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Analytical Method Validation (AMV) Procedure / Guide

Standard Operating Procedure (SOP) and Guideline for Validation of Analytical Method (AMV). The objective of validation of an analytical Method (AMV) is to demonstrate that it is suitable for its intended purpose.

Procedure for Analytical Method Validation (AMV)

1.0   Objective :

    • To lay down procedure for validation of Analytical Method (AMV) to be followed in laboratory.
    • The objective of validation of an analytical Method (AMV)is to demonstrate that it is suitable for its intended purpose.

2.0   Scope :

    • This SOP is applicable in quality control for validation of analytical Method (AMV).

3.0   Responsibility :

    • Analyst:
    • To prepare the protocol and report of validation of analytical Method (AMV) and to follow the procedure during validation of analytical procedure.
    • Head QC/Executive QA:
    • To review the protocol and report of validation of analytical Method (AMV).
    • Head Quality Assurance:
    • To approve the protocol and report of validation of analytical Method (AMV).
    • Head QA:
    • Approval of the protocol and report of validation of analytical Method (AMV).

4.0   Procedure for Analytical Method Validation (AMV):

    • Validation of analytical Method (AMV) is the process by which it is established, by laboratory studies, that the performance characteristics of the method meet the requirements for the intended analytical applications through certain standards of accuracy and reliability.
    • Prepare validation protocol as per Annexure-I and get it approved it from authorize persons.
    • Before starting the analytical Method validation (AMV) experiments, system suitability shall be done to determine the suitability of the chromatographic system for the analysis as per the individual method.
    • If required theoretical plates, capacity factor, resolution factor and tailing factor shall also be calculated.
    • When the system meets the system suitability parameters mentioned in the method, validation experiments shall be started.
    • Bracketing standards shall be injected throughout the validation.
    • The mean, standard deviation and relative standard deviation shall be determined as per the individual method system suitability criteria.

    • Analytical Method parameters in the validation are explained in detail as follow:

    • Precision
      • System precisionAnalytical Method Validation (AMV)
      • Method precision (Repeatability)
      • Ruggedness (Intermediate Precision)
    • Specificity
    • Forced degradation
    • Linearity
    • Range
    • Accuracy (recovery)
    • Limit of detection (LOD)
    • Limit of Quantitation (LOQ)
    • Solution Stability
    • Robustness
    • Filter paper selection study
    • System suitability test (SST)

  • Precision – Analytical Method Validation (AMV)

    • System Precision:
    • The system precision is the closeness of agreement between the responses of detector.
    • Usually expressed as the standard deviation (SD) or the relative standard deviation (RSD).
    • Procedure :
    • Inject separately diluent (blank), standard solution (six replicates) and record the area / absorbance responses of main analyte peak/absorbance in standard solution.
    • Calculate the % RSD of responses and retention time of main analyte peak for six replicates.
    • Acceptance Criteria :
    • For dissolution and assay, relative standard deviation for values (response) is not more than 2% and relative standard deviation for retention time is not more than 1.0%.
    • Related substances, relative standard deviation for peak area counts is not more than 5% and relative standard deviation for retention time is not more than 1.0%.
    • Residual solvents, relative standard deviation for peak area counts is not more than 10%.
    • Note: Acceptance criteria can be varied depending up on the requirement of method.

    • Method Precision (Repeatability):

    • The precision of an analytical method is the degree of agreement among individual test results when the procedure is applied repeatedly to multiple sampling of homogeneous sample.
    • Usually expressed as the standard deviation or the relative standard deviation.
    • Procedure:
    • Prepare six samples of single batch and analyze as per the test Method.
    • Calculate the % RSD for six samples.
    • For related substances:
    • Prepare six individual sample of same batch and analyze as per test method.
    • In case any known impurities are not present in the specimen samples.
    • The method precision can be done by spiking the known impurities at specification level (if impurities available).
    • Calculate the %RSD of impurities for six samples.

  Note (Dissolution and Assay):

    1. For dose Proportional
    • If multiple strengths are available methods precision can be performed on higher strength.
    1. For pseudo proportional
    • If multiple strengths are available method precision should be performed on all strength.

