliptin.
（4）test result：
The concentration of vildagliptin, impurity A, impurity B, impurity C, impurity
D, impurity E, impurity F and impurity G was linear regression analysis. The slope,
the axis intercept of Y and the correlation coefficient were calculated, and the results
were found to be 3.2.S.4.3.1-17~25.
136
Table 3.2.S.4.3.1-5: the result of the linear range of the impurity A

Linear range of impurity A
Linear concentra
investigation tion peak area
solution （μg/ml）
L1 0.632 19006
L2 1.012 31272
L3 1.265 38453
L4 2.529 78235
L5 5.059 157523
y=31276.9091x-766.3
Linear Equation
035
Correlation
1.0000
Coefficient（r2）
Table 3.2.S.4.3.1-5: the result of the linear range of the impurity B

Linear range of impurity B
Linear concentra
investigation tion peak area
L1 0.675 31966
L2 1.080 53043
L3 1.350 65159
L4 2.700 131048
L5 5.399 266303
y=49500.0709x-1409.
Linear Equation
0141
Correlation
0.9999
Coefficient（r2）
137
Table 3.2.S.4.3.1-5: the result of the linear range of the impurity C

Linear range of impurity C
Linear concentra
investigation tion Peak area
L1 0.669 10853
L2 1.071 18042
L3 1.339 22105
L4 2.677 44491
L5 5.355 89869
y=16827.0595x-320.0
Linear equation
525
Correlation
1.0000
Coefficient（r2）
Table 3.2.S.4.3.1-5: the result of the linear range of the impurity D

Linear range of impurity D
Linear
concentration
investigation Peak area
（μg/ml）
solution
L1 0.627 9194
L2 1.002 15499
L3 1.253 19639
L4 2.506 44719
L5 5.012 89976
Linear
y=18572.2216x-2828.1492
equation
Correlation
Coefficient 0.9996
（r2）
138
Table 3.2.S.4.3.1-5: the result of the linear range of the impurity E

Linear range of impurity E
Linear
concentration
（μg/ml）
solution
L1 0.594 14550
L2 0.951 23789
L3 1.189 32516
L4 2.378 60984
L5 4.756 124057
Linear
y=26151.5552x-436.0853
Equation
Correlation
Coefficient 0.9994
2
（r ）
Table 3.2.S.4.3.1-6: the result of the linear range of the impurity F

Linear range of impurity F
Linear
concentration
（μg/ml）
solution
L1 0.646 18906
L2 1.034 29709
L3 1.292 34734
L4 2.585 66773
L5 5.170 134991
Linear
y=25578.3979x+2144.2483
Equation
Correlation
Coefficient 0.9995
（r2）
139
Table 3.2.S.4.3.1-6: the result of the linear range of the impurity G

Linear range of impurity G
Linear
concentration Peak
investigation
（μg/ml） area
solution
L1 0.659 9501
L2 1.054 16868
L3 1.317 20671
L4 2.635 43442
L5 5.269 88316
Linear Equation y=17055.2108x-1537.0353
Correlation
0.9999
Coefficient（r2）
Table 3.2.S.4.3.1-6: vildagliptin linear range results

Vildagliptin linear range
Linear
concentration
（μg/ml）
solution
L1 0.589 8613
L2 0.942 12128
L3 1.178 15986
L4 2.355 34485
L5 4.711 71485
Linear
y=15494.5368x-1752.3013
Equation
Correlation
0.9991
Coefficient（r2）
140
Table 3.2.S.4.3.1-8: correction factor and correction factor for each impurity
Linear relation
Substance Correction factor Correction factor
slope
ImpurityA 31276.9091 0.5 2.0
ImpurityB 49500.0709 0.3 3.2
ImpurityC 16827.0595 0.9 1.1
ImpurityD 18572.2216 0.8 1.2
ImpurityE 26151.5552 0.6 1.7
ImpurityF 25578.3979 0.6 1.7
ImpurityG 17055.2108 0.9 1.1
Vildagliptin 15494.5368 1.00 1.00
（6）conclusion
The test results show that the response values of Vildagliptin, impurity A,
impurity B, impurity C, impurity D, impurity E, impurity F and impurity G are linear
in the concentration range.
The correction factors of impurity A, impurity B, impurity C, impurity D,
impurity E, impurity F and impurity G are 0.5, 0.3, 0.9, 0.8, 0.6, 0.6 and 0.9,
respectively. When the correction factor is between 0.9~1.1, the correction factor is
not used in the calculation, and the accuracy of the response factor will be verified
again under the recovery and durability examination items.
3.2.S.4.3.1.3.6 Precision
（1）Repeatability
An experimenter take the same batch of Vildagliptin to prepare 6 samples of test
solution and a reference substance, and determined the related substances.
Table 3.2.S.4.3.1-6: repeatability test results
Average
Peak Attribution 1 2 3 4 5 6 RSD（%）
Value
ImpurityA（%） ND ND ND ND ND ND / /
ImpurityB（%） ND ND ND ND ND ND / /
ImpurityC（%） ND ND ND ND ND ND / /
141
0.07
ImpurityD（%） 0.076 0.078 0.080 0.082 0.073 0.077 4.19
5
ImpurityE（%） ND ND ND ND ND ND / /
ImpurityF（%） ND ND ND ND ND ND / /
ImpurityG（%） ND ND ND ND ND ND / /
Unknown Impurity1 0.03
0.030 0.031 0.032 0.032 0.034 0.032 3.70
（%） 1
Unknown Impurity2 0.04
0.045 0.045 0.047 0.046 0.045 0.046 2.15
（%） 7
0.15
Total Impurity（%） 0.152 0.155 0.159 0.160 0.152 0.155 2.42
3
Number of impurities 3 3 3 3 3 3 3 /
Conclusion: the results of the test showed that the number of impurities in the 6
samples was the same, and the RSD of impurities was less than 10%.
（2）Intermediate precision
Another laboratory technician at different times and different instruments used
the same batch of Vildagliptin to prepare 6 samples of test solution and 1 reference
substances for determination of related substances.
Table 3.2.S.4.3.1-6: results of intermediate precision test
Average
Peak Attribution 1 2 3 4 5 6 RSD（%）
Value
ImpurityA（%） ND ND ND ND ND ND / /
ImpurityB（%） ND ND ND ND ND ND / /
ImpurityC（%） ND ND ND ND ND ND / /
ImpurityD（%） 0.066 0.065 0.064 0.066 0.068 0.066 0.066 1.92
ImpurityE（%） ND ND ND ND ND ND / /
ImpurityF（%） ND ND ND ND ND ND / /
ImpurityG（%） ND ND ND ND ND ND / /
Unknown impurity1
0.028 0.033 0.034 0.033 0.032 0.032 0.032 6.51
（%）
142
Unknown impurity2
0.036 0.039 0.037 0.037 0.042 0.041 0.039 5.93
（%）
Total impurity（%） 0.130 0.137 0.135 0.136 0.141 0.139 0.137 2.75
Number of
3 3 3 3 3 3 3 /
impurities
Table 3.2.S.4.3.1-6: comparison of repeatability and intermediate precision test

results
imp
Unkno Total Number
impuri impur urit impur Unknow n
impurity impurity impurit wn impur of
Peak attribution tyB ityC yF ityG impurity1
A（%） D（%） yE（%） impurit ity impuritie
（%）（%）（（%）（%）
y2（%）（%） s
%）
1 ND ND ND 0.075 ND ND ND 0.031 0.047 0.153 3

2 ND ND ND 0.076 ND ND ND 0.030 0.045 0.152 3
3 ND ND ND 0.078 ND ND ND 0.031 0.045 0.155 3
Operator1
4 ND ND ND 0.080 ND ND ND 0.032 0.047 0.159 3
5 ND ND ND 0.082 ND ND ND 0.032 0.046 0.160 3
6 ND ND ND 0.073 ND ND ND 0.034 0.045 0.152 3
1 ND ND ND 0.066 ND ND ND 0.028 0.036 0.130 3
2 ND ND ND 0.065 ND ND ND 0.033 0.039 0.137 3
3 ND ND ND 0.064 ND ND ND 0.034 0.037 0.135 3
Operator2
4 ND ND ND 0.066 ND ND ND 0.033 0.037 0.136 3
5 ND ND ND 0.068 ND ND ND 0.032 0.042 0.141 3
6 ND ND ND 0.066 ND ND ND 0.032 0.041 0.139 3
average value / / / 0.072 / / / 0.032 0.042 0.146 3
RSD（%） / / / 9.01 / / / 5.09 9.80 7.04 /
Conclusion: the results of the test showed that the number of impurities in the 12
samples was the same, and the RSD of impurities was less than 10%, which
conformed with the verification requirements.
3.2.S.4.3.1.3.7Accuracy (Recovery)
143
The accuracy is tested by the recovery rate of impurity addition method. The
recoveries of the impurities at 3 concentrations were 0.025% (limit of quantification),
0.1% and 0.16%, respectively, with three parallel operations on each concentration.
Solution preparation:
The preparation of the reference solution:
Take Vildagliptin reference substance 25mg, precisely weigh, put in 100ml bottle,
add diluent moderate shake to dissolve, dilute the diluent to the scale, shake well;
precision take 1ml, put in 100ml bottle, dilute the diluent to the scale, shake, get it.
(2.5 mu g/ml)
The preparation of various impurities storage liquid:
Take impurities A, impurity B, impurity C, impurity D, impurity E, impurity F,
and impurity G reference substance each 10mg, precisely weigh, separately put in
100ml bottles, add diluent to dissolved, use diluent diluted to the scale, shake up. (100
mu g/ml)
Preparation of recovery storage liquid:
Take the 25ml from the above storage solution and place it in the same 500ml
volumetric flask. Dilute the diluent to the scale and shake it well. (5 mu g/ml)
The preparation of the recovery solution:
According to table 3.2.S.4.3.1-29, take appropriate amount of this product, put in
the corresponding measuring bottle, add the amount of recovery storage liquid, dilute
the diluent to the scale, shake well, and make up 3 solutions in parallel.
Table 3.2.S.4.3.1-6: preparation of recovery solution
Volume of
Weigh recovery Impurity
The percentage Theoretical
this storage storage Volume of
relative to the Number concentration of
product liquid liquid added bottle
concentration of of copies recovery sample
quantity added to to volume （ml）
the sample solution（μg/ml）
（mg） volume （ml）
（ml）
0.025% 500 3 25 / 200 0.625
0.1% 125 3 25 / 50 2.5
0.16% 125 3 / 2 50 4.0
144
operation：
The recovery rate of impurities was determined by the method of determination of
related substances in Vildagliptin (principal component external standard method plus
correction factor) and impurity external standard method.
Results:
Table 3.2.S.4.3.1-6: recovery rate determination result impurity A (principal
component external standard plus correction factor)
Theoretic Amount
Backgroun Rate of Average
Concentration Peak al of Measurem RSD
d quantity recovery recovery
level area quantity addition ent（μg）（%）
（μg）（%） rate（%）
（μg）（μg）
0.025%-1 18997 -- 0.632 0.643 101.61
0.025%-2 18762 0.632 -- 0.632 0.635 100.36
0.025%-3 19056 -- 0.632 0.645 101.93
0.1%-1 75086 -- 2.529 2.540 100.41
0.1%-2 75200 2.529 -- 2.529 2.544 100.56 98.50 3.19
0.1%-3 73085 -- 2.529 2.472 97.73
0.16%-1 112554 -- 4.047 3.807 94.07
0.16%-2 114137 4.047 -- 4.047 3.861 95.39
0.16%-3 113006 -- 4.047 3.822 94.45
Table 3.2.S.4.3.1-6: recovery rate determination result impurity B (principal

Backgrou Amount
Theoretic Rate of Average
Concentration Peak nd of Measureme RSD
al quantity recovery recovery
level area quantity addition nt（μg）（%）
（μg）（μg）
0.025%-1 33046 -- 0.675 0.706 104.65
0.025%-2 32902 0.675 -- 0.675 0.703 104.19
0.025%-3 32680 -- 0.675 0.698 103.49 101.39 2.95
0.1%-1 129867 -- 2.700 2.776 102.81
2.700
0.1%-2 130147 -- 2.700 2.781 103.03
145
0.1%-3 128114 -- 2.700 2.738 101.42
0.16%-1 195717 -- 4.319 4.183 96.84
0.16%-2 199063 4.319 -- 4.319 4.254 98.50
0.16%-3 197132 -- 4.319 4.213 97.54
Determination of recovery rate impurity C (principal component external standard
plus correction factor)
Theoret Backgro Amount
Measu Rate of Average
Concentration Peak ical und of
rement recove recovery RSD（%）
level area quantity quantity addition
（μg） ry（%） rate（%）
（μg）（μg）（μg）
0.025%-1 11954 -- 0.669 0.752 112.29
0.025%-2 11740 0.669 -- 0.669 0.738 110.27
0.025%-3 11959 -- 0.669 0.752 112.33
0.1%-1 45704 -- 2.677 2.873 107.33
0.1%-2 45542 2.677 -- 2.677 2.863 106.95 106.39 4.34
0.1%-3 44629 -- 2.677 2.806 104.80
0.16%-1 68598 -- 4.284 4.313 100.68
0.16%-2 69349 4.284 -- 4.284 4.360 101.78
0.16%-3 68898 -- 4.284 4.332 101.12
Table 3.2.S.4.3.1-6: recovery rate determination result impurity D (principal

Averag
Theoretic Backgrou Amou
Measu Rate of e
Concentration Peak al nd nt of
rement recovery recover RSD（%）
level area quantity quantity additio
（μg）（%） y rate
（μg）（μg） n（μg）
（%）
0.025%-1 44746 1.8650 2.492 2.549 102.30
0.025%-2 43178 0.627 1.8663 2.493 2.460 98.66
0.025%-3 43601 1.9065 2.533 2.484 98.05
0.1%-1 76145 1.9262 4.432 4.337 97.85 99.12 1.95
0.1%-2 76117 2.506 1.9121 4.418 4.336 98.13
0.1%-3 73620 1.8658 4.372 4.194 95.92
0.16%-1 10338 4.010 1.8733 5.883 5.889 100.09
146
0
10534
0.16%-2 1.9292 5.939 6.000 101.03
1
10395
0.16%-3 1.9088 5.919 5.921 100.04
0
Determination of recovery rate impurity E (principal component external standard

plus correction factor)
Backgr Amount
Theoretica Measu Rate of Average
Concentration Peak ound of RSD
l quantity rement recovery recovery
level area quantity addition （%）
（μg）（μg）（%） rate（%）
（μg）（μg）
0.025%-1 14427 -- 0.594 0.584 98.17
0.025%-2 14314 0.594 -- 0.594 0.579 97.40
0.025%-3 14093 -- 0.594 0.570 95.90
0.1%-1 58906 -- 2.378 2.383 100.21
0.1%-2 58810 2.378 -- 2.378 2.379 100.05 97.49 1.75
0.1%-3 56788 -- 2.378 2.297 96.61
0.16%-1 89637 -- 3.805 3.626 95.31
0.16%-2 90895 3.805 -- 3.805 3.677 96.64
0.16%-3 91315 -- 3.805 3.694 97.09
Table 3.2.S.4.3.1-6: recovery rate determination result impurity F (principal

Theoretic Backgr Amount
Measur Rate of Average
Concentration Peak al ound of
ement recovery recovery RSD（%）
level area quantity quantit addition
（μg） y（μg）（μg）
0.025%-1 14254 -- 0.646 0.590 91.23
0.025%-2 15262 0.646 -- 0.646 0.631 97.68
0.025%-3 14180 -- 0.646 0.586 90.75 93.28 4.68
0.1%-1 60741 -- 2.585 2.512 97.19
2.585
0.1%-2 61633 -- 2.585 2.549 98.61
147
0.1%-3 61010 -- 2.585 2.523 97.62
0.16%-1 89239 -- 4.136 3.691 89.24
0.16%-2 88835 4.136 -- 4.136 3.674 88.84
0.16%-3 88377 -- 4.136 3.655 88.38
148
Table 3.2.S.4.3.1-5: recovery rate determination result impurity G (principal

Averag
Theore Backgr Amou
Measu Rate of e
Concentration Peak tical ound nt of
rement recove recove RSD（%）
level area quantit quantit additio
（μg） ry（%） ry rate
y（μg） y（μg） n（μg）
（%）
0.025%-1 9716 -- 0.659 0.603 91.50
0.025%-2 9781 0.659 -- 0.659 0.607 92.11
0.025%-3 9583 -- 0.659 0.594 90.24
0.1%-1 41354 -- 2.635 2.565 97.36
0.1%-2 41953 2.635 -- 2.635 2.602 98.77 94.13 3.60
0.1%-3 42177 -- 2.635 2.616 99.30
0.16%-1 62979 -- 4.216 3.907 92.67
0.16%-2 63039 4.216 -- 4.216 3.910 92.76
0.16%-3 62820 -- 4.216 3.897 92.44
Table 3.2.S.4.3.1-5: recovery rate determination result impurity A (impurity external

standard method)
Theoret Backg Amou Rate
Measu Average
Concentration Peak ical round nt of of
rement recovery RSD（%）
level area quantit quantit additio recove
（μg） rate（%）
y（μg） y（μg） n（μg） ry（%）
0.025%-1 18997 -- 0.632 0.614 97.13
0.025%-2 18762 0.632 -- 0.632 0.607 95.93
0.025%-3 19056 -- 0.632 0.616 97.43
0.1%-1 75086 -- 2.529 2.428 95.97
0.1%-2 75200 2.529 -- 2.529 2.431 96.12 94.15 3.19
0.1%-3 73085 -- 2.529 2.363 93.42
0.16%-1 112554 -- 4.047 3.639 89.92
0.16%-2 114137 4.047 -- 4.047 3.690 91.18
0.16%-3 113006 -- 4.047 3.654 90.28
149
Table 3.2.S.4.3.1-5: recovery rate determination result impurity B (impurity

external standard method)
Theore Backg Amou
Concentration tical round nt of
peak area rement recovery recovery RSD（%）
level quantit quantit additio
0.025%-1 33046 -- 0.675 0.681 100.87
0.025%-2 32902 0.675 -- 0.675 0.678 100.43
0.025%-3 32680 -- 0.675 0.673 99.75
0.1%-1 129867 -- 2.700 2.675 99.10
0.1%-2 130147 2.700 -- 2.700 2.681 99.31 97.72 2.95
0.1%-3 128114 -- 2.700 2.639 97.76
0.16%-1 195717 -- 4.319 4.032 93.34
0.16%-2 199063 4.319 -- 4.319 4.101 94.94
0.16%-3 197132 -- 4.319 4.061 94.02
Table 3.2.S.4.3.1-5: recovery rate determination result impurity C (impurity external

standard method)
Theore Backgr Amou
Concentration Peak tical ound nt of
rement recovery recovery RSD（%）
level area quantit quantit additio
0.025%-1 11954 -- 0.669 0.719 107.47
0.025%-2 11740 0.669 -- 0.669 0.706 105.55
0.025%-3 11959 -- 0.669 0.720 107.52
0.1%-1 45704 -- 2.677 2.750 102.73
0.1%-2 45542 2.677 -- 2.677 2.741 102.36 101.83 4.34
0.1%-3 44629 -- 2.677 2.686 100.31
0.16%-1 68598 -- 4.284 4.128 96.36
0.16%-2 69349 4.284 -- 4.284 4.173 97.42
0.16%-3 68898 -- 4.284 4.146 96.79
150
Table 3.2.S.4.3.1-5: recovery rate determination result impurity D (impurity external

standard method)
Theore Backg Amou
Concentration Peak tical round nt of RSD
rement recovery recovery
level area quantit quantit additio （%）
0.025%-1 44746 1.8445 2.471 2.508 101.49
0.025%-2 43178 0.627 1.8457 2.472 2.420 97.88
0.025%-3 43601 1.8855 2.512 2.444 97.27
0.1%-1 76145 1.9050 4.411 4.267 96.74
0.1%-2 76117 2.506 1.8910 4.397 4.266 97.01 98.06 1.97
0.1%-3 73620 1.8452 4.351 4.126 94.82
0.16%-1 103380 1.8526 5.863 5.794 98.83
0.16%-2 105341 4.010 1.9079 5.918 5.904 99.76
0.16%-3 103950 1.8877 5.898 5.826 98.78
Table 3.2.S.4.3.1-5: recovery rate determination result impurity E (impurity

external standard method)
Averag
Theore Backgr Amou
Measu Rate of e
Concentra Peak tical ound nt of RSD
rement recove recove
tion level area quantit quantit additio （%）
（μg） ry（%） ry rate
（%）
0.025%-1 14427 -- 0.594 0.563 94.63
0.025%-2 14314 0.594 -- 0.594 0.558 93.89
0.025%-3 14093 -- 0.594 0.550 92.44
0.1%-1 58906 -- 2.378 2.297 96.59
0.1%-2 58810 2.378 -- 2.378 2.293 96.44 93.97 1.75
0.1%-3 56788 -- 2.378 2.214 93.12
0.16%-1 89637 -- 3.805 3.495 91.87
0.16%-2 90895 3.805 -- 3.805 3.544 93.15
0.16%-3 91315 -- 3.805 3.561 93.58
151
Table 3.2.S.4.3.1-5: recovery rate determination result impurity F (impurity external

standard method)
Theoret Backgr Amou
Concentratio Peak ical ound nt of
n level area quantit quantit additio
0.025%-1 14254 -- 0.646 0.552 85.39
0.025%-2 15262 0.646 -- 0.646 0.591 91.43
0.025%-3 14180 -- 0.646 0.549 84.94
0.1%-1 60741 -- 2.585 2.351 90.97
0.1%-2 61633 2.585 -- 2.585 2.386 92.30 87.31 4.68
0.1%-3 61010 -- 2.585 2.362 91.37
0.16%-1 89239 -- 4.136 3.455 83.53
0.16%-2 88835 4.136 -- 4.136 3.439 83.15
0.16%-3 88377 -- 4.136 3.421 82.72
Table 3.2.S.4.3.1-5: recovery rate determination result impurity G (impurity external

standard method)
Theore Backgr Amou
Concentratio Peak tical ound nt of
n level area quantit quantit additio
0.025%-1 9716 -- 0.659 0.589 89.46
0.025%-2 9781 0.659 -- 0.659 0.593 90.06
0.025%-3 9583 -- 0.659 0.581 88.24
0.1%-1 41354 -- 2.635 2.508 95.19
0.1%-2 41953 2.635 -- 2.635 2.544 96.57 92.03 3.60
0.1%-3 42177 -- 2.635 2.558 97.09
0.16%-1 62979 -- 4.216 3.820 90.61
0.16%-2 63039 4.216 -- 4.216 3.823 90.69
0.16%-3 62820 -- 4.216 3.810 90.38
Conclusion: the method of determination of related substances in vildagliptin is

accurate.
152
3.2.S.4.3.1.3.8 Durability
Durability is the condition of fine-tuning chromatographic conditions, and the
tolerance degree of sample determination is not affected.
The chromatographic conditions for the fine-tuning are shown in table

