Validation methodology from HKMA CA-G-4

合集下载

化学分析方法确认指南(翻译)

化学分析方法确认指南(翻译)

化学分析方法确认指南(翻译)2.对已确认方法的验收(Verification for previously validated methods)对于像ASTM,US EPA,ISO和IP这些已通过合作研究(collaborative studies)进行确认的方法,且能够适用于方法所要求的范围之内,对于这类的方法则不需要向内部方法(in-house method)那样的严格验证,同样适用于出版的带有实验数据的科学文献。

鉴于此,对于使用这些方法的实验室,大体上能够验证(verify)操作者按照方法中要求操作且能够获得相同的结果即可。

方法的验收(verification of method)必须包括对现有已验证方法使用前所进行统计纠正(statistical correlation)。

但需要注意的是,由于不同标准机构或公认的技术组织对于方法标准的文件化(documentation)差异,如果组织在所发布的方法中没有验证报告或缺乏部分性能(performance characteristics)或仅对部分性能确认(verify),则实验室不能够直接使用方法,需要确认。

在使用条件下的方法验收不仅仅是要验收体系符合于方法的要求,同样也验收了方法准确度、精确度或其它参数的符合性。

常用以下方式证明方法性能(method performance):l 空白,评定污染l 实验控制样品(e.g.化学分析加标),评定准确性l 重复测定,评定精确度l 定量的、批次的分析,需要定期验证校准曲线标准溶液l 监控质量控制样品,常通过使用控制表(SPC)l 参与能力验证,这些组织可提供与方法要求一致的样品,具有选择代表性的浓度值、基体、分析参数等。

如对原内部方法(in-house method)做了微小的改变(换成不同生产商生产的色谱柱,选择不同的生长基质等),则也需要对方法做验收。

验收程序中关键参数依赖于方法的性质和所可能遇到样品的类型。

V alidation of high-performance liquid chromatography methods for

V alidation of high-performance liquid chromatography methods for

2. Step by step to establish method validation plan The first step in the development of a method validation protocol is to determine the objective of the method. How will the method be used? What is the method intended to demonstrate? Based on the response to these questions, there will be at least two main choices. For example, if the method is intended to monitor patients, release final product, or determine potency, level of impurity, or contaminants in a human drug product, the method is considered a Level I (quantitative assay). If the method is intended to serve as a qualitative evaluation for identity, the method is considered a Level II. Is the method to be used for establishing a limit of impurity (less than or greater than a standard)? Are the results visual? In all cases, the following additional questions will also need to be answered: What sampletypes will be tested using the method? Will the samples be whole blood, serum, plasma, purified protein, unpurified protein, chemical agents, etc.? Based on the sample-type, what interferences are expected? Is it likely that those interfering substances will impact the results? Is the method cell-based, chemical-based or enzyme-based? What level of accuracy, precision, sensitivity and limit of detection is required? Analyte concentration range for these validation parameters are given in Sections 4.1–4.4, respectively. The goal of the questions and the preliminary evaluation is to determine how best to meet the objective of the method validation so that it