    • Acceptance Criteria:

    • For dissolution method, relative standard deviation (RSD) of the six analysis shall be not more than 5% for release above 85% and not more than 10% for release below 85%.
    • For assay method, relative standard deviation (RSD) of the six analysis shall not be more than 2%.
    • Incase,  known impurities in related substances method, relative standard deviation of six analysis shall not be more than 10 % for the impurity level greater than 1.0% and relative standard deviation shall not be more than 15% for the impurity level between 0.11% and 0.99%.
    • This criterion is applicable for known impurities above LOQ level and for unknown impurities more than 0.1%.
    • For known residual solvents in residual solvents method, relative standard deviation of six analysis shall be not more than 10%.
    • Note: Acceptance criteria can be varied depending up on the requirement of method.

  • Ruggedness (Intermediate Precision) – Analytical Method Validation (AMV):

    • Intermediate precision expresses within laboratory variation with different analysts of equipment within the same laboratory using same lot of drug product as specified under precision.
    • Procedure:
    • The analysis of the same batch shall be done in six replicate analysis by using columns of the same make having different serial numbers by different analysts, by different systems on different day.
    • The mean, standard deviation and relative standard deviation shall be calculated.
    • For dose proportional and for pseudo proportional follow the same procedure as per method precision.

    • Acceptance Criteria:

    • For dissolution,
    • Relative standard deviation (RSD) of six analysis shall be not more than 5% for release above 85%, not more than 10% for release below 85% and relative standard deviation for retention time of main analyte in standard solution should not be more than 1.0%.
    • Overall relative standard deviation shall be not more than 5% for release above 85% and not more than 10% for release below 85% when compared with method precision results.

    • For assay

    • Relative standard deviation (RSD) of the six analysis shall be not more than 2% and relative standard deviation for retention time of main analyte in standard solution should not be more than 1.0%.
    • Overall relative standard deviation shall be not more than 2% when compared with method precision results.

    • For related substances

    • Relative standard deviation (RSD) of the six analysis shall be not more than 10% for the impurity level greater than 1.0% and relative standard deviation shall not be more than 15% for the impurity level between 0.11% and 0.99%.
    • Overall relative standard deviation shall be not more than 10% for the impurity level greater than 1.0% and relative standard deviation shall not be more than 15% for the impurity level between 0.11% and 0.99% when compared with method precision results.
    • These criteria applicable for known impurities above LOQ level and for unknown impurities more than 0.1%.
    • For residual solvents
    • Relative standard deviation (RSD) of the six analysis shall be not more than 10% and overall relative standard deviation shall be not more than 10% when compared with method precision results.

Note: Acceptance criteria may be varied depending up on the requirement of method with justification.

  • Specificity – Analytical Method Validation (AMV):

    • Specificity of analytical method is its ability to assess unequivocally the analyte in presence of components that may be expected to be present in the sample matrix.
    • Procedure:
    • For assay, inject blank (diluent), placebo, identification solution (one or more active components), standard, sample solution and spike sample.
    • Check the peak purity of main peak.
    • For dissolution by HPLC inject blank (Dissolution media/diluent), placebo and for dissolution by UV, scan the blank, placebo solution.
    • For related substances inject blank (diluent) and placebo in duplicate (for drug products).
    • Prepare one sample solution spiked with known impurities at specification level, inject and check the peak purity of main peak and known impurities.
    • Inject known impurities individually.

Note: The sample can be diluted for peak purity.

    • For residual solvents, inject blank (diluent) and placebo in duplicate (for drug products).
    • Inject the sample solution spiked with known residual solvents at specification level and known residual solvents individually.

Note: In case of multiple strengths, placebo solution can be prepared wherever placebo concentration is high.

    • Acceptance Criteria:

    • For Assay:
    • No peaks shall be eluted at the retention time of the main peak in the blank and placebo solution.
    • Peak purity of main peak shall be passed.
    • All the peak of blank, placebo and main analyte shall be well separated from each other in spiked sample solution.
    • For dissolution by HPLC:
    • No peaks shall be eluted at the retention time of main peak in the blank (diluent) and placebo absorbance.
    • For dissolution by UV:
    • The interference from blank and placebo shall not be more than 2% of standard absorbance.
    • For related substances:
    • No peaks shall be eluted at the retention time of main peak and known impurities in the blank (diluent) and placebo solution.
    • Peak purity of main peak and unknown impurities shall be passed.
    • For residual solvents:
    • Observe the elution of pattern of peak elution in blank and placebo solution.
    • Note:  Acceptance criteria can be varied depending up on the requirement of method.