3.2.S.4.3.1-44.
Table 3.2.S.4.3.1-5: the chromatographic conditions of the fine-tuning
Project Fine-tuning conditions

Standard condition Based on the test method
pH（±0.1） The fine-tuning is 2.7 and 2.9, others are acorrding to standard conditions
Flow rate The fine-tuning is 0.9ml/min and 1.1ml/min, others are acorrding to standard
conditions
Buffer salt concentration Take sodium dihydrogen phosphate 3.0g and 3.2g ， others are acorrding to
standard conditions
Ratio of mobile phase A 97:3/95:5
Column temperature 30 C and 40 C
wavelength 208nm and 212nm
Chromatographic column of
Replacing the same brand column with the same type
the same factory
Chromatographic columns
Change the same type of chromatographic column with different brands
of different manufacturers
Blank solvent:
0.2% (v/v) hydrochloric acid solution- acetonitrile (90:10)
The preparation of the reference solution:
Take Vildagliptin reference substance 25mg, precisely weighed, set in 100ml
bottle, use diluent to dissolve and dilute to the scale, shake well; precisely measured
1ml, set in 100ml bottle, dilute the diluent to the scale, shake well, got it. (2.5 g/ml)
Take the raw material of vildagliptin (batch number: 140601) 25mg, weigh it
accurately, put it in 10ml volumetric flask, dilute it with diluent and dilute it to scale,
153
shake well. (2.5mg/ml)
Preparation of sample solution:
Take the raw material of Vildagliptin (batch number: 140601) 125mg, precisely
weigh, set in 50ml bottle, add the diluent appropriately and shake to dissolve, add
linear term impurity storage liquid ② 25ml, dilute the diluent to the scale, shake well,
got it. (0.1%)
Operation：
Under the above chromatographic conditions, precisely weigh blank solvent,
reference solution and adding sample solution each 20μl, separately inject into the
liquid chromatograph and record the chromatograms.
Results：
Blank solvents do not interfere with the determination of related substances in
each condition. The recovery rate of each impurity is calculated by using the principal
component external standard method with correction factor.
Table 3.2.S.4.3.1-5: durability test results - Test Results for test solution
Peak Attribution
Impur Impur Impur Impur Unknow Unknow
condition Impurit Impurity Impurit
ity A ityB ityC ityD impurity impurity
yE（%） F（%） yG（%）
（%）（%）（%）（%） 1（%） 2（%）
Standard condition ND ND ND 0.075 ND ND ND 0.032 0.044
pH2.7 ND ND ND 0.068 ND ND ND 0.025 0.030
pH2.9 ND ND ND 0.067 ND ND ND 0.031 0.037
Flow speed 0.9 ND ND ND 0.076 ND ND ND 0.032 0.033
Flow speed 1.1 ND ND ND 0.076 ND ND ND 0.030 0.035
Sodium dihydrogen
phosphate quantity ND ND ND 0.066 ND ND ND 0.026 0.035
3.0g
Sodium dihydrogen
phosphate quantity ND ND ND 0.060 ND ND ND 0.026 0.035
3.2g
The ratio of Mobile ND ND ND 0.087 ND ND ND 0.026 0.041
154
phase A is97-3
The ratio of Mobile
ND ND ND 0.069 ND ND ND 0.025 0.035
phase A is 95-5
Column
ND ND ND 0.054 ND ND ND 0.031 0.041
temperature30℃
column
ND ND ND 0.069 ND ND ND 0.030 0.043
temperature40℃
Wavelength208nm ND ND ND 0.064 ND ND ND 0.040 0.018
Wavelength212nm ND ND ND 0.070 ND ND ND 0.059 0.015
Different
Chromatographic ND ND ND 0.064 ND ND ND 0.028 0.038
columns1
Different
Chromatographic ND ND ND 0.086 ND ND ND 0.022 0.037
columns2
Table 3.2.S.4.3.1-5: durability test results - sample test results 1

Degree of Rate of recovery
Condition Peak attribution
separation （%）
Standard condition
vildagliptin 0.595 /
impurityE 11.636 100.21
pH2.7 impurityC 3.845 97.17
155
impurityF 38.074 109.48
pH2.9
impurityF 42.703 106.53
Flow speed 0.9
impurityF 40.931 101.30
Flow speed 1.1
impurityF 47.838 104.62
impurityE 10.873 120.72
Sodium dihydrogen impurityD 2.121 95.86
phosphate impurityC 3.709 92.40
quantity3.0g impurityG 2.995 83.47
156
impurityE 10.100 110.11
Sodium dihydrogen
phosphate
quantity3.2g
The ratio of
mobile phase A is
97-3
The ratio of
mobile phase A is
95-5
impurityF 44.886 109.95
impurityE 10.580 140.37
Column
temperature 30℃
157
impurityF 34.280 104.07
Column impurityG 3.290 106.17
temperature 40℃ vildagliptin 0.559 /
impurityF 45.950 110.82
Wavelength208nm
impurityF 43.005 107.71
Wavelength212nm
impurityF 41.101 117.11
Different
chromatographic
columns 1
（phenomenex
158
Gemini C18 impurityB 4.000 94.25
4.6×250mm,5μm） impurityF 36.619 106.23
Different impurityD 1.842 118.61
chromatographic impurityC 3.667 92.58
columns 2 impurityG 1.979 94.01
（Agilent Extend vildagliptin 0.743 /
C18 impurityB 4.404 101.98
4.6×250mm,5μm） impurityF 41.602 112.69
Table 3.2.S.4.3.1-5: durability test results - sample test results 2
Degree of
Degree of
separation
separation Unknown Unknown impurity2
Condition Unknown
Unknown impurity impurity1（%）（%）
impurity2an
1 and impurity A
d impurity E
Standard
5.900 1.615 0.034 0.034
condition
pH2.7 5.365 1.636 0.037 0.020
pH2.9 5.661 2.458 0.041 0.035
Flow speed0.9 6.002 1.634 0.043 0.021
Flow speed1.1 5.670 / 0.040 ND
Sodium
Dihydrogen
5.641 / 0.041 ND
phosphate
quantity is 3.0g
Sodium
dihydrogen
5.771 3.151 0.033 0.037
phosphate
quantity is 3.2g
The ratio of
6.193 1.402 0.039 0.017
mobile phase A
159
is 97-3
The ratio of
mobile phase A 3.621 / 0.040 ND
is 95-5
Column
4.901 5.278 0.048 0.039
temperature30℃
Column
5.953 1.585 0.048 0.017
temperature40℃
Wavelength208n
5.571 3.685 0.031 0.038
m
Wavelength212n
5.429 3.503 0.040 0.036
m
Different
chromatographic 5.111 2.389 0.034 0.033
columns1
Different
chromatographic 5.445 3.339 0.031 0.039
columns2
Note: the condition of rough font is unqualified.
conclusion：
（1）When the buffer solution pH is adjusted for +0.1, the flow rate is adjusted
for -0.1ml/ minutes, the buffer salt concentration is adjusted to +5%, the column
temperature is + 5 C, the wavelength is adjusted to 2nm, the same type
chromatographic column is replaced by the same brand. After the replacement of the
same type chromatographic column, the order of the peaks of each component has not
changed, and the separation degree accords with the requirement, and the recovery
rate of each impurity is all in 80%~120%. In accordance with the requirements of
durability;
（2）When the PH value is -0.1, the separation degree between the impurity G and the
main peak is less than 0.5, which does not meet the durability requirement. Therefore,
the pH value of the mobile phase must be strictly controlled.
（ 3 ） When the flow rate is +0.1, the unknown impurity is contained in the
160
impurity E, the recovery rate of impurity E is not between 80%~120%, and does not
meet the durability requirement. Therefore, the flow rate must be strictly controlled.
（4）When the ratio is -1%, the separation degree between the impurity G and
the main peak is less than 0.5, the separation degree of the unknown impurity and the
impurity E is less than 1.5, the recovery rate of the impurity B and G is not between
the 80%~120%; when the proportion +1%, the separation degree of the impurity A
and the impurity D is less than 1.5, the unknown impurity is contained in the impurity
E, the impurity A, G, and recoveries are not between them. It is not consistent with
durability. Therefore, it is necessary to strictly control the proportion.
（ 5 ） When the buffer salt concentration is -5% and the unknown impurity is
contained in the impurity E, the buffer salt concentration must be strictly controlled.
3.2.S.4.3.1.3.9 Trial product test results
The test results of vildagliptin raw material show in table 3.2.S.4.3.1-48
Table 3.2.S.4.3.1-5: results of raw material examination of vildagliptin
A small Listed products
Sample information Verification batch
test (preparations)
Batches 140401 140601 140701 140702 S0020
impurityA（%） ND ND ND ND ND
impurityB（%） ND ND ND ND ND
impurityC（%） ND ND ND ND 0.145
impurityD（%） 0.004 0.077 0.017 0.020 0.144
impurityE（%） ND ND ND ND 0.019
impurityF（%） 0.04 ND 0.022 0.033 ND
impurityG（%） ND ND ND ND ND
Max unknown impurity
0.014 0.046 0.076 0.050 0.015
（%）
Total impurity（%） 0.058 0.155 0.114 0.103 0.322
Number of impurities 3 3 3 3 4
3.2.S.4.3.2Methodological Verification of enantiomer checking

The results of verification methods for enantiomers are summarized in table
3.2.S.4.3.2-1
161
Table 3.2.S.4.3.2-1: results of validation of enantiomers inspection methods
Project Verification result
System The separation degree of the enantiomer peak and vildagliptin peak is more than
applicabilit 0.9. The relative standard deviation of the main peak area of the 6 needle is 2.02%,
y less than 5%, which is in line with the system suitability requirements.
There was no interference peak in the vacancy solvent at the peak position of
vildagliptin and enantiomers. In the chromatogram of additive solution, the
Specificity separation degree between the enantiomer peak and the main peak is greater than
0.9.
correlation
Component Linear range linear equation coefficient
R2
linear vildagliptin 2.49μg/ml~29.84μg/ml y=3790.099x+522.621 0.999
Enantiomer 2.48μg/ml~29.71μg/ml y=3635.939x+788.350 1.000
The response value of each component has a linear relationship with the
concentration.
The enantiomers were not detected in 6 samples prepared by the same batch of
Precision
vildagliptin
component Rate of recovery（%） RSD(%)
Enantiomer 96.23 3.68
Accuracy
At the 3 level, the recoveries of enantiomers were between 80%~120%, which
(recovery)
conformed to the validation standard.
LOQ LOD
Equivalent to the Equivalent to
component concentrati percentage of concentrati the percentage
on（μg/ml） working on（μg/ml） of working
Limit of
concentration concentration
quantity
vildagliptin 2.487 0.025% 0.746 0.007%
and limit of
Enantiomer 2.476 0.025% 0.743 0.007%
detection
The maximum daily dose of Vildagliptin Tablets is 100mg. According to ICH, the
reporting limit of vildagliptin is 0.05%. The limit solution concentration of
vildagliptin and enantiomers is lower than the reported limit concentration, about
50% of the reported limit concentration.
Solution
The enantiomers of the solution were stable at room temperature for 22h.
stability
162
When the column temperature is + 5 C, the detection wavelength is adjusted for +
2nm, the flow rate is 0.1ml/ minutes, the proportion of two ethylamine is + 0.02,
durability the separation of the enantiomer peak and the vildagliptin peak is more than 0.9,
and the recovery rate of the enantiomer is between 80%~120%, which is in line
with the requirement of durability.

Table 3.2.S.4.3.2-1: instrument and test medicine
Name Type Source
HPLC Agilent 1200 Agilent, USA
Instrument Shanghai Jinghai Instrument Co.,
Electronic balance FA1004N
Ltd.
Ultraviolet visible Shanghai Precision Scientific
752N
spectrophotometer Instruments Co., Ltd.
Enantiomer of Anhui Haikang Pharmaceutical
20150418
vildagliptin Co., Ltd.
Anhui Haikang Pharmaceutical
vildagliptin 20150227
Co., Ltd.
Trial Vildagliptin Anhui Haikang Pharmaceutical
150601
medicine (verification batch) Co., Ltd.
Vildagliptin Anhui Haikang Pharmaceutical
150602
(verification batch) Co., Ltd.
Vildagliptin Anhui Haikang Pharmaceutical
150603
(verification batch) Co., Ltd.
3.2.S.4.3.2.2 Establishment of enantiomer method

（1）Selection of wave length
（a）Solution preparation
Blank solvent：
Anhydrous ethanol.
Test product solution:
Take this product about 10mg, and put it in a 200ml volumetric flask. Add the
amount of absolute alcohol, shake it and dissolve it, dilute it to scale with absolute
alcohol, shake it well. (50 mu g/ml)
Enantiomer solution：
163
Take Vildagliptin enantiomer reference substance about 10mg, put in 200ml bottle,
add moderately ethanol, vibrate as dissolved, and the alcohol was diluted with
anhydrous alcohol to the scale and shake well. (50 mu g/ml)
（b）Operation：
The above solutions were scanned at the wavelength of 400nm~200nm, record
chromatograms.
（c） Test results
Table 3.2.S.4.3.2-1: test results
Maximum absorption A value
Peak attribution
wavelength（nm）
Vildagliptin 207.00 0.663
Enantiomer 206.00 0.596
（d）conclusion
From the above results, it is known that both the product and the enantiomer have the
maximum absorption at the end wavelength, and the anhydrous ethanol as the mobile
phase has a large baseline fluctuation at the end wavelength, so 220nm is selected as
the wavelength for the detection of the enantiomer.
（2）Selection of sample concentration
（a）Solution preparation
Blank solvent:
Anhydrous ethanol.
Mixed solution:
Take this product and vildagliptin enantiomer reference substance each 20mg,
precisely weigh, set in the same 10ml bottle, add water alcohol moderately, shake to
dissolve, using anhydrous ethanol dilute to the scale, shake well. (2mg/ml)
（b）Chromatography operation：
chromatographic column：CHIRALPAK® IF（250×4.6mm，5µm）
Flow rate：0.5ml
column temperature：35℃
Sample volume：10μl
164
（c）Operation：
The above 10 L solution was accurately measured and injected into the liquid
chromatograph to record the chromatogram.
（d）Test result
Table 3.2.S.4.3.2-4: Test results
Peak attribution Retention time（min） Degree of separation S/N
Enantiomer 15.443 -- 8762
Vildagliptin 18.601 3.116 6455
（e）conclusion
According to the above results, the separation of enantiomeric peaks and main
peaks is in accordance with the regulations.
The maximum daily dose of Vildagliptin Tablets is 100mg. According to the
"guidelines for the study of chemical drug impurities", the report of the impurity
detection for the raw materials of vildagliptin is limited to 0.05%. The limit of
quantification should be about 50% of the reporting limit, as the limit of
quantification should about 0.03%.
According to the minimum response value, when the signal to noise ratio (SNR)
is 10, the corresponding concentration is 3.1 g/ml（），and the
concentration is about 0.03% of the working concentration of the test product. The
concentration of the test product is about 10mg/ml（），so the
concentration of the test product is about 10mg/ml.

Table 3.2.S.4.3.2-1: test results of additive solution (principal component 10 mg/ml,
enantiomer 0.15%)
Retention
Chromatographic Relative Degree of
time Trailing factor
peak retention time separation
（minutes）
Enantiomer 16.544 0.89 -- 1.220
Vildagliptin 18.544 1.00 1.132 3.874
The peak order of the added solution is: the peak of enantiomer, the main
component, the main peak. The peak of the isomer and the main component can be
165
separated but the separation degree is less than 1.5. It is proved that the separation of
the isomer peak and the main peak is more than 0.9, so the application of the solution
to the isomer peak can be determined. The degree of separation from the principal
component peak should not be less than 0.9.
3.2.S.4.3.2.3 Enantiomer method verification

3.2.S.4.3.2.3.1 System applicability
（1）Operation
The system application solution is taken to record the chromatogram at the
wavelength of 220nm. The separation degree between the peak of the enantiomer and
the peak of the enantiomer should be less than 0.9. (remark: the peak of the isomer,
the main component peak, the tail of the main peak, the peak of the isomer and the
main peak can be separated but the separation degree is less than 1.5, and the
separation degree is less than 1.5. When the separation degree of the enantiomer peak
and the main peak is greater than 0.9, the baseline separation can be reached.
Therefore, the separation of the enantiomer peak and the principal component peak of
the applicable solution should not be less than 0.9). The chromatogram was recorded
at the wavelength of 220nm at the wavelength of the reference substance, and the
relative standard deviation of the peak area of the continuous 6 needles should not
exceed 5%.
（2）results
166
Retention time Relative retention Degree of
Chromatographic peak
(minutes) time separation
Enantiomer 16.544 0.89 --
Results of repeated injection of 6 needles in a control solution

Retention time of main
Sampling Main peak area
peak
1 60405 20.248
2 63131 20.303
3 62519 20.299
4 63683 20.288
5 63857 20.293
6 62222 20.276
Mean Value 62636 20.285
RSD（%） 2.02 0.10
（3）conclusion
The test results show that the system meets the requirements of system applicability.
3.2.S.4.3.2.3.2Specificity
（1）Preparation of sample solution
Blank solvent:
Anhydrous ethanol.
Take this product 100mg, weigh it accurately, put it in a 10ml volumetric flask,
add an amount of absolute alcohol, shake it to dissolve it, dilute it to scale with
absolute alcohol, shake it well. (10mg/ml)
Control solution:
Vildagliptin control product 15mg, precise and fixed, set 100ml bottle, add water
alcohol proper amount, shake to dissolve, use anhydrous ethanol dilution to scale,
shake well; precision measure the solution 1ml, 10ml bottle, use anhydrous ethanol
dilution to scale, shake well. (15 mu g/ml)
Enantiomer positioning solution:
Vildagliptin was about 15mg of the enantiomer control, and in the 50ml bottle,
167
the amount of ethanol was added, the vibration was dissolved, and the alcohol was
diluted with anhydrous alcohol to the scale and shake well. (0.3mg/ml)
Sample solution:
Take this product about 200mg, set up 20ml bottle, add the amount of water ethanol,
shake to dissolve, add the enantiomer positioning solution 1ml, use anhydrous ethanol
dilution to the scale, shake well. (15 mu g/ml, 0.15%)
（2）Chromatography operation
The chromatographic peaks caused by all blank solvents were identified by blank
solvent injection.
The retention time of enantiomers was determined by injection of enantiomers.
The relative retention time of the enantiomer peak relative to the main peak was
determined by adding sample solution into the sample.
（2）test results
The results are shown in table 3.2.S.4.3.2-8~9.
Table 3.2.S.4.3.2-1: positioning solution test results
Component Retention time (minutes)
Enantiomer 16.698
Vildagliptin 18.346
Table 3.2.S.4.3.2-1: sample solution test results

Retention time Relative retention Degree of
Chromatographic peak
(minutes) time separation
Enantiomer 16.544 0.89 --
（4）conclusion
There was no interference between the blank solvents and the peak positions of the
enantiomers and the main peaks. The separation of the peaks of the enantiomers and
the main peaks was more than 0.9 in the sample solution.
3.2.S.4.3.2.3.3linear
The range of linearity was 0.025% to 0.30% (limit 0.15%), equivalent to 2.5 g/ml
to 30 g/ml.
Preparation of linear storage liquid:
vildagliptin to the isomer control products, the vildagliptin control products
168
25mg, respectively, respectively, and set the same 100ml bottle, adding a proper
amount of ethanol vibration to dissolve, with anhydrous ethanol dilution to the scale,
shake well. (250 mu g/ml)
Preparation of linear sample solution:
According to table 3.2.S.4.3.2-10, the linear reserve liquid was diluted to
different concentrations with anhydrous alcohol, mixed and labeled.
Table 3.2.S.4.3.2-1: preparation of linear sample solution
The percentage of
Volume extraction
Linear sample Volume of bottle the working
of linear storage
solution （ml） concentration of
liquid volume（ml）
enantiomers
L1 1 100 0.025%
L2 3 100 0.075%
L3 1 25 0.10%
L4 3 50 0.15%
L5 3 25 0.30%
（2）Chromatography operation
Determination of sample solution according to enantiomer determination
method.
（3）test results
The density of the peak area and the peak area of vildagliptin isomer were
analyzed by linear regression analysis. The slope, the Y axis intercept and the
correlation coefficient were calculated. The results were shown in table
3.2.S.4.3.2-11~13.
Table 3.2.S.4.3.2-1: results of vildagliptin linear investigation
The percentage of the
Concentration
Linear solution Peak area working concentration
（μg/ml） relative to the principal
component（%）
1 2.49 10316 0.025
2 7.46 29157 0.075
3 9.95 38495 0.099
4 14.92 55480 0.149
5 29.84 114193 0.298
169
Linear Equation y=3790.099x+522.621
Correlation
R2=0.999
Coefficient
Table 3.2.S.4.3.2-1：vildagliptin linear relation curve
Table 3.2.S.4.3.2-1：vildaliptin Linear investigation results of enantiomers

The percentage of
the working
Concentration
Linear solution Peak area concentration
（μg/ml） relative to the
principal component
（%）
1 2.48 9331 0.025
2 7.43 27923 0.074
3 9.90 37317 0.099
4 14.86 54765 0.149
5 29.71 108672 0.297
Linear Equation y=3635.939x+788.350
Correlation Coefficient R2=1.000
170
Table 3.2.S.4.3.2-2：vildagliptin Linear relation curve of enantiomer
Table 3.2.S.4.3.2-1: correction factor calculation results

