Verification of applicability of the validated/compendial API analytical method for the final formulation

Assay, dissolution test and degradants

Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Guidelines
ICH Q2A
Validation of Analytical Methods: Definitions and Terminology (CPMP/ICH/381/95)

ICH Q2B
Validation of Analytical Procedures: Methodology (CPMP/ICH/281/95)

ICH Q6A
Specifications: Test Procedures and Acceptance Criteria for New Drug Substances and New Drug Products: Chemical Substances (CPMP/ICH/367/96 corr)
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

API
Assay
Validation with respect to:
Specificity, linearity/range, accuracy, precision, robustness

Impurities
Validation with respect to:
Specificity, linearity/range, accuracy, precision, limit of detection (LOD), limit of quantitation (LOQ), robustness
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

FPP
Formulation of the drug product
Presence of further APIs Presence of excipients (individual formulation) Presence of known impurities/degradants of all APIs and potential new degradants or incompatibility products

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Requirements
Capability of the analytical method(s):
Assay of each API in the presence of the other APIs and all impurities/degradants Assay of each degradant in the presence of all APIs and all other degradants/impurities Influence of formulation components should be excluded/controlled

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Revalidation I
Revalidation of analytical methods with respect to:
Specificity
presence of new API(s) and impurities/degradants/formulation components

Range
test concentrations of API(s) versus FPP

Accuracy
influence of formulation components

Precision
influence of formulation and sample preparation

LOD/LOQ
test concentrations of API(s) versus FPP)

Robustness
change of column material, column parameters, solvents)
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Revalidation II
Revalidation reflected by ICH Q2A:
Revalidation may be necessary in the following circumstances:
Changes in the synthesis of the drug substance Changes in the composition of the finished product Changes in the analytical procedure
(e.g. robustness)

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Specificity
Identification
Discrimination between compounds of closely related structures
positive results (from samples containing the analyte) negative results (from samples that do not contain the analyte) components structurally similar to the analyte do not give positive results

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Specificity II
Assay and impurities
Chromatographic procedures
Representative chromatograms with appropriate labelling of individual components Investigation at an appropriate level

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Specificity III
Chromatogram with retention times and chemical structures of: (1) arteannuin B (2) artemisitene (3) artemisinin (4) artemisinic acid (5) artemether (IS)
Analytical standard containing 1.2g/ml of each analyte and 0.4 g/ml IS From: F.C.W. Van Nieuverburgh et al., J Chromatogr. A 1118 (2006) 180-187
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Specificity IV
Assay and impurities/degradants
Discrimination of analytes where impurities/degradants are available
Assay
Demonstration of discrimination of the analyte in the presence of all impurities/degradants and/or excipients f. ex. assay result unaffected by presence of spiked impurities/degradants: - Injection of pure API - Injection of API plus impurities/degradants

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Specificity V
Assay and impurities/degradants
Discrimination of analytes where impurities/degradants are available
Impurities/Degradants
Demonstration of separation of impurities/degradants individually and/or from excipients f. ex. spiking of API with appropriate levels of impurities/degradants and/or excipients: Chromatographic profiles of API with and without impurities/degradants/excipients

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Specificity VI
Assay and impurities/degradants
Discrimination of analytes where impurities/degradants are not available
Comparison of the test procedure to a second well-characterized (independent) procedure
Samples Test samples containing impurities/degradants Test samples stored under relevant stress conditions (potential degradants arising during shelf life)

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Specificity VII
Assay and impurities/degradants
Discrimination of analytes where impurities/degradants are not available
Assay
Comparison of test results by the two independent procedures

Impurities/Degradants
Comparison of impurity profiles

Peak purity assessment

Demonstration that the analyte peak is attributable to only one component
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Specificity VIII
Peak purity Overlapping peaks in HPLC (simulation)
A B