20140219_Analytical_Procedures_and_Methods_Validation_for_Drugs_and_Biologics

20140219_Analytical_Procedures_and_Methods_Validation_for_Drugs_and_Biologics

Analytical Procedures and Methods Validation for Drugsand BiologicsDRAFT GUIDANCEThis guidance document is being distributed for comment purposes only. Comments and suggestions regarding this draft document should be submitted within 90 days of publication in the Federal Register of the notice announcing the availability of the draft guidance. Submit electronic comments to . Submit written comments to the Division of Dockets Management (HFA-305), Food and Drug Administration, 5630 Fishers Lane, rm. 1061, Rockville, MD 20852. All comments should be identified with the docket number listed in the notice of availability that publishes in the Federal Registe r.For questions regarding this draft document contact (CDER) Lucinda Buhse 314-539-2134, or (CBER) Office of Communication, Outreach and Development at 800-835-4709 or 301-827-1800.U.S. Department of Health and Human ServicesFood and Drug AdministrationCenter for Drug Evaluation and Research (CDER)Center for Biologics Evaluation and Research (CBER)February 2014CMCAnalytical Procedures and Methods Validation for Drugsand BiologicsAdditional copies are available from:Office of CommunicationsDivision of Drug Information, WO51, Room 2201Center for Drug Evaluation and ResearchFood and Drug Administration10903 New Hampshire Ave., Silver Spring, MD 20993Phone: 301-796-3400; Fax: 301-847-8714druginfo@/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htmand/orOffice of Communication, Outreach andDevelopment, HFM-40Center for Biologics Evaluation and ResearchFood and Drug Administration1401 Rockville Pike, Rockville, MD 20852-1448ocod@/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/Guidances/default.htm(Tel) 800-835-4709 or 301-827-1800U.S. Department of Health and Human ServicesFood and Drug AdministrationCenter for Drug Evaluation and Research (CDER)Center for Biologics Evaluation and Research (CBER)Febr uary 2014CMCTABLE OF CONTENTSI.INTRODUCTION (1)II.BACKGROUND (2)III.ANALYTICAL METHODS DEVELOPMENT (3)IV.CONTENT OF ANALYTICAL PROCEDURES (3)A.Principle/Scope (4)B.Apparatus/Equipment (4)C.Operating Parameters (4)D.Reagents/Standards (4)E.Sample Preparation (4)F.Standards Control Solution Preparation (5)G.Procedure (5)H.System Suitability (5)I.Calculations (5)J.Data Reporting (5)V.REFERENCE STANDARDS AND MATERIALS (6)VI.ANALYTICAL METHOD VALIDATION FOR NDA, ANDAs, BLAs, AND DMFs (6)A.Noncompendial Analytical Procedures (6)B.Validation Characteristics (7)pendial Analytical Procedures (8)VII.STATISTICAL ANALYSIS AND MODELS (8)A.Statistics (8)B.Models (8)VIII.LIFE CYCLE MANAGEMENT OF ANALYTICAL PROCEDURES (9)A.Revalidation (9)B.Analytical Method Comparability Studies (10)1.Alternative Analytical Procedures (10)2.Analytical Methods Transfer Studies (11)C.Reporting Postmarketing Changes to an Approved NDA, ANDA, or BLA (11)IX.FDA METHODS VERIFICATION (12)X.REFERENCES (12)Guidance for Industry11Analytical Procedures and Methods Validation for Drugs and2Biologics345This draft guidance, when finalized, will represent the Food and Drug Administration’s (FDA’s) current 6thinking on this topic. It does not create or confer any rights for or on any person and does not operate to 7bind FDA or the public. You can use an alternative approach if the approach satisfies the requirements of 8the applicable statutes and regulations. If you want to discuss an alternative approach, contact the FDA9staff responsible for implementing this guidance. If you cannot identify the appropriate FDA staff, call 10the appropriate number listed on the title page of this guidance.11121314I. INTRODUCTION1516This revised draft guidance supersedes the 2000 draft guidance for industry on Analytical17Procedures and Methods Validation2,3 and, when finalized, will also replace the 1987 FDA18guidance for industry on Submitting Samples and Analytical Data for Methods Validation. It19provides recommendations on how you, the applicant, can submit analytical procedures4 and20methods validation data to support the documentation of the identity, strength, quality, purity,21and potency of drug substances and drug products.5It will help you assemble information and 22present data to support your analytical methodologies. The recommendations apply to drug23substances and drug products covered in new drug applications (NDAs), abbreviated new drug 24applications (ANDAs), biologics license applications (BLAs), and supplements to these25applications. The principles in this revised draft guidance also apply to drug substances and drug 26products covered in Type II drug master files (DMFs).2728This revised draft guidance complements the International Conference on Harmonisation (ICH) 29guidance Q2(R1)Validation of Analytical Procedures: Text and Methodology(Q2(R1)) for30developing and validating analytical methods.3132This revised draft guidance does not address investigational new drug application (IND) methods 33validation, but sponsors preparing INDs should consider the recommendations in this guidance.34For INDs, sufficient information is required at each phase of an investigation to ensure proper35identity, quality, purity, strength, and/or potency. The amount of information on analytical36procedures and methods validation will vary with the phase of the investigation.6 For general371 This guidance has been prepared by the Office of Pharmaceutical Science, in the Center for Drug Evaluation andResearch (CDER) and the Center for Biologics Evaluation and Research (CBER) at the Food and DrugAdministration.2 Sample submission is described in section IX, FDA Methods Verification.3 We update guidances periodically. To make sure you have the most recent version of a guidance, check the FDADrugs guidance Web page at/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/default.htm.4Analytical procedure is interchangeable with a method or test procedure.5The terms drug substance and drug product, as used in this guidance, refer to human drugs and biologics.6 See 21 CFR 312.23(a)(7).guidance on analytical procedures and methods validation information to be submitted for phase 38one studies, sponsors should refer to the FDA guidance for industry on Content and Format of39Investigational New Drug Applications (INDs) for Phase 1 Studies of Drugs, Including40Well-Characterized, Therapeutic, Biotechnology-Derived Products. General considerations for 41analytical procedures and method validation (e.g., bioassay) before conduct of phase three42studies are discussed in the FDA guidance for industry on IND Meetings for Human Drugs and 43Biologics, Chemistry, Manufacturing, and Controls Information.4445This revised draft guidance does not address specific method validation recommendations for46biological and immunochemical assays for characterization and quality control of many drug47substances and drug products. For example, some bioassays are based on animal challenge48models, and immunogenicity assessments or other immunoassays have unique features that49should be considered during development and validation.5051In addition, the need for revalidation of existing analytical methods may need to be considered 52when the manufacturing process changes during the product’s life cycle. For questions on53appropriate validation approaches for analytical procedures or submission of information not54addressed in this guidance, you should consult with the appropriate FDA product quality review 55staff.5657If you choose a different approach than those recommended in this revised draft guidance, we58encourage you to discuss the matter with the appropriate FDA product quality review staff before 59you submit your application.6061FDA’s guidance documents, including this guidance, do not establish legally enforceable62responsibilities. Instead, guidances describe the Agency’s current thinking on a topic and should 63be viewed only as recommendations, unless specific regulatory or statutory requirements are64cited. The use of the word should in Agency guidances means that something is suggested or65recommended, but not required.666768II.BACKGROUND6970Each NDA and ANDA must include the analytical procedures necessary to ensure the identity, 71strength, quality, purity, and potency of the drug substance and drug product.7 Each BLA must 72include a full description of the manufacturing methods, including analytical procedures that73demonstrate the manufactured product meets prescribed standards of identity, quality, safety,74purity, and potency.8 Data must be available to establish that the analytical procedures used in 75testing meet proper standards of accuracy and reliability and are suitable for