Verification of applicability of the Verification of applicability of the validated/compendialvalidated/compendialAPI analytical methodAPI analytical methodfor the final formulationfor the final formulation

(assay, dissolution test and (assay, dissolution test and degradants)degradants)

GuidelinesGuidelines

ICH Q2AICH Q2A Validation of Analytical Methods: Definitions and Validation of Analytical Methods: Definitions and

Terminology (CPMP/ICH/381/95)Terminology (CPMP/ICH/381/95)

ICH Q2BICH Q2B Validation of Analytical Procedures: Methodology Validation of Analytical Procedures: Methodology

(CPMP/ICH/281/95)(CPMP/ICH/281/95)

ICH Q6AICH Q6A Specifications: Test Procedures and Acceptance Specifications: Test Procedures and Acceptance

Criteria for New Drug Substances and New Drug Criteria for New Drug Substances and New Drug Products: Chemical SubstancesProducts: Chemical Substances(CPMP/ICH/367/96 corr)(CPMP/ICH/367/96 corr)

APIAPI

AssayAssay Validation with respect to:Validation with respect to:

Specificity, linearity/range, accuracy, precision, Specificity, linearity/range, accuracy, precision, robustnessrobustness

ImpuritiesImpurities Validation with respect to:Validation with respect to:

Specificity, linearity/range, accuracy, precision, Specificity, linearity/range, accuracy, precision, limit of detection (LOD), limit of quantitation (LOQ)limit of detection (LOD), limit of quantitation (LOQ), , robustnessrobustness

FPPFPP

New situation arising from the formulation New situation arising from the formulation of the drug productof the drug product Presence of Presence of further APIsfurther APIs Presence of Presence of excipientsexcipients (individual formulation) (individual formulation) Presence of Presence of knownknown impurities/degradants of impurities/degradants of

all APIs and potential all APIs and potential newnew degradants or degradants or incompatibility productsincompatibility products

RequirementsRequirements

Capability of the analytical method(s):Capability of the analytical method(s): Assay of Assay of each APIeach API in the presence of the in the presence of the

other APIs and all impurities/degradantsother APIs and all impurities/degradants Assay of Assay of each degradanteach degradant in the presence of in the presence of

all APIs and all other degradants/impuritiesall APIs and all other degradants/impurities Influence of Influence of formulation componentsformulation components should should

be excluded/controlledbe excluded/controlled

Revalidation IRevalidation I

Revalidation of analytical methods with respect to:Revalidation of analytical methods with respect to: SpecificitySpecificity

presence of new API(s) and impurities/degradants/formulation presence of new API(s) and impurities/degradants/formulation componentscomponents

RangeRangetest concentrations of API(s) versus FPPtest concentrations of API(s) versus FPP

AccuracyAccuracyinfluence of formulation componentsinfluence of formulation components

PrecisionPrecisioninfluence of formulation and sample preparationinfluence of formulation and sample preparation

LOD/LOQLOD/LOQtest concentrations of API(s) versus FPP)test concentrations of API(s) versus FPP)

RobustnessRobustnesschange of column material, column parameters, solvents)change of column material, column parameters, solvents)

Revalidation IIRevalidation II

Revalidation reflected by ICH Q2A:Revalidation reflected by ICH Q2A: Revalidation may be necessary in the Revalidation may be necessary in the

following circumstances:following circumstances: Changes in the synthesis of the drug substanceChanges in the synthesis of the drug substance

Changes in the composition of the finished Changes in the composition of the finished productproduct

Changes in the analytical procedureChanges in the analytical procedure (e.g. robustness)(e.g. robustness)

SpecificitySpecificity

IdentificationIdentification Discrimination between compounds of closely Discrimination between compounds of closely

related structuresrelated structures positive results (from samples containing the positive results (from samples containing the analyte)analyte)

negative results (from samples that do not contain negative results (from samples that do not contain the analyte)the analyte)

components structurally similar to the analyte do components structurally similar to the analyte do not give positive resultsnot give positive results

Specificity IISpecificity II

Assay and impuritiesAssay and impurities Chromatographic proceduresChromatographic procedures

Representative chromatograms with Representative chromatograms with appropriate appropriate labellinglabelling of individual components of individual components

Investigation at an Investigation at an appropriate levelappropriate level

Specificity IIISpecificity III

Assay and impurities/degradantsAssay and impurities/degradants Discrimination of analytes where Discrimination of analytes where

impurities/degradants are availableimpurities/degradants are available AssayAssay