Component K value Correction factor Relative response factor
Vildagliptin 3790.099 1.00 1.00
Enantiomer 3635.939 1.04 0.96
（4）conclusion
According to the experimental results, the response value and the concentration of
vildagliptin and enantiomers are linear in the concentration range.
3.2.S.4.3.2.3.4 Quantitative limit and detection limit
（1）operation
According to the test results of the concentration of the sample in the test process,
0.025% of the concentration of the sample was set to limit the concentration, and the
sample solution containing vildagliptin and enantiomers was 0.025% as a quantitative
limiting solution, and 3.3 times the dilution of the quantitative limit solution as the
detection limit solution. The limit limit solution was repeated injection 6 times, the
detection limit was repeated, and the injection was 3 needles.
（2）test results
Table 3.2.S.4.3.2-1: quantitative limit results -vildagliptin enantiomers
quantitative limit
Sampling Peak Peak area Area RSD Signal-to- Quantitative The percentage
area mean （%） noise ratio limit equivalent to the
171
concentration concentration of
（μg/ml） the sample（%）
1 9422
2 9483
3 9511
9331 4.62 37.71 2.476 0.025
4 8544
5 9213
6 9812
Table 3.2.S.4.3.2-1: quantitative limit results - vildagliptin
quantitative limit
The
percenta
ge
Quantitati
equivale
Signal- ve limit
Sampling Peak area Area nt to the
Peak area to-nois concentrat
mean RSD（%） concentr
e ratio ion
ation of
（μg/ml）
the
sample
（%）
1 9993
2 10556
3 10599 1031
2.46 25.20 2.487 0.025
4 10402 6
5 10300
6 10045
Table 3.2.S.4.3.2-1 limit of detection - vildagliptin enantiomer
quantitative limit
Detection limit Equivalent to
Sampling Peak area
Peak area concentration the sample size
mean
（μg/ml）（%）
1 1672 7.73 0.743 0.007
172
2 1817
3 2163
Table 3.2.S.4.3.2-1: test limit results - vildagliptin
quantitative limit
Detection limit Equivalent to
Sampling Peak area
Peak area concentration the sample size
mean
（μg/ml）（%）
1 2236
2 2267 7.96 0.746 0.007
3 3070
（3）conclusion
The maximum daily dose of Vildagliptin Tablets is 100mg. According to ICH, the
reporting limit of veglage is 0.05%. The limit solution concentration of vildagliptin
and enantiomers is lower than the reported limit concentration, about 50% of the
reporting limit.
3.2.S.4.3.2.3.5 Rate of recovery
（1） The recovery rates of vildagliptin enantiomers were investigated at 3
concentrations, 0.025% (quantitative limit), 0.15% (limited concentration),
and 0.30%, and three solutions were prepared in each concentration.
（2） preparation of sample solution
Control solution:
Vildagliptin control product 15mg, precision set, 100ml bottle, add water ethanol,
shake to dissolve, use anhydrous ethanol dilution to the scale, shake well; precision to
take 1ml, 10ml bottle, add anhydrous ethanol dilution to scale, shake well. (15 mu
g/ml)
Take this product 100mg, weigh it accurately, put it in 10ml volumetric flask,
add absolute alcohol to dissolve and dilute it to scale, shake well. (10mg/ml)
Vildagliptin enantiomer storage liquid:
Vildagliptin's enantiomeric reference substance 25mg was precisely weighed and
placed in a 100ml volumetric flask, dissolved in absolute alcohol and diluted to scale.
173
Shake well. (250 mu g/ml)
The preparation of the recovery sample solution:
According to table 3.2.S.4.3.2-18, appropriate amount of the sample was given
respectively. In the corresponding measuring bottle, a proper amount of the storage
liquid was added to the enantiomer, and the alcohol was diluted to the scale with
anhydrous ethanol. 3 solutions were prepared in parallel.
Table 3.2.S.4.3.2-18: preparation method of sample solution for recovery rate
Volume
The percentage extraction of
Recovery rate equivalent to Injection vildagliptin Volume of Theoretical
investigation the volume enantiomer bottle concentration
solution concentration （mg） storage （ml）（μg/mL）
of the sample liquid
volume（ml）
R1 0.025% 1000 1 100 2.5
R2 0.15% 500 3 50 15
R3 0.30% 250 3 25 30
（3） test results

The enantiomers were not detected in the solution of the product. The results of
the recoveries of enantiomers were shown in table 3.2.S.4.3.2-19.
Table 3.2.S.4.3.2-19: test results of enantiomer recovery
Additive
Theoretical Measured Rate of Average
concentrat RSD
Number Peak area concentration concentration recovery recovery
ion （%）
（μg/ml）（μg/ml）（%） rate（%）
（μg/ml）
1 9284 2.528 2.322 91.84
2 10013 2.528 2.528 2.504 99.05
3 9562 2.528 2.392 94.59
4 57930 15.171 14.489 95.51
5 54916 15.171 15.171 13.735 90.54 96.23 3.68
6 61450 15.171 15.369 101.31
7 117389 30.342 29.360 96.77
8 117608 30.342 30.342 29.415 96.95
9 120775 30.342 30.207 99.56
174
（4） Note: the background concentration of enantiomers was not detected.
（5） conclusion
The results showed that the recoveries of the enantiomers were 96.23% and RSD%
3.68% at 3 levels, which conformed to the validation standard.
（1）operation
6 enantiomers were prepared from the same batch of test solution and a reference
solution.
（2）test results
Nominal quantity
Sampling Peak area Enantiomer（%）
（mg）
1 99.64 39857606 ND
2 101.25 40502365 ND
3 100.56 40226077 ND
4 99.43 39771480 ND
5 101.85 40632959 ND
6 101.33 40531819 ND
（3）conclusion
The results showed that enantiomers were not detected in 6 samples from the same
batch of vildagliptin
（1）operation
In order to investigate the extent of the determination of the results, the determination
of the results is not affected. After the proper adjustment of the enantiomer
determination method, the separation degree and the recovery rate of the enantiomer
are investigated by using the control solution and the sample solution. The condition
of the investigation is shown in table 3.2.S.4.3.2-21.
Table 3.2.S.4.3.2-21: durability test conditions
Column Flow rate
Test
temperature （ml/min）
175
（℃）
initial 35℃ 0.5 Based on the test method

Test1 35℃ 0.4 Based on the test method
Test3 35℃ 0.5 Detection wavelength: 218nm
Test4 35℃ 0.5 Detection wavelength：222nm
Mobile phase: anhydrous ethanol -
Test5 35℃ 0.5
two ethylamine (100:0.08)
Mobile phase: anhydrous ethanol -
Test6 35℃ 0.5
two ethylamine (100:0.12)
（2）test result
Table 3.2.S.4.3.2-21: results of durability inspection
Recoveries of Separation of enantiomer
Condition
enantiomers（%） peak and main peak
Initial condition 95.47 1.132
Flow speed0.4 97.08 1.201
Flow speed0.6 99.28 1.121
Wavelength218nm 85.56 1.125
Wavelength222nm 87.39 1.129
Anhydrous
92.45 1.141
ethanol-diethylamine（0.08%）
Anhydrous ethanol-diethylamine
100.83 1.117
（0.12%）
Column temperature 30℃ 100.91 1.065
Column temperature40℃ 94.13 1.215
（3）conclusion
176
When the column temperature is + 5 C, the detection wavelength is adjusted for +
2nm, the flow rate is 0.1ml/ minutes, the proportion of diethylamine is + 0.02, the
separation of the enantiomer peak and the vildagliptin peak is more than 0.9, and the
recovery rate of the enantiomer is between 80%~120%, which is in line with the
requirement of durability.
（1）operation
The test solution was placed at room temperature for a period of time and determined
according to the enantiomer method.
（2）test results
Table 3.2.S.4.3.2-21: solution stability test results
Time Initial 8h 22h
Enantiomer（%） ND ND ND
（3）conclusion
Stability of the sample solution at room temperature for 22h.
3.2.S.4.3.2.4Sample test results
The results of examination of raw materials for vildagliptin 3.2.S.4.3.2-24 were
shown in table.
Table 3.2.S.4.3.2-21: results of raw material examination of vildagliptin
Sample Information Batches Enantiomer（%）
150601 ND
Verification batch 150602 ND
150603 ND
Listed products
S0020 0.12
(preparations)
3.2.S.4.3.3 Methodology validation of residual solvents and 3- amino -1-

amantadine
The results of validation of residual solvents are summarized in table
3.2.S.4.3.3-1.
Table 3.2.S.4.3.3-1: summary of methodological verification results of residual
solvents
177
The results of 6 consecutive observations of the illumination solution are as
follows：
Degree of
Component Peak area RSD（%）
separation
methanol 0.59 /
ethanol 0.29 5.837
System Isopropanol 0.46 3.838
applicability dichloromethane 1.60 2.093
Triethylamine 1.69 19.824
DMF 0.70 16.867
Blank solvent（DMSO） / 5.512
3- amino -1- amantadol 1.28 10.470

The separation degree between the peaks is greater than 1.5, and the system has
good applicability.
The blank solvent did not interfere with the determination of residual solvents, L-
Specificity
proline amide and 3- amino -1- amantadine.
LOQ LOD
Equivalent Equivalent
to the to the
Component concentratio percentage concentratio percentage
n（μg/ml） of working n（μg/ml） of working
concentratio concentratio
n（%） n（%）
Quantitative
methanol 0.090 0.090 0.027 0.027
limit and
ethanol 0.149 0.149 0.045 0.045
detection limit
Isopropanol 0.150 0.150 0.045 0.045
dichloromethan
0.018 0.018 0.005 0.005
e
Triethylamine 0.010 0.010 0.003 0.003
DMF 0.027 0.027 0.008 0.008
3- amino -1-
0.029 0.029 0.009 0.009
amantadol
correl
ation
Linear range Component Linear range linear equation
coeffi
cient
178
R2
0.090mg/ml~0.3
methanol y=724046.7123x-6020.8115 0.9942
61mg/ml
0.149mg/ml~0.5
ethanol y=1036797.7713x-27152.0943 0.9979
97mg/ml
0.150mg/ml~0.5
Isopropanol y=1142857.7527x-25907.1803 0.9981
99mg/ml
dichlorometh 0.018mg/ml~0.0
y=256327.1477x-438.3689 0.9976
ane 71mg/ml
Triethylamin 0.010mg/ml~0.0
y=1126638.0635x-988.0861 0.9918
e 38mg/ml
0.027mg/ml~0.1
DMF y=673608.8470x-6053.2336 0.9950
07mg/ml
3- amino -1- 0.029mg/ml~0.1
y=1244667.5419x+12908.3811 0.9978
amantadol 18mg/ml
There is a linear relationship between the peak area and the concentration of each
component.
3- amino
dichlorom
Compone methanol ethanol Isopropa Triethyla DMF -1-
ethane
nt （%）（%） nol（%） mine（%）（%） amantadol
（%）
（%）
1 0.004 0.003 0.105 0.005 ND ND 0.015
2 0.005 0.002 0.100 0.004 ND ND 0.015
3 0.005 0.002 0.100 0.004 ND ND 0.015
4 0.004 0.002 0.100 0.004 ND ND 0.015
5 0.005 0.002 0.101 0.004 ND ND 0.014
Repeatability
6 0.005 0.002 0.100 0.004 ND ND 0.014
average
value 0.005 0.002 0.101 0.004 / / 0.015
（%）
RSD（%） 3.18 14.43 2.02 11.14 / / 5.55
The detectable amount of other residual solvents except isopropanol was low, so
the RSD of other residual solvents was relaxed to 15%. The results showed that the
precision accords with the requirements.
Rate of Component Rate of recovery（%） RSD(%)
recovery methanol 88.78 4.25
179
ethanol 89.03 4.29
Isopropanol 97.02 0.57
dichloromethane 91.39 2.85
Triethylamine 94.53 9.40
DMF 96.10 4.22
3- amino -1-
104.81 8.83
amantadol
At 3 levels, the recoveries were between 80%~120% and RSD% were less than
10%, which conformed to the validation standard.
When the temperature of the column is changed to 5 degrees C, the pressure is
changed to 2kPa, the blank solvent does not interfere with the determination of all
durability components. The separation degree of each component is in accordance with the
regulations, and the recovery rate of each component is between 80%~120%,
which is in line with the requirement of durability.
table 3.2.S.4.3.3-2 for the instrument and the trial medicine.
Name type source
Shanghai Jinghai Instrument Co.,
Electronic balance FA1004N
Ltd.
Shanghai equitable instrument
instrument Electronic balance FA1104
and instrument factory
Gas
GC2010 Shimadzu
chromatography
Name batches source
Anhui Hai Kang Pharmaceutical
vildagliptin 150601
Co., Ltd.
Anhui Hai Kang Pharmaceutical
vildagliptin 150602
Trial Co., Ltd.
medicine Anhui Hai Kang Pharmaceutical
vildagliptin 150603
Co., Ltd.
Tianjin Chuang pharmaceutical
3- amino -1-
20150701 intermediate technology
amantadol
productivity promotion Co., Ltd.
180
3.2.S.4.3.3.2 Establishment of residual solvent and starting material method
（1）Residual solvent analysis
The solvent used in the starting material 1 synthesis process is methanol, and the
solvent used in the starting material 2 synthesis process is dichloromethane; the
solvents used in the vildagliptin synthesis process are chloroethyl chloride, 、
Triethylamine 、dichloromethane、Trifluoroacetic anhydride、isopropanol、DMF and
ethanol.
Therefore, the residual solvents to be investigated in the finished products are
methanol, ethanol, isopropanol, dichloromethane,Triethylamine , DMF, chloroacetyl
chloride.
Of which trifluoroacetic acid is acidic, and it is reacted with Triethylamine, so
trifluoroacetic acid is established separately by ion chromatography. Chloroacetic
chloride is unstable, when water becomes chloroacetic acid, ethanol is used in the
synthetic process of this product, and ethyl chloroacetate can be reacted to produce
ethyl chloroacetate. Our company entrusts Beijing medical science and Technology
Co., Ltd. of Li an in Beijing to control the content of Ethyl Chloroacetate.
The starting material 2 (3- amino -1- amantanol) has no obvious ultraviolet
absorption in the wavelength range of 200~400nm, so the residue of vildagliptin is
investigated under gas chromatography.
（3）Method and optimization
（4）the process of establishing the method is listed in the form below, and the
results are shown in table 3.2.S.4.3.3-3.
Table 3.2.S.4.3.3-1: the establishment of residual solvent (1) method
Chromatographic
Rsults
conditions
181
Chromatographic
conditions：
KB-624chromatographic
column
（30m×0.53mm,3.00μm）
Column temperature:
Blank solvent test results：
maintained at 3 for 45
Peak attribution Rt(min)
minutes and heated to 220
DMSO 9.531
degrees at 25 C / min for 10
minutes. unknown 14.802
Inlet temperature：250℃； Test result of control solution

Degree of
Test port temperature： Peak attribution Rt(min) Peak area
separation
280℃
methanol 2.388 -- 169811
conditio Shunt ratio：10:1；
ethanol 3.182 6.219 422517
n1 Column flow speed：4.0ml
Isopropanol 3.758 4.651 500048
Blank solvent：DMSO dichloromethane 4.095 3.021 13029
Concentration of control triethylamine 6.127 18.930 40490
solution： DMF 8.457 19.460 47947
Methanol :0.3mg/ml； 3- amino -1-
16.758 11.901 60220
Ethanol: 0.5mg/ml； amantadol
Isopropanol: 0.5mg/ml； Conclusion: the unknown peak in the blank solvent interferes
Dichloromethane :0.06mg/ with the detection and optimization of L- proline amide.
ml；
triethylamine :0.032mg/ml；
DMF 0.088mg/ml；
3- amino -1-
amantadol:0.1mg/ml。
182
Chromatographic
conditions：
KB-624chromatographic
column Chromatogram of the result of the control solution
（30m×0.53mm,3.00μm） test:
Column temperature:
minutes and heated to 220
degrees at 20 C / min for 10
minutes.
Inlet temperature：250℃；
Test port temperature：
280℃ The result of the control solution test:
Shunt ratio：10:1； Peak Degree of
Rt(min) Peak area
conditio Pressure control model： attribution separation
n2 Methanol 1.574 -- 153270
30kPa
Column flow speed：6.43ml Ethanol 2.105 4.678 418753
Blank solvent：DMSO Isopropanol 2.579 3.607 485025

Dichlorometh
Concentration of control 2.894 2.366 11885
ane
solution：
Triethylamine 5.794 20.241 35776
Methanol: 0.3mg/ml；
DMF 8.780 18.185 47968
Ethanol: 0.5mg/ml；
3- amino -1-
Isopropanol: 0.5mg/ml； 16.152 7.949 79521
amantadol
Dichloromethane: From the chromatogram, the unknown peaks in the blank
0.06mg/ml； solvents still interfere with the detection of L- proline amide and
Triethylamine: 0.032mg/ml； continue to optimize the chromatographic conditions.
DMF 0.088mg/ml；
3- amino -1-
amantadol :0.1mg/ml。
Chromatographic
Test result of control solution：
conditions：
Degree of
KB-624chromatographic Peak attribution Rt(min) Peak area
conditio separation
column
n3 methanol 1.623 -- 184277
（30m×0.53mm,3.00μm）
ethanol 2.288 5.411 424502
Column temperature:
Isopropanol 2.898 3.676 477972
183
minutes and heated to 220 dichloromethane 3.269 2.067 10712
degrees at 25 C / min for 10 Triethylamine 7.411 12.213 30080
minutes. DMF 9.516 6.857 42289
Inlet temperature：250℃； 3- amino -1-
16.289 10.342 82709
Test port temperature： amantadol
280℃ Test product solution results:
Shunt ratio：10:1； Peak Degree of Residual

Rt(min)
attribution separation solvent（%）
pressure：30kPa
methanol 1.651 -- 0.006
Pressure control model：
ethanol 2.314 4.033 0.003
30kPa
Isopropanol 2.905 3.106 0.111
Column flow speed：6.61ml
dichlorometh
Blank solvent：DMSO 3.265 2.023 0.002
ane
Concentration of control
3- amino -1-
solution： 16.293 9.893 0.016
amantadol
Methanol: 0.3mg/ml； Sample solution test results:
Ethanol: 0.5mg/ml； Peak Degree of Residual
Rt(min)
Isopropanol: 0.5mg/ml； attribution separation solvent（%）
Dichloromethane: methanol 1.625 -- 88.95
0.06mg/ml； ethanol 2.290 5.356 92.22
Triethylamine:0.032mg/ml； Isopropanol 2.899 3.606 97.18
DMF 0.088mg/ml； dichlorometh
3.268 2.024 91.91
3- amino -1- amantadol: ane
0.1mg/ml。 Triethylamin
7.208 14.742 88.90
e
Concentration of Trial
DMF 9.515 9.842 91.42
product solution：100mg/ml
Unknown
Concentration of sample 14.929 3.127 /
peak
solution：
3- amino -1- 106.12
Main 16.284 7.342
amantadol
component100mg/ml，
Additive solution chromatogram：
methanol:0.3%，
ethanol:0.5%，
Isopropanol:0.5%，
dichloromethane:0.06%，
Triethylamine:0.032%，
DMF0.088%，3- amino -1-
184
amantadol:0.1%。 The chromatographic diagram shows that the blank solvent does
not interfere with the determination of the residual solvents and
impurities in this product. The separation degree of the residual
solvent peaks and impurities is in accordance with the
regulations. The recovery rate is between 80%~120%. The
condition of this chromatographic condition is the condition for
the determination of the residual solvent in this product.
（ 3 ） Investigation of unknown peaks in residual solvents (RT about 15

minutes)
① The impurity is initially degraded at high temperature of 250 C, so the
unknown inlet peak temperature is reduced.
Blank solvent:
DMSO
Take this product 500mg, weigh it accurately, put it in a 5ml measuring bottle,
add solvent to shake it properly, dissolve it, dilute it to scale with solvent, shake well.
(100mg/ml)
Chromatographic conditions 1:
Chromatographic column: 6% cyanopropyl phenyl -94% Dimethyl Polysiloxane
(or nearly the same polarity) as a stationary liquid chromatographic column （KB-624
30m0.53mm，3.0μm）
Column temperature: the initial temperature is 40 degrees, maintain for 5
minutes, heat up to 220 degrees per minute at 25 degrees Celsius, maintain 10
minutes.Inlet temperature: 250℃
Test port temperature：280℃
Shunt ratio：10:1
Pressure control model：30kPa
Chromatographic conditions 2:
Chromatographic column: 6% capillary column with cyanopropyl phenyl -94%
Dimethyl Polysiloxane (or similar polarity) as stationary liquid. （ KB-624
30m0.53mm，3.0μm）
Column temperature: the initial temperature is 40 degrees, maintain for 5
minutes, heat up to 220 degrees per minute at 25 degrees Celsius, maintain 10
185
minutes.
Inlet temperature：150℃
Test port temperature：280℃
Shunt ratio：10:1
Pressure control model：30kPa
Chromatography operation:
The samples were collected for 1 L, respectively, and then injected under the
above two chromatographic conditions, and the chromatograms were recorded.
Test results：
Table 3.2.S.4.3.3-1: test results of the test product solution
Condition Unknown peak area
Injection port 250℃ 522262
Injection port 150℃ 2248
conclusion：
When the inlet temperature is lowered, the area of the unknown peak area
decreases obviously, which is in line with the expectation (the impurity is heated at
the inlet).
② The degradation conditions of unknown impurities were further verified
by high temperature degradation experiments.
Take this product 1.0g, weigh it accurately, put it in a 10ml measuring bottle, add
solvent to shake it properly, dissolve it, dilute it to scale with solvent,shake well.
(100mg/ml)
Table 3.2.S.4.3.3-2: preparation of high temperature degradation samples
Degradation conditions
The sample solution was 2ml and placed at about 100 4H in water bath.
The sample solution was 2ml and placed at about 250 30s.
The undegraded samples and the above two high temperature degradation
samples were collected for 1 L, respectively, and the chromatograms were recorded
by residual solvents.
186
Test results：
Table 3.2.S.4.3.3-2: test results under gas phase conditions
Condition Unknown peak area
Undegraded 51728
100℃ 57221
250℃ 216703
conclusion：
Under the condition of 100 C, there is no obvious change in the area of the peak
peak and peak area compared with the undegraded condition. The area of the
unknown peak peak is obviously increased under the condition of 250 C. Therefore, it
is speculated that the impurity is produced only at high temperature and will not
produce under the condition of storage and transportation of this product.
③Investigation of stability samples
The raw materials were collected for 0 days, and the raw materials were
accelerated to test the unknown peaks in June.
Blank solvent:
DMSO
Take the raw 500mg of vildagliptin, determine it accurately, put 5ml in the
measuring bottle, add solvent to shake it properly, dissolve it, dilute it to scale with
solvent, shake well. (100mg/ml)
The above solutions were 1 l each, and then injected into the residual solvents to
record the chromatograms.
Test results：
Table 3.2.S.4.3.3-2: results of stability test

