Raja Sundararajan et al / Int. J.

of Pharmacy and Analytical Research Vol-2(4) 2013 [194-203]

IJPAR |Volume 2 | Issue 4 | Oct - Dec-2013 ISSN: 2320-2831

Available Online at: www.ijpar.com

[Research article]
Analytical method development and validation for residual solvent of
Diltiazem hydrochloride extended release capsule by Gas chromatography
*
Raja Sundararajan, Christopher Vasanth Kumar, Jayaveera.
Gitam institute of Pharmacy, Gitam university, visakhapatnam -530045, Andrapradesh, India.

ABSTRACT
Residual solvents are organic volatile chemicals that are either used or produced during the manufacture of
active pharmaceutical ingredients, excipients, and drug products. Organic solvents such as di ethyl ether, N
heptane, acetone, methyl acetate, tertiary - butyl alcohol, iso-propyl alcohol, ethanol , toluene, Nbutanol, and n-
butyl acetate frequently used in pharmaceutical industry for the manufacturing of drug product Gas
Chromatography method for the determination of residual solvent of Diltiazem Hydrochloride Extended
Release Capsulesfor iso-propyl alcoholhas been carried out using this method.this method utilizedPerkin Elmer
clarus 580 GC Turbomatrix 40 Headspace sampler and DB-624, 30 m x 0.530 mm ID, 3.0 µm column was used
with flame ionizationdetector (FID). The GC method for the determination of residual solvent of Diltiazem
Hydrochloride Extended Release Capsules was validated. The method was found to be specific, precise, linear,
accurate, rugged, robust and suitable for its intended use.
Keywords: Diltiazem Hydrochloride, iso-propyl alcohol, ICH, validation.

INTRODUCTION
Diltiazem is a calcium ion influx inhibitor (slow Harmonization (ICH) [1-2]. General chapter in USP
[3]
channel blocker or calcium antagonist). .Gas chromatography method for the
Chemically, diltiazem hydrochloride is 1, 5- determination of residual solvents[4-5]. Analytical
Benzothiazepin-4(5H) one, 3-(acetyloxy)-5-[2- Methods for Residual Solvents Determination in
(dimethylamino)ethyl]-2,3-dihydro-2-(4-methoxy Pharmaceutical Products and excipients[6-7].
phenyl)-, mono hydrochloride, (+)-cis-. Its Chromatographic methods for residual solvents
molecular formula is C22H26N2O4S HCl and its analysis for iso propyl alcohol[8-9]. A review on
molecular weight is 450.99. Diltiazem produces its GC-MS and Method Development and
antihypertensive effect primarily by relaxation of Validation[10]. Since this drug is official in
vascular smooth muscle with a resultant decrease in pharmacopoeia only in dissolution method.No
peripheral vascular resistance. The magnitude of previous GC-FID method for the determination and
blood pressure reduction is related to the degree of quantification of Diltiazem Hydrochloride
hypertension; thus hypertensive individuals Extended Release Capsules in literature. Therefore,
experience an antihypertensive effect, whereas the purpose of this investigation was to develop
there is only a modest fall in blood pressure in and validate a method using a simple, rapid,
normotensives. International Conference on sensitive, specific, precise, linear, accurate, rugged,
robust GCFID method.

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*Corresponding author: Raja Sundararajan
E-mail address: sraja61@gmail.com
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MATERIALS AND METHOD

Reagents/chemicals
Isopropyl alcohol : GC/HPLC/GR
Dimethyl sulphoxide : HPLC Grade

GC conditions
Instrument : Gas chromatography with Headspace
Column : DB624, 30m x 0.530 mm ID x 3.0 µm
Carrier gas : Nitrogen
Linear velocity : 35 cm/sec
Detector : 240°C, FID
Range : 1
Attenuation : 4
Injector temperature : 200 °C
Oven temperature program :
Temperature : 40°C
Hold time ‘1’ : 0°C/min
Rate temp : 10°C/min
Hold temp : 220°C
Hold Time ‘2’ : 5 min

Run time : 23 min

Gas flow : Constant velocity
Split : 1:20
Hydrogen : 45 mL/min
Zero Air : 450 mL/min

