Method validation protocol for Dissolution by HPLC

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This article contains information about Method validation protocol for Dissolution by HPLC. It includes validation parameters, it’s procedure and Acceptance criteria.

Method validation protocol for Dissolution by HPLC

Protocol Content

Sr. No  

Contents Name

 
1.0 Purpose
2.0 Scope
3.0 Reference
4.0 Site of the Study
5.0 Responsibility
6.0 Materials and instrument
7.0 Validation Procedure
8.0 Acceptance Criteria
9.0 Results
10.0 Conclusion
11.0 Attachments

 1.0: Purpose :

To validate the analytical test method for Dissolution of Drug products of film coated tablets

2.0: Scope :

This method validation study is applicable for Dissolution of drug products which is film coated tablets.

3.0: Reference :

1) ICH Guideline

2) Analytical Method Validation SOP

4.0: Site of the study   :

Pharmaceutical Analytical Development laboratory

5.0: Responsibility   :

Analyst / Executive / Head Quality Control /Quality Assurance.

6.0: Materials, Columns, STP and Instrument used:

 Instrument:

Sr. no Instrument Name Make
1 Analytical Balance Mettler Toledo
2 Analytical Balance Mettler Toledo
3 pH Meter LABINDIA
4 HPLC SHIMADZU

 Reagent / Chemicals :

Sr. no Reagents/Chemicals Grade
1 Sodium Hydroxide AR Grade
2 Acetonitrile HPLC Grade
3 Trifluoroacetic acid AR Grade
4 Hydrochloric acid AR Grade
5 Hydrogen peroxide AR Grade

Working Standard/Impurity Standard

Sr. No. Standard
1 API working standard

STP

Sr. No. STP NO. Version
1 As per respective Test STP to be Choose

HPLC Column

Sr. No. Column Description Column No.
1 As per respective Drug product

7.0: Validation procedure:

  • Perform the method validation by following the procedure designed under following validation parameters.
  • Chromatographic conditions, Mobile phase, diluents, System suitability preparation and Standard preparation shall be prepared according to respective STP .
  • Injection sequence shall be modified when two or more parameters are merged together while performing analytical method validation.

Validation Parameters:  

  1. Specificity and System suitability
  2. Linearity
  3. Precision
  4. Accuracy
  5. Solution stability
  6. Robustness
  7. Filter study

8.0: Acceptance criteria:

   System suitability and Acceptance criteria for above validation parameters are specified in STP and individual parameters respectively.

9.0: Results:     

1) Observations and results shall be recorded in individual method validation test data sheet and excel sheet.

2)  Summarize the findings of the method validation study to draw Conclusion.

10.0: Conclusion:     

Based on the interpretation of the results in method validation report, draw the conclusion.

11.0: Attachments:    

1) Annexure I: Method of Analysis.

2) Annexure II: Method validation test data sheet.

 

Validation Parameter and it’s procedure in details:

7.1: Specificity and System suitability

Placebo preparation:

Weigh placebo equivalent to one tablet and transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’.

After 30 minutes, collect 10 mL of the solution from each vessel and filter using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

API for Identification:

Weigh 20 mg of API and transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’.

After 30 minutes, collect 10 mL of the solution from each vessel and filter using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

Sample preparation:

Take one tablet and transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’.

After 30 minutes, collect 10 mL of the solution from each vessel and filter using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

Injection Sequence:
Sr.No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6.  API solution (For ID) 1
7. Sample solution 1
8. Bracketing standard solution 1
 System suitability criteria:
Sr. No. System suitability parameter Acceptance criteria
1. % RSD %RSD of peak area response and retention time for six replicate injections of the standard solution B should not be more than 2.0 %.
2. Theoretical plates Theoretical plates for the peak due to API obtained from Standard Solution should not be less than 1500.
3. Tailing Factor Tailing factor for the peak due to API obtained from Standard Solution should not be more than 2.0.
4. Similarity factor The Similarity factor between standard solutions should be between 0.98 to 1.02.
Acceptance criteria:
Sr.No. Parameter Acceptance criteria
1. Interference Interference from blank, placebo peaks should not be more than 0.1%.
2. Peak Purity The peak purity index for Standard solution, Identification solution and Sample solution should not be less than 0.999.

