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Premium member Presentation Transcript Validation of Analytical Method : Validation of Analytical Method Ashok Suthar Regulatory Associate Regulatory Affairs Amneal Pharmaceuticals Slide 2: Definition: Method validation is the process of proving that ananalytical method is acceptable for its intended purposes. METHOD VALIDATION = ERROR ASSESSMENT Why Method Validation is Important? : Why Method Validation is Important? Develops confidence in using the method & Proof thatMethod is suitable for its intended purpose, The purpose of analytical measurement is to get consistent, reliable and accurate data. Incorrect measurement results can lead to tremendous costs. Regulatory requirement, Equal importance for those working in a regulated and in an accredited environment. U.S. FDA, ISO etc. Slide 4: When to be validated? Partial validation after development of method. Complete validation after manufacturing formula is finalized. Which methods are to be validated Compendial: Pharmacopoeial method Verification of suitability of method Non compendial methods: Laboratory developed methods. Pharmacopoeial methods used outside its scope. Validation: Prior ConsiderationsSuitability of Instrument Status of Qualification and Calibration Suitability of Materials Status of Reference Standards, Reagents, Placebo Lots Suitability of Analyst Status of Training and Qualification Records Suitability of Documentation Written and approved standard test procedure and proper approved protocol with pre-established acceptance criteria : Validation: Prior ConsiderationsSuitability of Instrument Status of Qualification and Calibration Suitability of Materials Status of Reference Standards, Reagents, Placebo Lots Suitability of Analyst Status of Training and Qualification Records Suitability of Documentation Written and approved standard test procedure and proper approved protocol with pre-established acceptance criteria Validation Activity Including the Complete Analytical Procedure : Validation Activity Including the Complete Analytical Procedure Sampling Sample Preparation Analysis Data Evaluation Reporting Validation Step : Validation Step Define the application, purpose and scope of the method. Analytes? Concentration? Develop a analytical method. Develop a validation protocol. Qualification of instrument. Qualify/train operator Slide 8: Qualification of material. Perform pre-validation experiments. Adjust method parameters and/or acceptance criteria if necessary. Perform full validation experiments. Develop Procedures for executing the method in routine analysis. Document validation experiments and results in the validation report. Validation Step Verification vs. Validation : Verification vs. Validation Compendial vs. Non-compendial Methods Compendial methods-Verification Regulatory analytical procedure in USP/NF Non- compendial methods-Validation Alternative analytical procedure proposed by the applicant for use instead of the regulatory analytical procedure Slide 10: BACKGROUND-LAB METHOD FLOW ICH/USP Validation Requirements : ICH/USP Validation Requirements Precision Repeatability Intermediate Precision Reproducibility Limit of Detection Limit of Quantitation Robustness System suitability Specificity Linearity Range Accuracy ImpuritiesSpecificityLOD & LOQLinearity and RangeAccuracy PrecisionRobustness : Validation Parameters ImpuritiesSpecificityLOD & LOQLinearity and RangeAccuracy PrecisionRobustness DissolutionSpecificityLinearity and RangeAccuracy PrecisionRobustness Assay / CU Specificity Linearity and Range Accuracy Precision Robustness Specificity : Specificity Ability of an analytical method to measure the analyte free from interference due to other components. Selectivity Bias Specificity: ICH/USP : Specificity: ICH/USP The ability to measure accurately and specifically the analyte in the presence of components that may be expected to be present in the matrix The degree of interference Active Ingredients Excipients Impurities (synthetic precursors, enantiomers) Degradation Products Placebo Ingredients Slide 16: Combination of 2 or more analytical procedures may be required to achieve necessary level of discrimination (i.e. LC-MS, etc.) Stability indicating analytical methods should always be specific Analysts should ascertain whether the peaks within a sample chromatogram are pure or consist of more than one compound. Therefore should know how many compounds are in the sample or use procedures to detect peak purity Linearity : Linearity Ability of an assay to elicit a direct and proportional response to changes in analyte concentration. Contd…. : Contd…. The Ability of the method to obtain test results that are directly proportional to concentration within a given range Method: dilution of stock solution/separate weightings Expressed as the variance of the slope of the regression line Correlation coefficient, y-intercept, slope of regression line and residual sum of squares should be presented together with plot of the data Range : Range The interval between the upper and lower concentrations of analyte in the sample that have been demonstrate to have a suitable level of precision, accuracy, and linearity. Range : Range Normally derived from Linearity studies. Established by confirming that the method provides acceptable degree of linearity, accuracy, and precision. Specific range dependent upon intended application of the procedure. Accuracy : Accuracy Closeness of the test results obtained by the method to the true value. Precision : Precision The closeness of agreement (degree of scatter) between a series of measurements obtained from multiple samplings of the same homogeneous sample. Should be investigated using homogeneous, authentic samples. Strike Strike Strike Strike Strike Strike Ball Ball Ball Ball Ball Ball Ball Ball Accuracy Vs