Condition for Head Space Sampler

Vial temperature : 100 °C
Needle temperature : 110 °C
Transfer line temperature : 120 °C
Injection volume : 0.06 mL
Thermostat : 15 min
High pressure inject : on
Head space mode : constant
Carrier gas pressure : 15 psi
Vial pressurization : 4 min
Withdraw time : 0.2 min
GC cycle time : 35 min

Solution preparation mg of Isopropyl alcohol (IPA) dilute to volume

Blank solution
Transfer 4.0 mL of Dimethyl sulphoxide in
headspace vial and seal it using vial crimper.
Standard stock solution
To a 100.0 mL volumetric flask containing 20.0
mL of Dimethyl sulphoxide, transfer about 250.0
with Dimethyl sulphoxide and mix well.
Standard solution
Transfer 25.0 mL of standard stock solution into
100 mL volumetric flask, dilute to volume with
Dimethyl sulphoxide and mix.
Transfer 4.0 mL of the standard solution into a
head space vial, and seal it using vial crimper.

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Test solution mL of Dimethyl sulphoxide and seal it using vial

Crush the pellets in to fine powder. Transfer 500 crimper.
mg of powdered sample in a headspace vial. Add 4

LOD, LOQ and retention time data*

Name LOD (ppm) LOQ (ppm)

Isopropylalcohol
25 80
alcohol

*LOD, LOQ data incorporated after method validation.

Calculation standard solution.

Isopropyl CS : Concentration of standard solution
AT CS P in mg/mL
alcohol content = x x x 106
CT : Concentration of sample solution in
in ppm AS CT 100
mg/mL
Where, P : Percentage purity of standard.
AT : Area of Isopropyl alcohol peak in
the chromatogramof sample
solution.
AS : Average area of Isopropyl alcohol
peak in the chromatogram of

RESULTS AND CONCLUSIONS Ruggedness of the method was verified by

PRECISION
SYSTEM PRECISION
Six replicate injections of standard solution were
injected. The mean and percentage relative
standard deviation (% RSD) for peak areas were
calculated. The results are tabulated in table - 1.
Acceptance criteria
Percentage relative standard deviation (% RSD) for
peak areas is not more than 10.0
METHOD PRECISION
Six samples of 360 mg capsuleswere prepared by
spiking the residual solvent at specification level
and analysed as per test method. The residual
solvent content in six samples and percentage
relative standard deviation (% RSD) for six results
were calculated and the results are tabulated in
table-2.
analysing the six samples spiked with residual
solvent at specification level from the same batch
which was used for method precision as per test
method by different analyst, different lot no. of
column, and different instrumentation different
day. The content of residual solventswas
determined. Calculated percentage relative standard
deviation (% RSD) for residual solventin six
samples and also calculated overall percentage
relative standard deviation (% RSD) for ruggedness
results with the method precision results. The
results are tabulated in table-3.
Acceptance criteria
The percentage relative standard deviation (%
RSD) for residual solvent contentfrom six samples
is not more than 15.0. Overall percentagerelative
standard deviation (%RSD) for method and
intermediate precision results is not more than 15.0

Acceptance criteria SPECIFICITY

Blank, placebo, standard, sample solution and
The percentage relative standard deviation (%
sample solution spiked with residual solvent at
RSD) for content of residual solvent from six
specification level were injected. The elution
samples is not more than 15.0.
pattern in blank and placebo observed. The
retention time of analyte peak from standard
INTERMEDIATE PRECISION solution and sample (spiked) are compiled in table-
(RUGGEDNESS)

