Analytical method validation raaj gprac [compatibility mode]

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Analytical method validation raaj gprac [compatibility mode]

  1. 1. Analytical Method Validation Rajashri Ojha Director, Raaj GPRAC Analytical Method Validation 1
  2. 2. Why Method Validation is Important?1. The purpose of analytical measurement is to get consistent, reliable and accurate data. Incorrect measurement results can lead to tremendous costs.2. Equal importance for those working in a regulated and in an accredited environment. U.S. FDA, EMEA, EPA, AOAC, ISO Analytical Method Validation 2
  3. 3. BackgroundNDA and ANDA must include the analytical procedures necessaryto ensure: Identity, Strength, Quality, Purity, and Potency of the Drug Substances and Drug product [21CFR 314.50(d)(l) and 314.94(a)(9)(i)] Data to establish and reliability [21CFR 211.169(e) and 211.194(a)(2)] Analytical Method Validation 3
  4. 4. Validation is an Old Concept But There are Many ProblemsLack of documented procedures and documented validation resultsSampling or Sample preparation step contribute to overall error.Accessories and materials used for equipment qualification are notqualified.Limits for Operational QualificationLack of software validation and computer system validationQualification and validation are done at just one particular point in time.Adaptation of acceptance criteria for qualification of new system Analytical Method Validation 4
  5. 5. Validation Activity Includingthe Complete Analytical Procedure Sampling Sample Preparation Analysis Data Evaluation Reporting Analytical Method Validation 5
  6. 6. Optimization of ValidationAdditional value and Cost VS. Completeness of validation Analytical Method Validation 6
  7. 7. Considerations Prior to Method ValidationSuitability of Instrument Status of Qualification and CalibrationSuitability of Materials Status of Reference Standards, Reagents, Placebo LotsSuitability of Analyst Status of Training and Qualification RecordsSuitability of Documentation Written analytical procedure and proper approved protocol with pre-established acceptance criteria Analytical Method Validation 7
  8. 8. Validation StepDefine the application, purpose and scope of themethod.Analytes? Concentration? Sample matrices?Develop a analytical method.Develop a validation protocol.Qualification of instrument.Qualify/train operator Analytical Method Validation 8
  9. 9. Validation StepQualification of material.Perform pre-validation experiments.Adjust method parameters and/oracceptance criteria if necessary.Perform full validation experiments.Develop SOP for executing the method in routine analysis.Document validation experiments and results in thevalidation report. Analytical Method Validation 9
  10. 10. System Suitability Validation CalibrationPump InjectorDetector Data System Analyst Method Sample Analytical Method Validation 10
  11. 11. Verification vs. ValidationCompendial 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 Analytical Method Validation 11
  12. 12. Regulations and Guidelines of ValidationUS FDA 21 CFR (Code of Federal Regulations) Part 210 and 211 Part 210: cGMP in Manufacturing, Processing, Packaging, or Holding of Drugs; General Part 211: cGMP for Manufacturing Practice for Finished PharmaceuticalsICH Guidelines Q2A, Text on Validation of Analytical procedures (March 1995) Q2B, Validation of Analytical Procedures: Methodology (May 1997)USP Chapter <1225> Validation of Compendial Methods Analytical Method Validation 12
  13. 13. The accuracy, sensitivity, specificity, and reproducibility of test methodsemployed by the firm shall be established and documented. Suchvalidation and documentation may be accomplished in accordance with211.194(a)(2). 21 CFR PART 211 - CURRENT GOOD MANUFACTURING PRACTICE FOR FINISHED PHARMACEUTICALS Subpart I-Laboratory Controls 211.165 Testing and release for distribution (e)Methods validation means establishing, through documented evidence,a high degree of assurance that an analytical method will consistentlyyield results that accurately reflect the quality characteristics of theproduct tested. 21 CFR PART 210 - CURRENT GOOD MANUFACTURING PRACTICE IN MANUFACTURING, PROCESSING, PACKING, OR HOLDING OF DRUGS 210.3 Definitions (b) (25) Analytical Method Validation 13