Batches Condition Unknown peak
150601 0 day 140348
Material
150601 Speed up 6 month 138303
187
150602 0 day 125409
150603 0 day 114263
conclusion：
The peak area of the unknown peak in the sample solution of the stable sample has no
obvious increase over 0 days, indicating that the peak will not be degraded under the
possible storage conditions of the sample, and the impurity should be produced only
at a very high temperature.
3.2.S.4.3.3.3 Methodology validation of residual solvents and 3- amino -1-
amantadine
3.2.S.4.3.3.3.1 System applicability test
The application of the system to establish the GC method is to repeat the sample
with the control solution and record the chromatogram. In the end, the RSD of the
peak area of each component of the 6 needles must not exceed 10%. The degree of
separation between the peaks of each component should be in accordance with the
regulations. During the validation process, the applicability test was carried out and
passed. The test results are shown in table 3.2.S.4.3.3-8~10.
Table 3.2.S.4.3.3-2: results of repeated injection of reference needle 6 needles -
retention time
Mean RSD
Component 1 2 3 4 5 6
Value （%）
Methanol 1.593 1.595 1.595 1.598 1.595 1.595 1.595 0.10
Ethanol 2.246 2.248 2.249 2.253 2.249 2.249 2.249 0.10
Isopropanol 2.848 2.848 2.849 2.854 2.850 2.850 2.850 0.09
Dichloromethane 3.213 3.213 3.213 3.217 3.214 3.213 3.214 0.05
Triethylamine 6.885 6.881 6.880 6.875 6.875 6.872 6.878 0.07
DMF 9.464 9.464 9.465 9.466 9.466 9.466 9.465 0.01
3- amino -1- 16.18 16.18 16.18 16.18 16.18 16.18 16.18
0.01
amantadol 5 2 2 2 1 2 2
188
Table 3.2.S.4.3.3-2: results of repeated injection of reference solution 6 needles - peak

area
Mean RSD
value （%）
21819 21971 21985 22127 22159 22125 22031
Methanol 0.59
8 9 4 1 1 5 5
49577 49842 49751 49917 49979 49873 49823
Ethanol 0.29
1 7 6 9 0 1 6
54486 54743 54766 55095 55119 55030 54873
Isopropanol 0.46
8 1 8 4 8 4 7
Dichloromethan
16851 16996 17370 17366 16875 16741 17033 1.60
e
Triethylamine 38078 38865 39486 39628 38954 39917 39155 1.69
DMF 52985 53544 53470 53806 53095 53922 53470 0.70
3- amino -1- 13321 12939 12854 12946 13017 13113 13032
1.28
amantadol 9 6 0 8 7 1 2
Table 3.2.S.4.3.3-2: results of repeated injection of reference needle 6 needles -

separation degree
Methanol -- -- -- -- -- --
Ethanol 5.988 5.957 5.954 5.532 5.775 5.814
Isopropanol 3.915 3.888 3.887 3.722 3.803 3.810
Dichloromethane 2.128 2.112 2.102 2.047 2.084 2.086
Triethylamine 19.986 19.860 19.785 19.602 19.871 19.839
DMF 5.591 5.585 5.513 5.474 5.473 5.434
3- amino -1-
10.643 10.593 10.533 10.389 10.338 10.321
amantadol
Blank solvent：
DMSO
189
Residual solvent location solution:
Methanol 300mg, ethanol 500mg, isopropanol 500mg, dichloromethane 60mg,
three ethylamine 32mg, N, N- two methyl formamide (DMF) 88mg, precision called,
respectively, in the 100ml measuring bottle, respectively, with two methyl sulfoxide
dilution to the scale, shake well. (methanol 3mg/ml, ethanol 5mg/ml, isopropanol
5mg/ml, dichloromethane 0.6mg/ml, three ethylamine 0.32mg/ml, DMF 0.88mg/ml)
3-amino -1- amantadol positioning solution:
Take 3- amino -1- amantadine 50mg, precise, set in 50ml volumetric flask,
dissolve and dilute to scale with two methyl sulfoxide, shake well. (1mg/ml)
Control solution:
Precise positioning of each component of the solution 5ml, placed in the same
50ml volumetric flask, diluted with DMSO to scale, shake well. (methanol 0.3mg/ml,
ethanol 0.5mg/ml, isopropanol 0.5mg/ml, dichloromethane 0.06mg/ml, Triethylamine
0.032mg/ml, DMF 0.088mg/ml, L- prolyamide 0.1 mg/ml, 3- amino -1- adamantol
0.1 mg/ml)
Take this product 0.5g, weigh it accurately, put it in 5ml volumetric flask, add
DMSO, shake it, dissolve it, dilute it to scale with DMSO, shake it well. (100mg/ml)
Sample solution:
Take this product 0.5g, precision, set, 5ml bottle, precision measure each
component positioning solution 0.5ml, shake to dissolve vildagliptin, use DMSO
dilution to the scale, shake well. (principal component 100mg/ml, methanol 0.3mg/ml,
ethanol 0.5mg/ml, isopropanol 0.5mg/ml, dichloromethane 0.06mg/ml, Triethylamine
0.032mg/ml, DMF 0.088mg/ml, 3- amino -1-, 0.1 mg/ml)
Operation:
The blank solvent, reference substance solution, each component location
solution and the sample solution 1 ul were injected into the gas chromatograph
respectively, and the chromatogram was recorded.
Results：
Table 3.2.S.4.3.3-2: results of a single residual solvent localization solution test
Peak attribution Retention time (minutes)
methanol 1.607
190
ethanol 2.273
Isopropanol 2.886
dichloromethane 3.260
Triethylamine 6.798
DMF 9.489
3- amino -1- amantadol 16.256

Retention time Rate of recovery
Peak attribution Degree of separation
(minutes) （%）
methanol 1.619 -- 86.28
ethanol 2.282 5.764 87.29
Isopropanol 2.891 3.835 92.37
dichloromethane 3.259 2.091 82.75
Triethylamine 6.876 19.478 97.44
DMF 9.499 5.691 87.43
3- amino -1-
16.270 7.333 94.76
amantadol
Conclusion: the blank solvent has no interference at the peak position of each
component, and the separation degree of the impurities in the sample solution is in
line with the requirements.
3.2.S.4.3.3.3.3 Sensitivity
Operation：
The signal-to-noise ratio of each component in reference solution under special
conditions is shown in table 3.2.S.4.3.3-13.
Table 3.2.S.4.3.3-2: the signal-to-noise ratio of the control solution
Methan Isopropa Dichlorome Triethylami

Component Ethanol DMF 3- amino -1- amantadol
ol nol thane ne
S/N 2523 4107 3463 117 208 532 727
According to the signal to noise ratio and solution concentration, the response
factor of dichloromethane in the residual solvent is the lowest, with the concentration
of dichloromethane S/N more than 30 (in order to take into account the difference of
191
the response of different detectors) as the quantitative limit concentration and the
quantitative limit concentration dilution 3.3 times as the detection limit concentration;
the quantitative limit repeat sample is 6 times and the detection limit is limited.
Repeat the sample 3 times.
Results：
The test limits and quantitation limits are shown in table 3.2.S.4.3.3-14~15.
Table 3.2.S.4.3.3-2: quantitative limit test results
Signal to
noise ratio A percentage
Concentrati Mean Area RSD
Component (first equivalent to a
on（mg/ml） area （%）
needle limit（%）
result)
Methanol 0.090 419.25 60510 4.94 30
Ethanol 0.149 642.27 136394 4.76 30
Isopropanol 0.150 808.10 158163 3.03 30
Dichloromethane 0.018 30.36 5223 4.41 30
Triethylamine 0.010 128.64 15030 3.40 30
DMF 0.027 53.51 10364 9.88 30
3- amino -1-
0.029 151.97 51955 8.96 30
amantadol
Table 3.2.S.4.3.3-2: test limit test results

The amount
Signal to
of the A percentage
Concentrati noise ratio
Component Mean area principal equivalent to a
on（mg/ml） (first needle
component limit（%）
result)
（%）
Methanol 0.027 134.86 20923 0.027 9
Ethanol 0.045 194.66 45247 0.045 9
Isopropanol 0.045 269.00 54807 0.045 9
Dichloromet
0.005 9.04 1680 0.005 9
hane
192
Triethylami
0.003 45.54 5071 0.003 9
ne
DMF 0.008 17.51 3059 0.008 9
3- amino -1-
0.009 86.75 30879 0.009 9
amantadol
Conclusion: the detection sensitivity of methanol, ethanol, isopropanol,
dichloromethane,Triethylamine, DMF and 3- amino -1- amantadine is in line with the
regulations.
3.2.S.4.3.3.3.4Linear range
The contents of methanol, ethanol, isopropanol, dichloromethane,Triethylamine ,
DMF, 3- amino -1- adamantanol were limited to the limit concentration of 120%, and
the linearity of each component was investigated. The requirement is linear in the
concentration range, and the correlation coefficient r is more than 0.99.
Preparation of solution:
The preparation of linear reserve liquid for residual solvent:
Methanol 300mg, ethanol 500mg, dichloromethane 60mg, DMF
88mg,Triethylamine 32mg and isopropanol 500mg respectively, are niced and
determined respectively. In the same 100ml bottle,DMSO is dissolved and diluted to
the scale and shake well. (methanol 3mg/ml, ethanol 5mg/ml, isopropanol 5mg/ml,
dichloromethane 0.6mg/ml,Triethylamine 0.32mg/ml, DMF 0.88mg/ml)
Preparation of L- proline amide and 3- amino -1- adamantane linear storage
solution:
Take 3- amino -1- amantadine 100mg each, precise, set the same 100ml volume
bottle, use DMSO to dissolve and dilute to scale, shake well. (1mg/ml)
The preparation of linear solutions of various concentrations is shown in table
3.2.S.4.3.3-16.
Table 3.2.S.4.3.3-2: preparation of linear solution
Volume of linear Volume of linear
reserve liquid of storage liquid of 3- Volume of a measuring
Concentration level
residual solvent amino -1- bottle（ml）
（ml） amantadol（ml）
limit of
3 3 100
quantitation
193
60% 3 3 50
80% 2 2 25
100% 5 5 50
120% 3 3 25
operation：
The linear solution was measured at 1 L, injected into gas chromatograph and
recorded chromatogram.
Results：
Table 3.2.S.4.3.3-2: the results of linear investigation of methanol
Linear range of methanol
C（mg/ml） Peak area
0.090 60386
0.181 118447
0.241 171659
0.301 219113
0.361 249967
linear y=724046.7123x-6020.811
equation 5
correlation
coefficient R2=0.9942
（r2）
194
Table 3.2.S.4.3.3-2: results of linear investigation of ethanol

Linear range of ethanol
C（mg/ml） peak area
0.149 128102
0.298 272761
0.398 394176
0.497 495314
0.597 583916
linear y=1036797.7713x-27152.09
equation 43
correlatio
n
R2=0.9979
coefficient
（r2）
Table 3.2.S.4.3.3-2: results of linear investigation of isopropanol

linear range of Isopropanol
0.150 145467
0.300 306886
0.399 441301
0.499 550572
0.599 651245
linear y=1142857.7527x-25907.18
equation 03
correlation
（r2）
195
Table 3.2.S.4.3.3-2: results of linear investigation of Triethylamine

Linear range of Triethylamine
0.010 9197
0.019 20522
0.026 28805
0.032 36364
0.038 40776
linear y=1126638.0635x-988.086
equation 1
correlation
（r2）
Linear results of table 3.2.S.4.3.3-2:DMF

linear range of DMF
0.027 10648
0.054 30751
0.072 44224
0.090 54480
0.107 64754
linear
y=673608.8470x-6053.2336
equation
correlation
（r2）
196
Table 3.2.S.4.3.3-2: linear investigation results of dichloromethane

linear range of dichloromethane
0.018 4047
0.035 8479
0.047 11929
0.059 14997
0.071 17437
y=256327.1477x-438.368
linear equation
9
correlation
R2=0.9976
coefficient（r2）
Table 3.2.S.4.3.3-2:3- linear investigation results of amino -1- amantadine

Linear range of 3- amino -1- amantadol
0.029 51163
0.059 85707
0.079 108820
0.098 133403
0.118 162171
linear y=1244667.5419x+12908.381
equation 1
correlation
（r2）
Conclusion: the experimental results show that the peak area and concentration
of methanol, ethanol, isopropanol, dichloromethane, Triethylamine, DMF, 3- amino
-1- and amantadol have a linear relationship with the concentration range.
197
3.2.S.4.3.3.3.5 Repeatability
6 samples were prepared in parallel and determined by residual solvent method.
dichloro
methano ethanol Isopropa Triethyla DMF 3- amino -1-
component methane
l（%）（%） nol（%） mine（%）（%） amantadol（%）
（%）
1 0.004 0.003 0.105 0.005 ND ND 0.015

2 0.005 0.002 0.100 0.004 ND ND 0.015
3 0.005 0.002 0.100 0.004 ND ND 0.015
4 0.004 0.002 0.100 0.004 ND ND 0.015
5 0.005 0.002 0.101 0.004 ND ND 0.014
6 0.005 0.002 0.100 0.004 ND ND 0.014
mean value
0.005 0.002 0.101 0.004 / / 0.015
（%）
RSD(%) 3.18 14.43 2.02 11.14 / / 5.55

Conclusion: the detection amount of other residual solvents except isopropanol is
low, so the RSD requirements for other residual solvents are relaxed to 15%, and the
number of detection requirements is consistent. It can be seen from the test results that
the precision is in accordance with the requirements.
3.2.S.4.3.3.3.6 Accuracy (recovery)
The recovery rate was determined by selecting 3 concentrations of 150%, 100%
(limit concentration) and quantitation limit, and 3 samples were prepared in parallel
with each concentration.
Preparation of solution:
Preparation of residual solvent storage liquid:
Methanol 300mg, ethanol 500mg, dichloromethane 60mg,DMF 88mg,
Triethylamine 32mg, and isopropanol 500mg were respectively given, respectively. In
the same 100ml bottle, DMSO was diluted to the scale and shake well. (methanol
3mg/ml, ethanol 5mg/ml, dichloromethane 0.6mg/ml, DMF0.88mg/ml, Triethylamine
0.32mg/ml, isopropanol 5mg/ml)
Preparation of 3- amino -1- amantadol storage liquid:
198
Take 3- amino -1- amantadine 50mg, each of which is precisely defined, put it in
the same 50ml volumetric flask, dissolve and dilute it to scale with DMSO, shake well.
(3- amino -1- amantadol 1mg/ml)
The 5ml of the above storage solution was accurately measured and placed in the
same 50ml volumetric flask. Diluted with DMSO to scale, shake well. (methanol
0.3mg/ml, ethanol 0.5mg/ml, dichloromethane 0.06mg/ml, DMF 0.088mg/ml,
Triethylamine 0.032mg/ml, isopropanol 0.5mg/ml, 3- amino -1- of amantadol
0.1mg/ml)
Take this product 0.5g, precision weighing, put 5ml measuring bottle, add
DMSO to shake it properly, dilute it to scale with DMSO, shake well. (100mg/ml)
The preparation of the recovery solution:

According to table 3.2.S.4.3.3-27, take appropriate amount of the test product,
set up the corresponding volume bottle, add the amount of recovery storage liquid,
add DMSO to shake to dissolve, use DMSO to dilute the scale, shake well. 3 solutions
are prepared at each level.
Table 3.2.S.4.3.3-2: preparation of recovery sample solution
The
Volume
percentage
extraction of
Recovery rate content of Volume of 3- amino -1-
Sample residual Volume of
investigation the limit adamantanol storage
size（g） solvent bottle（mL）
solution concentratio liquid（mL）
volume
n
（mL）
（%）
limit of
R1 10 3 3 100
quantitation
R2 100 1 1 1 10
R3 150 2.5 3 3 25
Operation：
The precision recovery rate was determined by 1 ul of each solution, injected
into gas chromatograph and recorded chromatogram.
Results：
199
Table 3.2.S.4 the results of the recovery of methanol
Backgrou Theoretic
Recovery Measured Average
nd al Concentra Rate of
rate Peak concentrat recover RSD
concentrat concentrat tion recover
investigatio area ion y rate （%）
ion ion
（mg/ml） y（%）
n solution （mg/ml）（%）
（mg/ml）（mg/ml）
30%-1 52932 0.004 0.094 0.079 84.05
30%-2 53233 0.004 0.090 0.094 0.080 84.53
30%-3 53382 0.004 0.094 0.080 84.78
100%-1 187363 0.004 0.305 0.281 92.14
100%-2 183516 0.004 0.301 0.305 0.275 90.25 88.78 4.25
100%-3 175719 0.004 0.305 0.264 86.42
150%-1 225967 0.004 0.365 0.339 92.82
150%-2 225937 0.004 0.361 0.365 0.339 92.80
150%-3 222204 0.004 0.365 0.333 91.27
Table 3.2.S.4.3.3-2: the results of the determination of the recovery of ethanol

Backgroun Theoretica Averag
Recovery Measured
d l Concentrat Rate of e
rate Peak concentrat
concentrat concentrat ion recover recover RSD（%）
investigatio area ion
ion ion
（mg/ml） y（%） y rate
n solution （mg/ml）
（mg/ml）（mg/ml）（%）
30%-1 119376 0.002 0.151 0.126 83.26
30%-2 121359 0.002 0.149 0.151 0.128 84.64
30%-3 120657 0.002 0.151 0.127 84.15
100%-1 436086 0.002 0.499 0.460 92.13
100%-2 428728 0.002 0.497 0.499 0.452 90.57 89.03 4.29
100%-3 430044 0.002 0.499 0.453 90.85
150%-1 523721 0.002 0.598 0.552 92.26
150%-2 523023 0.002 0.596 0.598 0.551 92.14
150%-3 518085 0.002 0.598 0.546 91.27
Table 3.2.S.4.3.3-2: the results of the recovery of isopropanol
200
Recovery rate Background Theoretical Concentrat Measured Rate of Average
Peak RSD
investigation concentrati concentrati ion concentration recovery recovery
area （%）
solution on（mg/ml）on（mg/ml）（mg/ml）（mg/ml）（%） rate（%）
30%-1 257045 0.101 0.251 0.245 97.57
30%-2 256952 0.101 0.150 0.251 0.245 97.48
30%-3 256132 0.101 0.251 0.244 97.28
100%-1 614649 0.101 0.600 0.585 97.56
100%-2 605796 0.101 0.499 0.600 0.577 96.17 97.02 0.57
100%-3 607056 0.101 0.600 0.578 96.38
150%-1 714336 0.101 0.700 0.680 97.16
150%-2 714690 0.101 0.599 0.700 0.681 97.20
150%-3 708715 0.101 0.700 0.675 96.40
Table 3.2.S.4.3.3-2: Determination results of dichloromethane recovery

Backgrou Theoretica
nd l Measured Average
Recovery rate Concentrati Rate of
RSD
Peak area concentrat concentrat concentrati recovery
investigation on（mg/ml） recovery（%）（%）
ion ion on（mg/ml） rate（%）
（mg/ml）（mg/ml）
30%-1 4321 0.005 0.022 0.019 87.60
30%-2 4480 0.005 0.017 0.022 0.020 90.80
30%-3 4467 0.005 0.022 0.020 90.59
100%-1 13574 0.005 0.063 0.061 96.78
100%-2 12938 0.005 0.058 0.063 0.058 92.25 91.39 2.85
100%-3 13109 0.005 0.063 0.059 93.47
150%-1 14990 0.005 0.074 0.067 90.14
150%-2 15189 0.005 0.070 0.074 0.068 91.34
150%-3 14887 0.005 0.074 0.067 89.53
Table 3.2.S.4.3.3-2: Triethylamine of ethylamine recovery
201
Backgrou Theoretica Averag
Recovery Measured
nd l Concentra Rate of e
rate Peak concentrat RSD
concentrat concentrat tion recovery recover
investiga area ion （%）
ion ion
（mg/ml）（%） y rate
tion （mg/ml）
30%-1 8117 - 0.009 0.007 82.07
30%-2 8025 - 0.009 0.009 0.007 81.14
30%-3 8484 - 0.009 0.008 85.78
100%-1 33040 - 0.030 0.030 100.22
100%-2 32820 - 0.030 0.030 0.030 99.55 94.53 9.40
100%-3 31852 - 0.030 0.029 96.61
150%-1 40140 - 0.036 0.037 101.46
150%-2 40288 - 0.036 0.036 0.037 101.83
150%-3 40405 - 0.036 0.037 102.13
Table 3.2.S.4.3.3-2:DMF recovery rate determination result

Backgrou Theoretica Averag
Recovery Measured
nd l Concentra Rate of e
rate Peak concentrat RSD
concentrat concentrat tion recovery recover
investiga area ion （%）
ion ion
（mg/ml）（%） y rate
tion （mg/ml）
/30%-1 14039 - 0.026 0.025 95.80
30%-2 13982 - 0.026 0.026 0.025 95.41
30%-3 12960 - 0.026 0.023 88.43
100%-1 45998 - 0.088 0.082 94.16
100%-2 47078 - 0.088 0.088 0.084 96.37 96.10 4.22
100%-3 45468 - 0.088 0.081 93.08
150%-1 59169 - 0.105 0.106 100.94
150%-2 58847 - 0.105 0.105 0.105 100.39
150%-3 58798 - 0.105 0.105 100.30
Table 3.2.S.4.3.3-2:3- determination of amino -1- amantadine recovery
202
Backgrou
Recovery nd Theoretical
Concentrat Measured Rate of Average
rate Peak concentra concentrati RSD
ion concentrati recover recovery
investigatio area tion on （%）
（mg/ml）on（mg/ml）y（%） rate（%）
n （mg/ml （mg/ml）
）
30%-1 61543 0.018 0.050 0.056 112.42
30%-2 63730 0.018 0.031 0.050 0.058 116.36
30%-3 63201 0.018 0.050 0.057 115.52
100%-1 151741 0.018 0.123 0.138 112.16
100%-2 124299 0.018 0.104 0.123 0.113 91.88 104.81 8.83
100%-3 135743 0.018 0.123 0.123 100.36
150%-1 152368 0.018 0.144 0.138 96.19
150%-2 155834 0.018 0.125 0.144 0.141 98.38
150%-3 158493 0.018 0.144 0.144 100.06
Conclusion: the test results are in accordance with the validation standard.
3.2.S.4.3.3.3.7 durability
Durability refers to the degree of tolerance under the condition of fine tuning
chromatography and the determination of residual solvents. The durability of the
method was investigated according to the interference of the blank solvent, the
separation of the peaks and the recovery rate of the residual solvents. It is required
that the blank solvent does not interfere with the determination. The separation peak
between the peaks is greater than 1.5, and the recovery rate of each residual solvent is
between 80%~120%.
The chromatographic conditions for the fine-tuning are shown in table
3.2.S.4.3.3-33.
Table 3.2.S.4.3.3-2: conditions for the durability of the micro adjustment
chromatography
Test condition
Standard
Unadjusted
condition
Column
±5℃
temperature
203
Pressure ±2kPa
Blank solvent:
DMSO
Preparation of residual solvent storage liquid:
Methanol 300mg, ethanol 500mg, dichloromethane 60mg,DMF88mg,
Triethylamine 32mg, and isopropanol 500mg were respectively given, respectively.
In the same 100ml bottle, DMSO was diluted to the scale and shake well. (methanol
3mg/ml, ethanol 5mg/ml, dichloromethane 0.6mg/ml, DMF 0.88mg/ml,
Triethylamine 0.32mg/ml, isopropanol 5mg/ml)
Preparation of 3- amino -1- amantadol storage liquid:
Take 3- amino -1- amantadine 50mg each, precise, set in 50ml volumetric flask,
dissolve and dilute to scale with DMSO, shake well. (3- amino -1- amantadol
1mg/ml)
The 5ml of the above storage solution was accurately measured and placed in the
same 50ml volumetric flask. Diluted with DMSO to scale, shake well. (methanol
0.3mg/ml, ethanol 0.5mg/ml, dichloromethane 0.06mg/ml, DMF 0.088mg/ml,
Triethylamine 0.032mg/ml, isopropanol 0.5mg/ml, 3- amino -1- of amantadol
0.1mg/ml)
Preparation of sample solution:
Take this product 1g, precision and set, in the 10ml bottle, the precision of the
residual solvent storage liquid 1ml, and the diluent to shake to dissolve, dilute the
diluent to the scale, shake well.
operation：
The blank solvent, reference substance solution and sample sample solution were
accurately separated by 1 L, respectively, and injected into gas chromatograph
respectively, and the chromatogram was recorded.
Results：
Table 3.2.S.4.3.3-2: durability test results - separation degree
Standard Pressure Pressure Starting Starting
condition
condition 28kPa 32kPa temperature temperature
204
35℃ 45℃
methanol -- -- -- -- --
ethanol 5.764 5.647 5.309 6.030 5.086
Isopropanol
3.835 3.816 3.599 3.987 3.424
dichloromethane 2.091 2.103 1.984 1.959 2.111