From: Prof. Siegfried Ebel, University of Wuerzburg, in: Stavros Kromidas, Validierung in der Analytik, Wiley-VCH
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Specificity IX
Peak purity
Fatty acids were reacted with ethylene oxide and separated by HPLC (Fractions 1-6)

Fraction 5 was analysed by MALDI

From: Dr. Michael Schmitt, Henkel KGaA, Dsseldorf, in: Stavros Kromidas, Validierung in der Analytik, Wiley-VCH
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Specificity with FDCs

FDC (e.g. artesunate and amodiaquine)
One analytical method for both APIs
Capability of one method to quantify both APIs and to separate/discriminate one API and its impurities/degradants and potential incompatibility products from the other API and its impurities/degradants/incompatibility products
Some reference material for impurities/degradants will be available (spiking experiments applicable) Other degradants are not available as reference material (stress testing necessary to generate in situ degradants)
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Range
Minimum specified ranges
Assay
80 120% of the test concentration
Content uniformity 70 130% of the test concentration Dissolution Q-20% - 120%

Impurities/Degradants
Reporting level to 120% of specification limit

Revalidation is necessary, if the ranges covered during validation of the API-methods are different from those of the FPP-methods (different test concentrations)
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Accuracy
Assay
Application of the analytical procedure to synthetic mixtures of the product components (placebo mixture) to which known quantities of the analyte have been added In case certain product components are unavailable:
Application of the analytical procedure to the product to which known quantities of the analyte have been added Comparison of results obtained by a second (independent) procedure with defined accuracy
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Accuracy II
Impurities/Degradants
Assessment of samples spiked with known amounts of impurities/degradants In case certain impurities/degradation products are unavailable
Comparison of results obtained by a second (independent) procedure with defined accuracy

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Precision
Assay and impurities/degradants
Repeatability
9 determinations (3 x 3) covering the specified range or 6 determinations at 100% of the test concentration

Intermediate precision
Effects of random events on the precision of the procedure, e.g.
Days Analysts Equipment

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

Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Detection Limit
Determination based on
Visual evaluation (non-instrumental and instrumental methods) Signal to Noise (baseline noise) Standard deviation of response (s) and slope (S)
DL=3.3s/S
Estimation of S from the calibration curve of the analyte Estimation of s from the standard deviation of the blank from the standard deviation (regression line or y-intercept) of a calibration curve in the range of the DL
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Quantitation Limit
Determination based on
Visual evaluation (non-instrumental and instrumental methods) Signal to Noise (baseline noise) Standard deviation of response (s) and slope (S)
QL=10s/S
Estimation of S from the calibration curve of the analyte Estimation of s from the standard deviation of the blank from the standard deviation (regression line or y-intercept) of a calibration curve in the range of the QL
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Robustness I
Reliability of an analysis with respect to deliberate variations in method parameters
Susceptibility to variations in analytical conditions?
control of analytical conditions or precautionary statement establishment of system suitability parameters

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Robustness II
Examples of variations
Stability of analytical solutions Extraction time

In the case of liquid chromatography

Influence of variations of pH in a mobile phase Influence of variations in mobile phase composition Influence of columns (different lots and/or suppliers) Influence of temperature Influence of flow rate

In the case of gas chromatography

Influence of columns (different lots and/or suppliers) Influence of temperature Influence of flow rate
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Robustness III
Influence of pH of elution on separation of amino acids by RP-HPLC

From: Waters, in: Stavros Kromidas, Validierung in der Analytik, Wiley-VCH

Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Robustness
Electropherograms under identical conditions by different analytical equipment

From: Dr. Michael Krmer, NOVARTIS, Basel, in: Stavros Kromidas, Validierung in der Analytik, Wiley-VCH
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Dissolution
Applicability of the analytical method used for assay and impurities/degradants
Sample preparation Range

Applicability of the dissolution method

Appropriateness of drug release acceptance criteria
Solubility criteria of the APIs

Appropriateness of test conditions and acceptance criteria

Dissolution affecting bioavailability Changes in formulation or manufacturing variables affecting dissolution
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Dissolution II
Applicability of the analytical method used for assay and impurities/degradants
Potential parameters for revalidation
Sample preparation
Stability of analytes in the dissolution medium? Preparation of an injectable sample volume according to the analytical method? Precision of analysis of the prepared dissolution sample?