their intended76purpose.9 For BLAs and their supplements, the analytical procedures and their validation are77submitted as part of license applications or supplements and are evaluated by FDA quality78review groups.79807 See 21 CFR 314.50(d)(1) and 314.94(a)(9)(i).8 See 21 CFR 601.2(a) and 601.2(c).9 See 21 CFR 211.165(e) and 211.194(a)(2).Analytical procedures and validation data should be submitted in the corresponding sections of 81the application in the ICH M2 eCTD: Electronic Common Technical Document Specification.108283When an analytical procedure is approved/licensed as part of the NDA, ANDA, or BLA, it84becomes the FDA approved analytical procedure for the approved product. This analytical85procedure may originate from FDA recognized sources (e.g., a compendial procedure from the 86United States Pharmacopeia/National Formulary (USP/NF)) or a validated procedure you87submitted that was determined to be acceptable by FDA. To apply an analytical method to a88different product, appropriate validation studies with the matrix of the new product should be89considered.909192III.ANALYTICAL METHODS DEVELOPMENT9394An analytical procedure is developed to test a defined characteristic of the drug substance or95drug product against established acceptance criteria for that characteristic. Early in the96development of a new analytical procedure, the choice of analytical instrumentation and97methodology should be selected based on the intended purpose and scope of the analytical98method. Parameters that may be evaluated during method development are specificity, linearity, 99limits of detection (LOD) and quantitation limits (LOQ), range, accuracy, and precision.100101During early stages of method development, the robustness of methods should be evaluated102because this characteristic can help you decide which method you will submit for approval.103Analytical procedures in the early stages of development are initially developed based on a104combination of mechanistic understanding of the basic methodology and prior experience.105Experimental data from early procedures can be used to guide further development. You should 106submit development data within the method validation section if they support the validation of 107the method.108109To fully understand the effect of changes in method parameters on an analytical procedure, you 110should adopt a systematic approach for method robustness study (e.g., a design of experiments 111with method parameters). You should begin with an initial risk assessment and follow with112multivariate experiments. Such approaches allow you to understand factorial parameter effects 113on method performance. Evaluation of a method’s performance may include analyses of114samples obtained from in-process manufacturing stages to the finished product. Knowledge115gained during these studies on the sources of method variation can help you assess the method 116performance.117118119IV.CONTENT OF ANALYTICAL PROCEDURES120121You should describe analytical procedures in sufficient detail to allow a competent analyst to 122reproduce the necessary conditions and obtain results within the proposed acceptance criteria. 123You should also describe aspects of the analytical procedures that require special attention. An 124analytical procedure may be referenced from FDA recognized sources (e.g., USP/NF,12510 See sections 3.2.S.4 Control of Drug Substance, 3.2.P.4 Control of Excipients, and 3.2.P.5 Control of DrugProduct.Association of Analytical Communities (AOAC) International)11 if the referenced analytical126procedure is not modified beyond what is allowed in the published method. You should provide 127in detail the procedures from other published sources. The following is a list of essential128information you should include for an analytical procedure:129130A.Principle/Scope131132A description of the basic principles of the analytical test/technology (separation, detection, etc.); 133target analyte(s) and sample(s) type (e.g., drug substance, drug product, impurities or compounds 134in biological fluids, etc.).135136B.Apparatus/Equipment137138All required qualified equipment and components (e.g., instrument type, detector, column type, 139dimensions, and alternative column, filter type, etc.).140141C.Operating Parameters142143Qualified optimal settings and ranges (allowed adjustments) critical to the analysis (e.g., flow144rate, components temperatures, run time, detector settings, gradient, head space sampler). A145drawing with experimental configuration and integration parameters may be used, as applicable. 146147D.Reagents/Standards148149The following should be listed:150151•Grade of chemical (e.g., USP/NF, American Chemical Society, High152Performance or Pressure Liquid Chromatography, or Gas153Chromatography and preservative free).154•Source (e.g., USP reference standard or qualified in-house reference material). 155•State (e.g., dried, undried, etc.) and concentration.156•Standard potencies (purity correction factors).157•Storage controls.158•Directions for safe use (as per current Safety Data Sheet).159•Validated or useable shelf life.160161New batches of biological reagents, such as monoclonal antibodies, polyclonal antisera, or cells, 162may need extensive qualification procedures included as part of the analytical procedure.163164E.Sample Preparation165166Procedures (e.g., extraction method, dilution or concentration, desalting procedures and mixing 167by sonication, shaking or sonication time, etc.) for the preparations for individual sample tests. 168A single preparation for qualitative and replicate preparations for quantitative tests with16911 See 21 CFR 211.194(a)(2).appropriate units of concentrations for working solutions (e.g., µg/ml or mg/ml) and information 170on stability of solutions and storage conditions.171172F.Standards Control Solution Preparation173174Procedures for the preparation and use of all standard and control solutions with appropriate175units of concentration and information on stability of standards and storage conditions,176including calibration standards, internal standards, system suitability standards, etc.177178G.Procedure179180A step-by-step description of the method (e.g., equilibration times, and scan/injection sequence 181with blanks, placeboes, samples, controls, sensitivity solution (for impurity method) and182standards to maintain validity of the system suitability during the span of analysis) and allowable 183operating ranges and adjustments if applicable.184185H.System Suitability186187Confirmatory test(s) procedures and parameters to ensure that the system (equipment,188electronics, and analytical operations and controls to be analyzed) will function correctly as an 189integrated system at the time of use. The system suitability acceptance criteria applied to190standards and controls, such as peak tailing, precision and resolution acceptance criteria, may be 191required as applicable. For system suitability of chromatographic systems, refer to CDER192reviewer guidance on Validation of Chromatographic Methods and USP General Chapter <621> 193Chromatography.194195I.Calculations196197The integration method and representative calculation formulas for data analysis (standards,198controls, samples) for tests based on label claim and specification (e.g., assay, specified and199unspecified impurities and relative response factors). This includes a description of any200mathematical transformations or formulas used in data analysis, along with a scientific201justification for any correction factors used.202203J.Data Reporting204205A presentation of numeric data that is consistent with instrumental capabilities and acceptance 206criteria. The method should indicate what format to use to report results (e.g., percentage label 207claim, weight/weight, and weight/volume etc.) with the specific number of significant figures 208needed. The American Society for Testing and Materials (ASTM) E29 describes a standard209practice for using significant digits in test data to determine conformance with specifications. For 210chromatographic methods, you should include retention times (RTs) for identification with211reference standard comparison basis, relative retention times (RRTs) (known and unknown212impurities) acceptable ranges and sample results reporting criteria.213214215V.REFERENCE STANDARDS AND MATERIALS216217Primary and secondary reference standards and materials are defined and discussed in the218following ICH guidances: Q6A Specifications: Test Procedures and Acceptance Criteria for 219New Drug Substances and New Drug Products: Chemical Substances (ICH Q6A), Q6B220Specifications: Test Procedures and Acceptance Criteria for Biotechnological/Biological221Products, and Q7 Good