Demonstration of discrimination of the analyte in the Demonstration of discrimination of the analyte in the presence of all impurities/degradants and/or excipientspresence of all impurities/degradants and/or excipients

f. ex. assay result unaffected by presence of spiked f. ex. assay result unaffected by presence of spiked impurities/degradantsimpurities/degradants

Specificity IVSpecificity IV

Assay and impurities/degradantsAssay and impurities/degradants Discrimination of analytes where Discrimination of analytes where

impurities/degradants are availableimpurities/degradants are availableImpurities/DegradantsImpurities/Degradants

Demonstration of separation of impurities/degradants Demonstration of separation of impurities/degradants individually and/or from excipientsindividually and/or from excipients

f. ex. spiking of API with appropriate levels of f. ex. spiking of API with appropriate levels of impurities/degradants and/or excipientsimpurities/degradants and/or excipients

Specificity VSpecificity V

Assay and impurities/degradantsAssay and impurities/degradants Discrimination of analytes where Discrimination of analytes where

impurities/degradants are impurities/degradants are notnot available available Comparison of the test procedure to a second Comparison of the test procedure to a second well-characterized (independent) procedurewell-characterized (independent) procedure

SamplesSamples

Test samples containing impurities/degradantsTest samples containing impurities/degradants

Test samples stored under relevant stress conditions Test samples stored under relevant stress conditions (potential degradants arising during shelf life)(potential degradants arising during shelf life)

Specificity VISpecificity VI

Assay and impurities/degradantsAssay and impurities/degradants Discrimination of analytes where Discrimination of analytes where

impurities/degradants are impurities/degradants are notnot available available AssayAssay

Comparison of Comparison of test resultstest results by the two independent by the two independent proceduresprocedures

Impurities/DegradantsImpurities/Degradants Comparison of Comparison of impurity profilesimpurity profiles

Peak purity assessmentPeak purity assessment Demonstration that the analyte peak is attributable to Demonstration that the analyte peak is attributable to

only one componentonly one component

ExampleExample

FDC (artesunate and amodiaquine)FDC (artesunate and amodiaquine) One analytical method for both APIsOne analytical method for both APIs

Capability of one method to quantify both APIs and Capability of one method to quantify both APIs and to separate/discriminate the other API and its to separate/discriminate the other API and its impurities/degradants and potential incompatibility impurities/degradants and potential incompatibility products from the existing API and its products from the existing API and its impurities/degradantsimpurities/degradants

Some reference material for impurities/degradants will be Some reference material for impurities/degradants will be available (spiking experiments applicable)available (spiking experiments applicable)

Other degradants are not available as reference material Other degradants are not available as reference material (stress testing necessary to generate in situ degradants)(stress testing necessary to generate in situ degradants)

RangeRange

Minimum specified rangesMinimum specified ranges AssayAssay

80 – 120% of the 80 – 120% of the test concentrationtest concentration Content uniformityContent uniformity

70 – 130% of the 70 – 130% of the test concentrationtest concentration DissolutionDissolution

Q-20% - 120%Q-20% - 120% Impurities/DegradantsImpurities/Degradants

Reporting level to 120% of specification limitReporting level to 120% of specification limit

Revalidation is necessary, if the ranges covered Revalidation is necessary, if the ranges covered during validation of the API-methods are during validation of the API-methods are different from those of the FPP-methodsdifferent from those of the FPP-methods(different test concentrations)(different test concentrations)

AccuracyAccuracy

AssayAssay Application of the analytical procedure to Application of the analytical procedure to synthetic synthetic

mixturesmixtures of the product components (placebo of the product components (placebo mixture) to which known quantities of the analyte mixture) to which known quantities of the analyte have been addedhave been added

In case certain product components are In case certain product components are unavailable:unavailable:

Application of the analytical procedure to the Application of the analytical procedure to the productproduct to to which known quantities of the analyte have been addedwhich known quantities of the analyte have been added

Comparison of results obtained by a Comparison of results obtained by a secondsecond (independent) (independent) procedureprocedure with defined accuracy with defined accuracy

Accuracy IIAccuracy II

Impurities/DegradantsImpurities/Degradants Assessment of samples Assessment of samples spikedspiked with known with known

amounts of impurities/degradantsamounts of impurities/degradants In case certain impurities/degradation In case certain impurities/degradation

products are unavailableproducts are unavailableComparison of results obtained by a second Comparison of results obtained by a second (independent) (independent) procedureprocedure with defined accuracy with defined accuracy

PrecisionPrecision

Assay and impurities/degradantsAssay and impurities/degradants RepeatabilityRepeatability

9 determinations (3 x 3) covering the specified range9 determinations (3 x 3) covering the specified rangeoror6 determinations at 100% of the test concentration6 determinations at 100% of the test concentration

Intermediate precisionIntermediate precisionEffects of random events on the precision of the procedure, Effects of random events on the precision of the procedure, e.g.e.g.

