《仪器与方法验证专著》（英文版） Analytical Method Validation and Instrument Performance veriticatig

POTENCY METHOD VALIDATION three concentration levels covering the specified range (e.g, three concentra tions/three replicates) For a drug substance, the common method of determining accuracy is to apply the analytical procedure to the drug substance and to quantitate it against reference standard of known purity. For the drug product, accuracy is usually determined by application of the analytical procedure to synthetic mixtures of the drug product components or placebo dosage form to which known quantities of drug substance of known purity have been added. The range for the accuracy imit should be within the linear range. Typical accuracy of the recovery of the drug substance in the mixture is expected to be about 98 to 102%0. Values of accuracy of the recovery data beyond this range need to be investigated 2. 4.3 Precision The precision of an analytical procedure expresses the closeness of agreement legree of scatter) between a series of measurements obtained from multiple samples of the same homogeneous sample under prescribed conditions. Preci- sion is usually investigated at three levels: repeatability, intermediate precision and reproducibility. For simple formulation it is important that precision be deter- mined using authentic homogeneous samples. A justification will be required if homogeneous sample is not possible and artificially prepared samples or sample solutions are used Repeatability(Precision ). Repeatability is a measure of the precision under the e operating conditions over a short interval of time. It is sometimes referred to as intraassay precision. Two assaying options are allowed by the ich fo A minimum of nine determinations covering the specified range for the procedure (e.g, three concentrations/three replicates as in the accuracy ), 2. A minimum of six determinations at 100%o of the test concentration The standard deviation, relative standard deviation(coefficient of variation), nd confidence interval should be reported as required by the ICh Tables 2.2 and 2.3 are examples of repeatability data. Table 2.2 shows repeatability data. However, note that the data show a slight bias below (all data between 97.5 and 99. 1%0). This may not be an issue, as the true of the samples and the variation of the assay may be between 97.5 and 99.1%0 Table 2 3 shows two sets of data for a formulation at two dose strengths that were performed ets of six determinations at 100%o test concentration The data indicate a definite bias and high variability for the low-strength dose formulation It may call into question the appropriateness of the low-dose samples for the validation experiment

18 POTENCY METHOD VALIDATION three concentration levels covering the specified range (e.g., three concentra￾tions/three replicates). For a drug substance, the common method of determining accuracy is to apply the analytical procedure to the drug substance and to quantitate it against a reference standard of known purity. For the drug product, accuracy is usually determined by application of the analytical procedure to synthetic mixtures of the drug product components or placebo dosage form to which known quantities of drug substance of known purity have been added. The range for the accuracy limit should be within the linear range. Typical accuracy of the recovery of the drug substance in the mixture is expected to be about 98 to 102%. Values of accuracy of the recovery data beyond this range need to be investigated. 2.4.3 Precision The precision of an analytical procedure expresses the closeness of agreement (degree of scatter) between a series of measurements obtained from multiple samples of the same homogeneous sample under prescribed conditions. Preci￾sion is usually investigated at three levels: repeatability, intermediate precision, and reproducibility. For simple formulation it is important that precision be deter￾mined using authentic homogeneous samples. A justification will be required if a homogeneous sample is not possible and artificially prepared samples or sample solutions are used. Repeatability (Precision). Repeatability is a measure of the precision under the same operating conditions over a short interval of time. It is sometimes referred to as intraassay precision. Two assaying options are allowed by the ICH for investigating repeatability: 1. A minimum of nine determinations covering the specified range for the procedure (e.g., three concentrations/three replicates as in the accuracy experiment), or 2. A minimum of six determinations at 100% of the test concentration. The standard deviation, relative standard deviation (coefficient of variation), and confidence interval should be reported as required by the ICH. Tables 2.2 and 2.3 are examples of repeatability data. Table 2.2 shows good repeatability data. However, note that the data show a slight bias below 100% (all data between 97.5 and 99.1%). This may not be an issue, as the true value of the samples and the variation of the assay may be between 97.5 and 99.1%. Table 2.3 shows two sets of data for a formulation at two dose strengths that were performed using sets of six determinations at 100% test concentration. The data indicate a definite bias and high variability for the low-strength dose formulation. It may call into question the appropriateness of the low-dose samples for the validation experiment