Range of test concentrations of API / impurities / degradants according to the validated ranges?
Test concentration of prepared dissolution sample versus test concentration of FPP sample
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Dissolution III
Applicability of the dissolution method
Appropriateness of drug release acceptance criteria
Solubility of the APIs (ICH Q6A Definitions)
Rapidly dissolving products Not less than 80% of the label amount of the drug substance dissolves within 15 minutes in each of the following media: pH 1.2, pH 4.0, pH 6.8 Highly water soluble drugs Drugs with a dose/solubility volume of less than or equal to 250 ml over a pH range of 1.2 to 6.8 Low solubility drugs Drugs with a dose/solubility volume of more than 250 ml
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Dissolution IV
Appropriateness of drug release acceptance criteria
Solubility of the APIs
Problem with low solubility drugs:
Solution of the drugs may become a time-limiting step Dissolution also dependent on the strength of the drug product Dissolution test cannot reflect batch to batch consistency

Possible solution of the problem

Extending the dissolution volume and Validation of the dissolution procedure with extended volume (applicability of the pharmacopoeial procedure)

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Dissolution V
Sink conditions
Ph. Eur 2.9.3: ..the material already in solution does not exert a significant modifying effect on the rate of dissolution of the remainder Sink conditions normally occur in a volume of dissolution medium that is at least 3 to 10 times the saturation medium Consequently: the amount of API contained in the dosage form should be soluble in NMT 300 ml of dissolution medium

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Dissolution V
Applicability of the dissolution method
Appropriateness of test conditions and acceptance criteria (ICH Q6A)
Dissolution significantly affecting bioavailability
Have relevant developmental batches exhibited unacceptable bioavailability? Development of test conditions and acceptance criteria which can distinguish batches with unacceptable bioavailability

Changes in formulation or manufacturing variables affecting dissolution

Control of these changes by another procedure and acceptance criterion or Development of test conditions and acceptance criteria which can distinguish these changes
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Major problems
Solubility of Artemisinins
Sink condition cannot be established
(+) Addition of solubilizers could help establish a (dis)solution test (-) The test would disconnect dissolution and bioavalability and could only serve as parameter for batch to batch consistency Disintegration could be considered as additional parameter

Stability of Artemisinins
Artesunate decomposes (to DHA) in buffers required for dissolution testing (e.g. pH 1.2, pH 4.5)
Dissolution could only be performed at a neutral pH (~ 7.0)
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Deficiencies from PQ
Validation of precision
Precision of the drug substance solution lower than precision of the drug product solution Acceptance criteria for precision of the drug substance solution wider than for precision of the drug product solution Acceptance criteria much wider than real values assessed Acceptance criteria of assay specifications and precision do not match
(3 x RSD outside the specification range)
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

Deficiencies from PQ II
Assay of API and impurities/degradants
No acceptable mass balance found between assay of API and impurities/degradants Quantitation limit of impurities too high
ICH requirement on threshold for identification and qualification of unknown impurities cannot be fulfilled

Dar es Salaam, August, 21-25, 2006

Dr. Birgit Schmauser, BfArM, Bonn

Deficiencies from PQ III

Dissolution
Necessary information on development of dissolution test not presented Dissolution method (pharmacopoeial) not presented along with development of dissolution test and/or validation of applicability of analytical methods Test conditions and acceptance criteria of the dissolution test not justified
Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn

THANK YOU FOR YOUR ATTENTION

Dar es Salaam, August, 21-25, 2006 Dr. Birgit Schmauser, BfArM, Bonn