Manufacturing Practice Guidance for Active Pharmaceutical222Ingredients. For all standards, you should ensure the suitability for use. Reference standards for 223drug substances are particularly critical in validating specificity for an identity test. You should 224strictly follow storage, usage conditions, and handling instructions for reference standards to225avoid added impurities and inaccurate analysis. For biological products, you should include226information supporting any reference standards and materials that you intend to use in the BLA 227and in subsequent annual reports for subsequent reference standard qualifications. Information 228supporting reference standards and materials include qualification test protocols, reports, and 229certificates of analysis (including stability protocols and relevant known impurity profile230information, as applicable).231232Reference standards can often be obtained from USP and may also be available through the233European Pharmacopoeia, Japanese Pharmacopoeia, World Health Organization, or National 234Institute of Standards and Technology. Reference standards for a number of biological products 235are also available from CBER. For certain biological products marketed in the U.S., reference 236standards authorized by CBER must be used before the product can be released to the market.12 237Reference materials from other sources should be characterized by procedures including routine 238and beyond routine release testing as described in ICH Q6A. You should consider orthogonal 239methods. Additional testing could include attributes to determine the suitability of the reference 240material not necessarily captured by the drug substance or product release tests (e.g., more241extensive structural identity and orthogonal techniques for purity and impurities, biological242activity).243244For biological reference standards and materials, we recommend that you follow a two-tiered 245approach when qualifying new reference standards to help prevent drift in the quality attributes 246and provide a long-term link to clinical trial material. A two-tiered approach involves a247comparison of each new working reference standard with a primary reference standard so that it 248is linked to clinical trial material and the current manufacturing process.249250251VI.ANALYTICAL METHOD VALIDATION FOR NDA, ANDAs, BLAs, AND 252DMFs253254A.Noncompendial Analytical Procedures255256Analytical method validation is the process of demonstrating that an analytical procedure is257suitable for its intended purpose. The methodology and objective of the analytical procedures 258should be clearly defined and understood before initiating validation studies. This understanding 25912 See 21 CFR 610.20.is obtained from scientifically-based method development and optimization studies. Validation 260data must be generated under an protocol approved by the sponsor following current good261manufacturing practices with the description of methodology of each characteristic test and262predetermined and justified acceptance criteria, using qualified instrumentation operated under 263current good manufacturing practices conditions.13 Protocols for both drug substance and264product analytes or mixture of analytes in respective matrices should be developed and executed. 265266ICH Q2(R1) is considered the primary reference for recommendations and definitions on267validation characteristics for analytical procedures. The FDA Reviewer Guidance: Validation of 268Chromatographic Methods is available as well.269270B.Validation Characteristics271272Although not all of the validation characteristics are applicable for all types of tests, typical273validation characteristics are:274275•Specificity276•Linearity277•Accuracy278•Precision (repeatability, intermediate precision, and reproducibility)279•Range280•Quantitation limit281•Detection limit282283If a procedure is a validated quantitative analytical procedure that can detect changes in a quality 284attribute(s) of the drug substance and drug product during storage, it is considered a stability285indicating assay. To demonstrate specificity of a stability-indicating assay, a combination of286challenges should be performed. Some challenges include the use of samples spiked with target 287analytes and all known interferences; samples that have undergone various laboratory stress288conditions; and actual product samples (produced by the final manufacturing process) that are289either aged or have been stored under accelerated temperature and humidity conditions.290291As the holder of the NDA, ANDA, or BLA, you must:14 (1) submit the data used to establish292that the analytical procedures used in testing meet proper standards of accuracy and reliability, 293and (2) notify the FDA about each change in each condition established in an approved294application beyond the variations already provided for in the application, including changes to 295analytical procedures and other established controls.296297The submitted data should include the results from the robustness evaluation of the method,298which is typically conducted during method development or as part of a planned validation299study.1530013 See 21 CFR 211.165(e); 21 CFR 314.50 (d), and for biologics see 21 CFR 601.2(a), 601.2(c), and 601.12(a).14 For drugs see 21 CFR 314.50 (d), 314.70(d), and for biologics see 21 CFR 601.2(a), 601.2(c), and 601.12(a). For aBLA, as discussed below, you must obtain prior approval from FDA before implementing a change in analyticalmethods if those methods are specified in FDA regulations15 See section III and ICH Q2(R1).pendial Analytical Procedures302303The suitability of an analytical procedure (e.g., USP/NF, the AOAC International Book of304Methods, or other recognized standard references) should be verified under actual conditions of 305use.16 Compendial general chapters, which are complex and mention multiple steps and/or306address multiple techniques, should be rationalized for the intended use and verified. Information 307to demonstrate that USP/NF analytical procedures are suitable for the drug product or drug308substance should be included in the submission and generated under a verification protocol.309310The verification protocol should include, but is not limited to: (1) compendial methodology to 311be verified with predetermined acceptance criteria, and (2) details of the methodology (e.g.,312suitability of reagent(s), equipment, component(s), chromatographic conditions, column, detector 313type(s), sensitivity of detector signal response, system suitability, sample preparation and314stability). The procedure and extent of verification should dictate which validation characteristic 315tests should be included in the protocol (e.g., specificity, LOD, LOQ, precision, accuracy, etc.). 316Considerations that may influence what characteristic tests should be in the protocol may depend 317on situations such as whether specification limits are set tighter than compendial acceptance318criteria, or RT or RRT profiles are changing in chromatographic methods because of the319synthetic route of drug substance or differences in manufacturing process or matrix of drug320product. Robustness studies of compendial assays do not need to be included, if methods are 321followed without deviations.322323324VII.STATISTICAL ANALYSIS AND MODELS325326A.Statistics327328Statistical analysis of validation data can be used to evaluate validation characteristics against 329predetermined acceptance criteria. All statistical procedures and parameters used in the analysis 330of the data should be based on sound principles and appropriate for the intended evaluation.331Reportable statistics of linear regression analysis R (correlation coefficient), R square332(coefficient of determination), slope, least square, analysis of variance (ANOVA), confidence 333intervals, etc., should be provided with justification.For information on statistical techniques 334used in making comparisons, as well as other general information on the interpretation and335treatment of analytical data, appropriate literature or texts should be consulted.17336337B.Models338339Some analytical methods might use chemometric and/or multivariate models. When developing 340these models, you should include a statistically adequate number and range of samples for model 341development and comparable samples for model validation. Suitable software should be used for 342data analysis. Model parameters should be deliberately varied to test model robustness.34334416 See 21 CFR 211.194(a)(2) and USP General Chapter <1226> Verification of Compendial Procedures.17 See References section for examples including USP <1010> Analytical Data – Interpretation and Treatment.。