7.2: Linearity and Range

Preparation of Linearity standard stock solution:

Weigh accurately about 25 mg of API standard and transfer to a 100 mL volumetric flask. Add 60 mL of diluent. Sonicate to dissolve. Dilute to volume with diluent and mix well.

Preparation of linearity levels:
Levels Volume of Solution A to be taken in mL Dilute to volume Concentration (ppm)
Level – 1 (50%) 1.0 25 10
Level – 2 (80%) 1.6 25 16
Level – 3 (100%) 2.0 25 20
Level – 4 (120%) 2.4 25 24
Level – 5 (150%) 3.0 25 30
Injection Sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Linearity Level – 1 (50%) 6
6. Bracketing standard solution 1
7. Linearity Level – 2 (80%) 3
8. Linearity Level – 3 (100%) 3
9. Bracketing standard solution 1
10. Linearity Level – 4 (120%) 3
11. Bracketing standard solution 1
12. Linearity Level – 5 (150%) 6
13. Bracketing standard solution 1
 Acceptance criteria:
Sr. No. Parameter Acceptance criteria
1. Correlation coefficient Not less than 0.995 over the working range
2. % RSD at each level Not more than 2.0%
3. Slope Report the value
4. Residual sum of squares Report the value
5. Y-Intercept Report the value
6. % Y-Intercept NMT + 2.0%
7. Range Report the values

7.3: Precision:

System precision
Injection Sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
Acceptance criteria:
Sr. No. System suitability parameter Acceptance criteria
1. % RSD %RSD of peak area response and retention time for six replicate injections of the standard solution B should not be more than 2.0 %.
2. Theoretical plates Theoretical plates for the peak due to API obtained from Standard Solution should not be less than 1500.
3. Tailing Factor Tailing factor for the peak due to API obtained from Standard Solution should not be more than 2.0.
4. Similarity factor The Similarity factor between standard solutions should be between 0.98 to 1.02.
 Method precision

Placebo solution: For placebo preparation refer above procedure.

Sample Solution : For sample preparation refer above procedure.

Injection Sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6. Sample solution – 1 1
7. Sample solution – 2 1
8. Sample solution – 3 1
9. Sample solution – 4 1
10. Sample solution – 5 1
11. Sample solution – 6 1
12. Bracketing standard solution 1
 Acceptance Criteria:
Sr.No. Parameter Acceptance criteria
1. % Assay of each Sample and mean of all samples Not less than 80% (Q) in 30 minutes (S1, S2 & S3 applicable)
2. %RSD %RSD of % Release of six sample preparations should not be more than 5.0.
3. 95% Confidence interval Report the value
Intermediate precision

Placebo solution: For placebo preparation refer above procedure

Sample Solution: For sample preparation refer above procedure

Injection Sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6. Sample solution – 1 1
7. Sample solution – 2 1
8. Sample solution – 3 1
9. Sample solution – 4 1
10. Sample solution – 5 1
11. Sample solution – 6 1
12. Bracketing standard solution 1
Acceptance criteria:
Sr.No. Parameter Acceptance criteria
1. % Assay of each Sample and mean of all samples Not less than 80% (Q) in 30 minutes (S1, S2 & S3 applicable)
2. %RSD %RSD of % Release of six sample preparations should not be more than 5.0.
3. Absolute Difference of mean values (Six Method precision and six Intermediate precision) Absolute difference of mean values (n=12) should not be more than 5.0%.
4. 95% Confidence interval Report the value

 7.4 : Accuracy

Placebo solution: For placebo preparation refer above procedure

Accuracy Level I (50%):

Weigh placebo equivalent to one tablet and API 10 mg, transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’.

After 30 minutes, collect 10 mL of the solution from each vessel and filter using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

Accuracy Level II (100%):

Weigh placebo equivalent to one tablet and API 20 mg, transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’.

After 30 minutes, collect 10 mL of the solution from each vessel and filter using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

Accuracy Level III (150%) :

Weigh placebo equivalent to one tablet and API 30 mg, transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’.

After 30 minutes, collect 10 mL of the solution from each vessel and filter using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

Prepare all three levels in triplicate.