  • Forced Degradation – Analytical Method Validation (AMV) :

    • For a stability indicating method, it is essential to perform forced degradation studies by applying appropriate accelerated stress conditions to the sample.

    • The proposed stress conditions are:

    • Photolytic (Sunlight and ultraviolet light for 2-4 hours)
    • Thermal (At 105°C or below melting point)
    • Acid degradation
    • Base degradation
    • Oxidation
    • Water Hydrolysis
    • Humidity (25°C/90%RH)*
    • Prepare super saturated solution of Potassium Nitrate and keep this solution in a well closed Desiccator.
    • Keep the sample and Placebo in the chamber.
    • If the drug product is in dose proportionate then forced degradation study will be performed on higher strength only, but if the drug product is not in dose proportionate then force degradation study will be performed based on qualitative composition of the drug product.
    • Procedure:
    • For assay, prepare the untreated sample solution and treated samples solution (heat, acid, base, hydrolysis, humidity and oxidation).
    • Inject and calculate the %assay.
    • Calculate the %difference between the untreated sample and treated samples results.
    • Check the peak purity of main peak.
    • For related substances, prepare the untreated sample solution and treated samples solution (light, heat, acid, base, hydrolysis and oxidation).
    • Inject and calculate the % impurities.
    • Calculate the %difference between the untreated sample and treated samples results.
    • Check the peak purity of main peak.
    • Known impurities are spiked wherever necessary.

Note: For peak purity, stress sample shall be diluted, wherever required.

    • Acceptance Criteria:

    • For Assay:
    • The treated sample results shall be compared with untreated sample and the difference shall be not more than 15%.
    • Peak purity of main peak shall be passed.
    • For related substances:
    • Degrade to the extent up to 15% is performed to evaluate the interference.
    • Peak purity for main peak shall be required.
    • Mass Balance determination:
    • Mass balance has to be verified by demonstrating that the decrease the concentration of the substance exposed to stress conditions corresponds to an equivalent increased amount of degradation products.

    • General guidelines as follows:

    • For all degraded samples, both assay and related substances have to be tested.
    • If for some reason the official assay test cannot be performed, the decrease of the main peak signal in the related substances test could be evaluated and to justify mass balance.
    • The following data should be reported both for stressed and unstressed samples (control).
    • Amount of individual degraded substances found, total amount of degraded substances found, assay and some of assay and degraded substances.
    • An example is provided in the below table:
Impurity 1

(% w/w)

 Impurity 2

(% w/w)

 Total

(% w/w)

 Assay Total Impurities + Assay
Control 0.1 0.1 0.2 99.8 100.0
Stressed Sample 5.0 4.0 9.0 90.5 99.5
    • The mass balance should be in the range of 95 % – 105 % of the control sample.
    • However, if the decrease in the assay value due to degradation is less than 5%, tighter criteria may be more appropriate.
    • Mass balance is confirmed when the concentration of the parent drug found in the stressed sample is consistent with the amount of degraded compounds formed after the stress test.
    • The following examples are intended to represent potential real case scenarios:
Impurity 1

(% w/w)

 Total

(% w/w)

 Assay Total Impurities + Assay Comment
Control 0.1 0.2 99.8 100.0
Stressed sample 1 8.0 9.0 90.5 99.5 Good mass balance
Stressed Sample 2 0.1 0.3 55.0 55.3 No Mass Balance

(Some degradation products are not detected)
Stressed sample 3 10.0 15.0 95 110 No Mass Balance

(Some degradation products are overestimated)
    • Mass balance cannot be accurately evaluated during early method development.
    • However, the balance may be a useful tool to ensure that there is no significant degradation products unaccounted.
    • In case mass balance is not achieved, the degradation should be scientifically evaluated and justified.

  • Linearity – Analytical Method Validation (AMV):

    • Its ability (within a given range) to obtain test results which are directly proportional to the concentration levels shall be prepared.
    • Procedure:
    • For assay, prepare and inject the standard solution in the range of 80% to 120% concentration level and
    • Calculate the correlation coefficient “r” by calculation of a regression line by the least square method.
    • Also determine the Residual sum of square.
    • For content uniformity, prepare and inject the standard solution in the range of 70% to 130% concentration level and
    • Calculate the correlation coefficient “r” by calculation of a regression line by the least square method.
    • Also determine the Residual sum of squares.
    • For dissolution testing, ± 20% over the specified range (e.g., if the acceptance criteria for a controlled- release product cover a region from 30% , after 1 hour, and up to 90%, after 24 hours, the validated range would be 10% to 110% of the label claim.
    • For related substances, prepare and inject the known impurities solution and standard solution in the range of LOQ to 200% concentration level calculate the correlation coefficient “r” by calculation of a regression line by the least square method and calculate the response factor for known impurities by using the below given formula:

                            Slope of active substance

Response Factor   = ——————————————–

                            Slope of impurity

 

                                                 Slope of impurity

Relative Response Factor (RRF) = —————————————–

                                                    Slope of active substance

    • Also determine residual sum of squares
    • For residual solvents, prepare a solution of known residual solvents and standard solution in the range of LOQ to 150% concentration level,
    • Inject and calculate the correlation coefficient “r” by calculation of a regression line by the least square method.
    • Also determine the residual sum of squares.

    • Acceptance Criteria:

    • The value of correlation coefficient “r” for dissolution and assay shall be not less than 0.999, for related substances and residual solvents not less than 0.99.

Note: Acceptance criteria can be varied depending up on the requirement of method.

  • Range – Analytical Method Validation (AMV):

    • The range of an analytical Method is the interval between the upper and lower concentration (amounts) of analyte in the sample (including these concentrations) for which it has been demonstrated that the analytical Method has a suitable level of precision, accuracy and linearity.
    • The range is normally expressed in the same units as test results (e.g. percent, parts per million) obtained by the analytical method.

  • Accuracy (Recovery) –Analytical Method Validation (AMV) :

    • Procedure :
    • For assay, prepare the sample solution by spiking the drug substance to the placebo at about 70%, 100% and 130% of test concentration level in triplicate in each level and calculate the RSD for recovery obtained at each level separately and overall RSD.
    • Incase, Dissolution prepare the sample solution by spiking the drug substance to the placebo at about ± 20% specified range in triplicate in each level and calculate the % overall average recovery.
    • For related substances, prepare the sample solution without spiking known impurities in triplicates and inject, prepare the sample solution in triplicate by spiking with known impurities at LOQ level to 150% of specification limit (as per shelf life specification limit) and calculate the % overall average recovery for known impurities.
    • For residual solvents, prepare the sample solution without spiking known residual solvents in triplicate and inject,
    • Prepare the sample solution in triplicate by spiking with known residual solvents at LOQ level to 150% of specification limit and calculate the % overall average recovery for known residual solvents.

Note: Accuracy experiment for API to be inferred from the experiment data of specificity, linearity and precision.

    • Acceptance Criteria:

    • For assay,
    • % recovery of the drug at each level shall be between 98% and 102% and RSD shall be not more than 2%.
    • The overall average shall be between 98% to 102% with RSD of not more than 2%.
    • For dissolution,
    • % recovery of the drug at each spiking level shall be between 95% and105% and RSD shall be not more than 5%.
    • The overall average recovery shall be between 95% to 105% with RSD of not more than 5%.

Note: For less soluble drugs “In cases of poor drug solubility, if feasible, the stock solution may be prepared by dissolving the drug substance in a small amount of organic solvent and diluting to the final concentration with diluent”.

    • For related substances,
    • The recovery of known impurities of the drug at each spiking level shall be between 85% and 115% and RSD from replicate analysis shall be not more than 10%.
    • The overall average recovery shall be between 85% and 115% with RSD of not more than 10%.
    • For residual solvents, the recovery of known residual solvents of the drug at each spiking level shall be between 85% and 115% and RSD from replicate analysis shall be not more than 10%.
    • The overall average recovery shall be between 85% and 115% with RSD of not more than 10%.

  • Limit of Detection (LOD) – Analytical Method Validation – AMV:

    • It is the lowest concentration of analyte in a sample that can be detected but not necessarily quantitate under the stated experimental conditions.
    • The limit of detection is usually expressed as the concentration of analyte e.g. percentage, ppm, ppb etc.
    • Procedure:
    • Based on Signal-to-Noise Ratio
    • Determination of the signal-to-noise ratio is performed by comparing measured signals from samples with known low concentrations of analyte with those of blank samples and establishing the minimum concentration at which the analyte can be reliably detected.
    • A signal-to-noise ratio 3:1 is acceptable for estimating the detection limit.
    • Based on the Standard Deviation of the Response and the Slope
    • For related substances and residual solvents prepare and inject the known impurities solutions and standard solution in the range of LOD to 200% of specification level and calculate the limit of detection by using below formula.

