Dissolution by HPLC Validation method

1.0 OBJECTIVE:

To provide general procedure for Validation of Analytical test method for Dissolution by HPLC based on USP, ICH and In-house Guidelines. 

2.0 SCOPE

This SOP is Applicable for validation of Analytical test method for Dissolution by HPLC in XXX company.

3.0 BACKGROUND:

Not applicable

4.0 RESPONSIBILITY:

4.1 Analytical Research personnel to perform the validation of test Procedure.

4.2 Group Leader- Analytical Research to check the Validation of Test procedure.

4.3 Head - Analytical Research to review the data and to ensure the compliance.

5.0 PROCEDURE

5.1 System suitability:

5.1.1 System precision: The precision as measured by replicate injections of homogeneous standard solution indicates the performance of the HPLC instrument under the chromatographic conditions and the day tested.  As a part of method validation, a minimum of 10 injections of the standard preparation is recommended.

5.1.2 Acceptance criteria:

The % RSD for peak areas of ten replicate injections of standard should be not more than half the value given for system suitability.      

5.1.3 Other system suitability parameters:

Prepare Resolution solution preparation / any other system suitability preparations as per the test method and evaluate system suitability parameters as required by the test method.

5.1.4 Acceptance criteria:

The system suitability parameters should be within the specified limits in the test method.

Note : If system suitability parameters mentioned in the Test procedure are observed  to be inadequate or difficult to meet the requirements, New system suitability  criteria shall be evaluated with proper reason, and the same shall be followed during the Validation activity.

5.2 Linearity of detector response:

5.2.1  Demonstrate the Linearity in the range of 25% to 150% of the target concentration.

Note: For multiple dosage strength, conduct linearity from 25% of the lowest 

strength to 150% of the highest strength. If standard and test concentrations are 

different, establish linearity covering both the ranges and also linearity levels shall be altered based on the requirement.

5.2.2 Prepare a series of standard solutions (not less than six is recommended) using drug substance in the above concentration range and inject in duplicate into the HPLC system. Plot average peak area versus the actual concentration, in microgram/ ml or mg/ml.

5.2.3 Calculate the correlation coefficient (r).

Note : Linearity range shall be altered based on the requirement.

5.2.4 Acceptance criteria’s

The correlation coefficient (r) should be not less than 0.999.In case of discrepancy investigate and explain the reason. Repeat if necessary and report the range in which it is linear

5.3 Sink condition for non compendial Dissolution Methods:

Note: Sink conditions need not to be performed for compendial methods.

5.3.1 Determine solubility of active drug substances in specified volume of dissolution medium in triplicate at room temperature (25°C), as per test method except RPM at 250

5.3.2  Acceptance criteria:

NLT 1.5 times the active drug substance in one tablet or capsules (of the highest dosage strength) will completely dissolve in the designated volume of the    dissolution medium at room temperature.

5.3.3 If the above acceptance criteria is not met, repeat point 5.3.1 to 5.3.2 at 37.5°Ctemperature

5.3.4 If the acceptance criteria is not met at 37.5°C also, repeat point 5.3.1 to 5.3.2 with formulated blend at room temperature (25°C).

5.3.5 If the acceptance criteria is not met at 25°C for blend, repeat point 5.3.1 to 5.3.2 with formulated blend at 37.5°C temperature.

5.4 Method Precision:

5.4.1 Repeatability:

5.4.1.1  Prepare six test preparations as per the test method on Tablets/Capsules/Oral suspension analyze and calculate % dissolution, average % dissolution of six unit dosages and RSD.

5.4.1.2 The method is considered “PRECISE”, if the average % dissolution meets the specification limit and relative standard deviation for % dissolution of six-dosage unit is not more than 5.0%.If the method is used for over a range of dosage strength and test preparation is same for all dosage strength, then conduct method precision for highest and lowest  dosage strength.     

5.4.1.3 If the test preparation is different from strength to strength then conduct the    method precision for all dosage strength.

5.4.1.4 If the method is used for a range of dosage types, (i.e. Tablets/ Capsules /Suspension etc), then conduct method precision for each dosage type.  

5.5 Recovery / Accuracy:

5.5.1 Prepare homogeneous Tablet / Capsule blends having 25%, 50%, 75%, 100% and 150 % of the labeled amount of drug substances.

Note: Spiked levels of drug substances shall be altered based on the requirement.

5.5.2 Prepare blend by keeping average weight constant if the quantity of active ingredient is reduced, increase the weight of placebo or vice versa to keep the average weight constant or add the active ingredient in solid state or solution state and placebo individually for each test instead of using blend or compress the   blends to tablets and use (or) prepare blends by keeping placebo constant.