Triethylamine 19.478 19.481 18.796 19.036 18.935
DMF 5.691 5.695 5.517 5.426 5.604
3- amino -1-
7.333 7.482 7.114 7.438 7.170
amantadol
Table 3.2.S.4.3.3-2: durability test results - recovery rate

Starting Starting
Standard Pressure Pressure
condition temperature temperature
condition 28kPa 32kPa
35℃ 45℃
methanol（%） 86.28 96.37 92.35 94.03 93.96
ethanol（%） 87.29 94.74 92.61 93.26 94.58
Isopropanol
92.37 99.71 97.11 98.80 99.75
(%)
dichloromethane(%) 82.75 105.51 97.06 101.72 106.24
Triethylamine(%) 97.44 101.98 82.63 116.04 103.71
DMF(%) 87.43 97.41 96.62 90.30 98.33
3- amino -1-
94.76 113.44 117.00 103.64 110.65
amantadol(%)
Conclusion: when the temperature of the column is changed to 5 degrees C, the

pressure is changed to 2kPa, the blank solvent does not interfere with the
determination of all components. The separation degree of each component is in
accordance with the regulations, and the recovery rate of each component is between
80%~120%, which is in line with the requirement of durability.
3.2.S.4.3.3.3.8 Sample determination result

The results of the 3 batch of verification samples are shown in table
3.2.S.4.3.3-36.
The results of residual solvents in the samples of vildagliptin
Residual solvent Sample information
205
Small test
Verification batch
batch
140401 140601 140701 140702
methanol（%） 0.004 0.004 0.004 0.005
ethanol（%） 0.003 0.003 0.002 0.001
Isopropanol
0.111 0.105 0.103 0.102
(%)
dichloromethane(%) 0.017 0.005 0.004 0.008
Triethylamine(%) ND ND ND ND
DMF(%) ND ND ND ND
3- amino -1-
0.024 0.015 0.015 0.015
amantadol(%)
3.2.S.4.3.4 Methodology validation of content

A summary of the method validation results is shown in table 3.2.S.4.3.4-1.
Table 3.2.S.4.3.4-1: validation of analytical method for determination of raw material
content
Verification
Verification result
project
The degree of separation between the main peak and the main peak in the
oxidation degradation solution is 1.69, more than 1.5. The mean value of the peak
System
area of the main peak is 2234219, RSD is 0.23%, less than 1%, and the tailing
applicability
factor of the main peak is about 0.9 and less than 1.8, which is in line with the
application requirements of the system.
The blank solvent did not interfere with the determination of the content of the
product, and the separation degree of the impurities accords with the principal
component separation degree. The tailing factor of the main peak is less than 1.8.
Specificity
The spectra of the degradation samples show that the separation degree of the
wig lant peak and the adjacent degradation products are both greater than 1.5, and
the peak purity of the main peak is in conformity with each condition.
In the concentration range of 0.100mg/ml~1.002mg/ml, the response value is
Linearity
linear with the concentration, and the linear equation is
and range
y=3962930.777x+35569.122 R2=1.000.
Repeatabilit The average content of 6 samples is 100%, RSD% is 0.29%, which meets the
y requirements.
Intermediate The mean content of the 6 samples was 99.9%, RSD% was 0.33%, and the mean
206
precision difference between repeatability and intermediate precision was 0.1%, which
accords with the verification requirement.
Solution The reference solution remained stable at room temperature for 23 hours at room
stability temperature.
When the flow rate is adjusted for 0.2ml/ minutes, the pH of the buffer solution is
adjusted to 0.2, the organic phase ratio of the liquid phase is adjusted to 2, the
column temperature is adjusted to 5 C, the detection wavelength is adjusted to
durability 2nm, the chromatographic column is changed, the separation degree of the
impurities and the main component peaks of the oxidative degradation is more
than 1.5, and the content measurement is between 98.0%~102.0%, which is in line
with the durability.

Instrument and test medicine show in table 3.2.S.4.3.4-2。
Name Type Source
High performance liquid
LC-16 Shimadzu
chromatography
High performance liquid
1100 Agilent
chromatography
FA1004 Shanghai Jinghai Instrument
Instru balance
N Co., Ltd.
ment
Sardo Scientific Instruments
balance BS124S
Co., Ltd.
Shanghai electric science
pH meter PHS-3C instrument Limited by Share
Ltd
Name batche source
dipotassium hydrogen National drug group chemical
/
phosphate reagents Co., Ltd.
Medici
dipotassium hydrogen National drug group chemical
ne /
phosphate reagents Co., Ltd.
sodium hydroxide / Beijing chemical plant
vildagliptin 150601 Anhui Hai Kang
207
Pharmaceutical Co., Ltd.
Anhui Hai Kang
vildagliptin 150602
Anhui Hai Kang
vildagliptin 150603
2013121 Shenzhen Zhen Qiang Bio
Impurity A
9 Technology Co., Ltd.
Impurity B
TH11937 Shenzhen Zhen Qiang Bio
Impurity C
-018 Technology Co., Ltd.
Impurity D
1424-093 Shenzhen Zhen Qiang Bio
Impurity E
A3 Technology Co., Ltd.
Impurity F
Impurity G
3.2.S.4.3.4.2The establishment of analytical methods

Referring to the conditions under the content determination of Vildagliptin Tablets
import registration standard, the method for determining the content of the product is
established.
（1）preparation of sample solution
Phosphate buffer solution:
The potassium dihydrogen phosphate 1.3g, add 1000ml water to dissolve, with
two potassium hydrogen phosphate solution (three out of two potassium hydrogen
phosphate hydrate 1.96g, 10ml water to dissolve) to adjust the pH value to 6.5.
mobile phase：
A：Phosphate buffer- water -acetonitrile -methanol (400:600:15:15).
B：Phosphate buffer - acetonitrile - methanol (400:450:150).
Diluent solution:
0.2% (v/v) hydrochloric acid solution acetonitrile (90:10)
208
Trial product solution：
Take this product 12.5mg, precision weighed, put in 25ml measuring bottle, add
diluent to the right amount, shake to dissolve, dilute to scale, shake well. (0.5mg/ml)
Impurity location solution:
Taking impurities A, impurity B, impurity C, impurity D, impurity E,
impurity F, and impurities G each 10mg, respectively, in the 100ml measuring
bottle, adding the diluent to dissolve, dilute the diluent to the scale and shake well.
(0.1mg/ml)
Sample solution:
（2）Take this product 10mg, precision, set, 20ml bottle, add the diluent
appropriate amount, add the above positioning solution each 1ml, shake
to dissolve, dilute the diluent to the scale, shake well. (principal
component 0.5mg/ml, impurity 1%)
（3）Chromatographic conditions：
chromatographic column：Waters XTerra ®MS C18（50×4.6mm，3.5µm）
Flow speed：1.8ml
column temperature：35℃
Sample volume：10μl
（4）operation：
The 10 L of the above solutions were accurately injected into the liquid
chromatograph respectively, and the chromatograms were recorded.
（5）test results
Table 3.2.S.4.3.4-2: single impurity location solution results

Component Rt(min)
impurityA 1.432
impurityB 3.602
impurityG 3.827
impurityC 4.057
impurityD 4.496
209
vildagliptin 6.129
impurityE 7.089
impurityF 8.566
Table 3.2.S.4.3.4-1: test results of added sample solution

Peak Degree of Theoretic Trailing
Rt(min) Area RRT
attribution separation al tray# factor
impurityA 1.423 42498 -- 523 0.23 0.896
impurityB 3.578 75806 9.092 4041 0.58 1.026
impurityG 3.823 3820 1.254 8525 0.62 1.257
impurityC 4.094 29242 1.552 7902 0.67 1.176
impurityD 4.510 36130 2.103 7298 0.74 1.003
vildaglipti
6.132 2499358 8.785 24101 1.00 0.962
n
impurityE 7.096 37077 7.071 61250 1.16 1.232
impurityF 8.574 46952 11.599 59813 1.40 1.115
Additive solution chromatogram：
（6）Conclusion:
The results show that the separation degree of the impurities and the main peaks
is good, the number of theoretical plates of the main peak is 24101, the trailing factor
is 0.962, the peak is good, which shows that the method is suitable for the
determination of the content of this product.
210
3.2.S.4.3.4.3 Verification of analytical methods
3.2.S.4.3.4.3.1 System applicability
（1）operation
The separation of the degradation impurity peaks and the main peaks should be more
than 1.5 with the oxidation degradation solution, and the control solution is repeated
into the sample and the chromatogram is recorded at the wavelength of 210nm. The
relative standard deviation of the main peak area of the 5 needles must not exceed 1%.
The trailing factor of Vildagliptin may not be over 1.8.
（2）results
Table 3.2.S.4.3.4-1: experimental results of oxidative degradation solution
Rt（min） Peak attribution Degree of separation
2.905 Unknown --
3.477 ImpurityD 2.41
4.448 Unknown 4.35
4.793 Unknown 2.20
5.029 Unknown 1.75
5.241 Vildagliptin 1.69
Table 3.2.S.4.3.4-1: results of repeated injection of 5 needles in reference solution.

Sampling Peak area Retention time Trailing factor
1 2235698 5.231 0.91
3 2240845 5.218 0.90
4 2230457 5.234 0.90
5 2228045 5.229 0.89
mean value 2234219 5.229 --
RSD% 0.23 0.12 --
（3）conclusion
The test results show that the system meets the requirements of system
applicability.
（a）Sample solution preparation
Diluent：
211
0.2% (v/v) hydrochloric acid solution acetonitrile (90:10).
Take this product 12.5mg, precision weighed, put in 25ml measuring bottle, add
diluent to the right amount, shake to dissolve, dilute dilute to scale, shake well.
(0.5mg/ml)
Impurity location solution:
Taking impurities A, impurity B, impurity C, impurity D, impurity E,
impurity F, and impurities G each 10mg, respectively, in the 100ml measuring
bottle, adding the diluent to dissolve, dilute the diluent to the scale and shake
well. (0.1mg/ml)
Sample solution:
Take this product 10mg, precision, set, 20ml bottle, add the diluent appropriate
amount, add the above positioning solution each 1ml, shake to dissolve, dilute the
diluent to the scale, shake well. (principal component 0.5mg/ml, impurity 1%)
（b）Chromatography operation
The chromatographic peaks caused by all blank solvents were identified by blank
solvent injection.
Determination of retention time of impurities by injection of sample with
impurity.
The relative retention time of known impurities and other impurities relative to the
main peak was determined by adding sample solution into the sample.
（c）test results
The determination results of single impurity solution, sample solution and added
sample solution are shown in table 3.2.S.4.3.4-7~8.
Table 3.2.S.4.3.4-1: results of a single impurity solution test
Component Retention time (minutes)

vildagliptin 5.270
impurityA 1.451
impurityB 3.217
impurityC 3.342
impurityD 3.874
impurityE 5.974
212
impurityF 7.440
impurityG 3.210

Degree
Trailing
of Theoretical
Peak attribution Rt(min) Area factor RRT
separati tray#
on
impurityA 1.443 43.82317 1.25 -- 656 0.27
impurityB 3.216 80.89339 1.03 9.90 9501 0.61
impurityC 3.356 25.65315 0.91 1.15 14087 0.64
impurityD 3.861 34.93119 1.06 3.81 10359 0.73
2390.350
vildagliptin 5.257 1.23 10.43 33168 1.00
34
impurityE 5.960 34.63648 0.92 6.94 75777 1.13
impurityF 7.442 45.20078 1.00 14.95 71477 1.42
（d）conclusion
The blank solvent has no interference at the peak peak position, and the separation
degree between the impurity and the main peak in the added sample solution meets
the requirements.
（2）Coercive degradation test
（a） Under the conditions of acid, alkali, high temperature, oxidation and light, the
forced degradation test of this product was carried out to investigate the
separation of the degradation products from the main peak, and the purity of
the main peak was investigated.
（b） preparation of sample solution
Diluent:
0.2% (v/v) hydrochloric acid solution acetonitrile (90:10)
Undamaged sample solution：
Take this product 125mg, precision and set, in the 25ml bottle, use diluent to
dissolve and dilute to the scale, shake well; precision take 1ml, set 10ml bottle, dilute
the diluent to the scale, shake well.
Acid-base blank solution:
213
Take 1mol/LNaOH solution 0.9ml and 1mol/LHCl solution 1ml respectively, and
place it in the same 10ml volumetric flask, dilute the diluent to scale, shake well.
Solid light degradation solution:
Take this product 0.2g, put in the weighing bottle, put the exposure under the
ultraviolet lamp for 24 hours, the precision is called 25mg to set the 50ml bottle,
dilute the diluent to the scale, shake well.
Solid high temperature degradation solution:
Take this product 0.2g, put in a weighing bottle, at 105 C for 3 hours, take out
and put cold to room temperature, precision called 25mg 50ml bottle, diluent and
dilute to the scale, shake well.
Oxidation blank solution:
Take 3%H2O2 solution 1ml, and put it in 10ml bottle, dilute dilute to scale,
shake well.
Degrading storage liquid preparation:
Take this product 125mg, precise and set, in the 25ml bottle, dissolve and dilute
the diluent to the scale, shake well; the precision is 1ml, respectively, in the 10ml
bottle, according to the table 3.2.S.4.3.4-9, under the conditions of the compulsory
degradation.
Table 3.2.S.4.3.4-1: coercive degradation conditions
Degradation Condition of Destruction
Destruction of solution
model destruction time
1mol/L HCl solution 1ml（use
Acid
1mol/L NaOH solution 0.9ml 80℃ 2h
destruction
neutralization）
1mol/L NaOHsolution 0.9ml Immediate
Alkali room
（use 1mol/L HCl solution neutralizatio
destruction temperature
1mlneutralization） n
High
temperature NO 80℃ 2h
damage
Under the
Illumination
NO ultraviolet 2h
damage
lamp
214
Oxidative
3% H2O2 solution 1ml 80℃ 3h
damage
（b）Chromatography operation
The PDA detector was used to determine the sample solution and the purity of the
peak was determined according to the content detection method.
（c）test results
The results of the forced degradation test are shown in table 3.2.S.4.3.4-10.
Table 3.2.S.4.3.4-1: results of vildagliptin forced degradation test

The degree of
separation
Degradation Main peak
Degradation between the Main peak
of main peak purity
model main peak purity angle
（%） threshold
and the
adjacent peak
Undegraded -- -- 5.405 12.839
Acid
0.47 10.37 8.279 21.458
degradation
Alkali
6.57 9.75 7.673 20.461
degradation
High
temperature 0.25 -- 5.884 13.050
degradation
Photoperiod
1.12 -- 6.010 13.601
degradation
Oxidative
0.48 1.69 72.565 90.000
degradation
Solid
photoperiod 0.33 -- 6.252 13.476
degradation
High
0.17 -- 7.747 13.385
temperature
215
degradation
of solid
Note: the peak purity angle is smaller than the purity threshold, which means that the peak purity
meets the requirements.
（d）conclusion
The degradation peak is far away from the main peak, and the peak purity of the main
peak is in accordance with the requirements under various conditions; the impurities
produced by the oxidation degradation are close to the main peak, so the oxidation
degradation is used as the system inspection solution.
3.2.S.4.3.4.3.3 linear
（1）Preparation of sample solution
Linear storage liquid:
Take this product 250mg, precision weighing, set 50ml volume bottle, add
diluent, shake shake to dissolve, dilute dilute to scale, shake well (5mg/ml).
Linear sample solution:
According to table 3.2.S.4.3.4-11, the linear reserve liquid is diluted to dilute
diluent to different concentrations, mixed and labeled.
Table 3.2.S.4.3.4-1: preparation of linear sample solution
Linear Volume extraction of The percentage of
Volume of bottle
sample linear storage liquid working
（ml）
solution volume（ml） concentration（%）
L1 2 100 20
L2 5 100 50
L3 5 50 100
L4 3 20 150
L5 5 25 200
（2）operation
The linear sample solution was determined according to the content determination
method.
（3）test results
vildagliptin's area was used for linear regression analysis, and the slope, Y
intercept and correlation coefficient were calculated. The results are shown in table
3.2.S.4.3.4-11.
216
Table 3.2.S.4.3.4-1: vildagliptin's linear survey results
Linear investigation Concentration of Vildagliptin peak
solution vildagliptin(μg/ml) area
L1 0.100 424409
L2 0.250 1034188
L3 0.501 2028835
L4 0.751 3007289
L5 1.002 4004880
linear equation y=3962930.777x+35569.122
correlation coefficient R2=1.000
Table 3.2.S.4.3.5.3.3-1:3.2.S.4.3.5.3.3-1: vildagliptin's content linear relationship

curve
（4）conclusion
In the concentration range, the response value and the concentration of vegludin
were linear.
（a）operation
A test technician took the same batch of wickludin to prepare 6 samples of the test
solution in parallel and determined the content according to the content.
（b）test results
The results of the determination of 6 samples were shown in table 3.2.S.4.3.4-13.
217
Table 3.2.S.4.3.4-1: reproducible test results for sample solution
Main peak Average peak Average
sample Assaying（%） RSD%
area area content
sample1-
2365418
1
2365050 100.24
sample1-
2364682
2
sample2-
2253213
1
2250877 100.15
sample2-
2248541
2
sample3-
2255490
1
2256401 100.27
sample3-
2257312
2
100.0 0.29
sample4-
2207432
1
2205735 99.71
sample4-
2204038
2
sample5-
2181330
1
2192247 99.58
sample5-
2203163
2
sample6-
2197692
1
2197780 100.11
sample6-
2197868
2
（c）conclusion
The results showed that the repeatability of the content determination method met
the requirements.
（2）Intermediate precision
（a）operation
The other testers were prepared at different times with the same batch of
218
vildagliptin for 6 sample solutions, measured in accordance with the content
determination method, and each sample solution was measured two times.
（b）test results
The results of the determination of 6 samples were shown in table
3.2.S.4.3.4-14~15.
Table 3.2.S.4.3.4-1: results of intermediate precision test for sample solution
Main peak Average peak Average
sample Assaying（%） RSD%
area area content
sample1-
2066115
1
2069163 100.48
sample1-
2072210
2
sample2-
2063438
1
2050332 99.74
sample2-
2037226
2
sample3-
2082383
1
2080779 99.79
sample3-
2079175
2
99.9 0.33
sample4-
2083335
1
2081150 99.72
sample4-
2078965
2
sample5-
2080377
1
2078351 99.67
sample5-
2076325
2
sample6-
2078226
1
2059497 100.23
sample6-
2040768
2
Table 3.2.S.4.3.4-1: comparison of intermediate precision and repeatability test results
219
Test group I II
Date of analysis 2014-7-13 2014-7-14
Mean value of 6 sample
100.0 99.9
content determination（%）
Determination of
0.1
difference（%）
（c）conclusion
The results showed that the repeatability and intermediate precision of the method
were all in line with the requirements.
（1）Operation
Preparation of reference solution and sample solution of 1 copies, respectively, at
room temperature for a period of time, then determined according to the content
determination method.
The recoveries (%) of the main peak area in the chromatogram of reference
solution and sample solution at each time point were calculated.
（2）Test results
The results of the test are shown in table 3.2.S.4.3.4-16~17, and the
chromatogram of the sample solution is shown in table 3.2.S.4.3.4-66~69.
Table 3.2.S.4.3.4-1: stability test results of reference substance solution
Main peak area of the control
Time （h） Relative 0 - time recovery %
solution
0 2296817 --
11 98.84
24 2264678 98.60
Table 3.2.S.4.3.4-1: stability test results for sample solution

Main peak area of the sample
Time （h） Relative 0 - time recovery %
solution
0 2069163 --
11 99.84
23 2057724 99.45
（3）Conclusion
220
The reference solution remained stable at room temperature for 23 hours at
room temperature.
（1）Operation
In order to investigate the measurement conditions with small changes, the
measurement results are not affected, and the content determination method is
adjusted properly, and the oxidation degradation solution, the test product solution and
the control solution are used for sampling. The list of investigation conditions is
shown in table 3.2.S.4.3.4-18.
Table 3.2.S.4.3.4-18: durability test conditions
column
Flow speed
Test temperature Mobile phase
（ml/min）
（℃）
Initial 35℃ 1.8 Based on the test method