    • Calculate the limit of Detection by below formula:

3.3 x σ

LOD =—————–

S

Where,

σ = the standard deviation of the response.

S = the slope of the calibration curve.

    • The slope S may be estimated from the calibration curve of the analyte.
    • The estimation of σ may be carried out based on the calibration curve.
    • A specific calibration curve shall be studied using samples containing an analyte in the range of DL.
    • The residual standard deviation of a regression line or the standard deviation of y-intercepts of regression line may be used as the standard deviation (σ).

    • Verification at LOD:

    • Prepare the LOD solution using blank/placebo spiked with known impurities or known residual solvents at determined LOD level and inject in six replicates.
    • Calculate % RSD for six replicates of known impurities.
    • Acceptance Criteria:
    • % RSD for six replicates responses of known impurities or known residual solvent shall be not more than 30.

  • Limits of Quantitation (LOQ) – Analytical Method Validation (AMV):

    • It is the lowest concentration of analyte in a sample that can be quantitate with acceptable precision under the stated experimental condition.
    • The limit of quantification is usually expressed as the concentration of analyte. e.g. percentage, ppm , ppb etc.
    • Procedure :
    • Based on Signal-to-Noise Ratio
    • Determination of the signal-to-noise ratio is performed by comparing measured signals from samples with known low concentrations of analyte with those of blank samples and establishing the minimum concentration at which the analyte can be reliably quantified.
    • A typical signal-to-noise ratio is 10:1.

    • Based on the Standard Deviation of the Response and the Slope

    • For related substances and residual solvents prepare and inject the known impurities solutions and standard solution in the range of LOQ to 200% specification level calculate the limit of quantification by using below formula add perform the precision at LOQ analysis and calculate the %RSD.
    • Calculate the limit of Quantitation by below formula:

 

10 x σ

LOQ =—————–

S

Where,

σ = the standard deviation of the response.

S = the slope of the calibration curve.

    • The slope S may be estimated from the calibration curve of the analyte.
    • The estimation of σ may be carried out based on the calibration curve.
    • A specific calibration curve shall be studied using samples containing an analyte in the range of QL.
    • The residual standard deviation of a regression line or the standard deviation of y-intercepts of regression line may be used as the standard deviation (σ).

    • Precision at LOQ:

    • Prepare the LOQ solution using blank/placebo spiked with known impurities or known residual solvents at determined LOQ level and inject in six replicates.
    • Calculate % RSD for six replicates of known impurities.
    • Acceptance Criteria:
    • % RSD for six replicates responses of known impurities or known residual solvent shall be not more than 10.
    • Note:  Acceptance criteria can be varied depending up on the requirement of method.

  • Solution Stability – Analytical Method Validation (AMV):

    • It is essential when validating an analytical method to confirm that the analyte has adequate stability in both the standard and sample solution during analytical measurement stages of the testing.
    • Procedure :
    • For dissolution, prepare the standard solution and perform the dissolution on one tablet or capsule as per the test method.
    • Analyze the standard solution and sample solution at the different time intervals and calculate the % difference for the result.
    • For assay, prepare the standard solution and sample solution as per the test method,
    • Analyze the standard solution at the different time intervals and calculate the % difference for the result.
    • For related substances, prepare the standard solution and sample solution spiked with known impurities at the specification level as per the test method.
    • Analyze the standard solution and sample solution at the different time intervals and calculate the % cumulative RSD of peak area for known impurities and main peak.

    • Acceptance Criteria:

    • For dissolution, the difference in results shall not be more than 2.
    • Incase assay, the difference in results shall not be more than 2 for drug products and shall not be more than 1 for drug substance.
    • For related substances, Cumulative % RSD of peak area for known impurities and main peak shall not be more than 10.
    • Note: Acceptance criteria can be varied depending up on the requirement of method.
    • If acceptance criteria are not met then a time limit is set within which the analysis is to be completed.
    • This assumes that the precaution have been taken to minimize degradation.
    • For analyses where a small degree of degradation in unavoidable, automation of the assay ensures that exactly the same degree of degradation occurs with each sample and standard.