美国FDA分析方法验证指南中文译稿[1]

美国FDA分析方法验证指南中文译稿[1]

1II. 背景 (2)III. 分析方法的类型 (3)A. 法定分析方法 (3)B. 可选择分析方法 (3)3 C. 稳定性指示分析 (3)IV. 对照品……………………………………………………………………………4A. 对照品的类型 (4)B. 分析报告单 (4)C. 对照品的界定 (4)V. IND 中的分析方法验证 (6)VI. NDA, ANDA, BLA 和PLA 中分析方法验证的内容和格式 (6)A. 原则 (6)B. 取样 (7)C. 仪器和仪器参数 (7)D. 试剂 (7)E. 系统适应性实验 (7)F. 对照品的制备 (7)G. 样品的制备 (8)H. 分析方法 (8)L. 计算 (8)J. 结果报告 (8)VII. NDA,ANDA,BLA 和PLA 中的分析方法验证 (9)A.非法定分析方法 (9)1.验证项目 (9)2. 其它分析方法验证信息 (10)a. 耐用性 (11)b. 强降解实验 (11)c. 仪器输出/原始资料 (11)3.各类检测的建议验证项目 (13)B.法定分析方法 (15)VIII. 统计分析…………………………………………………………………………15A. 总则 (15)C. 统计 (16)IX. 再验证 (16)X. 分析方法验证技术包:内容和过程……………………………………………17A. 分析方法验证技术包 (17)B. 样品的选择和运输 (18)C. 各方责任 (19)XI. 方法………………………………………………………………………………20A. 高效液相色谱(HPLC) (20)B. 气相色谱(GC) (22)C. 分光光度法,光谱学,光谱法和相关的物理方法 (23)D. 毛细管电泳 (23)E. 旋光度 (24)F. 粒径相关的分析方法 (25)G. 溶出度 (26)H. 其它仪器分析方法 (27)附件A:NDA,ANDA,BLA 和PLA 申请的内容 (28)附件B:分析方法验证的问题和延误 (29)参考文献……………………………………………………………………………………30术语表………………………………………………………………………………………32This guidance provides recommendations to applicants on submitting analytical procedures, validation data, and samples to support the documentation of the identity, strength, quality, purity, and potency of drug substances and drug products.1. 绪论本指南旨在为申请者提供建议,以帮助其提交分析方法,方法验证资料和样品用于支持原料药和制剂的认定,剂量,质量,纯度和效力方面的文件。

生物医药 分析方法验证流程

生物医药 分析方法验证流程

生物医药分析方法验证流程Analytical method validation is a crucial step in the field of biopharmaceuticals, ensuring the reliability and accuracy of test results. 方法验证是生物制药领域中一个至关重要的步骤,它能够确保测试结果的可靠性和准确性。

Validation of analytical methods is necessary to demonstrate that the method is suitable for its intended use and that it meets predetermined acceptance criteria. 分析方法的验证是必要的,它能够证明该方法适用于既定用途,并满足预定的接受标准。

This process involves a series of experiments and assessments to determine the method's performance characteristics such as accuracy, precision, specificity, and linearity. 这个过程涉及一系列实验和评估,以确定方法的性能特征,如准确性、精密度、特异性和线性。