  14. 14. The objective of validation of an analytical procedure is todemonstrate that it is suitable for its intended purpose ICH Guideline for Industry Q2AIn practice, it is usually possible to design the experimental work suchthat the appropriate validation characteristics can be consideredsimultaneously to provide a sound, overall knowledge of the capabilitiesof the analytical procedure, for instance: Specificity, Linearity, Range,Accuracy, and Precision. ICH Guideline for Industry Q2B Analytical Method Validation 14
  15. 15. ICH/USP Validation Requirements PrecisionSpecificity RepeatabilityLinearity Intermediate PrecisionRange Limit of DetectionAccuracy Limit of Quantitation Robustness Analytical Method Validation 15
  16. 16. USP Data Elements Required For Assay Validation Analytical Assay Category 2 Assay Performance Assay Category 1 Quantitative Limit Tests Category 3 ParameterAccuracy Yes Yes * *Precision Yes Yes No YesSpecificity Yes Yes Yes *LOD No No Yes *LOQ No Yes No *Linearity Yes Yes No *Range Yes Yes * *Ruggedness Yes Yes Yes Yes * May be required, depending on the nature of the specific test. Category 1: Quantitation of major components or active ingredients Category 2: Determination of impurities or degradation products Category 3: Determination of performance characteristics Analytical Method Validation 16
  17. 17. ICH Validation Characteristics vs. Type of Analytical ProcedureType of Analytical Impurity testing Identification Assay Procedure Quantitative Limit TestsAccuracy No Yes No YesPrecision Repeatability No Yes No Yes Interm. Prec. No Yes No YesSpecificity Yes Yes Yes YesLOD No No Yes NoLOQ No Yes No NoLinearity No Yes No YesRange No Yes No Yes Analytical Method Validation 17
  18. 18. SpecificityAbility of ananalytical methodto measure theanalyte free frominterference due to Selectivityother components. Bias Analytical Method Validation 18
  19. 19. Specificity: ICH/USPAn investigation of specificity should be conductedduring the validation of an identification test, animpurities assay, and a potency assay.Procedures used will depend on the intendedobjective of the analytical procedure.If a method can not completely discriminate, twoof more procedures are recommended. Analytical Method Validation 19
  20. 20. Specificity: IdentificationShould be able to discriminate between compoundsclosely related in structure.Confirmed by obtaining negative results for sampleswith spiked related compounds and positive resultsfor samples with analyte.Choice of potential interfering substances should bebased on sensible scientific judgment consideringsubstances that could likely occur. Analytical Method Validation 20
  21. 21. Specificity: Impurities/AssayChromatographic Methods Demonstrate ResolutionImpurities/Degradants Available Spike with impurities/degradants Show resolution and a lack of interferenceImpurities/Degradants Not Available Stress Samples For assay, Stressed and Unstressed Samples should be compared. For impurity test, impurity profiles should be compared. Analytical Method Validation 21
  22. 22. Pure and Impure HPLC peaksPeak purity tests can also be evaluated with The spectra of Photodiode array detectors Mass spectrometry Analytical Method Validation 22
  23. 23. Specificity: Potential Interference Placebo Drug Substance Degradants Drug Product Degradants Related Substances Packaging Extractables Analytical Method Validation 23
  24. 24. Forced Degradation StudiesHeat High Temperature (50 to 60 oC)Humidity Humidity (70 to 80%)Acid Hydrolysis Acid Hydrolysis (0.1 N)Base Hydrolysis Base Hydrolysis (0.1 N)Oxidation Peroxide Oxidation (3 to 30%)Light Intense UV/Visible LightIntent is to create 10 to 30 % Degradation Analytical Method Validation 24
  25. 25. Specificity Study DEG DEG DEG DEG ActiveCondition #1 #2 #3 #4 ingredients2 N HCl 0 0 17.23 4.71 95.17% 0.1 N 0 0 0 0 100% NaOH 30% 1.12 1.41 1.65 1.2 99.98% H2O2 UV/Vis 0 0 4.68 0 94.67% Analytical Method Validation 25
  26. 26. LinearityAbility of an assayto elicit a direct andproportionalresponse tochanges in analyteconcentration. Analytical Method Validation 26