  • Robustness – Analytical Method Validation (AMV) :

    • The method shall show reliability of an analysis with respect to deliberate variations in method parameters.
    • If measurements are susceptible to variations in analytical conditions, mention the same in test method.
    • Under the variable conditions in method parameters and system suitability parameters shall be established to ensure that the validity of the analytical Method and the conditions shall be suitability controlled or a precautionary statement shall be included in the procedure.
    • Following deliberate variations shall be done in case of dissolution, assay and related substances:
      • pH of mobile phase (± 0.2 units)
      • Column oven temperature (± 5°C)
      • Flow rate (±10%)
      • Mobile phase composition (organic composition) (up to ±10% absolute)
    • RPM (for dissolution) (±5RPM)
    • System suitability parameters shall be performed as per the test method for each deliberate variation.

    • Procedure:

    • For dissolution, prepare the standard solution and perform the dissolution on three tablets or capsules by deliberate variations made in the method for each condition as mentioned in protocol and analyze.
    • Calculate the results and compare with method precision results.
    • For assay, prepare the standard solution and sample solution in triplicate as per the test method by deliberate variations made in the method for each condition as mentioned in protocol and analyze.
    • Calculate the RRT’s for known impurities and compare with RRT’s from specificity experiment.
    • For Related substances, prepare the standard solution and sample solution spiked with known impurities at the specification level as per the test method by deliberate variations made in the method for each condition as mentioned in protocol and analyze.
    • Calculate the RRT’s for known impurities and compare with RRT’s from specificity experiment.
    • For residual solvent, prepare the standard solution and sample solution spiked with known residual solvents at the specification level per the test method by deliberate variation made in the method for each condition as mentioned in protocol and analyze.
    • Calculate the RRT for known residual solvents and compare with RRT from specificity experiment.
    • Acceptance Criteria:
    • For dissolution, overall % RSD shall be not more than 5 with of the method precision data for individual experiments.
    • For assay, overall % RSD shall be not more than 2 with of the method precision data for individual experiments.
    • Related substance and residual solvent, method shall meet the system suitability criteria.
    • Note:
    • To meet acceptance criteria, robustness parameters can be changed as per method requirement.

  • Filter paper selection study – Analytical Method Validation (AMV):

    • Procedure
    • For Assay/Dissolution/UOD:
    • Prepare sample solution in triplicate as per test method.
    • A portion of sample solution shall be centrifuged and other portion of sample solution shall be filtered with filters (e.g. PVDF/Nylon).
    • Inject all the samples into HPLC as per method.
    • Calculate the % Assay or % drug release difference for each sample and calculate the % difference between centrifuged vs. filtered samples.
    • For Related Substance:
    • Prepare sample solution (spiked sample solution).
    • A portion of sample solution shall be centrifuged and other portion of sample solution shall be filtered with filters (e.g. PVDF/Nylon).
    • Inject all the samples into HPLC as per method.
    • Calculate the % impurity for each sample and calculate the % impurity difference between centrifuged vs. filtered samples.
    • Acceptance Criteria:
    • Difference shall not be more than 2 % for Assay.
    • Difference shall not be more than 5% for dissolution.
    • If the impurity is less than 0.1%, no comparison shall be made.
    • For known impurities   and total impurities % difference shall be less than ±10%.

  • System Suitability Test (SST) – Analytical Method Validation – (AMV):

    • System suitability tests are based on concept that the equipment, electronics, analytical operations and sample to be analyzed,
    • Test of System suitability provide the added assurance that on specific occasion the method is given accurate and precise results.
    • System suitability test shall be run before an experiment is initiated and whenever there is change in the environment analysis.
    • The nature of the test and acceptance criteria shall be based upon the data generated during method development, optimization and validation experiments.
    • System suitability parameters, which are generally required to be monitored including number of theoretical plates (efficiency of the column), internal precision (repeatability), reproducibility, tailing factor (peak asymmetry), resolution, relative retention time and capacity factor depending upon the requirement of analytical method being validated. Some other parameters may be included in system suitability test, shall be justified in validation report.
    • Each experiment conducted as part of method validation exercise should have a corresponding system suitability test.
    • After completion of validation experiment as part of protocol, prepare the report for the same as per annexure II.
    • Summary – Analytical Method Validation (AMV):
Analytical Performance Characteristics Assay Related Substances Dissolution Residual Solvents
Accuracy   √ √ * √
Precision Applicable √ Applicable √
Specificity √ √ * Applicable
Linearity Applicable √ * √
Robustness √ Applicable *   √
Forced Degradation    √ √ –  –
Detection Limit  – √ – √
Quantitation Limit  – Applicable – √
Solution Stability  √ √  *   –

 √: Applicable                             – : Not applicable

*: Required, depending on the nature of the specific test.