STRATEGIES AND VALIDATION PARAMETERS Table 2.2. Repeatability at Different Concentration Concentration (Nominal Concentration 75ug/mL) Replicate14.829.644.574.51491223.7 97998.397.698.398.7986 98098497598.298298 23456 98.2 99.699 198.199 97998899.1988977988 98498.597.899098.099.2 99.198698498398.198.7 Mean98398.598.398698.1 %RSD0.470.180.870.420.33 Table 2.3. Repeatability at High and Low Concen trations Replicate Low Dose High Dose 100.6 1.8 23456 94.1 100.5 93.8 1014 94.7 94.1 %O RSD Intermediate Precision. Intermediate precision is defined as the variation within the laboratory. The extent to which intermediate pred needs to be stablished depends on the circumstances under which the procedure is intended to be used. Typical parameters that are investigated include day-to-day varia- tion, analyst variation, and equipment variation. Depending on the extent of the study, the use of experimental design is encouraged. Experimental design will minimize the number of experiments that need to be performed. It is important to note that the ICh allows exemption from doing intermediate precision when reproducibility is proven. It is expected that the intermediate precision should show variability that is in the same range or less than repeatability variation. The ICH recommended the reporting of standard deviation, relative standard deviation (coefficient of variation), and confidence interval of the data. Reproducibility. Reproducibility measures the precision between laboratories as in collaborative studies. This parameter should be considered in the standardiza tion of an analytical procedure(e.g, inclusion of procedures in pharmacopoeias

STRATEGIES AND VALIDATION PARAMETERS 19 Table 2.2. Repeatability at Different Concentration Concentration (Nominal Concentration 75µg/mL) Replicate 14.8 29.6 44.5 74.5 149.1 223.7 1 97.9 98.3 97.6 98.3 98.7 98.6 2 98.0 98.4 97.5 98.2 98.2 98.5 3 98.2 98.5 99.6 99.1 98.1 99.0 4 97.9 98.8 99.1 98.8 97.7 98.8 5 98.4 98.5 97.8 99.0 98.0 99.2 6 99.1 98.6 98.4 98.3 98.1 98.7 Mean 98.3 98.5 98.3 98.6 98.1 98.8 % RSD 0.47 0.18 0.87 0.42 0.33 0.28 Table 2.3. Repeatability at High and Low Concen￾trations Replicate Low Dose High Dose 1 94.8 100.6 2 91.8 102.1 3 94.1 100.5 4 93.8 99.4 5 95.3 101.4 6 94.7 101.1 Mean 94.1 100.9 % RSD 1.30 0.90 Intermediate Precision. Intermediate precision is defined as the variation within the same laboratory. The extent to which intermediate precision needs to be established depends on the circumstances under which the procedure is intended to be used. Typical parameters that are investigated include day-to-day varia￾tion, analyst variation, and equipment variation. Depending on the extent of the study, the use of experimental design is encouraged. Experimental design will minimize the number of experiments that need to be performed. It is important to note that the ICH allows exemption from doing intermediate precision when reproducibility is proven. It is expected that the intermediate precision should show variability that is in the same range or less than repeatability variation. The ICH recommended the reporting of standard deviation, relative standard deviation (coefficient of variation), and confidence interval of the data. Reproducibility. Reproducibility measures the precision between laboratories as in collaborative studies. This parameter should be considered in the standardiza￾tion of an analytical procedure (e.g., inclusion of procedures in pharmacopoeias

POTENCY METHOD VALIDATION and method transfer between different laboratories). To validate this character istic, similar studies need to be performed at other laboratories using the same homogeneous sample lot and the same experimental design. In the case of method transfer between two laboratories, different approaches may be taken to achieve the successful transfer of the procedure. However, the most common approach is the direct method transfer from the originating laboratory to the receiving labo- ratory. The originating laboratory is defined as the laboratory that has developed and validated the analytical method or a laboratory that has previously been cer- tified to perform the procedure and will participate in the method transfer studies The receiving laboratory is defined as the laboratory to which the analytical pro cedure will be transferred and that will participate in the method transfer studies In direct method transfer it is recommended that a protocol be initiated with details of the experiments to be performed and acceptance criteria(in terms of the difference between the means of the two laboratories) for passing the method transfer. Table 2. 4 gives a set of sample data where the average results obtained between two laboratories were within 0.5%0 2. 4. 4 Robustness The robustness of an analytical procedure is a measure of its capacity to remain unaffected by small but deliberate variations in the analytical procedure param- eters. The robustness of the analytical procedure provides an indication of its reliability during normal use. The evaluation of robustness should be considered during development of the analytical procedure. If measurements are susceptible to variations in analytical conditions, the analytical conditions should be suitably controlled or a precautionary statement should be included in the procedure. For example, if the resolution of a critical pair of peaks was very sensitive to the percentage of organic composition in the mobile phase, that observation would have been observed during method development and should be stressed in the procedure. Common variations that are investigated for robustness include filter effect, stability of analytical solutions, extraction time during sample prepara tion, pH variations in the mobile-phase composition, variations in mobile-phase composition, columns, temperature effect, and flow rate Table 2.5 shows examples of sample and standard stability performed on an analytical procedure. The two sets of data indicate that the sample and standard Table 2, 4. Results from method Transfer between two Laboratories 100.7 laboratory Receiving