  27. 27. Linearity Should be EvaluatedBy Visual Inspection of plot of signals vs. analyteconcentrationBy Appropriate statistical methods Linear Regression (y = mx + b) Correlation Coefficient, y-intercept (b), slope (m), residual sum of squares Requires a minimum of 5 concentration levels Analytical Method Validation 27
  28. 28. Linearity ExampleR square = 0.999Slope = 0.97y-intercept = 0.233Line Eq.: Y = 0.97X + 0.233Std. Error = 1.319Std. Deviation of Slopes = 0.0079 Analytical Method Validation 28
  29. 29. RangeThe interval between theupper and lowerconcentrations of analytein the sample that havebeen demonstrate to havea suitable level ofprecision, accuracy, andlinearity. Analytical Method Validation 29
  30. 30. RangeNormally derived from Linearity studies.Established by confirming that the method providesacceptable degree of linearity, accuracy, andprecision.Specific range dependent upon intendedapplication of the procedure. Analytical Method Validation 30
  31. 31. Minimum Specified Range:For Drug Substance & Drug product Assay 80 to 120% of test ConcentrationFor Content Uniformity Assay 70 to 130% of test ConcentrationFor Dissolution Test Method +/- 20% over entire Specification RangeFor Impurity Assays From Reporting Level to 120% of Impurity Specification for Impurity Assays From Reporting Level to 120% of Assay Specification for Impurity/Assay Methods Analytical Method Validation 31
  32. 32. AccuracyCloseness of the testresults obtained by themethod to the true value. Analytical Method Validation 32
  33. 33. AccuracyShould be established across specified range of analyticalprocedure.Should be assessed using a minimum of 3 concentrationlevels, each in triplicate (total of 9 determinations)Should be reported as: Percent recovery of known amount added (reference material) or The difference between the mean assay result and the accepted value Analytical Method Validation 33
  34. 34. Accuracy Data Set (1 of 3) % Recovery % Recovery Amount Amount Percent 99.2 98.9 Found RecoveryAdded (mg) (mg) 99.3 99.3 0.0 0.0 --- 99.4 99.7 50.2 50.4 100.5 Mean 99.3 99.3 79.6 80.1 100.6 Std.dev. 0.1 0.4 99.9 100.7 100.8 95%C.I 99.3±0.25 99.3±0.99 Analytical Method Validation 34
  35. 35. Analyte recovery at different concentrationAnalyte Ingred. (%) Analyte ratio Unit Mean recovery (%) 100 1 100 % 98-102 ≥ 10 10-1 10 % 98-102 ≥1 10-2 1% 97-103 ≥ 0.1 10-3 0.1% 95-105 0.01 10-4 100 ppm 90-107 0.001 10-5 10 ppm 80-110 0.0001 10-6 1 ppm 80-110 0.00001 10-7 100 ppb 80-110 0.000001 10-8 10 ppb 60-115 0.0000001 10-9 1 ppb 40-120AOAC manual for the Peer-Verified Methods program Analytical Method Validation 35
  36. 36. Precision Ball Ball Strike Strike Ball Strike Ball Ball Ball StrikeStrikeThe closeness of agreement Ball Ball Strike(degree of scatter) between aseries of measurements obtainedfrom multiple samplings of thesame homogeneous sample.Should be investigated usinghomogeneous, authentic samples. Analytical Method Validation 36
  37. 37. Precision… Considered at 3 Levels Repeatability Intermediate Precision Reproducibility Analytical Method Validation 37
  38. 38. RepeatabilityExpress the precision Should be assessedunder the same using minimum of 9operating conditions determinationsover a short interval (3 concentrations/ 3of time. replicates) orAlso referred to as Minimum of 6 determinations at theIntra-assay precision 100% level. Analytical Method Validation 38
  39. 39. Intermediate PrecisionExpress within-laboratory Depends on thevariations. circumstances under whichExpressed in terms of standard the procedure is intendeddeviation, relative standard to be used.deviation (coefficient of variation) Studies should includeand confidence interval. varying days, analysts,Known as part of Ruggedness in equipment, etc.USP Analytical Method Validation 39
  40. 40. Repeatability & Intermediate Precision Day 1 Day 2 100.6 99.5 100.8 99.9 100.1 98.9 100.3 99.2 100.5 99.7 100.4 99.6 Mean = 100.5 Mean = 99.5 RSD = 0.24% RSD = 0.36% CI = 100.5 ± 0.24 CI = 99.5 ± 0.36 Grand Mean = 100.0 RSD = 0.59% Analytical Method Validation 40
  41. 41. ReproducibilityDefinition: Ability reproduce data within thepredefined precisionDetermination: SD, RSD and confidence interval Repeatability test at two different labs. Note: Data not required for BLA/NDA Analytical Method Validation 41