Test 1 35℃ 1.6 Based on the test method

Test 7 35℃ 1.8 Buffer salt pH value 6.7
Test 8 35℃ 1.8 Buffer salt pH value6.3
Test 9 Buffer salt -water- acetonitrile- methanol
35℃ 1.8
(400:600:13:13)
Test 10 Buffer salt -water- acetonitrile- methanol
35℃ 1.8
(400:600:17:17)
Test 11 Replacement chromatography column
35℃ 1.8 （ Welch Ultimate® XB-C18，
4.6mm×50mm，5μm）
（2）Test results
221
Table 3.2.S.4.3.4-18: results of durability test results of sample solution
Test Content（%）
Initial 100.0
Test 1 100.1
Test 2 100.5
Test 3 100.1
Test 4 100.6
Test 5 99.7
Test 6 100.2
Test 7 100.9
Test 8 101.3
Test 9 99.4
Test 10 99.6
Test 11 99.5
Table 3.2.S.4.3.4-18: results of durability study: the peak of oxidation and the
separation of the main peak adjacent to the main peak.
Test Degree of separation
Initial 1.69
Test 1 1.70
Test2 1.65
Test3 1.55
Test4 1.65
Test5 1.64
Test6 1.62
Test7 3.88
Test8 1.56
Test9 1.81
Test10 1.74
Test11 1.81
（3）Conclusion
When the flow rate is adjusted for 0.2ml/ minutes, the buffer liquid pH is
adjusted to 0.2, the organic phase ratio of the mobile phase is adjusted to 2, the
222
column temperature is adjusted to 5 degrees C, the detection wavelength is adjusted to
2nm. After the replacement of the chromatographic column, the range of content
determination is between 98%~102%, the separation degree between the oxidation
degradation impurity and the main peak is more than 1.5, so, It meets the
requirements of durability.
3.2.S.4.3.4.4 Sample test results
vildagliptin's content test results are shown in table 3.2.S.4.3.4-20.
Table 3.2.S.4.3.4-18: results of vildagliptin's content test
Sample information Batches Content（%）
150601 99.9
Verification batch 150602 99.7
150603 99.7
Listed products
S0020 99.8
(preparations)
3.2.S.4.3.5 Verification of content determination by potentiometric titration
The results of verification of the method for determination of content are

summarized in table 3.2.S.4.3.5-1.
Table 3.2.S.4.3.5-1: results of validation of the method for determination of content
Titration The content of the product was determined by nonaqueous potentiometric
curve titration, and the titration end point of the potentiometric titration curve was
obvious.
Specificity As each test is corrected by blank test, no interference of blank solvent is
necessary.
linear In the range of investigation (151.03mg~400.51mg), the consumption of titrin
titrating liquid was linear with the sample size, and the linear regression equation
was y=0.0324x+0.2196, and the correlation coefficient R = 0.9999.
Accuracy The recovery rate of this product is 99%~101%, and RSD% is 0.16%, which
meets the requirements.
Repeatability The average content of 6 samples was 99.7% and RSD was 0.12%, which met the
requirements.
Intermediate The average content of 6 samples was 99.8%, RSD was 0.10%, the difference
precision between repeatability and intermediate precision test results was 0.1%, which
223
accords with the verification requirements.
Solution The solution is stable at room temperature for 4 hours.
stability

Instrument：
Electronic balance Manufacturer: Shanghai Jinghai Instrument Co., Ltd.
type：FA1004N
Potentiometric Titrant Manufacturer : Shanghai Precision Scientific
Instruments Co., Ltd. type：ZDJ-4A
Sample：
vildagliptin batch：150601 source：Anhui Hai Kang Pharmaceutical Co.,
Ltd.
Ltd.
Ltd.
3.2.S.4.3.5.2 Method verification

3.2.S.4.3.5.2.1 Drawing of titration curve
（1）Operation
Blank operation:
Glacial acetic acid 25ml, acetic anhydride 25ml, titration with potentiometric
titration, and perchloric acid titration solution (0.1mol/L) titration.
Trial product operation：
This product is about 0.25g, precision named, icing acetic acid 25ml, acetic anhydride
25ml, shake to dissolve, potentiometric titration, perchloric acid titrate (0.1mol/L)
titration, record titration volume (V) and corresponding E (MV) values.
（2）Test results
The test results are shown in table 3.2.S.4.3.5-2~3, and the titration curve is
shown in table 3.2.S.4.3.5-1~3.
Table 3.2.S.4.3.5-1: vildagliptin content determination titration curve data
V E ΔV ΔE ΔE/ΔV △2E/△V2
224
0.000 284.0 -- -- --
3.000 301.5 3.000 17.5 5.8
5.019 317.7 2.019 16.2 8.0 1.1
7.018 348.2 1.999 30.5 15.3 3.6
7.219 354.4 0.201 6.2 30.8 77.6
7.419 362.1 0.200 7.7 38.5 38.3
7.620 372.9 0.201 10.8 53.7 75.8
7.821 391.7 0.201 18.8 93.5 198.0
8.022 459.2 0.201 67.5 335.8 1205.4
8.223 668.0 0.201 208.8 1038.8 3497.4
8.302 682.2 0.079 14.2 179.7 -10874.2
8.382 690.6 0.080 8.4 105.0 -934.3
8.521 700.3 0.139 9.7 69.8 -253.4
8.720 709.5 0.199 9.2 46.2 -118.4
9.118 720.0 0.398 10.5 26.4 -49.9
9.417 725.4 0.299 5.4 18.1 -27.8
10.510 735.3 1.093 9.9 9.1 -8.2
11.803 740.0 1.293 4.7 3.6 -4.2
12.500 741.4 0.697 1.4 2.0 -2.3
13.096 742.0 0.596 0.6 1.0 -1.7
Table 3.2.S.4.3.5-1: vildagliptin content determination titration data processing

V0 mean Content
V（ml） C（mol/L） W（mg） Water （%）
value（ml）（%）
0.179 8.223 0.1010 247.4 0.09 99.72
Table 3.2.S.4.3.5-1：Titration curve -0 - order curve for the determination of vildagliptin

content
225

content

content
（3）Conclusion
The content of the product was determined by nonaqueous potentiometric
titration, and the titration end point of the potentiometric titration curve was obvious.
As each test is corrected by blank test, no interference of blank solvent is
necessary.
3.2.S.4.3.5.2.3 linear
The concentration range of linearity is 60% to 160% of the working
concentration.
（1） Operation
226
150mg, 200mg, 250mg, 300mg and 400mg were precise weighed, with glacial
acetic acid 25ml, acetic anhydride 25ml to dissolve, potentiometric titration, titration
with perchloric acid titrant (0.1mol/L), and the results of titration were corrected by
blank test. The volume of the corrected volume (ML) is used to map vildagliptin’s
injection volumn (mg).
（2） Test results
Table 3.2.S.4.3.5-1: linear test results of vildagliptin content determination
The
Actual percentage
Titrant
consumpt equivalent to
W（mg） concentrati V0（ml） V1（ml） Content（%）
ion V the working
on（mol/L)
（ml） concentration
（%）
151.03 5.107 4.928 60 100.08
200.55 6.694 6.515 80 99.64
250.31 0.1010 0.179 8.374 8.195 100 100.42
300.02 9.965 9.786 120 100.04
400.51 13.183 13.004 160 99.58
Table 3.2.S.4.3.5-4：The linear relationship curve of the content determination of

vildagliptin
227
（3） Conclusion
In the concentration range, the consumption of perchloric acid titration solution
(0.1mol/L) is linearly related to the sample size.
A test was carried out in parallel with 6 batches of the same batch of vildaliptin.
The results were shown in table 3.2.S.4.3.5-5.
Water of
Titrant
Determ Trial V0
W（mg） V（ml） concentration Content（%）
ination product S （ml）
C（mol/L）
（%）
1 251.21 8.369 99.86
2 249.95 8.323 99.80
3 247.84 8.231 99.51
0.1010 0.09 0.190
4 250.55 8.332 99.67
5 251.11 8.355 99.73
6 249.08 8.289 99.73
Mean
-- -- -- -- -- 99.7
value
RSD% -- -- -- -- -- 0.12
（c） Intermediate precision
Another analyst, in parallel with the French method, used the same batch of
Vildagliptin for 6 times on different days, and the results were shown in table
3.2.S.4.3.5-6~7.
Table 3.2.S.4.3.5-1: results of intermediate precision test
Titrant
Water of
concentrati Content
Determination W（g） V（ml） Trial product V0（ml）
onC （%）
S（%）
（mol/L）
1 244.90 8.157 0.1010 0.09 0.179 99.92
228
2 250.64 8.332 99.77
3 243.19 8.079 99.63
4 252.84 8.409 99.83
5 246.17 8.189 99.80
6 245.08 8.147 99.72
Mean value -- --- -- -- -- 99.8
RSD% -- -- -- -- -- 0.10
Table 3.2.S.4.3.5-1: comparison of intermediate precision and repeatability test results

Mean value of Content
Test Date of analysis content determination
determination（%） difference（%）
Repeatability 2014-4-18 99.7
Intermediate 0.1
2014-4-19 99.8
precision
（2） Conclusion
The results showed that the repeatability and intermediate precision of the method
were all in line with the requirements.
3.2.S.4.3.5.2.5 Accuracy (recovery)
（1）Operation
The amount of the product was weighed separately and determined according to
the content determination method.
Table 3.2.S.4.3.5-1: the sample size of the recoverable sample
Number Nominal Quantity（mg）
1 199.28
2 200.42
3 202.44
4 251.17
5 254.34
6 249.28
7 300.47
8 298.94
229
9 302.48
Remarks: the water content of these tablets was 0.09 and the content was 99.7% (the
mean value of the repeatability test results).
（2）Test result
Table 3.2.S.4.3.5-1: recovery test results
Amount of Real
Rate of recovery
Level V0（ml） V（ml） addition measurement
（%）
（mg）（mg）
6.624 198.54 198.32 99.89
80% 6.681 199.67 200.07 100.20
6.733 201.68 201.66 99.99
8.347 250.23 251.12 100.36
100% 0.152 8.419 253.39 253.33 99.98
8.263 248.35 248.55 100.08
9.944 299.35 300.06 100.24
120% 9.877 297.82 298.01 100.06
9.978 301.35 301.10 99.92
Mean value -- -- -- -- 100.08
RSD% -- -- -- -- 0.16
（3）Conclusion
According to the experimental results, the accuracy of the content determination
method meets the requirements.
3.2.S.4.3.5.2.6 Stability of sample solution
（1）Operation
3 sample solutions were prepared and placed at room temperature for a period of time.
The difference between the content of WLG and the initial sample measured at each
time point was calculated.
（2）Test results
The stability of the sample solution is shown in table 3.2.S.4.3.5-10.
Table 3.2.S.4.3.5-1: stability test results of sample solution
230
Storage Difference of
V0（ml） V（ml） W（mg） Assaying（%）
time content （%）
0h 8.329 250.57 99.75 --
2h 0.180 8.301 249.43 99.86 0.11
4h 8.274 248.76 99.80 0.05
（3）Conclusion
The solution remained stable at room temperature for 4 hours.
The results of the test of vildagliptin are shown in table 3.2.S.4.3.5-11.
Table 3.2.S.4.3.5-1: Determination of the content of vildagliptin.
Content determination result
Sample Information Batches
（%）
150601 99.9
150603 99.8
3.2.S.4.3.6 Verification of Ethyl Chloroacetate Method

Chloroacetic chloride and absolute alcohol were used in the synthetic process，
Chloroacetic chloride is unstable. When water becomes chloroacetic acid,
chloroacetic acid can react with ethanol to form ethyl chloroacetate.
Chloroacetic acid, toxic, half lethal dose (rat through the mouth) 76mg/kg, the
maximum daily dose of 100mg, reference ICH Guideline For Residual Solvents Q3C
(R5) daily maximum intake of intake calculation formula:
Formula: PDE: maximum daily intake

Weight adjustment: default 50kg
F1：Conversion coefficients between species (deduced from the rat dose,
the coefficient of human dose is 5).
F2：The coefficient of variation among individuals, the value of which is
10
F3：Short term toxicity related variability coefficient, upper limit value 10
F4：The coefficient associated with the production of severe toxicity, the
231
upper limit of 10
F5 ： If conversion coefficient is not established when NOEL is not
established, if it is only LOEL data, the value may be 10.
In the formula, 70kg is the average weight of the human body.

Chloroacetic acid LD50:76mg/kg (rat via the mouth)
NOEL=76×70/2000=2.66mg/day
PDE=2.66×50/（5×10×10×10×10）=0.00266mg/day
Limit (%) =PDE/ maximum daily dose =0.00266/100 * 100%=0.003%
The limit of Ethyl Chloroacetate should not be controlled by 15ppm to control
the quality of the product. The company entrusts Beijing Beijing medical science and
Technology Co., Ltd. to control the residue of Ethyl Chloroacetate in this product and
to verify the method.
The method validation results are outlined in table 3.2.S.4.3.6-1.
Table 3.2.S.4.3.6-1: summary of method validation results
Specificity The blank solvent did not interfere with the determination of ethyl
chloroacetate, and the separation degree between the peaks was
greater than 1.5, which met the requirements.
Detection line The limit of quantification of Ethyl Chloroacetate was 300ng/ml,

and quantitative and the detection limit was 100ng/ml.
limit
3.2.S.4.3.6.1Sample test results

The results of the test of vildagliptin are shown in table 3.2.S.4.3.6-2.
Table 3.2.S.4.3.6-1: test results of vildagliptin
Results of determination
Sample Information Batches
（%）
150601 ND
150603 ND
232
3.2.S.4.3.7 Impurity I
According to the synthetic route, the 3- amino -1- amantadine reacts with
chloroacetic acid to form impurity I. The structure of the impurity I shows that it has
almost no UV absorption. The maximum daily dose of this product is 100mg.
According to the guiding principles of chemical drug impurity research technology,
the identification limit is 0.1% and the quality control limit is 0.15%. In order to better
control the quality of the product, the limit is not over 0.1%. The reference substance
was prepared according to the limit 0.1%. When the sample was injected under the
condition of residual solvent, the boiling point was initially judged to be high and the
gas phase could not be determined. The residue of impurity I in the product was
detected by HPLC-MS method, and the method was verified by the method.
The method validation results are outlined in table 3.2.S.4.3.7-1.

Table 3.2.S.4.3.7-1: summary of method validation results
Specificity The blank solvent did not interfere with the determination of
impurity I, and the separation degree between the peaks was
greater than 1.5, which met the requirements.
Detection line The limit of quantification for impurity I is 1ng/ml, and the
and quantitative detection limit is 0.5ng/ml.
limit

The results of the test of wig are shown in table 3.2.S.4.3.7-2.
Table 3.2.S.4.3.7-1: test results of vildagliptin
Sample information batches Results of determination
233
（%）
150601 ND
150603 ND
3.2.S.4.3.8 Verification of trifluoroacetic acid method

Trifluoroacetic anhydride is used in the synthetic process of this product, and
trifluoroacetic anhydride reacts with water to form trifluoroacetic acid. 。 Reference
ICH Guideline For Residual Solvents Q3C(R5) Daily Acceptable Maximum Intake
Calculation Formula：
In the formula：PDE：Daily Acceptable Maximum Intake

Weight adjustment：Default 50kg
F1：Conversion coefficient between species（The coefficient of human dose
calculated from rat dose was 5）
F2：Coefficient of variation between individuals，the value is 10
F3：Short-term toxicity-related coefficient of variation，Upper limit value 10
F4：Coefficient related to the production of severe toxicity，Upper limit value 10
F5：Conversion factor used when NOEL is not established，If only LOEL data，
Its value may be an upper limit of 10
In the formula: 70kg as the people’s average weight。

Due to the MSDS，trifluoroacetic acid LD50：200mg/kg（Rat oral）
NOEL=200×70/2000=7mg/day
Due to the MSDS，Trifluoroacetic acid has no serious toxicity，F4 is set to 1，
F2, F3 and F5 are set to 10
PDE=7×50/（5×10×10×1×10）=0.07mg/day
Limited （%）=PDE/Maximum daily dose=0.07/100×100%=0.07%
Therefore ，The limit of trifluoroacetic acid is set at 0.07%。
The solubility of this product is the largest in methanol ， about 500mg/ml ，
however , Trifluoroacetic acid is unstable with methanol as a solvent，Replaced with
234
dim-ethyl sulfoxide as solvent，This product is dissolved in dim-ethyl sulphide up to
200mg/ml，The preparation of trifluoroacetic acid as a control solution，no peak in
the control solution.So FID detector cannot accurately determine the residual amount
of trifluoroacetic acid in this product. Our company commissioned Beijing Ming Bo
Fei Technology Co., Ltd.，used the Ion chromatography，Determination of residues of
the trifluoroacetic acid in this product，And the method is verified by method.
Trifluoroacetic acid method validation results summary as the chart show
3.2.S.4.3.8-1.
Table 3.2.S.4.3.8-1：Trifluoroacetic acid method validation results summary
Project Validation results
Specificity Blank solvent does not interfere with the determination of
trifluoroacetic acid；The degree of separation between each peak
is greater than 1.5，Comply with the regulations.
Linear Within the measured concentration range（0.56μg/ml~3.38
μg/ml），Regression equation of the Trifluoroacetic Acid is
y=0.079x-0.003，R2=0.998，Linear relationship between response
value and concentration。
Detection The limit of quantitation of trifluoroacetic acid is 0.56 μ
line and The limit g/ml，The detection limit was 0.17 μg/ml.
of implantation
Recovery On the three levels ，the average recovery of trifluoroacetic
rate acid is 105.39%，RSD% is 3.54%，Less than 10%, in line with
verification standards。
Durability When the flow rate is ± 0.1ml per minute,Mobile phase
concentration adjustment ± 0.5mmol/L ， The recoveries of
trifluoroacetic acid are all 80% to 120% and meet the durability
requirements.

The test of Vildagliptin show in table 3.2.S.4.3.8-2
Table 3.2.S.4.3.8-1：Test results of vildagliptin sample
Sample information Batch Results of determination
235
（%）
150601 ND
150603 ND
3.2.S.4.3.9 Hygroscopicity
A summary of the results of the hygroscopicity survey is given in Table 3.2.S.4.3.9-1.
Table 3.2.S.4.3.9-1：Hygroscopicity ‘s analysis method validation
Project Validation results
Hygroscopicity This product is no Hygroscopicity。
3.2.S.4.3.9. 1 Instrument and test drug

Balance：FA1004N manufacture ：Shanghai Jinghai Instrument Co., Ltd.
Vildagliptin：Batch：150601、150602、150603
3.2.S.4.3.9.2 Results
Following the instructions for hygroscopicity，Inspected batches of small test
samples and validation batch samples.
The test results are shown in Table 3.2.S.4.3.9-2。
Table 3.2.S.4.3.9-1：Hygroscopicity test result
Sample Sample W1 Weight Average value

Sample information W2（g） W3（g）
batch number （g） gain（%）（%）
30.22
1 31.1878 31.1887 0.094
78
150601 0.090
28.65
2 29.7027 29.7036 0.086
78
30.68
1 31.6328 31.6336 0.085
68
32.15
2 33.1633 33.1642 0.089
66
29.69
1 30.6620 30.6628 0.082
16
150603 0.085
31.24
2 32.3956 32.3966 0.087
86
236
Conclusion ：This product no hygroscopicity
3.2.S.4.5 The basis of quality standard formulation

3.2.S.4.5.1 Character
Reference to information Vildagliptin issued by EMEA（Vildagliptin is a white to
slightly yellowish or slightly greyish crystalline powder and no polymorphs）and the
Results of multiple batches of small samples and verified samples，and the stability
test sample trait test results，Formulated as a white to slightly yellow or slightly gray
crystalline powder. Through the X-ray diffraction inspection，This product has crystal
shape，and it is obviously crystallinity。Therefore ，The trait standard for this product
is defined as white to yellowish or slightly gray crystalline powder。
Reference to the Chinese Pharmacopoeia 2010 Edition, Part II Annex XIX J ，
examine the hygroscopicity of this product，it’s no hygroscopicity of the product。
3.2.S.4.5.2 Solubility
Reference to information Vildagliptin issued by EMEA （ Vildagliptin is
non-hygroscopic and freely soluble in water and polar organic solvents），This product
is soluble in water or polar organic solvents.Organic solvent used in the mobile phase
of this product is methyl alcohol、Ethanol、acetonitrile；The medium for dissolution
measurement is 0.01 mol/L hydrochloric acid ；The solvent for the determination of
residual solvent is dimethyl sulfoxide；The organic solvent used for recrystallization is
isopropyl alcohol，so the Solubility investigation of the Organic solvents is the methyl
alcohol、Ethanol、iso-Propyl alcohol 、acetonitrile、Dimethyl sulfoxide(DMSO).
Through the more batch of the sample testing ，The limitation of solubility is ：
This product is soluble in methanol, ethanol, dimethyl sulfoxide, water or 0.01mol/L
hydrochloric acid solution, slightly soluble in acetonitrile or isopropanol.
3.2.S.4.5.3 Melting Point

Taking the product ,according to the Melting point method （ Chinese
Pharmacopoeia 2010 Edition II Appendix IIIC）testing，the result of the chart show
3.2.S.4.5-1，so making this product ‘melting point is 146℃~151℃.
Table 3.2.S.4.5-1：Melting point measurement results
237
Sample information batches Melting point
150301 147.0℃~148.0℃
Small test 150302 147.5℃~148.5℃
150303 146.5℃~147.5℃
150601 147.5℃~148.5℃
Verification batch 150602 146.5℃~147.5℃
150603 146.5℃~147.5℃
150601 147.0℃~148.5℃
Accelerated in June 150602 146.0℃~147.5℃
150603 147.0℃~148.5℃
3.2.S.4.5.4 Specific rotation

Using methanol as solvent to measure the optical rotation of lots of samples of
this product ， Concentration is 97mg/ml ， Optical rotation readings are around
-8.5 ° .Based on the results of multiple batches and stability measurements ， The
specific rotation of this product is -84° to -90°。The specific rotation results are
shown in Table 3.2.S.4.5-2.
Table 3.2.S.4.5-1: results of specific rotation test

Sample optical specific
Batch
information rotation rotation
150301 -8.42° -86.92°
Small test 150302 -8.46° -87.12°
150303 -8.49° -87.53°
150601 -8.40° -86.70°

Verification
150602 -8.46° -87.31°
batch
150603 -8.44° -87.22°
Accelerated in 150601 -8.43° -87.09°

June 150602 -8.39° -86.56°
238
150603 -8.39° -86.58°
3.2.S.4.5.5 Identification
（1）HPLC method
Identification by HPLC method，the standard is：In the chromatogram recorded
under the determination, the retention time of the main peak of the test solution is the
same as the retention time of the main peak of the reference solution.
（2）IR method
Using IR method to identify，the standard is ：This product's infrared absorption
spectrum should be consistent with the reference map
3.2.S.4.5.6 Moisture
This product is soluble in methanol，so it’s suitable for determining the moisture
content of the product with Fisher's moisture method。Due to the more batch of small
testing and the verification batch sample ‘s moisture measurement results
show :Water content less than 0.5%，so the moisture limit of this product is not over
0.5%。
More batch of the sample testing result as the chart show 3.2.S.4.5-3。
Table 3.2.S.4.5-1：Moisture measurement results
Sample information Batch Moisture （%）