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4. Refer figure 2 to 6 for the chromatograms of
blank, placebo, standard, sample and spiked sample
Acceptance criteria
Observe the elution of peak pattern in the blank and
placebo.
$- Refer retention time for standard and sample
solution from method precision experiment.
LIMIT OF DETECTION AND LIMIT OF
QUANTITATION
(LOD and LOQ)
The limit of detection (LOD) and limit of
quantitation (LOQ) was determined for Isopropyl
alcoholby signal to noise ratio method by using the
formula.
Signal to noise ratio (S/N) = 2H/h
H - Height of analyte
h - Height of noise
LOQ value was verified by giving six replicate
injections of solution containing Isopropyl alcohol.
LOD and LOQ values are summarized in table -5a.
The percentage relative standard deviation (%
RSD) calculated for peak areas and tabulated in
table-5b
Acceptance criteria
Signal to noise ratio 10:1 at the level of LOQ and
2:1 or 3:1 at the level of LOD.
The percentage relative standard deviation (%
RSD) for peak areas at LOQ level is not more than
15.0
LINEARITY
Linearity for Isopropyl alcohol was performed
from about LOQ to 150 % level. A graph was
plotted with concentration (in ppm) on x-axis and
peak areas on y-axis. Slope, y-intercept, correlation
coefficient and residual sum of squares (RSS) were
determined. The results are tabulated in table -6
graphically represented in figure- 1
Acceptance criteria
The correlation coefficient (r-value) value is not
less than 0.97
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ACCURACY (RECOVERY)
Known amount of residualsolvent spiked with
sample at about LOQ, 100% and 150% of
specification limit. The percentage recovery was
calculated from the amount found and actual
amount added. The results are tabulated in table - 7.
Acceptance criteria
Percentage recovery at each level is in between
85.0 and 115.0
Percentage relative standard deviation (%RSD) is
not more than 15.0 at each level.
RANGE
Range inferred from the data of linearity, accuracy
and precision experiments.
ROBUSTNESS
Robustness of the method was verified by
deliberately varying the following condition.
i) By changing the flow rate by  10%.
System suitability was evaluated and residual
solvent spiked sample was analysed in triplicate.
Calculated cumulative percentage relative standard
deviation (% RSD) for each condition and method
precision data. The results tabulated in table-8. The
system suitability parameters are tabulated in table-
9.
Acceptance criteria
Overall percentage relative standard deviation (%
RSD) for each change condition and method
precision results is not more than ± 15.0
SUMMARY OF SYSTEM SUITABILITY
System suitability was evaluated by injecting
standard solution during various days of validation.
The percentage relative standard deviation (%
RSD) for the peak areas were verified. The results
are tabulated in table – 9
Acceptance Criteria
The percentage relative standard deviation (%
RSD) for the six injections of standard is not more
than 10.0
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TABLES
TABLE – 1-SYSTEM PRECISION

Injection No. Peak area

1 187952
2 191760
3 189915
4 188706
5 188366
6 192441
Mean 189857
% RSD 1.0

TABLE – 2-METHOD PRECISION

Sample No. Isopropyl alcohol (ppm)

1 6125
2 6284
3 6171
4 6097
5 6145
6 6118
Mean 6157
% RSD 1.1

TABLE – 3-INTERMEDIATE PRECISION

Sample No. Isopropyl alcohol (ppm)

Analyst 1 Analyst 2
1 6125 6280
2 6284 6101
3 6171 6266
4 6097 6205
5 6145 6345
6 6118 6487

Mean 6157 6281

Percentage relative 1.1 2.1
standard deviation (%
RSD)
Overall average 6219

Over all Percentage 1.9

relative standard deviation
(% RSD)

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TABLE – 4-SPECIFICITY

Peak Name Sample type RT (min.) $

Isopropyl alcohol Standard 3.159

Spiked sample 3.157

TABLE -5a-SUMMARY OF LOD AND LOQ VALUES

Name LOD LOQ

ppm S/N ppm S/N

Isopropyl alcohol 25 27 80 68

TABLE -5b-PRECISION AT LOQ

Injection No. Peak area

1 3018
2 2924
3 2976
4 2949
5 2966
6 2994
Mean 2971
% RSD 1.1
S/N - Signal to noise ratio

TABLE –6-LINEARITY OF ISOPROPYL ALCOHOL

Level (%) Concentration in ppm Peak area

LOQ 80.0384 2956
25 1250.6000 45189
50 2501.2000 90831
75 3751.8000 138527
100 5002.4000 185430
125 6253.0000 237433
150 7503.6000 285464
Slope 38.1514
y-intercept -2739.8875
Correlation coefficient 0.99978
Residual sum of squares 17614806.9169

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TABLE –7-ACCURACY OF ISOPROPYL ALCOHOL

Level Amount found in µg Actual amount added in µg % Recovery
Mean % RSD

Level - 1 70.7097 79.9584 88.4 95.7 12.9

(LOQ)
70.9501 79.9584 88.7
87.8531 79.9584 109.9
Level - 2 5184.0542 4997.3976 103.7 105.3 1.9
(100%)
5377.5980 4997.3976 107.6