20 POTENCY METHOD VALIDATION and method transfer between different laboratories). To validate this character￾istic, similar studies need to be performed at other laboratories using the same homogeneous sample lot and the same experimental design. In the case of method transfer between two laboratories, different approaches may be taken to achieve the successful transfer of the procedure. However, the most common approach is the direct method transfer from the originating laboratory to the receiving labo￾ratory. The originating laboratory is defined as the laboratory that has developed and validated the analytical method or a laboratory that has previously been cer￾tified to perform the procedure and will participate in the method transfer studies. The receiving laboratory is defined as the laboratory to which the analytical pro￾cedure will be transferred and that will participate in the method transfer studies. In direct method transfer it is recommended that a protocol be initiated with details of the experiments to be performed and acceptance criteria (in terms of the difference between the means of the two laboratories) for passing the method transfer. Table 2.4 gives a set of sample data where the average results obtained between two laboratories were within 0.5%. 2.4.4 Robustness The robustness of an analytical procedure is a measure of its capacity to remain unaffected by small but deliberate variations in the analytical procedure param￾eters. The robustness of the analytical procedure provides an indication of its reliability during normal use. The evaluation of robustness should be considered during development of the analytical procedure. If measurements are susceptible to variations in analytical conditions, the analytical conditions should be suitably controlled or a precautionary statement should be included in the procedure. For example, if the resolution of a critical pair of peaks was very sensitive to the percentage of organic composition in the mobile phase, that observation would have been observed during method development and should be stressed in the procedure. Common variations that are investigated for robustness include filter effect, stability of analytical solutions, extraction time during sample prepara￾tion, pH variations in the mobile-phase composition, variations in mobile-phase composition, columns, temperature effect, and flow rate. Table 2.5 shows examples of sample and standard stability performed on an analytical procedure. The two sets of data indicate that the sample and standard Table 2.4. Results from Method Transfer between Two Laboratories Runs Average % Originating 12 100.7 laboratory Receiving 4 100.2 laboratory

STRATEGIES AND VALIDATION PARAMETERS Table 2.5. Stability of Sample and Standard Solutions Sample Standard 100.2 Table 26. effect of Filter Replication Unfiltered Filter 1 Filter 2 1010 101.1 100.5 101.1 97.1 101.2 96.4 verage 100.8 96.8 solutions were stable for 3 and 4 days respectively. Table 2. 6 gives some data on the effect of a filter on the recovery of the analytical procedure. In a filter study it is common to use the same solution and to compare a filtered solution to an unfiltered solution. For the unfiltered solution, it is common to centrifuge the sample solution and use the supernatant liquid for the analysis. The data set indicated that filter I would be recommended for the final analytical procedure 2.4.5 Specificity Specificity is the ability to assess unequivocally an analyte in the presence of components that may be expected to be present. In many publications, selectivity and specificity are often used interchangeably. However, there are debates over he use of specificity over selectivity [6]. For the purposes of this chapter, the definition of specificity will be consistent with that of the ICH The specificity of a test method is determined by comparing test results from an analysis of samples containing impurities, degradation products, or placebo ingredients with those obtained from an analysis of samples without impurities, degradation products, or placebo ingredients. For the purpose of a stability- indicating assay method, degradation peaks need to be resolved from the drug substance. However, they do not need to be resolved from each other Critical separations in chromatography should be investigated at the appro priate level. Specificity can best be demonstrated by the resolution of two chro- the peaks would be the analyte peak. Figure 2. 4 illustrates the selectivity of a method to resolve known degradation peaks from the parent peak. Based on the