By validating an analytical method, researchers can have confidence in the results obtained from their experiments, leading to more reliable and reproducible scientific data. 通过验证分析方法,研究人员可以对来自实验中获得的结果充满信心,从而获得更可靠和可重复的科学数据。

methodology 的缩写

methodology 的缩写

methodology 的缩写METHODOLOGY的缩写为标题METHODOLOGY(方法学)是指研究和应用一定方法和技术的原理、规范和程序,用于解决特定问题或达到特定目标。

在科学研究和实践应用中,方法学起着重要的作用。

下面将围绕METHODOLOGY的每个字母进行阐述,探讨方法学的重要性和应用。

M - Method(方法)方法是指达到特定目标或解决特定问题的步骤和技术。

在科学研究中,正确的方法可以保证研究结果的准确性和可靠性。

无论是实验研究还是调查研究,选择合适的方法是至关重要的。

方法的选择应该根据研究目的、研究对象和研究资源等因素进行综合考虑。

E - Evaluation(评估)评估是方法学中的重要环节,它用于衡量方法和技术的有效性和可行性。

通过评估,我们可以了解到方法的优点和不足之处,进而对方法进行改进和提高。

评估可以基于实验数据、统计分析和专家意见等多种方式进行,旨在为决策提供科学依据。

T - Theory(理论)理论是方法学的基础,它提供了解释现象和推导原理的框架和原则。

在研究过程中,理论为研究者提供了思路和指导,帮助他们构建研究模型和假设。

理论与方法相互依存,理论的验证和发展需要借助于方法,而方法的设计和应用也需要建立在理论的基础上。

H - Hypothesis(假设)假设是研究中的重要概念,它是对现象或问题的解释和预测。

假设可以从理论推导出来,也可以根据实际观察得出。

在研究中,假设需要经过验证和检验,以确定其是否成立。

通过假设的提出和验证,我们可以了解现象背后的原理和机制。

O - Observation(观察)观察是方法学中收集信息和数据的手段之一。

通过观察,我们可以直接获得现象的特征和规律。

观察可以分为主动观察和被动观察,可以通过肉眼观察、仪器观察和记录观察等多种方式进行。

观察的结果可以作为研究的依据和证据。

D - Data(数据)数据是方法学中非常重要的资源,它是研究的基础和依据。

方法确认(Method Validation)-黄亨建

方法确认(Method Validation)-黄亨建
Random Error (imprecision)
.重复测定同一样本 .误差有正或负 .误差的方向和大小不可预测 .随机误差的表达:SD and CV
随机误差的确认
Determination of Random error
随机误差的测定 20份相同的样本 批内、日间、批间。 样本:基质相同的血清、尿液、脑脊液、标 准液、质控液。 浓度在医学决定水平,批内、日内< 0.25 Tea
准确度Accuracy
The Comparison of Methods Experiment
–– 每份样本测定二次 –– 足够的样本量 –– 时间周期
5 天或20 天 2小时内完成
准确度Accuracy
The Comparison of Methods Experiment
准确度的统计学包括: --图形分析Graph analysis
Data Analysis
回归分析Regression analysis
准确度Accuracy The Comparison of Methods Experiment–
Data Analysis
回归分 析
绘制比 较数据
(Y - Axis) Accu-Chek Glucose, mg/dL
速试验, 潜血试验, etc. ––方法确认不要求 ––按生产厂家说明书
管理机构的要求
requirements by regulation
Non-waived,未更改的中等和高度复杂性 e.g. 化学、血液学 etc . 要求进行四个方面
的方法确认 1.1. 线性实验Linearity experiment
总误差 Total Error ( TEa )

MIQE Guidelines

MIQE Guidelines

/pcr
AMPLIFICATION
Nucleic Acid Extraction (核酸提 取)
/pcr
RNA 纯度和完整性
AMPLIFICATION
Experion Virtual Gel
L C 3’ 5’ 10’ 15’ 1h 2h 4h
Primer B
Forward Primer
Reverse Primer A
1
110
/pcr
Amplicon Secondary Structures
AMPLIFICATION
/mfold/applications
编辑们犯难了“定量PCR数据可信吗?”
/pcr
What are the MIQE guidelines?
AMPLIFICATION
qPCR的国际标准:就评价qPCR实验和发表文章时所必需的实验 信息提出了最低限度的标准。
/pcr
MIQE 指南好处
AMPLIFICATION
/pcr
Reverse Transcription 反转录
AMPLIFICATION
/pcr
Reverse Transcription
AMPLIFICATION
RNA
cDNA
Reality Ideal ?
Reproducible Data Not Reproducible
/pcr
qPCR Target Information
AMPLIFICATION
/pcr
序列同源性分析(BLAST)
AMPLIFICATION
http://www.ncbi.nlm.nih. gov/BLAST/Blast.cgi

Validation of Analytical Procedures- Methodology, Harmonized Tripartite Guideline