150301 0.09
Small test 150302 0.10
150303 0.10
150601 0.10
150603 0.08
150601 0.12
Accelerated in June 150602 0.10
150603 0.12
3.2.S.4.5.7 Sulfate
It ‘s used Sodium sulfate in the Synthetic process of the product, therefore
making the standard of sulphate examination：no more than 0.02%. The test method is
239
the same as the Chinese Pharmacopoeia 2010 edition II Appendix VIII B 。 The
company's multiple batches of small test and verification batch samples meet this
requirement.
3.2.S.4.5.8 Chloride
Chloroacetyl chloride has been used in the synthesis of this product.，Therefore, a
chloride inspection standard was established：no more than 0.02%。The testing method
is the same as the Chinese Pharmacopoeia 2010 edition II Appendix VIII A 。 The
company's multiple batches of small test and verification batch samples meet this
requirement.
3.2.S.4.5.9 Related substances
According to the synthetic route of this product is known Possible impurities
include L-prolinamide, 3-amino-1-adamantanol (starting material 2), impurity A
(VGT-1), impurity B (VGT-2/starting material 1), impurities C (cyclic quin-one),
impurity D (synthetic amide), impurity E (dike-tone piperazine), impurity F
(disubstituted), impurity G (carboxylic acid metabolite)。
Original material 2 is no UV rays to be absorbed ，can not be detected with the
relevant material method of this product. So the product is controlled when residual
solvent is detected.；impurities A、impurities B、impurities F as the process impurities；
impurities C、D、E is the Process impurities and it also is degradation impurities。
Our company established HPLC method to test the related impurities of the
Vildagliptin
Detection of related substances in vildagliptin，Chromatographic conditions are shown
in Table 3.2.S.4.5-4
There are 1 chiral centers in the molecular structure of this product，our company
has made its own enantiomers and established a method for the detection of isomers
using chiral columns.The maximum daily dose of this product is 100mg，According to
"Technical guidelines for chemical drug impurity research"，the identification limit is
0.1% ， the limit of quality control is 0.15% ， So a single specific impurity limit is
0.15%，A single unknown impurity limit is not to exceed 0.10%，the total impurity
limit is 0.5%. The method of testing the related substances established by this method
is verified by method，It proved that have a good specificity, linear range, sensitivity,
precision, accuracy and durability.。
240
The chromatographic conditions for the material are shown in table3.2.S.4.5-4.
Table 3.2.S.4.5-1：Related material chromatographic conditions
Reference standard method（Vildagliptin

parameter Establish method
Pill ）
Phosphate Buffered Saline[taking Potassium
Phosphate Monobasic 1.3g，add water 1000ml to
Phosphate Buffered Saline（take Sodium
dissolve，used the Potassium hydrogen
phosphate dibasic dihydrate 3.12g，Dissolve with
phosphate anhydrous solution （taking
Mobile phaseA 1000ml of water，Add triethylamine 1ml，mixing，
Potassium hydrogen phosphate anhydrous 1.5g，
Using phosphoric acid to regulate pH value to 2.8) -
add water 10ml to dissolved）adjust pH to 6.5]-
acetonitrile (96:4)
water- acetonitrile- methyl alcohol
（400:600:15:15）
Phosphate Buffered Saline（Take sodium Phosphate Buffered Salin-
dihydrogen phosphate dihydrate 3.12g，Dissolve acetonitrile -methyl alcohol（400:450:150）
Mobile phaseB with 1000ml of water，add Triethylamine 1ml，

mixing，Using phosphoric acid to regulate pH value
to 2.8) - acetonitrile (75:25)
mobile mobile phase
Time（minute）
mobile phase mobile phase phase A（%） B（%）
Time（minute）
A（%） B（%） 0.0 100 0
0.0 100 0 1.0 100 0
Gradient 25.0 100 0 3.0 90 10

procedure 50.0 0 100 8.0 30 70
50.01 100 0 8.1 100 0
65 100 0 11.0 100 0
chromatographic
Agilent Extend C18（250×4.6mm，5µm） Waters XTerra MS C18，50×4.6mm，3.5μm
column
1.0ml 1.8ml
Velocity of flow
Column
35℃ 35℃
temperature
Detection
210nm 210nm
wavelength
241
Solution
concentration of 2.5mg/ml 0.5mg/ml
the sample
20 mu l of system suitability solution ， Injection
liquid chromatography The chromatograms were
recorded. The order of peaks was impurity A, D, C,
and G.、Vildagliptin, B, F, E, impurity G and main 209-01 （ impurity D ） peak and 202-01peak
peak separation degree greater than 0.5 ， The （ impurity C ） ’s resolution should not be less
degree of separation between other impurities and than 2.5 ， Vildagliptin peak ‘s Trailing factor
System suitability main peaks and impurities should be greater than should not be greater than 1.8 ； 5 times of
1.5。Precisely take 20μl of the reference solution
continuous injection of the control solution，The
and inject it into the liquid chromatography，Record relative standard deviation of the area of the
the chromatogram ， The RSD of the continuous Vildagliptin peak should not exceed 2.0%.
6-pin main peak area must not exceed 5.0%，The
signal-to-noise ratio of the main peak should not be
less than 40。
Injection volume 20μl 10μl

Impurity
Adding principal component external standard Adding principal component external standard
quantitative
method of correction factor method of correction factor
method
Impurity A could not exceed 0.15%； 202-01（impurity D）not exceed 2.0%；
Impurity B could not exceed 0.15%； 209-01（impurity C）not exceed 0.3%；
Impurity C could not exceed 0.15%； 207-01（impurity E）not exceed 0.5%；
Impurity D could not exceed 0.15%； Other single impurities not exceed 0.2%；
Impurity E could not exceed0.15%； Total impurities not exceed 3.0%
Impurity limit
Impurity F could not exceed0.15%；
Impurity G could not exceed 0.15%；
The other single impurities could not exceed
0.1%；
Total impurities n could not exceed 0.5%
Advantages of self-designed standards relative to reference standards：

⑴Self-made standard impurity G（Carboxylic acid metabolites）The peak shape is
better than the reference standard
Self-designed standards ：0.861

Impurity G Tailing factor
Reference standards ：0.565
242
Self-design standard impurity G positioning solution results
Refer to the standard under the impurity G positioning solution results

⑵In the Self-design standard ,the degree of separation between impurity G and
impurity C is better than under the reference standard。
Self-designed system suitability results under criteria

Peak Area
Peak # Rt（min） peak area Resolution Tailing factor
ownership （%）
1 Impurity A 7.465 76065 0.205 -- 0.974
2 Impurity D 8.628 45927 0.124 2.061 0.874
3 Impurity C 10.524 48924 0.132 3.723 1.087
4 Impurity G 13.461 45389 0.122 2.582 0.848
Vildagliptin 3664471
5 14.791 98.634 0.604 6.603
8
6 Impurity B 21.422 132377 0.356 4.215 1.111
7 Impurity F 44.354 91454 0.246 44.942 0.982
243
8 Impurity E 47.617 62055 0.167 10.793 1.139
Reference standards project ‘s system suitability results

Peak Area
Peak # Rt（min） peak area Resolution Tailing factor
ownership （%）
1 Impurity A 1.423 42498 1.534 -- 0.896
2 Impurity B 3.578 75806 2.736 9.092 1.026
3 Impurity G 3.823 3820 0.138 1.254 1.257
4 Impurity C 4.094 29242 1.055 1.552 1.176
5 Impurity D 4.510 36130 1.304 2.103 1.003
6 Vildagliptin 6.132 2499358 90.201 8.785 0.962
7 Impurity E 7.096 37077 1.338 7.071 1.232
8 Impurity F 8.574 46952 1.695 11.599 1.115
244
Table 3.2.S.4.5-1：Related substances test results

Sample
Verification batch Accelerated in June
information
Batch 150601 150602 150603 150601 150602 150603
Impurity A（%） ND ND ND ND ND ND
Impurity B（%） ND ND ND ND ND ND
Impurity C（%） ND ND ND ND ND ND
Impurity D（%） 0.077 0.017 0.020 0.103 0.037 0.059
Impurity E（%） ND ND ND 0.006 0.003 0.010
Impurity F（%） ND 0.022 0.033 ND 0.019 0.026
Impurity G（%） ND ND ND ND ND ND
Maximum
unknown 0.046 0.076 0.050 0.036 0.068 0.044
impurity（%）
Total impurity
0.155 0.114 0.103 0.172 0.126 0.140
（%）
245
3.2.S.4.5.10 Residual solvent and 3-amino-1-adamantanamine
The synthesis of 3- amino -1- amantadine did not use a class of solvents.，The second
type of solvent used is dichloromethane. The solvents involved in this product are
dichloromethane, triethylamine, isopropanol, N,N-dimethylformamide, ethanol,
trifluoroacetic anhydride. So The residual solvents to be controlled in this product
are methanol, dichloromethane, triethylamine, ethanol, N,N-dimethylformamide,
isopropanol, trifluoroacetic acid 。 Among them, dichloromethane, triethylamine,
ethanol, methanol, N,N-dimethylformamide and isopropyl alcohol are used for
residual solvent detection. ， Trifluoroacetic acid was established by ion
chromatography to be controlled
Residual solvent method validated by method，The proofs all have specificity, linear
range, sensitivity, precision, accuracy and durability
Table 3.2.S.4.5-1：Residual solvent chromatographic conditions

Parameter Proposed method
Detector Hydrogen flame ionization detector
Carrier gas Nitrogen
Mainte
Heating Tempe
nance time
rate(℃/min) rature（℃）
Gradient program （minute ）
0 40 5
25 220 10
chromatographic column KB-624 30m×0.53mm， 3.00μm
Control mode Pressure control mode（30KPa）

Split ratio 10:1
Inlet temperature 250℃
Detection port temperature 280℃
Repeat injection with control solution ， record

elution profile ， The RSD of the peak area of the last 6
System suitability consecutive needles should not exceed 10.0%.The degree of
separation between the peaks of each component should
meet the requirements。
246
Impurity quantitative method External standard method
Pharmacopoeia 2010 Edition II Appendix VII P，The limit of residual solvents is

shown in table 3.2.S.4.5-7，Triethylamine limit refers to EDQM triethylamine limit，
The test results of the company's multiple batches of samples are shown in Table
3.2.S.4.5-8.
Table 3.2.S.4.5-1：Residual solvent limit
Residual solvent Limit

Methyl alcohol Not to exceed 0.3%
Ethanol Not to exceed 0.5%
Iso-Propyl alcohol Not to exceed 0.5%
Dichloromethane Not to exceed 0.06%
Triethylamine Not to exceed 0.032%
N,N-Dimethylformamide Not to exceed 0.088%
3-AMINO-1-ADAMANTANOL Not to exceed 0.1%
Table 3.2.S.4.5-1：Sample test results

Verification batch
Residual solvent
150601 150602 150603
Methyl alcohol 0.004 0.004 0.005
Ethanol 0.003 0.002 0.001
Iso-Propyl alcohol 0.105 0.103 0.102
Dichloromethane 0.005 0.004 0.008
Triethylamine ND ND ND
N,N-Dimethylformamid
ND ND ND
e
3-AMINO-1-ADAMANTA
0.015 0.015 0.015
NOL
247
3.2.S.4.5.11 Trifluoroacetic acid
This product is used in the synthesis of trifluoroacetic anhydride，Trifluoroacetic
anhydride reacts with water to give trifluoroacetic acid. Our company commissioned
Beijing Ming Bofei Technology Co., Ltd. ， The trifluoroacetic acid residue of the
product was detected by ion chromatography and the method was verified by the
method. ， it to be proven to have specificity, linear range, sensitivity, accuracy and
durability.
Table 3.2.S.4.5-1：Chromatographic conditions
Parameter Proposed method
Detector Conductivity detector
Mobile phase 8mmol/L Potassium hydroxide solution
Chromatographic
Anion chromatography column（AS19，4.0×250mm）
column
Electricity 25mA
Velocity of flow 1.0ml
Impurity quantitative
External standard method
method
limit Not to exceed 0.07%
Table 3.2.S.4.5-1:Sample test results

Verification batch
Residual solvent
150601 150602 150603
Trifluoroacetic acid ND ND ND
Trifluoroacetic acid was not detected in the 3 batches of samples of the verification
batch of this product, and it was not determined as a quality standard.
3.2.S.4.5.12 Impurity I
According to the synthetic route, 3-amino-1-adamantanol reacts with
chloroacetic acid to generate impurity I。The maximum daily dose of this product is
100mg，According to "Technical guidelines for chemical drug impurity research"，The
identification limit is 0.1% and the quality control limit is 0.15%. In order to better
control the quality of this product, the impurity I limit is set to not exceed 0.1%..Our
248
company commissioned Zhongli An Beijing Pharmaceutical Technology Co., Ltd. ，
using the HPLC-MS method to detect the residue of impurity I of this product，and
method validation for this method，Proven to have specificity and sensitivity.
Table 3.2.S.4.5-11：Sample test results

Verification batch
Residual solvent
150601 150602 150603
Impurity I ND ND ND
No impurity I was detected in the 3 batches of samples of the verification batch of the
product, and it was not determined as a quality standard.
3.2.S.4.5.13 Ethyl chloroacetate
Chloroacetyl chloride, anhydrous ethanol are used in the synthesis of this

product.，Chloroacetic chloride is unstable and becomes chloroacetic acid in water，
Chloroacetic acid and ethanol can be esterified to produce ethyl chloroacetate。
The quality of this product should be controlled by the limit of Ethyl
Chloroacetate 15ppm. Our company commissioned Zhongli An Beijing
Pharmaceutical Technology Co. Ltd.，use the GC-MS method to control the residue of
ethyl chloroacetate，And this method has been verified by the method and proved to
have specificity and sensitivity.
Table 3.2.S.5-12：Test results of vildagliptin sample
Results of determination
Sample information Batch
（%）
150601 ND
Verification Batch 150602 ND
150603 ND
249
Ethyl chloride ethyl acetate was not detected in the 3 batches of this product in the
verification batch, and it was not determined as a quality standard.
3.2.S.4.5.14 Enantiomers
There are 1 chiral centers in the molecular structure of this product，our company's
own enantiomers ， The detection method of isomers was controlled by chiral
chromatographic column. ， Chromatographic conditions are shown in Table
3.2.S.4.5-13.The method is verified by method，Proven with specificity, linear range,
sensitivity, precision, accuracy and durability.
According to the stability study results of this product, we know ， none of the
enantiomers were detected ， For details, see Table 3.2.S.4.5-13 。 In addition ，
According to "Technical guidelines for chemical drug impurity research"，The limit of
enantiomers is: no more than 0.15%.
Table 3.2.S.4.5-1：Enantiomeric chromatography conditions
Parameter Formulation method
Mobile phase Anhydrous ethanol-diethylamine（100:0.1）
Cromatographic column CHIRALPAK ®IF 4.6×250mm，5.0µm

Velocity of flow 0.5ml/min
Solution concentration of the
10mg/ml
sample
Application of system solution ， Recording a
chromatogram at the wavelength of 220nm ， The
separation of the peak of the Vildagliptin peak from
the adjacent isomer should not be less than 0.9.
System suitability
Repeat injection with control solution ， Record the
chromatogram at a wavelength of 220nm，The relative
standard deviation of the main peak area of 6
consecutive needles must not exceed 5.0%.
Impurity quantitative method Main component external standard method
Limit Not to exceed 0.15%
The test results of the company's multiple batches of samples are shown in Table
250
3.2.S.4.5-14.
Table 3.2.S.4.5-14：Enantiomeric results of a sample of vildagliptin
Sample information Batch Enantiomers（%）
150601 ND
150603 ND
150601 ND
Verification batch accelerated
150602 ND
in June
150603 ND
3.2.S.4.5.15 Ignition Residue

Draw up the standard of burning residue：Take the product 1.0g，According to the
Chinese Pharmacopoeia 2010 Edition II Appendix VIII N Determination ， Residual
residue must not exceed 0.1%. The results of the inspection of multiple batches of
samples are shown in Table 3.2.S.4.5-15.
Table 3.2.S.4.5-1：Results of Ignition Residue Determination of Vildagliptin Sample
Sample information Batch Residue on ignition（%）

150601 0.06
150603 0.06
3.2.S.4.5.16 Heavy Metal

Draw up the standard of heavy metal：Residual residue from burning residue，
According to the Chinese Pharmacopoeia 2010 Edition II Appendix VIII H second
method，the limit is not to exceed twenty millionths。The company's multiple batches
of small test and verification batch samples meet this requirement.
3.2.S.4.5.17 Determination of content（potentiometric titration）
Taking potentiometric titration，Establish a method for determining the content of
this product：Take this product about 0.25g，Precision weighed，add glacial acetic acid
25ml，acetic anhydride 25ml，Shaking to dissolve，According to the potentiometric
titration method (Appendix VII A of the Chinese Pharmacopoeia 2010 Edition Part II),
251
titrate with perchloric acid titrant (0.1mol/L) and correct the results of the titration
with a blank test. ， Each 1 ml of perchloric acid titrant (0.1 mol/L) corresponds to
30.34 mg of C17H25N3O2.
The method was validated by the method and proved to have specificity,
precision, accuracy and solution stability。The company did not use this method as the
content determination method in this product standard, but used this method as the
calibration method for the vildagliptin working standard.
Table 3.2.S.4.5-1：Test results of multiple batch of samples
Sample information Batch Content （%）

150601 99.9
150603 99.8
3.2.S.4.5.18 Determination of content（HPLC method ）

Determine the content of the product by referring to the method for determining
the content of the vildagliptin tablet import registration standard (standard number:
JX20100243). ， Chromatographic conditions are shown in Table 3.2.S.4.5-17 ， The
method has been verified by the method and proved to have good specificity, linear
range, sensitivity, precision, accuracy and durability.
Table 3.2.S.4.5-1：Determination of chromatographic conditions

Parameter Proposed method Reference standard method
A Phase ： Phosphate Buffered Saline[Take potassium
dihydrogen phosphate 1.3g, add water 1000ml to dissolve,
use dipotassium hydrogen phosphate solution （ Take
anhydrous potassium hydrogen phosphate 1.5g, add water
Mobile phase
Same reference standard method 10ml to dissolve ） adjust pH to 6.5]-water-acetonitrile-
methylalcohol（400:600:15:15）。
B Phase ： Phosphate Buffered Saline- acetonitrile- methyl

alcohol（400:450:150）
Waters XTerra ®MS C18（50×4.6mm，3.5µm）
252
chromatographic
column
Velocity of flow 1.8ml
Solution
concentration of the 0.5mg/ml
sample
Take the system suitability solution 10μ Precisely take 10μl of the separation test solution and the
l, into the liquid chromatograph, record reference solution, inject them into the liquid
the chromatogram, The separation of the chromatograph, and record the chromatograms. 。 The
Vildagliptin peak from adjacent impurity
degree of separation of impurity D and impurity C should
peaks should not be less than 1.5;Repeat
not be less than 2.5, and the tailing factor of the
the injection with the control solution
System suitability and record the chromatogram at a vildagliptin peak should not be greater than 1.8. ； The
wavelength of 210 nm，The relative reference standard solution was continuously injected 5
standard deviation of the continuous times. The relative standard deviation of the area of the
5-pin main peak area must not exceed vildagliptin peak should not exceed 2.0%
1.0%，The tailing factor of the
Vildagliptin peak should not be greater
than 1.8.
Injection volume 10μl 10μl
In terms of anhydrous, C17H25N3O2 With vildagliptin (C17H25N3O2) should be labeled
Limit
should be 98.0% ~102.0%. amount of 95.0% ~105.0%
Determine the content of this product by referring to the method for determining
the content of the vildagliptin tablet's import registration standard (standard number:
JX20100243).，but the system suitability solution has been adjusted.
Registration of the system for the determination of the suitability of the system
for the determination of the content of the vildagliptin tablets (standard number:
JX20100243)：Separately take 25 mg of vildagliptin control, 0.25 mg of impurity D,
0.25 mg of impurity C, and 0.25 mg of impurity E. Place them in the same 50 ml
volumetric flask and add the solvent to dissolve and dilute to the mark.
Additive solution chromatogram under specific content：
253
According to the chromatogram, impurity C, impurity D, and impurity E are far

away from the main peak.；In the degradation test, the unknown impurity peak in the
oxidative degradation is closer to the main peak (the chromatogram is as follows), so
the system suitability solution is replaced with the oxidative degradation solution.
Requirement ： The separation peak between the oxidation peak and the main
peak adjacent to the main peak is greater than 1.5.
System suitability solution preparation method ： Take about 50mg of
Vildagliptin reference substance, place it in a 10ml volumetric flask, dissolve it
with thinner and dilute to the mark, shaking (5mg/ml) ；Precisely take 1ml, set it
into a 10ml volumetric flask, add 1ml of 3% H2O2 solution, place it in a water bath at
80°C for about 3 hours, take it out, cool to room temperature, dilute to the mark with
diluent, and shake.
System Suitability Solution Chromatogram：
System applicability solution results：
Peak# Rt（min） Peak area Area（%） Resolution

1 0.956 16927 0.58 --
254
2 4.228 12087 0.42 19.507
3 4.914 27704 0.95 2.954
4 5.373 27652 0.95 2.124
5 5.897 39600 1.36 2.761
6 6.313 33151 1.14 2.363
7 6.522 21596 0.74 1.420
8 6.799 2734013 93.86 1.807
The test results of the batches of samples are shown in Table 3.2.S.4.5-18.
Table 3.2.S.4.5-1：Determination of Vildagliptin Sample Content
Potentiometric titration HPLC assay
Sample information Batch
assay results（%） results（%）
150601 99.9 99.9
Verification batch 150602 99.8 99.7
150603 99.8 99.7
3.2.S.4.5.19 Microbial Limit

This product is vildagliptin API ， It is intended to be used for preparing
vildagliptin tablets。Therefore, please refer to Appendix XI J of the 2010 edition of the
Chinese Pharmacopoeia.，Formulate microbial limit criteria for this product。Limit in
accordance with the limits of solid oral preparations：The number of bacteria can not
exceed 1000cfu per 1g，1100cfu per 1ml；No more than 100 cfu per 1g for mold and
yeast；Escherichia coli can not be detected every 1g
The microbial limit detection method has been fully validated and is suitable for
the detection of microbial limit of vildagliptin
3.2.S.5 Packaging materials and containers

（1）Package type、Sources are shown in Table 3.2.S.6-1，
Table 3.2.S.6-1：Package type, source and related certification documents
Project Packaging container
Package type Medicinal low density polyethylene bags
Packaging material manufacturers Shandong Xinhua Packing Co., Ltd.
255
Package Registration No. Chinese medicine package word 20140469
Package Registration Validity 2019.5.25
Packaging material quality standard
YBB00072005
number
（2） The choice of packaging materials

After inspection, this product cited wet weight gain of about 0.08%，According
to Appendix XIX J of the 2010 edition of the Chinese Pharmacopoeia, this product
has no humectant properties.。The product was placed under RH92.5%±5% for 30
days. The number of detected impurities increased from 3 to 10, and the total
impurities increased from 0.155 to 0.175.No other indicators have changed
significantly. It can be seen that the related substances have grown slightly within the
limits，This product is unstable under high humidity conditions.
This product was placed at 60°C for 30 days. The number of detected
impurities increased from 3 to 8；The largest single miscellaneous rose from 0.077%
to 0.092%；Total impurities increased from 0.155% to 0.210%. Enantiomers increased
from undetected to 0.03%, with no significant changes in other indicators。It can be
seen that the related substances and enantiomers have increased slightly within the
limits and the product is unstable under high temperature conditions.The product was
placed in a lighting condition of 4500 lx±500 lx for 30 days. The number of detected
impurities increased from 3 to 6；The largest single miscellaneous rose from 0.077%
to 0.082%；Total impurities increased from 0.155% to 0.198%。There are no
significant changes in other indicators。It can be seen that the related substances have
grown slightly within the limits.，This product is unstable under light conditions
Therefore, the packaging materials for this product are: medical low-density
polyethylene bags, and the outer packaging is cardboard drums.
Take 3 batches of verification batch samples and plan to market them after
packaging，Leave for 6 months under accelerated test conditions of 40°C±2°C and
relative humidity of 75%±5%.25℃±2℃、When placed at room temperature under
25°C±2°C and relative humidity 60%±10% for 12 months, the related substances
slightly increased within the limits, and other indicators did not change significantly.，
256
explain that this product has good stability under this packing condition.
3.2.S.6 Stability
3.2.S.6.1 Stability summary
The company conducted a stability inspection of the three batches of
vildagliptin produced
1）Stability test sample
See Table 3.2.S.7.1-1 for stability test samples.
Table 3.2.S.7.1-1：Stability test sample information
Batc Producti Inner packaging

Origin of production Batch Test type
h on Date material
2015-6-2
Medicinal low Influencing factors (bare
1506 ～ Anhui Haikang
115.0kg density polyethylene release), acceleration and
01 2015-6-1 Pharmaceutical Co., Ltd.
bags long-term testing
0
2015-6-1
Medicinal low
1407 1～ Anhui Haikang Accelerated and long-term
115.9kg density polyethylene
01 2015-6-1 Pharmaceutical Co., Ltd. testing
bags
9
2015-6-2
Medicinal low
1407 0～ Anhui Haikang Accelerated and long-term
120.0kg density polyethylene
02 2015-6-2 Pharmaceutical Co., Ltd. testing
bags
8
（2）Research content
The stability investigation items and methods are shown in Table 3.2.S.7.1-2.
Table 3.2.S.7.1-2：Stability check items and analysis methods
Detection project Analytical Whether verified Whether to include

method quality standards
Character Visual inspection General inspection, no yes
257
verification
General inspection, no yes
Moisture Fisher Method
verification
Melting point General inspection, no yes
Melting point
method verification
General inspection, no yes
Specific rotation Polarimetry
verification
Related substances HPLC Method Through verification yes
Enantiomers HPLC Method Through verification yes
Determination of Through verification yes
HPLC Method
content
Microbial limit Through verification yes
Microbial Limit
test
The content of stability study is shown in table 3.2.S.7.1-3.

