THINH A VETERINARY MEDICINE PRODUCTION AND TRADING

JOINT STOCK COMPANY (ASIFAC)

QUALITY CONTROL DEPARTMENT
VALIDATION OF ANALYTICAL PROCEDURES OF
ASI - ENROFLOXACIN 100 LIQUID
Version n 3 Number of documents: QC – AVP006-15

Written by Quality Control Manager Approved by Site Manager

Date: January 2022 Date: January 2022

Ta Thi Hao Nguyen Le Thanh Tuan

Distribution
 QC □ Stock
 QA □ Maintenance
□ R&D □ Marketing
□ IPC □ Production
□ Logistics

History of modifications

Amendment/addition
Edition N° Reason for amendment/addition
of page number
1 January 2020 Revision of the procedure

2 January 2022 Revision of the procedure

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 THINH A VETERINARY MEDICINE PRODUCTION AND TRADING
JOINT STOCK COMPANY (ASIFAC)
QUALITY CONTROL DEPARTMENT
VALIDATION OF ANALYTICAL PROCEDURES OF
ASI - ENROFLOXACIN 100 LIQUID
Version n 3 Number of documents: QC – AVP006-15

1. Objective
To prove that the analytical method of Enrofloxacin in ASI-ENROFLOXACIN 100 is suitable
for its intended use.

2. Scope
Analytical method of Enrofloxacin in ASI-ENROFLOXACIN 100.

3. Responsibility:
Methodology section head is responsible for implementation of the protocol.
QA manager is responsible for reviewing and approval of the protocol and the final report.

4. Composition:
Active Ingredients Quantity per 1ml
1 Enrofloxacin 100 mg

5. Documentation required:
Analytical monograph for ASI-ENROFLOXACIN 100
Equipment operation procedure.

6. Analytical performance characteristics and acceptance criteria:

STT Parameters Acceptance criteria
1 Linearity and range r2 ≥ 0,995
2 Accuracy and recovery 98% – 102%
3 Precision and repeatability RSD ≤ 2,0%
4 Specificity/Selectivity Pass the specificity/Selectivity test
Pass Accuracy and recovery test → Pass
5 Roughness
Roughness test

Page 2 of 6
 THINH A VETERINARY MEDICINE PRODUCTION AND TRADING
JOINT STOCK COMPANY (ASIFAC)
QUALITY CONTROL DEPARTMENT
VALIDATION OF ANALYTICAL PROCEDURES OF
ASI - ENROFLOXACIN 100 LIQUID
Version n 3 Number of documents: QC – AVP006-15

7. Revalidation:
It is necessary in the followwing circumstances:
- Changes in the composition of finished product.
- Changes in the analytical procedure.

8. Procedure:
8.1 Linearity and range:
- Linearity shows the correlation between sample concentration and meter absorbance.
- Preparation of solutions: Weigh about 50 mg of soluble standard and complete to 100 ml
with 0.1% NaOH solution. Accurately aspirate 2 ml of this solution, dilute and make up to
50 ml with 0.1% NaOH.
- Prepare a straight line: The standard solution was transferred in turn to a 10 ml volumetric
flask as shown in the following table and diluted to 10 ml with the same solvent. Measure
the absorbance.
- Blank sample: NaOH 0.1%.
- Measure the absorbance at 271 nm .

The standard solutions (ml) Volume (ml)

30% 3 10
40% 4 10
50% 5 10

60% 6 10

70% 7 10

- From the obtained statistics, determine the correlation coefficient r2, build a regression
equation and plot a linear line. Coefficient r2 ≥ 0,995.

Page 3 of 6
 THINH A VETERINARY MEDICINE PRODUCTION AND TRADING
JOINT STOCK COMPANY (ASIFAC)
QUALITY CONTROL DEPARTMENT
VALIDATION OF ANALYTICAL PROCEDURES OF
ASI - ENROFLOXACIN 100 LIQUID
Version n 3 Number of documents: QC – AVP006-15

- For each concentration, conduct 2 times → the average result.

8.2 Precision and repeatability:

- Determination of content in 6 independently phased test samples of the finished lot.

+ Standard: Weigh about 50 mg of the standard solution and complete to 100 ml with
0.1% NaOH solution. Take 1 ml of this solution, dilute and complete to 50 ml with 0.1%
NaOH.

+ Sample: Dissolve exactly 1 ml of product and complete to 100 ml with 0.1% NaOH.
Take 1 ml of this solution, dilute and complete to 100 ml with 0.1% NaOH.

+ Blank sample: NaOH 0.1%.

+ Measure the absorbance at 271 nm .

Calculate the quantity in mg/ml, of Enrofloxacin by the formula:

Absmau m T
X =  chuan  k
Abs chuan Vmau 100

Where:
mstd: Mass of standard substance (mg)
Vsmp: Volumn of sample (ml)
Astd: Absorbance of standard.
Asmp: Absorbance of sample.
T: Titer of standard substance.
k: Dilution factor of sample and standard

- Request: Precision and repeatability RSD ≤ 2,0%

Page 4 of 6
 THINH A VETERINARY MEDICINE PRODUCTION AND TRADING
JOINT STOCK COMPANY (ASIFAC)
QUALITY CONTROL DEPARTMENT
VALIDATION OF ANALYTICAL PROCEDURES OF
ASI - ENROFLOXACIN 100 LIQUID
Version n 3 Number of documents: QC – AVP006-15

8.3 Accuracy and recovery:

- Accuracy and recovery is performed on 5 independently phased test samples of a batch of
finished product.
- Sample: Dissolve exactly 1 ml of product in the 0.1% NaOH and complete to 100.0 ml and
add an amount of the standard not less than 10% of the analyte content in the sample. Dilute
1/100.0 ml with the 0.1% NaOH.
- The recovery coefficient is calculated according to the formula:
Abs T
Y =  mchuan  k
Abs chuan 100

Y − (Vmau  X )
A% = 100
mi
- Trong đó:
➢ Y – Total content of standard addition sample (mg).
➢ Abs – Absorbance of sample and added standard.
➢ Abschuan – Absorbance of the standard.
➢ mchuan – Standard mass (mg).
➢ Vmau – Sample volume (ml).
➢ mi – Standard mass added (mg).
➢ k – Dilution factor.
➢ T – Standard content (%)

8.4 Specificity/Selectivity
- The specificity/Selectivity of the method is evaluated to ensure that there is no influence on
the quantitative results from impurities and excipients in the formulation.

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 THINH A VETERINARY MEDICINE PRODUCTION AND TRADING
JOINT STOCK COMPANY (ASIFAC)
QUALITY CONTROL DEPARTMENT
VALIDATION OF ANALYTICAL PROCEDURES OF
ASI - ENROFLOXACIN 100 LIQUID
Version n 3 Number of documents: QC – AVP006-15

- Determine the absorbance of self-made samples including solvents, excipients similar to the
test solution but without Enrofloxacin active ingredient. From that, the specificity of the analytical
method can be concluded.

8.5 Roughness:
- Pass Accuracy and recovery test → Pass Roughness test.

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