Validation of Analytical Procedures- Methodology, Harmonized Tripartite Guideline

bioprocess development & outsource management for 25 yearsINTERNATIONAL CONFERENCE ON HARMONISATION OF TECHNICAL REQUIREMENTS FOR REGISTRATION OF PHARMACEUTICALS FOR HUMAN USEICH H ARMONISED T RIPARTITE G UIDELINEV ALIDATION OF A NALYTICAL P ROCEDURES:T EXT AND M ETHODOLOGYQ2(R1)Current Step 4 versionParent Guideline dated 27 October 1994(Complementary Guideline on Methodology dated 6 November 1996incorporated in November 2005)This Guideline has been developed by the appropriate ICH Expert Working Group and has been subject to consultation by the regulatory parties, in accordance with the ICH Process. At Step 4 of the Process the final draft is recommended for adoption to the regulatory bodies of the European Union, Japan and USA.Q2(R1) Document HistoryFirstCodification History DateNew Codification November2005Parent Guideline: Text on Validation of Analytical ProceduresQ2 Approval by the Steering Committee under Step 2 and release for public consultation.26October1993Q2Q2A Approval by the Steering Committee under Step 4 and recommendation for adoption to the three ICHregulatory bodies.27October1994Q2Guideline on Validation of Analytical Procedures: Methodology developed to complement the Parent GuidelineQ2B Approval by the Steering Committee under Step 2 and release for public consultation.29November1995in Q2(R1)Q2B Approval by the Steering Committee under Step 4 and recommendation for adoption to the three ICHregulatory bodies.6November1996in Q2(R1)Current Step 4 versionQ2A and Q2B The parent guideline is now renamed Q2(R1) as theguideline Q2B on methology has been incorporated tothe parent guideline. The new title is “Validation ofAnalytical Procedures: Text and Methodology”.November2005Q2(R1)VALIDATION OF ANALYTICAL PROCEDURES:T EXT AND M ETHODOLOGYICH Harmonised Tripartite GuidelineTABLE OF CONTENTSPART I:TEXT ON VALIDATION OF ANALYTICAL PROCEDURES (1)1.Introduction (1)2.Types of Analytical Procedures to be Validated (1)TABLE (3)GLOSSARY (4)PART II:VALIDATION OF ANALYTICAL PROCEDURES: METHODOLOGY (6)INTRODUCTION (6)1.SPECIFICITY (6)1.1.Identification (7)1.2.Assay and Impurity Test(s) (7)2.LINEARITY (8)3.RANGE (8)4.ACCURACY (9)4.1.Assay (9)4.2.Impurities (Quantitation) (10)4.3.Recommended Data (10)5.PRECISION (10)5.1.Repeatability (10)5.2.Intermediate Precision (10)5.3.Reproducibility (10)5.4.Recommended Data (10)6.DETECTION LIMIT (11)6.1.Based on Visual Evaluation (11)6.2.Based on Signal-to-Noise (11)6.3Based on the Standard Deviation of the Response and the Slope (11)6.4Recommended Data (11)Validation of Analytical Procedures: Text and Methodology7.QUANTITATION LIMIT (12)7.1.Based on Visual Evaluation (12)7.2.Based on Signal-to-Noise Approach (12)7.3.Based on the Standard Deviation of the Response and the Slope (12)7.4Recommended Data (13)8.ROBUSTNESS (13)9.SYSTEM SUITABILITY TESTING (13)PART I:T EXT ON V ALIDATION OF A NALYTICAL P ROCEDURESICH Harmonised Tripartite GuidelineHaving reached Step 4 of the ICH Process at the ICH Steering Committee meeting on27 October 1994, this guideline is recommended for adoptionto the three regulatory parties to ICH1. IntroductionThis document presents a discussion of the characteristics for considerationduring the validation of the analytical procedures included as part of registrationapplications submitted within the EC, Japan and USA. This document does notnecessarily seek to cover the testing that may be required for registration in, orexport to, other areas of the world. Furthermore, this text presentation serves asa collection of terms, and their definitions, and is not intended to providedirection on how to accomplish validation. These terms and definitions are meantto bridge the differences that often exist between various compendia andregulators of the EC, Japan and USA.The objective of validation of an analytical procedure is to demonstrate that it issuitable for its intended purpose. A tabular summation of the characteristicsapplicable to identification, control of impurities and assay procedures isincluded. Other analytical procedures may be considered in future additions tothis document.2. Types of Analytical Procedures to be ValidatedThe discussion of the validation of analytical procedures is directed to the fourmost common types of analytical procedures:- Identification tests;- Quantitative tests for impurities' content;- Limit tests for the control of impurities;- Quantitative tests of the active moiety in samples of drug substance or drugproduct or other selected component(s) in the drug product.Although there are many other analytical procedures, such as dissolution testingfor drug products or particle size determination for drug substance, these have not been addressed in the initial text on validation of analytical procedures.Validation of these additional analytical procedures is equally important to thoselisted herein and may be addressed in subsequent documents.A brief description of the types of tests considered in this document is providedbelow.- Identification tests are intended to ensure the identity of an analyte in asample. This is normally achieved by comparison of a property of the sample(e.g., spectrum, chromatographic behavior, chemical reactivity, etc) to that ofa reference standard;Validation of Analytical Procedures: Text- Testing for impurities can be either a quantitative test or a limit test for theimpurity in a sample. Either test is intended to accurately reflect the puritycharacteristics of the sample. Different validation characteristics are requiredfor a quantitative test than for a limit test;- Assay procedures are intended to measure the analyte present in a givensample. In the context of this document, the assay represents a quantitativemeasurement of the major component(s) in the drug substance. For the drugproduct, similar validation characteristics also apply when assaying for theactive or other selected component(s). The same validation characteristicsmay also apply to assays associated with other analytical procedures (e.g.,dissolution).The objective of the analytical procedure should be clearly understood since thiswill govern the validation characteristics which need to be evaluated. Typical validation characteristics which should be considered are listed below: AccuracyPrecisionRepeatabilityIntermediate PrecisionSpecificityDetection LimitQuantitation LimitLinearityRangeEach of these validation characteristics is defined in the attached Glossary. Thetable lists those validation characteristics regarded as the most important for thevalidation of different types of analytical procedures. This list should beconsidered typical for the analytical procedures cited but occasional exceptions should be dealt with on a case-by-case basis. It should be noted that robustness