If analytical method of dissolution is by HPLC, precision shall be done.

If analytical method of dissolution is by UV, accuracy, precision, specificity, linearity, robustness and solution stability shall be covered.

Note: The above states will be protocol bound with respective projects.

  • Re-validation of Analytical Method :

    • Compendial methods are accepted as the official method.
    • However suitability of the method shall be checked from in-house product under actual conditions of use.
    • General tests and assays, which are already established, may also be validated to verify their accuracy when used for new drug products.
    • Revalidation may be required when there are changes in the drug product e.g. new source or synthesis of drug substance, product strength, new excipients, change in level of excipients, change in manufacturing process etc.
    • Evaluate the parameters of the validation depending on the change.
    • Revalidation of the method is required whenever significant changes in method are done, sample solution, chromatographic system components and chromatographic conditions.
    • The parameters of the validation shall be evaluated depending on the change.
    • Verification of validated analytical method shall be done at-least once in three years as per approved protocol.

  • Verification of Compendia Methods.

    • The verification process for compendial test procedures is the assessment of whether the procedure can be used for its intended purpose, under the actual conditions of use for a specified drugs substance and/or drug product matrix.
    • Compendial analytical Method are not required to validate these procedures, when first used in their laboratories, but documented evidence of suitability should be established under actual conditions of use.
    • In the United States, these requirements is established in 21CFR 211.194(9) (2) of the current GMP regulations which state that the “Suitability of all testing methods used shall be verified under actual conditions of use”.
    • Complete validation of a compendial method is not required to verify the suitability of a procedure under actual condition of use.
    • Analytical performance characteristics used in method verification of pharmacopoeial methods shall be (but not limited to) Specificity, Precision, Linearity and Range, System Suitability and Stability in Analytical Solutions.
    • Annexure-I and Annexure-II shall be used for the preparation of method verification protocol and report. Terminology “Validation” shall be replaced with “Verification”

5.0   REFERENCES :

ICH guidelines – Q2 (R1) “Validation of Analytical Procedure : Text And Methodology”

USP 38 chapter <1225> validation of compendial procedures.

USP 38 chapter <1226> verification of compendial procedures.

6.0   GLOSSARY :

SOP             : Standard Operating Procedure

AD                : Analytical Development

R & D           : Research and development

LOQ             : Limit of Quantitation

RSD             : Relative standard deviation

RPM             : Rotation per minute

LOD             : Limit of detection

SST              : System suitability test

SD                : Standard Deviation

7.0   ANNEXURES:

Annexure I: Template for Preparation of Analytical Method Validation (AMV) Protocol

Sr. No. Subject Page No.
1. Protocol Approval
2. Objective
3. Scope
4. Responsibility of validation team
5. Product profile
6. Methodology
7. Incident/Deviation
8. Summary
9. Revision history

Annexure II: Template for Preparation of Analytical Method Validation (AMV) Report

APPROVAL:

Prepared By:

Functional Area Name Designation Signature/Date
Quality Control    

 Reviewed By:

Functional Area Name Designation Signature/Date
Quality Assurance    
Head Quality Control    

 Approved By:

Functional Area Name Designation Signature/Date
Head QA      

 

Sr. No. Subject
  1 Approval
2 Introduction
3 Objective
4 Summary Table
5 Methodology
6 Equipment & Materials
7 Validated parameters
8 Summary of system suitability
9 Incident/Deviation
10 Final Conclusion
11 Annexure
12 Revision History

 

1.     INTRODUCTION :
2.    OBJECTIVE :
3.    SUMMARY TABLE :
4.    METHODOLOGY :
5.    EQUIPMENT & MATERIALS :
6.    VALIDATED PARAMETERS :
7.    SUMMARY OF SYSTEM SUITABILITY :
8.    INCIDENT/DEVIATION :
9.    FINAL CONCLUSION :
10.  ANNEXURE :
11.  REVISION HISTORY

 

 

 

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pharmabeginers

Janki Singh is experienced in Pharmaceuticals, author and founder of Pharma Beginners, an ultimate pharmaceutical blogging platform. Email: [email protected]

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