 Injection Sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6. Accuracy Level I (50%) – 1 1
7. Accuracy Level I (50%) – 2 1
8. Accuracy Level I (50%) – 3 1
9. Accuracy Level II (100%) – 1 1
10. Accuracy Level II (100%) – 2 1
11. Accuracy Level II (100%) – 3 1
12. Bracketing standard solution 1
13. Accuracy Level III (150%) – 1 1
14. Accuracy Level III (150%) – 2 1
15. Accuracy Level III (150%) – 3 1
16. Bracketing standard solution 1
 Acceptance Criteria:
Sr. No. Parameter Acceptance criteria
1. % Recovery at each level % Recovery should be between 95.0 and 105.0
2. Mean % Recovery of all level % Recovery should be between 95.0 and 105.0
3. %RSD at each level %RSD of % Recovery at each level should not be more than 5.0.
4. %RSD of all levels %RSD of % Recovery of all levels should not be more than 5.0.

7.4: Solution stability

Placebo solution: For placebo preparation, refer above procedure.

Standard solution (Initial): For standard preparation refer respective STP

Sample solution (Initial): For sample preparation, refer above procedure

Injection Sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6. Standard solution (Initial) 1
7. Sample solution (Initial) 1
8. Bracketing standard solution 1

 Note: Repeat the sequence for standard solution and sample solution for 24 and 48 hours, if at any point solution stability of standard solution or sample solution fails, discontinue the study of individual solution. Report the solution stability of standard solution and sample solution individually.

Acceptance Criteria:
Sr. No. Parameter Acceptance criteria
1. % Difference of assay for standard solution % Difference of assay should be not more than 2.0 at different time intervals from initial for the standard solution.
2. % Difference of release for sample solution % Difference of % Release should be not more than 2.0 at different time intervals from initial for the sample solution.

 7.5: Robustness

For Robustness follow variation in analytical method as per given table.
Sr.No. Parameter Variation Value after variation
1. Variation in column oven temperature (+2°C) from actual value. i) -2°C (38°C)

ii) +2°C (42°C)
2. Variation in rpm (+4% rpm) from actual value. i) -4% rpm (48 rpm)

ii) +4% rpm (52 rpm)
3. Variation in dissolution medium (+10 %) from actual value. i) -10 % (900)

ii) +10% (1100)

 Placebo solution: For placebo preparation, refer above procedure .

Sample solution: For sample preparation, refer above procedure .

Injection sequence:
Sr.No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6. Sample solution – 1 1
7. Sample solution – 2 1
8. Sample solution – 3 1
9. Sample solution – 4 1
10. Sample solution – 5 1
11. Sample solution – 6 1
12. Bracketing standard solution 1

For all the robustness parameters follow injection sequence as mentioned above.

Acceptance criteria:
Sr. No. System suitability parameter Acceptance criteria
1. % RSD %RSD of peak area response and retention time for six replicate injections of the standard solution B should not be more than 2.0 %.
2. Theoretical plates Theoretical plates for the peak due to API obtained from Standard Solution should not be less than 1500.
3. Tailing Factor Tailing factor for the peak due to API obtained from Standard Solution should not be more than 2.0.
4. Similarity factor The Similarity factor between standard solutions should be between 0.98 to 1.02.

For all robustness parameters, system suitability criteria should meet as per test method criteria.

Acceptance Criteria:
Sr. No. Parameter Acceptance criteria
1. Interference Interference from blank, placebo peaks should not be more than 0.1%.
2. % Difference of release for sample solution % Difference of % Release should be not more than 5.0 from Method precision for the sample solution.

 7.6 Filter Study

Placebo solution: For placebo preparation, refer procedure 7.1.1.

Sample solution (Unfiltered):

Take one tablet and transfer in individual jar containing the dissolution medium previously maintained at the temperature of 37°C ± 0.5 °C Operate the dissolution apparatus as specified under section ‘Dissolution Parameters’. After 30 minutes, collect 10 mL of the solution from each vessel. (Sample stock) Centrifuge the stock solution at 3000 rpm for 10 minutes.

Sample solution (Filtered):

Filter the sample stock solution using 0.45 µm Nylon filter, discarding the first few mL of filtrate. (Use separate filter for each jar)

Injection sequence:
Sr. No. Solution No. of Injections
1. Blank 1
2. Standard solution_A 1
3. Standard solution_B 6
4. Blank 1
5. Placebo solution 1
6. Sample solution (Unfiltered) 1
7. Sample solution (Filtered) 1
8. Bracketing standard solution 1
 Acceptance criteria:
Sr. No. Parameter Acceptance criteria
1. % Difference of % release for sample solution % Release difference should not be more than 2.0% between filtered and unfiltered sample solution.

 

 

 

 

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