Precision : Accuracy Vs Precision Precision… Considered at 3 Levels : Precision… Considered at 3 Levels Repeatability Intermediate Precision Reproducibility Repeatability : Repeatability Express the precision under the same operating conditions over a short interval of time. Also referred to as Intra-assay precision Intermediate Precision : Intermediate Precision Express within-laboratory variations. Expressed in terms of standard deviation, relative standard deviation (coefficient of variation) and confidence interval. Known as part of Ruggedness in USP (Different Analysts, Different Laboratories, Different Instruments, Different Reagents, Different Days) Depends on the circumstances under which the procedure is intended to be used. Studies should include varying days, analysts, equipment, etc. Reproducibility : Definition: Ability to reproduce data within the predefined precision Determination: SD, RSD and confidence interval Repeatability test at two different labs. Reproducibility Reproducibility Study : Reproducibility Study Detection Limit (DL) : Lowest amount of analyte in a sample that can be detected but not necessarily quantitated. Estimated by Signal to Noise Ratio of 3:1. Detection Limit (DL) Lowest amount of analyte in a sample that can be quantified with suitable accuracy and precision. Estimated by Signal to Noise Ratio of 10:1. Quantitation Limit (QL) LOD, LOQ and SNR : LOD, LOQ and SNR Limit of Quantitation (LOQ) Limit of Detection (LOD) Signal to Noise Ratio (SNR) noise Peak ALOD Peak BLOQ Baseline Slide 31: Definition: Capacity to remain unaffected by small but deliberate variations in method parameters Determination: Comparison results under differing conditions with precision under normal conditions Variations may include: stability of analytical solution, variation of pH in a mobile phase, different column (lot/supplier), temperature, flow rate. Robustness Slide 32: Solutes may readily decompose prior to chromatographicinvestigations e.g. during sample preparation, extraction,cleanup, phase transfer or storage of prepared vials(refrigerators or automatic sampler). Method development should investigate the stability of theAnalytes AND standards. Stability of analytical solution System stability Stability of the samples being analyzed in a sample solution. e.g. 1 – 48 hours using a single solution. should be determined by replicate analysis of the samplesolution. Slide 33: The checking of a system, before or during analysis of unknowns, to ensure system performance. “No sample analysis is acceptable unless the requirements for system suitability have been met.” (USP Chapter 621) Plate Count, Tailing, Resolution Determination of reproducibility (%RSD) For %RSD < 2.0%, Five replicates For %RSD > 2.0%, Six replicates System Suitability "Sample“ - A mixture of main components andexpected by-products utilized to determine system suitability “Whenever There is a Significant change in Equipment or ReagentsSystem Suitability Testing Should be Performed” (USP Chapter 621) SYSTEM SUITABILITY Confuse of Precision Terms : Confuse of Precision Terms Repeatability Intermediate Precision Reproducibility Ruggedness Robustness Precision Terms : Precision Terms Instrument Precision Repeatability Intermediate Precision Reproducibility Ruggedness Robustness - 6 Standard Injections - One Analysis (6 preps) Two Analyses - Two different Lab. Many Variables - Intentional Changes Slide 36: Robustness Variations All Assays HPLC Assays GC Assays -Sample Prep Manipulation -Extraction Time -Mobile Phase Composition -Different Columns -Temperature -Flow Rate -Different Columns -Temperature -Flow Rate Slide 37: Change in the analytical procedure, drug substance, drug product, the changes, may necessitate revalidation of the analytical procedures. “The degree of revalidation depends on the nature of the change.” “FDA intends to provide guidance in the future on post-approval changes in analytical procedures.” Revalidation should accompany formulation changes (new samples with new compounds or new matrices) manufacturing batch changes new analysts with different skills, new instruments with different characteristics, new location with different environmental conditions, new chemicals and/or reference standards and modification of analytical parameters. Revalidation Slide 38: Validation Report Objective and scope of the method (applicability, type). Summary of methodology. Type of compounds and matrix. All chemicals, reagents, reference standards, QC samples with purity, grade, their source or detailed instructions on their preparation. Procedures for quality checks of standards and chemicals used. Method parameters. Critical parameters taken from robustness testing. Listing of equipment and its functional and performance requirements, e.g., cell dimensions, baseline noise and column temperature range. Detailed conditions on conduct of experiments, including sample preparation Statistical procedures and representative calculations. Procedures for QC in routine analyses, e.g., system suitability tests. Representative plots, e.g., chromatograms, spectra and calibration curves. Method acceptance limit performance data and expected uncertainty of measurement results. Criteria for revalidation. The person's) who developed and validated the method. References (if any). Summary and conclusions. Slide 39: General requirements Qualified and calibrated instruments Documented methods Reliable reference standards Qualified analysts Sample integrity Change control (e.g., synthesis, FPP composition) Analytical methods should be used within GMP and GLP environments, and must be developed using the protocols and acceptance criteria set out in the ICH guidelines Q2 (R1) Thank you : Thank you You do not have the permission to view this presentation. In order to view it, please contact the author of the presentation.