STRATEGIES AND VALIDATION PARAMETERS 21 Table 2.5. Stability of Sample and Standard Solutions % Initial Day Sample Standard 1 100.2 99.8 2 100.0 99.8 3 99.9 100.2 4 — 99.5 Table 2.6. Effect of Filter Replication Unfiltered Filter 1 Filter 2 1 101.0 101.1 96.8 2 100.5 101.1 97.1 3 101.0 101.2 96.4 Average 100.8 100.2 96.8 solutions were stable for 3 and 4 days respectively. Table 2.6 gives some data on the effect of a filter on the recovery of the analytical procedure. In a filter study it is common to use the same solution and to compare a filtered solution to an unfiltered solution. For the unfiltered solution, it is common to centrifuge the sample solution and use the supernatant liquid for the analysis. The data set indicated that filter 1 would be recommended for the final analytical procedure. 2.4.5 Specificity Specificity is the ability to assess unequivocally an analyte in the presence of components that may be expected to be present. In many publications, selectivity and specificity are often used interchangeably. However, there are debates over the use of specificity over selectivity [6]. For the purposes of this chapter, the definition of specificity will be consistent with that of the ICH. The specificity of a test method is determined by comparing test results from an analysis of samples containing impurities, degradation products, or placebo ingredients with those obtained from an analysis of samples without impurities, degradation products, or placebo ingredients. For the purpose of a stability￾indicating assay method, degradation peaks need to be resolved from the drug substance. However, they do not need to be resolved from each other. Critical separations in chromatography should be investigated at the appro￾priate level. Specificity can best be demonstrated by the resolution of two chro￾mographic peaks that elute close to each other. In the potency assay, one of the peaks would be the analyte peak. Figure 2.4 illustrates the selectivity of a method to resolve known degradation peaks from the parent peak. Based on the

POTENCY METHOD VALIDATION 77.000 74.000 71.000 E品 68.000 65.000 56.000 Impurity standard solution 50.000 TTTTTTTTTTTTTTTTTTTTTTTTTT 2002803604405206006807608409201000 Figure 2.4. Overlay chromatogram of an impurity solution with a sample solution experience with the analyte and the chemistry of the analyte, the scientist will be able to identify which of the impurities may be used as the critical pair. 2.5 POTENCY METHOD REVALIDATION There are various situations during the life cycle of a potency method that require revalidation of the method 1. During optimization of the formulation or drug substance synthetic process significant changes may have to be introduced into the process. As a result, to ensure that the analytical method will still be able to analyze the poten- tially different profile of the drug substance or drug product, revalidation may be necessary. 2. The method was found to be deficient in some areas, such as precision and system suitability. This is especially important as the analytical laboratory gets more experience and more information as to the degradation profile of the sample as it progresses toward submission. If a new impurity is found that makes the method deficient this method will need to be revalidated 3. The composition and/or the final manufacturing process of a sample ana ed with the method have been modified after optimization 4. Changes in equipment or in suppliers of critical supplies at the time of anufacturing. This is important, as critical components of the manufac- turing process have the potential to change the degradation profile of the

22 POTENCY METHOD VALIDATION Analyte Impurities Sample solution Impurity standard solution Time (s) 200 280 360 440 520 600 680 760 840 920 1000 Response (mV) 50.000 80.000 77.000 74.000 71.000 68.000 65.000 62.000 59.000 56.000 53.000 Figure 2.4. Overlay chromatogram of an impurity solution with a sample solution. experience with the analyte and the chemistry of the analyte, the scientist will be able to identify which of the impurities may be used as the critical pair. 2.5 POTENCY METHOD REVALIDATION There are various situations during the life cycle of a potency method that require revalidation of the method. 1. During optimization of the formulation or drug substance synthetic process, significant changes may have to be introduced into the process. As a result, to ensure that the analytical method will still be able to analyze the poten￾tially different profile of the drug substance or drug product, revalidation may be necessary. 2. The method was found to be deficient in some areas, such as precision and system suitability. This is especially important as the analytical laboratory gets more experience and more information as to the degradation profile of the sample as it progresses toward submission. If a new impurity is found that makes the method deficient, this method will need to be revalidated. 3. The composition and/or the final manufacturing process of a sample ana￾lyzed with the method have been modified after optimization. 4. Changes in equipment or in suppliers of critical supplies at the time of manufacturing. This is important, as critical components of the manufac￾turing process have the potential to change the degradation profile of the product