5.5.3 Prepare test solutions in triplicate for each spike level and analyze as per the testmethod. Calculate the % recovery and average % recovery Note:If % Recovery not meeting the acceptance criteria continue the  dissolution at 200 RPM for 15minutes.   

5.5.4 Acceptance criteria:

The method is considered “ACCURATE”, if the average % recovery is not less  than 95.0%. Note: In case of discrepancy investigate the reason, repeat the experiment if   necessary and explain the reason for discrepancy

5.6 Linearity of the Method:

5.6.1 Calculate the average “mg” added and average “mg” found in recovery section and plot a graph to “average mg added” versus “average mg found”. Evaluate the coefficient of correlation.

5.6.2 Acceptance criteria:

The coefficient of correlation (r) shall be NLT 0.999. If the correlation coefficient (r)   is less than 0.999, report the range in which the method is linear.

5.7 Specificity (Placebo interference):

5.7.1 Perform the dissolution as per the test method on weight of placebo in triplicate, equivalent to the amount present in weight of tablet powder / Capsule content. In case of Capsule use filled capsules with placebo.         

5.7.2 Acceptance criteria:

No interference from placebo

5.8 Ruggedness:

5.8.1 System to system variability:

5.8.1.1 Conduct system to system variability study on two dissolution systems and HPLC systems (of the same or different manufacturer), by keeping analyst, column and sample same. Perform the dissolution on six-unit dosage as per the test method.

5.8.1.2  Calculate the % dissolution, average % dissolution of six dosage units and overall % RSD for both the systems

5.8.1.3  Acceptance criteria:

The method is considered rugged for system to system variability, if the average %dissolution meets the specification limit and overall RSD not more than 5.0%, for both the systems.

5.8.2 Column to Column variability:

5.8.2.1 Conduct column to column variability on two HPLC columns (of the same or different manufacturers), by the same analyst using the same HPLC system and    Dissolution system. Perform dissolution on six unit dosages as per the test method    using same sample.

5.8.2.2 Calculate the % dissolution, average % dissolution of six dosage units and %RSD for both the columns.

5.8.2.3 Acceptance criteria:

The method is considered rugged for column to column variability, if the average % dissolution meets the specification limit and overall RSD not more than 5.0%, for both the columns.

5.8.3 Analyst to Analyst variability:

5.8.3.1 Conduct analyst to analyst variability study by two analysts using the same HPLC system, same dissolution system and same column. Perform the dissolution on six dosage units as per the test method using same sample.

5.8.3.2 Calculate the % dissolution, average % dissolution of six unit dosages and %RSD

for both the analysts.

5.8.3.3 Acceptance criteria:

The method is considered rugged for analyst to analyst variability, if the average %dissolution meets the specification limit and overall RSD is not more than 5.0% for both the analysts.

Note: It is not considered necessary to study the variations (days, columns, equipment and analysts) individually.  The use of an experimental design (matrix) is encouraged          

5.8.4 Bench top stability of standard and test preparations:

5.8.4.1 Establish the stability of standard and test solutions on bench top for a period of 2days.

5.8.4.2 Prepare standard solution and prepare test solutions in “duplicate” as per the test method and keep them on bench top. Inject standard and test solutions into the HPLC system following the conditions described in the test method at initially, after  1 day and 2 days. Calculate % dissolution of test solution and % assay for standard against a fresh standard each time. Note: Keep potency of standard as initial Assay of the standard.

5.8.4.3 Acceptance criteria:

The solutions are considered “STABLE”, if the difference in % dissolution /% assay results from initial is not more than 2.0

5.8.4.4 If the solutions are found to be not stable for 1 day, inject and calculate the results at shorter time intervals. e.g at 1, 2, 3, hours etc, interval and establish the period of time during which the solutions are stable.

5.8.5 Refrigerator stability of standard and test preparations:

5.8.5.1 Establish the stability of standard and test solutions in refrigerator for a period of 2 days.

5.8.5.2 Prepare standard and test solutions in duplicate as per the test method and keep them in refrigerator. Inject standard and test solutions into the HPLC system following the conditions described in the test method at initially, 1 day and 2 days. Note: Allow the samples to come to room temperature before injecting.

5.8.5.3 Calculate % dissolution of test solution and % assay for standard against a fresh standard each time

5.8.5.4 Acceptance criteria:     

The solutions are considered “STABLE”, if the difference in % dissolution /% assay results from initial is not more than 2.0

5.8.5.5 If the solutions are found to be not stable for 1 day, inject and calculate the results at shorter time intervals. e.g. at 1, 2, 3, hours etc,  interval and establish the period of time during which the solutions are stable.