DaysDays AnalystsAnalysts EquipmentEquipment

To be performed with aTo be performed with a test solution prepared test solution prepared from the drug productfrom the drug product

Detection LimitDetection Limit

Determination based on Determination based on Visual evaluation (non-instrumental and instrumental Visual evaluation (non-instrumental and instrumental

methods)methods) Signal to Noise (Signal to Noise (baseline noisebaseline noise)) Standard deviation of response (Standard deviation of response () and) and

slope (S)slope (S)DL=3.3DL=3.3/S/S

Estimation of SEstimation of Sfrom the calibration curve of the analytefrom the calibration curve of the analyte

Estimation of Estimation of from the standard deviation of the from the standard deviation of the blankblankfrom the standard deviation (regression line or y-intercept) from the standard deviation (regression line or y-intercept) of a calibration curve in the range of the DLof a calibration curve in the range of the DL

Quantitation LimitQuantitation Limit

Determination based on Determination based on Visual evaluation (non-instrumental and instrumental Visual evaluation (non-instrumental and instrumental

methods)methods) Signal to Noise (Signal to Noise (baseline noisebaseline noise)) Standard deviation of response (Standard deviation of response () and) and

slope (S)slope (S)QL=10QL=10/S/S

Estimation of SEstimation of Sfrom the calibration curve of the analytefrom the calibration curve of the analyte

Estimation of Estimation of from the standard deviation of the from the standard deviation of the blankblankfrom the standard deviation (regression line or y-intercept) from the standard deviation (regression line or y-intercept) of a calibration curve in the range of the QLof a calibration curve in the range of the QL

Robustness IRobustness I

Reliability of an analysis with respect to Reliability of an analysis with respect to deliberate variations in method parametersdeliberate variations in method parameters Susceptibility to variations in analytical Susceptibility to variations in analytical

conditions?conditions?control of analytical conditionscontrol of analytical conditionsororprecautionary statementprecautionary statement

establishment of establishment of system suitability parameterssystem suitability parameters

Robustness IIRobustness II

Examples of variationsExamples of variations Stability of analytical solutionsStability of analytical solutions Extraction timeExtraction time

In the case of liquid chromatographyIn the case of liquid chromatography Influence of variations of pH in a mobile phaseInfluence of variations of pH in a mobile phase Influence of variations in mobile phase compositionInfluence of variations in mobile phase composition Influence of columns (different lots and/or suppliers)Influence of columns (different lots and/or suppliers) Influence of temperatureInfluence of temperature Influence of flow rateInfluence of flow rate

In the case of gas chromatographyIn the case of gas chromatography Influence of columns (different lots and/or suppliers)Influence of columns (different lots and/or suppliers) Influence of temperatureInfluence of temperature Influence of flow rateInfluence of flow rate

DissolutionDissolution

Applicability of the Applicability of the analytical methodanalytical method used for used for assay and impurities/degradantsassay and impurities/degradants Sample preparationSample preparation RangeRange

Applicability of the Applicability of the dissolution methoddissolution method Appropriateness of Appropriateness of drug releasedrug release acceptance criteriaacceptance criteria

Solubility criteria of the APIsSolubility criteria of the APIs Appropriateness of Appropriateness of test conditionstest conditions andand acceptance acceptance

criteriacriteriaDissolution affecting bioavailabilityDissolution affecting bioavailabilityChanges in formulation or manufacturing variables affecting Changes in formulation or manufacturing variables affecting dissolutiondissolution

Dissolution IIDissolution II

Applicability of the analytical method used Applicability of the analytical method used for assay and impurities/degradantsfor assay and impurities/degradants Potential parameters for revalidationPotential parameters for revalidation

SampleSample preparation preparation Stability of analytes in the dissolution medium?Stability of analytes in the dissolution medium? Preparation of an injectable sample volume according to Preparation of an injectable sample volume according to

the analytical method?the analytical method? Precision of analysis of the prepared dissolution sample?Precision of analysis of the prepared dissolution sample?

RangeRange of test concentrations of API / impurities / of test concentrations of API / impurities / degradants according to the validated ranges?degradants according to the validated ranges?