5233.5884 4997.3976 104.7

Level - 3 7710.7255 7496.0964 102.9 101.3 1.4
(150%)
7526.1539 7496.0964 100.4
7544.0716 7496.0964 100.6

TABLE –8-ROBUSTNESS
S.No. Method precision Flow rate (31.5 cm/sec) Flow rate
(38.5cm/sec)
1 6125 5654 5600

2 6284 5654 5605

3 6171 5795 5765

4 6097 - -
5 6145 - -
6 6118 - -
Mean 6157 - -
%RSD 1.1 - -
Overall Mean - 6005 5990

Overall %RSD - 4.0 4.3

TABLE -9-SUMMARY OF SYSTEM SUITABILITY

S.No Name of Experiment % RSD
1 System precision and 1.0
Method precision
2 Specificity and Robustness 0.6
3 Robustness 0.6
(Flow Rate – 38.5 cm/Sec)

4 Robustness 1.0
(Flow Rate – 31.5 cm/Sec)
5 LOQ, LOD, Linearity and Accuracy 0.8
6 Intermediate Precision 1.8

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FIGURES
Figure 1: Linearity plot for Isopropyl alcohol

300000 Isopropyl alcohol

250000
200000
Area

150000
100000
50000
0
0.0000 2000.0000 4000.0000 6000.0000 8000.0000
Concentration µg/mL

Figure - 2: Chromatogram of blank

Figure - 3: Chromatogram of placebo

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Figure - 4: Chromatogram of standard

Figure - 5: Chromatogram of sample (unspiked)

Figure - 6: Chromatogram ofspiked sample

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ACKNOWLEDGEMENTS chromatographic method for estimation of residual

The authors are grateful to piramal health care Ltd,
India. For providing Diltiazem hydrochloride as a
gift sample.
CONCLUSION
This study presents a simple and validated Gas
solvents in Diltiazem Hydrochloride Extended
Release Capsule drug product. The developed
method is specific, precise, linear, accurate, rugged,
robust and suitable for its intended use.

REFERENCES
[1] ICH Expert Working Group. Impurities: Guideline for Residual solvents. Q3C[R5],
2011.
[2] ICH. Validation of Analytical Procedures, Text and Methodology, Q2 (R1), 2005.
[3] USP <467> Residual solvents.
[4] Marie-JoséeRocheleau, MélanieTitley, Julie Bolduc, Measuring residual solvents in pharmaceutical
samples using fast gas chromatography techniques, Journal of Chromatography B, 805, 2004, 77–86.
[5] Chusena Narasimharaju Bhimanadhuni and Devala Rao Garikapati, Development and validation of gas
chromatography method for the determination of residual solvents in Mirabegron,Der PharmaChemica,
2013, 5(1):55-60.
[6] Katarzyna Grodowska and Andrzej Parczewski, Analytical Methods For Residual Solvents
Determination In Pharmaceutical Products, Acta Poloniae Pharmaceutica ñ Drug Research, Vol. 67
No.1, 2010:13-26.
[7] RohitkumarM.Yadav and KapilPatil, Development and Validation of Gas Chromatographic Analytical
Method for Estimation of Residual Solvents in Excipients (Klucell, Triacetan, HPMC), International
Journal of Research in Pharmaceutical and Biomedical Sciences,Vol. 4 (2) Apr– Jun 2013, 618-623.
[8] Praveen Kumar Baliyan, R.P. Singh and SaurabhArora, Simultaneous Estimation of Residual Solvents
(Isopropyl Alcohol and Dichloromethane) in Dosage Form by GC-HS-FID, Asian Journal of
Chemistry, Vol. 21, No. 3 (2009), 1739-1746.
[9] S.B. Puranik, Varun R. Pawar, N. Lalitha, P.N. Sanjay Pai and G.K. Rao, Gas chromatographic
methods for residual solvents analysis, Oriental Journal of Chemistry, Vol. 24(1), 2008, 529-536.
[10] Lakshmi Hima Bindu M.R, Angala Parameswari S,Gopinath C,A Review on GC-MS and Method
Development and Validation, International Journal of Pharmaceutical Quality Assurance 2013; 4 ( 3) ;
42-51.

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