5.8.6 Refrigerator stability of system suitability solutions other than standard preparations

5.8.6.1 Establish the stability of system suitability solutions in refrigerator for a period of about 1day to 4 weeks or more.

5.8.6.2 Prepare the system suitability solutions as per the test method and keep in refrigerator. Inject system suitability solution into HPLC system following the conditions described in the test method at initial, after 1, 2 days or more.

Note: Allow the samples to come to room temperature before injecting

5.8.6.3 Acceptance criteria:

The solutions are considered “STABLE”, if the system suitability criteria passes.

5.8.7 Bench top stability of mobile Phase:

5.8.7.1 Establish the bench top stability of mobile phase for a period of about 2 days.

5.8.7.2 Prepare the mobile phase as per the test method and keep them on Bench top in well-closed condition. Evaluate system suitability parameters using following    conditions described in the test method at initial, 1 day and 2 days. Prepare system suitability solutions as per the test method.

5.8.7.3 Acceptance criteria:

The mobile phase is considered to be “STABLE” if all the system suitability values      are within the acceptable criteria set forth in the test method.

5.9  Robustness:

5.9.1 Filter Validation:

5.9.1.1 To demonstrate that the filtration does not effect the analysis results, validate at least two types of filters before use.       

5.9.1.2 Prepare test solutions in triplicate and standard solution as per the test method  without filtration. Centrifuge a portion of test solution and filter portions of test solutions through individual filters. Inject unfiltered standard solution & centrifuged  and filtered test solutions into HPLC system under the test conditions. Calculate the % dissolution for centrifuged and filtered test solutions and difference of %  dissolution between centrifuged and filtered samples.

5.9.1.3 Acceptance criteria:

The filter is considered acceptable, if the difference in % dissolution between centrifuged and filtered test solution is not more than 2.0

5.9.2 Effect of variation in mobile phase composition:

5.9.2.1 Prepare two mobile phases having 90% to 110% method organic phase  composition. Prepare system suitability solution (e.g. Resolution and standard solution) as per the method and inject into the HPLC system using both mobile phases. Evaluate the system suitability parameters as required by the test method for both the mobile phases.

5.9.2.2 Acceptance criteria:

The method is considered robust for variation of organic phase composition, if all the system suitability parameters meet the acceptance criteria set forth in the test method.

5.9.2.3 If any of the system suitability parameter fails, narrow the organic phase composition and establish the allowable range of variation for organic phase composition.

5.10 Effect of variation of pH in mobile phase:     

5.10.1 Prepare two mobile phases having the buffer pH with +/- 0.2 of the method pH. Prepare system suitability solution (e.g. Resolution solutions, standard preparations) as per the test method and inject into the HPLC system using both mobile phases. Evaluate the system suitability values as required by the test  method for both the mobile phases.

5.10.2 Acceptance criteria:

The method is considered robust for variation of pH, if all the system suitability parameters meet the acceptance criteria set forth in the test method.

5.10.3 If any of the system suitability parameters fails, narrow the pH range and establish the allowable range of variation of pH in mobile phase.

5.10.4 Effect of variation in flow rate.

5.10.4.1 Prepare system suitability solutions (e.g. Resolution solution, standard preparation, etc), as per the test method and inject into the HPLC system with +/-  0.2ml of method flow. Evaluate the system suitability parameters as required by the test method for both the flow rates.

5.10.4.2 Acceptance criteria:

The method is considered robust for variation of flow rate, if all the system  suitability parameters meet the acceptance criteria set forth in the test method.

If any of the system suitability parameters fails, narrow the flow rate range and establish the allowable range of variation of flow rate.

5.10.5 Effect of variation in column temperature:

5.10.5.1 Prepare system suitability (e.g. Resolution solution, standard preparation, etc), as per the test method and inject into the HPLC system with +/-5°C of method column temperature. Evaluate the system suitability parameters as required by the test method for both the column temperatures.   

Note: If the column oven specification limit does not allow to conduct the above variability, then conduct possible variation within the column oven specification limit. Consider the ambient temperature as 25°C.                 

5.10.5.2 Acceptance criteria:

The method is considered robust for variation in column temperature, if all the system suitability parameters meet the acceptance criteria set forth in the test method

6.0 REFERENCES:

ICH guidelines - Validation of Analytical Procedures Text and Methodology Q2(R1)

7.0  VERSION HISTORY:

8.0 ANNEXURES

NIL