Test concentration of prepared dissolution sample Test concentration of prepared dissolution sample versus test concentration of FPP sampleversus test concentration of FPP sample

Dissolution IIIDissolution III

Applicability of the dissolution methodApplicability of the dissolution method Appropriateness of drug release acceptance criteriaAppropriateness of drug release acceptance criteria

Solubility of the APIs (ICH Q6A Definitions)Solubility of the APIs (ICH Q6A Definitions) Rapidly dissolving productsRapidly dissolving products

Not less than 80% of the label amountNot less than 80% of the label amount of the drug of the drug substance dissolves substance dissolves within 15 minuteswithin 15 minutes in each of the in each of the following media: following media: pH 1.2, pH 4.0, pH 6.8pH 1.2, pH 4.0, pH 6.8

Highly water soluble drugsHighly water soluble drugs

Drugs with a Drugs with a dose/solubility volumedose/solubility volume of of less than or equal to less than or equal to 250 ml250 ml over a over a pH range of 1.2 to 6.8pH range of 1.2 to 6.8

Low solubility drugsLow solubility drugs

Drugs with a dose/solubility volume of Drugs with a dose/solubility volume of more thanmore than250 ml250 ml

Dissolution IVDissolution IV

Appropriateness of drug release acceptance Appropriateness of drug release acceptance criteriacriteria Solubility of the APIsSolubility of the APIs

ProblemProblem with low solubility drugs: with low solubility drugs: Solution of the drugs Solution of the drugs may become a time-limiting stepmay become a time-limiting step

Dissolution also dependent on the strength of the drug Dissolution also dependent on the strength of the drug productproductDissolution test cannot reflect batch to batch consistencyDissolution test cannot reflect batch to batch consistency

Possible solution of the problemPossible solution of the problem Extending the dissolution volume beyond the time-limiting Extending the dissolution volume beyond the time-limiting

volumevolume Validation of the dissolution procedure with extended volume Validation of the dissolution procedure with extended volume

(applicability of the pharmacopoeial procedure) (applicability of the pharmacopoeial procedure)

Dissolution VDissolution V

Applicability of the dissolution methodApplicability of the dissolution method Appropriateness of test conditions and acceptance Appropriateness of test conditions and acceptance

criteria (criteria (ICH Q6AICH Q6A))Dissolution significantly affecting bioavailabilityDissolution significantly affecting bioavailability

Have relevant developmental batches exhibited unacceptable Have relevant developmental batches exhibited unacceptable bioavailability?bioavailability?

Development of test conditions and acceptance criteria Development of test conditions and acceptance criteria which can distinguish batches with unacceptable which can distinguish batches with unacceptable bioavailabilitybioavailability

Changes in formulation or manufacturing variables affecting Changes in formulation or manufacturing variables affecting dissolutiondissolution

Control of these changes by another procedure and acceptance Control of these changes by another procedure and acceptance criterioncriterionoror

Development of test conditions and acceptance criteria which Development of test conditions and acceptance criteria which can distinguish these changescan distinguish these changes

Deficiencies from prequalificationDeficiencies from prequalification

Validation of precisionValidation of precision Precision of the drug substance solution lower than Precision of the drug substance solution lower than

precision of the drug product solutionprecision of the drug product solution Acceptance criteria for precision of the drug Acceptance criteria for precision of the drug

substance solution wider than for precision of the drug substance solution wider than for precision of the drug product solutionproduct solution

Acceptance criteria much wider than real values Acceptance criteria much wider than real values assessedassessed

Acceptance criteria of assay specifications and Acceptance criteria of assay specifications and precision do not matchprecision do not match

(3 x RSD outside the specification range) (3 x RSD outside the specification range)

Deficiencies from prequalification IIDeficiencies from prequalification II

Assay of API and impurities/degradantsAssay of API and impurities/degradants No acceptable mass balance found between No acceptable mass balance found between

assay of API and impurities/degradantsassay of API and impurities/degradants Quantitation limit of impurities too highQuantitation limit of impurities too high

ICH requirement on threshold for identification and ICH requirement on threshold for identification and qualification of unknown impurities cannot be qualification of unknown impurities cannot be fulfilledfulfilled

Deficiencies from prequalification IIIDeficiencies from prequalification III

DissolutionDissolution Necessary information on development of Necessary information on development of

dissolution test not presenteddissolution test not presented Dissolution method (pharmacopoeial) not Dissolution method (pharmacopoeial) not

presented along with development of presented along with development of dissolution test and/or validation of dissolution test and/or validation of applicability of analytical methodsapplicability of analytical methods

Test conditions and acceptance criteria of the Test conditions and acceptance criteria of the dissolution test not justifieddissolution test not justified