Table 3.2.S.7.1-3: the content of stability research
Test Planned Completed
Test stage Inspection project
conditions sampling point sampling point
0day、5days、 0day 、 5days 、
40℃
10days、30days 10days、30days
Determination of
60℃ properties,
moisture, melting 10days、30days 10days、30days
Influence 4500lx±500l point, specific 0day、5days、 0day 、 5days 、

factor x curl, related 10days、30days 10days、30days
substances, 0day、5days、 0day 、 5days 、
RH75% enantiomers and
content
RH92.5%
Characteristics,
Acceleration 40℃±2℃， 0、1、2、3、6 0、1、2、3、6
moisture, melting
stability RH75%±5% months months
point, specific
258
rotation,
enantiomer, related
substances, content
determination,
microbial limit
(June)
Characteristics,
moisture, melting
point, specific curl,
25℃±2℃， related substances, 0、3、6 、9、
Long-term
RH60%±10 enantiomers, 12、18、24、 0、3、6、9、12
stability
% determination of 36
content and
microbial limit
(June)
The results of the routine stability inspection are shown in table 3.2.S.7.1-4.
Table 3.2.S.7.1-4: General stability inspection results
Storage Test conclusion
Test stage
conditions period
40℃ 30days It was verified that the samples were initially detected 3
impurities, the maximum single impurity 0.077%, the total
impurity 0.155%, 40 C for 30 days, 7 impurities, the largest
single impurity 0.060%, the total impurity 0.170%, 30 days for
High
30 days, 30 days, 0.077% impurities, the largest single impurity
temperat
60℃ 30days 0.092%, the total impurities 0.210%, and the enantiomers
ure test
increased from undetected to 0.03%, thus possible See, this
Influence product slightly increased under high temperature conditions.
factors There was no obvious change in all the other indexes. There is
no obvious change in the crystal form for 0 days.
The product was placed under strong light, 6 impurities were
Strong detected, the maximum single impurity was 0.082%, the total
light 4500lx±50 impurity was 0.198%, and the other indexes had no significant
30days
irradiatio 0lx changes. Thus, the impurity increased slightly under the light
n test conditions. There is no obvious change in the crystal form for 0
days.
259
RH75% 30days Under high humidity conditions, there was no significant
change in moisture and hygroscopic weight, and high humidity
75% was placed for 30 days, 6 impurities were detected, the
High maximum single impurity was 0.068%, and the total impurity
humidity was 0.173%. The high humidity 92.5% was placed for 30 days,
RH92.5% 30days
test and 6 impurities were detected, the largest single impurity was
0.073%, and the total impurity was 0.175%. Thus, the impurity
was slight under the condition of high humidity. Growth. There
is no obvious change in the crystal form for 0 days.
The company's products are placed for 6 months under the
40℃±2℃
conditions of the listed packaging, and the impurities in the
， 6
Accelerated test related substances increase slightly within the limits. There is
RH75%±5 months
no obvious change in the crystal form for 0 days.
%
25℃±2℃ The products of our company are placed for 12 months under
， 36 the conditions of being listed and packaged. The study of

Long term test
RH60%±1 months stability is still in progress.
0%
Note: ND is not detected
（3）Research conclusion
The conclusions are shown in table 3.2.S.7.1-5.
Table 3.2.S.7.1-5: conclusions of the study
Internal packing material Medicinal low density polyethylene bag
Storage conditions Seal, room temperature preservation
Term of validity 24 months
Hints of relevant contents in the
NO
instructions
3.2.S.6.2 Stability commitment & stability scheme after listing

The company promises to investigate the long-term retention stability of the first
three batches of the products produced after the market, and to examine the stability
of the long-term samples of at least one batch of products produced each year, and
notify the management authorities in a timely manner if there are abnormal
conditions.
Refer to 3.2.S.7.1, the acceleration of the three batch of validation batch
260
samples (40 C + 2 C, RH70% + 5%) and long term (25 C + + 2 C, RH60% + 10%)
stability are being studied. The company is now committed to continuing to study the
stability of the three products and to examine the stability of the long-term samples of
at least one batch of products produced each year (25 C + 2, RH60% + 10%), and
notify the administration in a timely manner if there is an abnormal situation.
Table 3.2.S.7.1-6: stability research program
Inspection Time of investigation(month)
project 0 1 2 3 6 9 12 18 24 36
Character X X X X X X X X X X
water X X X X X X X X X X
melting point X X X X X X X X X X
Specific rotation X X X X X X X X X X
Related
X X X X X X X X X X
substances
Enantiomer X X X X X X X X X X
Determination of
X X X X X X X X X X
content
microbial limit X X X X X
Note: X indicates that corresponding tests will be conducted at the corresponding time points.
Table 3.2.S.7.1-7: follow up annual batch stability test indicators and testing methods
Inspection project Standard Limit Test Method
white to yellowish or slightly grayish
Character
crystalline powder.
water ≤0.5%
Melting point 146℃~151℃

The specific rotation is -84 degree to
Specific rotation Show in table
-90 degree
3.2.S.4.2
impurityA≤0.15%，
impurityB≤0.15%，
Related substances impurityC≤0.15%，
impurityD≤0.15%，
impurityE≤0.15%，
261
impurityF≤0.15%，
impurityG≤0.15%，
Others single impurities≤ 0.1%，
Total impurities≤ 0.5%。
Enantiomer ≤0.15%
According to the calculation of
anhydrous substance, vildagliptin
Determination of content
(C17H25N3O2) should be 98% to
102%.
Number of bacteria：≤1000cfu/g
Mildew and yeast number ： ≤
microbial limit
100cfu/g
Escherichia coli：N.D /g
3.2.S.6.3 Stability data

Stability research data see the following table.
262
3.2.S.6.3.1 Influence factor test
Batch：150601 （First batch of verification batch） Batch Qty：115.0kg Specifications：API

High temperature test
High temperature test
Light test 4500Lx at 40 degrees RH92.5% high RH75% high humidity
Inspection Limit Initial at 60 degrees
(day) centigrade (day) humidity test (day) test (day)
project requirements centigrade (day)
0 5 10 30 5 10 30 5 10 30 5 10 30 5 10 30
Yello Yello Yello Yello Yello Yello Yello Yello Yello Yello Yello Yello
white to Yellowi Yellowi Yellowi
wish wish wish wish wish wish wish wish wish wish wish wish
yellowish or Yellowish sh sh sh
crysta crysta crysta crysta crysta crysta crysta crysta crysta crysta crysta crysta
Character slightly gray crystalline crystalli crystalli crystalli
lline lline lline lline lline lline lline lline lline lline lline lline
crystalline powder ne ne ne
powd powd powd powd powd powd powd powd powd powd powd powd
powder. powder powder powder
er er er er er er er er er er er er
146.0℃ 146.5℃ 146.5℃ 146.5℃ 146.0℃ 146.5℃ 146.0℃ 146.0℃ 146.5℃ 146.0℃ 146.5℃ 146.5℃ 146.5℃ 146.5℃ 146.5℃
147.5℃~
Melting point 146℃~151℃ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~ ~
148.5℃
147.0℃ 148.0℃ 147.5℃ 147.5℃ 147.5℃ 147.5℃ 147.5℃ 147.0℃ 147.5℃ 147.5℃ 147.5℃ 147.5℃ 147.5℃ 147.5℃ 147.5℃
Weight gain
and weight / / 0.03 0.06 0.14 -0.03 -0.10 -0.15 -0.03 -0.03 -0.07 0.11 0.12 0.29 0.03 0.03 0.13
loss（%）
263
Moisture
Water （%） content ≤ 0.10 0.13 0.13 0.18 0.09 0.08 0.06 0.10 0.09 0.07 0.34 0.37 0.47 0.24 0.25 0.33
0.5%
Specific
-84°至-90° -86.70° -86.39° -86.36° -86.40° -86.66° -87.34° -86.29° -86.70° -87.02° -86.21° -87.26° -87.02° -86.70° -86.48° -86.81° -86.88°
rotation
Impurity A≤
ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND ND
0.15%
ImpurityB≤
0.15%
Impurity C≤
ND ND ND ND ND ND 0.010 ND ND ND ND ND ND ND ND ND
0.15%
Impurity D
0.077 0.084 0.073 0.082 0.091 0.092 0.092 0.067 0.063 0.060 0.072 0.074 0.073 0.069 0.060 0.068
Related ≤0.15%
substances Impurity E≤
ND ND ND ND ND ND 0.005 ND ND ND ND ND ND ND ND ND
（%） 0.15%
Impurity F≤
0.15%
Impurity G
≤0.1%
Other single
impurities 0.046 0.039 0.041 0.045 0.045 0.041 0.041 0.043 0.049 0.043 0.043 0.038 0.043 0.039 0.038 0.042
≤0.1%
264
Number of
3 3 6 6 3 6 8 3 5 7 3 6 6 3 6 6
impurities
Total
impurities 0.155 0.153 0.180 0.198 0.169 0.191 0.210 0.142 0.159 0.170 0.144 0.175 0.175 0.138 0.156 0.173
≤0.5%
Assaying 98.0%~102.0
99.9 99.7 99.7 99.9 99.8 99.7 99.8 99.8 99.9 100.0 99.8 100.0 100.0 99.9 99.8 99.7
（%） %
Enantiomer
≤0.15% ND ND ND ND ND ND 0.031 ND ND ND ND ND ND ND ND ND
（%）
265
3.2.S.6.3.2 Accelerated test
batch：150601 batch Qty：115.0kg Specifications：API Condition of investigation：40℃±2℃，RH75%±5%
Inspection Time (month)

Limit requirements
project 0 1 2 3 6
white to yellowish or
Yellowish crystalline Yellowish crystalline Yellowish crystalline Yellowish Yellowish crystalline
Character slightly grayish
powder powder powder crystalline powder powder
crystalline powder.
Moisture content ≤
Water （%） 0.10 0.12 0.11 0.12 0.12
0.5%
Melting point 146℃~151℃ 147.5℃~148.5℃ 147.5℃~149.0℃ 147.5℃~148.5℃ 147.0℃~148.5℃ 147.0℃~148.5℃
Specific
-84° to -90° -86.70º -86.21º -86.98º -86.48º -87.09º
rotation
impurityA≤0.15% ND ND ND ND ND
impurityB≤0.15% ND ND ND ND ND
impurityC≤0.15% ND ND ND ND ND
impurityD≤0.15% 0.077 0.081 0.088 0.083 0.103
impurityE≤0.15% ND ND ND ND 0.006
Related
impurityF≤0.15% ND ND ND ND ND
substances（%）
impurityG≤0.15% ND ND ND ND ND
Other single impurity
0.046 0.039 0.038 0.035 0.036
≤ 0.1%
Number of impurity 3 3 3 3 4
Total impurity≤ 0.5% 0.155 0.150 0.157 0.151 0.172
Assaying（%） 98.0%~102.0% 99.9 99.9 99.9 99.7 99.8
Enantiomer（%） ≤0.15% ND ND ND ND ND
266
microbial limit Should be conforms / / / / conforms
267

Limit requirements
project 0 1 2 3 6
Yellowish Yellowish crystalline Yellowish crystalline Yellowish Yellowish crystalline
Character slightly grayish crystalline
crystalline powder powder powder crystalline powder powder
powder.
Water （%） 0.11 0.11 0.12 0.11 0.10
0.5%
Melting point 146℃~151℃ 146.5℃~147.5℃ 146.5℃~147.5℃ 146.5℃~147.5℃ 146.5℃~148.0℃ 146.0℃~147.5℃
Specific
-84° to -90° -87.31º -87.24º -87.41º -86.28º -86.56º
rotation
Impurity A≤0.15% ND ND ND ND ND
Impurity B≤0.15% ND ND ND ND ND
ImpurityC≤0.15% ND ND ND ND ND
ImpurityD≤0.15% 0.017 0.015 0.025 0.034 0.037
Related ImpurityE≤0.15% ND ND ND ND 0.003
substances ImpurityF≤0.15% 0.022 0.021 0.024 0.023 0.019
（%） ImpurityG≤0.15% ND ND ND ND ND
Other single impurity
0.076 0.071 0.061 0.066 0.068
≤0.1%
Total impurity≤ 0.5% 0.114 0.108 0.110 0.123 0.126
Assaying（%） 98.0%~102.0% 99.7 99.9 100.0 99.8 100.0
microbial limit Should be conforms / / / / conforms
268
Batch ：150603 batch Qty：120.0kg Specifications：API Condition of investigation：40℃±2℃，RH75%±5%

Limit requirements
project 0 1 2 3 6
Yellowish Yellowish crystalline Yellowish crystalline Yellowish Yellowish crystalline
Character slightly grayish crystalline
crystalline powder powder powder crystalline powder powder
powder.
Water （%） 0.08 0.11 0.12 0.14 0.12
0.5%
Melting point 146℃~151℃ 146.5℃~147.5℃ 147.5℃~149.0℃ 147.5℃~148.5℃ 147.5℃~149.0℃ 147.0℃~148.5℃
Specific
-84° to -90° -87.22º -86.89º -87.18º -87.01º -86.58º
rotation
Impurity A ≤ 0.15% ND ND ND ND ND
Impurity C≤0.15% ND ND ND ND ND
Impurity D≤0.15% 0.020 0.022 0.031 0.035 0.060
Related Impurity E≤0.15% ND ND ND ND 0.011
substances Impurity F≤0.15% 0.033 0.033 0.036 0.032 0.027
（%） Impurity G≤0.15% ND ND ND ND ND
Other single impurity≤
0.050 0.049 0.047 0.047 0.045
0.1%
Total impurity≤ 0.5% 0.103 0.104 0.114 0.114 0.143
Assaying （%） 98.0%~102.0% 99.7 99.8 99.9 99.9 100.0
microbial limit Should be conforms / / / / Conforms
269
3.2.S.6.3.3 Long term test

Inspection Time （month）

Limit requirements
project 0 3 6 9 12 18 24 36
white to yellowish or Yellowish Yellowish Yellowish Yellowish Yellowish
Character slightly grayish crystalline crystalline crystalline crystalline crystalline crystalline
powder. powder powder powder powder powder
Water （%） 0.10 0.12 0.11 0.11 0.13
0.5%
147.5℃~ 147.5℃~ 147.5℃~ 147.5℃~ 146.5℃~
148.5℃ 149.0℃ 149.0℃ 148.5℃ 147.5℃
Specific
-84° to -90° -86.70º -86.40° -86.78° -85.44° -85.65°
rotation
ImpurityB≤0.15% ND ND ND ND ND
Related ImpurityC≤0.15% ND ND ND ND ND
substances ImpurityD≤0.15% 0.077 0.077 0.070 0.072 0.073
（%） ImpurityE≤0.15% ND ND ND ND ND
ImpurityF≤0.15% ND ND ND ND ND
ImpurityG≤0.15% ND ND ND ND ND
270

0.046 0.039 0.037 0.036 0.044
0.1%
Total impurity ≤0.5% 0.155 0.147 0.136 0.138 0.156
Assaying（%） 98.0%~102.0% 99.9 100.0 99.9 100.1 100.1
Microbial limit Should be conforms / / conforms /
271

Limit requirements
project 0 3 6 9 12 18 21 36
Water （%） 0.11 0.12 0.12 0.12 0.14
0.5%
146.5℃~ 146.5℃~ 146.5℃~ 146.5℃~ 147.0℃~
147.5℃ 147.5℃ 147.5℃ 147.5℃ 148.0℃
Specific
-84° to -90° -87.31º -87.03° -86.97° -86.03° -85.59°
rotation
Impurity D≤0.15% 0.017 0.023 0.031 0.023 0.027
Related Impurity E≤0.15% ND ND ND ND ND
0.076 0.069 0.062 0.075 0.072
0.1%
Total impurity ≤0.5% 0.114 0.113 0.111 0.119 0.121
Assaying（%） 98.0%~102.0% 99.7 99.7 99.8 100.0 99.9
272
Microbial limit Should be conforms / / Conforms /
273

Limit requirements
project 0 3 6 9 12 18 24 36
Water （%） 0.08 0.14 0.14 0.13 0.17
0.5%
146.5℃~ 147.5℃~ 147.5℃~ 147.5℃~ 147.0℃~
147.5℃ 149.0℃ 149.0℃ 148.5℃ 148.0℃
Specific
-84° to -90° -87.22º -86.87° -87.05° -86.92° -86.86°
rotation
Impurity D≤0.15% 0.020 0.022 0.028 0.025 0.026
Related Impurity E≤0.15% ND ND ND ND ND
0.050 0.050 0.049 0.051 0.046
0.1%
Total impurity ≤0.5% 0.103 0.107 0.105 0.105 0.103
Assaying（%） 98.0%~102.0% 99.7 99.9 99.9 100.1 100.0
274
Microbial limit Should be conforms / / Conforms /
275
276
277
278
279
280
281

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3.2.S.4.5.11 Trifluoroacetic acid ....................................................................................... 245

Physical& chemical project Property description

Character White to yellowish or slightly gray crystalline powder

Melting point 146℃～151℃

Soluble in methanol, ethanol, DMSO, Water or 0.01mol/L hydrochloric acid solution,

Specific rotation -84° ～ -90°

3.2.S.2 Manufacture Information

起始原料1 起始物料2 粗品：维达列汀

isopropanol, purified water / 60~70 C, 1~1.5h, crystallization -5~0 C, 8~9h;

Step2：Absolute ethanol / solution temperature 60~70 C, 1~1.5h, crystallization temperature -5~0.,8~10h;

Note 2：SM-1：Starting material 1，(2S) -N- chloracetyl -2- cyanopyrrolidine

SM-2：Starting material 2，3- amino -1- amantadol

2. Process Flow Diagram

(2S) -N- chloracetyl

Heating up to 60～ Send to

filtrat Vacuum drying 10 ~

Step2：Vildagliptin Refining Medicinal

packing cleaning cleaning

Outer packing Outer packing

Table 3.2.S.2.2-1 The material input ratio of Initial vildagliptin

(2S) -N- chloracetyl -2-

Material name Molecular weight Inventory rating Material input ratio

Initial of vildagliptin 303.40 135.0kg 1.00

Anhydrous ethanol / 213.3kg 1.58(2V)

Medicinal activated carbon / 6.75 kg 0.05

4. list of major equipment Manufacture

Centrifuge LB1000 Stainless steel filter

Low temperature vacuum Initial of vildagliptin

Bag filter SUS304 Stainless steel Pressure filtration Refined

500LEnamel glass reaction

Centrifuge LBF1000 Stainless steel filter filter

Low temperature vacuum

Water Crusher YF3-1 Stainless steel Smash Smash

HD-50 Multi-motion mixer HD-50 Stainless steel Mixture Mixture

5. Large production scale range

Table 3.2.S.2.3-1 Summary of production materials of vildagliptin

material Quality standard

Jiangsu MAN TAI dazzling Biological

Supplier (producer) Ji'nan Ren Yuan Chemical Co., Ltd.

10 Isopropanol 1,2 GB/T 7814-2008

Solvent Supplier (producer) Wuhu Sanyou Chemical Trade Co., Ltd

NH2 NH2 NH2

B-1 B-2 B-3

In order to ensure the safety and controllability of pharmaceutical production,

(2S) -N- chloracetyl

l-prolinamide Starting material 0.125% ≤0.5%

VGT-1 Isomer By-product 0.030% ≤0.1%

VGT-2 Isomer By-product 0.069% ≤0.1%

Other unknown Single impurity ≤0.5%

Chemical name：(2S) -N- chloracetyl -2- cyanopyrrolidine

Detection project Acceptability limit Method index reference

Appearance White to yellow powder Visual measurement

Total impurities： ≤2.0%

Enantiomer: ≤0.5% Internal control standard

High performance liquid chromatography，Area normalization with correction factor

▪High performance liquid chromatograph equipped with ultraviolet detector

▪chromatographic column：Cyanic bond silica gel as a filling agent（such as Agilent ZORBAX

water acetonitrile (400:600:15).

▪Detection wavelength：210nm Flow rate：1.0ml/min

▪Injection volume：10μl Column temperature：35℃

dilute the solution containing about 1.0mg in each 1ml.

Determination method：Area normalization with correction factor