  42. 42. Reproducibility Study Lab 1 Lab 2 Lab 3Day 1 Day 2 Day 1 Day 2 Day 1 Day 2Analyst Analyst Analyst Analyst Analyst Analyst 1 2 1 2 1 23 Preps 3 Preps 3 Preps 3 Preps 3 Preps 3 Preps Analytical Method Validation 42
  43. 43. Analyte concentration versus precision Analyte % Analyte ratio Unit RSD (%) 100 1 100 % 1.3 ≥ 10 10-1 10 % 2.7 ≥1 10-2 1% 2.8 ≥ 0.1 10-3 0.1% 3.7 0.01 10-4 100 ppm 5.3 0.001 10-5 10 ppm 7.3 0.0001 10-6 1 ppm 11 0.00001 10-7 100 ppb 15 0.000001 10-8 10 ppb 21 0.0000001 10-9 1 ppb 30AOAC manual for the Peer-Verified Methods program Analytical Method Validation 43
  44. 44. Detection Limit (DL) Quantitation Limit (QL) Lowest amount of analyte in a Lowest amount of analyte in a sample that can be detected sample that can be quantified but not necessarily quantitated. with suitable accuracy and precision. Estimated by Signal to Noise Estimated by Signal to Noise Ratio of 3:1. Ratio of 10:1. Analytical Method Validation 44
  45. 45. Detection Limit (DL) and Quantitation Limit (QL) Estimated by1. Based in Visual Evaluations - Used for non-instrumental methods2. Based on Signal-to Noise-Ratio - 3:1 for Detection Limit - 10:1 for Quantitation Limit3. Based on Standard Deviation of the Response and the Slope Analytical Method Validation 45
  46. 46. Based on Signal-to Noise-Ratio Analytical Method Validation 46
  47. 47. Detection Limit (DL) and Quantitation Limit (QL) 3.3s 10s DL = QL = S SS = slope of calibration curves = standard deviation of blank readings or standard deviation of regression line Validated by assaying samples at DL or QL Analytical Method Validation 47
  48. 48. RobustnessDefinition: Capacity to remain unaffected by smallbut deliberate variations in method parametersDetermination: Comparison results under differingconditions 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. Analytical Method Validation 48
  49. 49. Confuse of Precision TermsRepeatability Reproducibility RuggednessIntermediatePrecision Robustness Analytical Method Validation 49
  50. 50. Precision TermsInstrument Precision - 10 Std. InjectionsRepeatability - One Analysis (6 preps)Intermediate Precision - Two AnalysesReproducibility - Two Lab.Ruggedness - Many VariablesRobustness - Intentional Changes Analytical Method Validation 50
  51. 51. Robustness VariationsAll Assays -Sample Prep Manipulation -Extraction TimeHPLC Assays -Mobile Phase Composition -Different Columns -Temperature -Flow RateGC Assays -Different Columns -Temperature -Flow Rate Analytical Method Validation 51
  52. 52. Robustness-Mobile Phase Change MeOH/ Retention Retention Resolution Water Time 1 Time 2 75:25 11.94 16.41 7.39 80:20 8.47 11.17 6.17 85:15 7.81 10.18 5.93 Analytical Method Validation 52
  53. 53. ICH/USP System SuitabilityICH USP 23 <621> System Suitability Requirements Definition: evaluation of equipment, electronic, Parameters Recommendations analytical operations and K’ In general k’ ≥ 2.0 samples as a whole R > 2, between the peak of Determination: repeatability, interest and the closest R potential interferent tailing factor (T), capacity (degradant, internal STD, impurity, excipient, etc…..) factor (k’), resolution (R), and T T≤2 theoretical Plates (N) N In general N > 2000 Repeatability RSD ≤ 2.0% (n ≥ 5) Analytical Method Validation 53
  54. 54. Guidance on Re-Validation“When sponsors make changes in the analyticalprocedure, drug substance, drug product, thechanges, may necessitate revalidation of theanalytical procedures.”“The degree of revalidation depends on the natureof the change.”“FDA intends to provide guidance in the future onpost-approval changes in analytical procedures.” Analytical Method Validation 54
  55. 55. References‘Analytical Methods Validation for FDA Compliance’ 교육교재 The Center forProfessional Advancement 2003. 3.12-14.Guideline for submitting samples and analytical data for kethods validation(Feb. 1987)ICH Q2AICH Q2B21 Code of Federal Registrations Part 210 and 211Michael E. Swatrz and Ira S. Krull, Analytical method development andvalidation. Mrcel Dekker, Inc. New York, 1997.USP 23 <1225>http://www.waters.comLudwig Huber, Validation and Qualification in Analytical Laboratories,Interpharm Press Inc. Buffalo Grove, Illinois, 1999. Analytical Method Validation 55

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