isnot listed in the table but should be considered at an appropriate stage in thedevelopment of the analytical procedure.Furthermore 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.The degree of revalidation required depends on the nature of the changes. Certainother changes may require validation as well.Validation of Analytical Procedures: Text TABLEType of analytical procedure I DENTIFICATION T ESTING FORIMPURITIESA SSAY- dissolution(measurement only)- content/potencycharacteristics quantitat. limitAccuracy - + - + PrecisionRepeatabilityInterm.Precision --+ -+ (1) -++ (1)Specificity (2) + + + + Detection Limit - - (3) + - Quantitation Limit - + - - Linearity - + - + Range - + - +- signifies that this characteristic is not normally evaluated+ signifies that this characteristic is normally evaluated(1) in cases where reproducibility (see glossary) has been performed, intermediateprecision is not needed(2) lack of specificity of one analytical procedure could be compensated by othersupporting analytical procedure(s)(3) may be needed in some casesValidation of Analytical Procedures: TextGLOSSARY1. ANALYTICAL PROCEDUREThe analytical procedure refers to the way of performing the analysis. It shoulddescribe in detail the steps necessary to perform each analytical test. This mayinclude but is not limited to: the sample, the reference standard and the reagents preparations, use of the apparatus, generation of the calibration curve, use of the formulae for the calculation, etc.2. SPECIFICITYSpecificity is the ability to assess unequivocally the analyte in the presence of components which may be expected to be present. Typically these might include impurities, degradants, matrix, etc.Lack of specificity of an individual analytical procedure may be compensated by other supporting analytical procedure(s).This definition has the following implications:Identification: to ensure the identity of an analyte.Purity Tests: to ensure that all the analytical procedures performed allow an accurate statement of the content of impurities of an analyte, i.e.related substances test, heavy metals, residual solvents content, etc.Assay (content or potency):to provide an exact result which allows an accurate statement on thecontent or potency of the analyte in a sample.3. ACCURACYThe accuracy of an analytical procedure expresses the closeness of agreement betweenthe value which is accepted either as a conventional true value or an accepted reference value and the value found.This is sometimes termed trueness.4. PRECISIONThe precision of an analytical procedure expresses the closeness of agreement (degreeof scatter) between a series of measurements obtained from multiple sampling of thesame homogeneous sample under the prescribed conditions. Precision may be considered at three levels: repeatability, intermediate precision and reproducibility.Precision should be investigated using homogeneous, authentic samples. However, ifit is not possible to obtain a homogeneous sample it may be investigated using artificially prepared samples or a sample solution.The precision of an analytical procedure is usually expressed as the variance, standard deviation or coefficient of variation of a series of measurements.Validation of Analytical Procedures: Text 4.1. RepeatabilityRepeatability expresses the precision under the same operating conditions over a short interval of time. Repeatability is also termed intra-assay precision .4.2. Intermediate precisionIntermediate precision expresses within-laboratories variations: different days, different analysts, different equipment, etc.4.3. ReproducibilityReproducibility expresses the precision between laboratories (collaborative studies, usually applied to standardization of methodology).5. DETECTION LIMITThe detection limit of an individual analytical procedure is the lowest amount of analyte in a sample which can be detected but not necessarily quantitated as an exact value.6. QUANTITATION LIMITThe quantitation limit of an individual analytical procedure is the lowest amount of analyte in a sample which can be quantitatively determined with suitable precision and accuracy. The quantitation limit is a parameter of quantitative assays for low levels of compounds in sample matrices, and is used particularly for the determination of impurities and/or degradation products.7. LINEARITYThe linearity of an analytical procedure is its ability (within a given range) to obtain test results which are directly proportional to the concentration (amount) of analyte in the sample.8. RANGEThe range of an analytical procedure is the interval between the upper and lower concentration (amounts) of analyte in the sample (including these concentrations) for which it has been demonstrated that the analytical procedure has a suitable level of precision, accuracy and linearity.9. ROBUSTNESSThe robustness of an analytical procedure is a measure of its capacity to remain unaffected by small, but deliberate variations in method parameters and provides an indication of its reliability during normal usage.PART II:VALIDATION OF ANALYTICAL PROCEDURES:METHODOLOGYICH Harmonised Tripartite GuidelineHaving reached Step 4 of the ICH Process at the ICH Steering Committee meeting on6 November 1996, and incorporated into the core guideline in November 2005, thisguideline is recommended for adoption to the three regulatory parties to ICH INTRODUCTIONThis document is complementary to the parent document which presents a discussionof the characteristics that should be considered during the validation of analyticalprocedures. Its purpose is to provide some guidance and recommendations on how to consider the various validation characteristics for each analytical procedure. In somecases (for example, demonstration of specificity), the overall capabilities of a numberof analytical procedures in combination may be investigated in order to ensure thequality of the drug substance or drug product. In addition, the document provides anindication of the data which should be presented in a registration application .All relevant data collected during validation and formulae used for calculatingvalidation characteristics should be submitted and discussed as appropriate. Approaches other than those set forth in this guideline may be applicable andacceptable. It is the responsibility of the applicant to choose the validation procedure and protocol most suitable for their product. However it is important to rememberthat the main objective of validation of an analytical procedure is to demonstrate thatthe procedure is suitable for its intended purpose. Due to their complex nature,analytical procedures for biological and biotechnological products in some cases maybe approached differently than in this document.Well-characterized reference materials, with documented purity, should be usedthroughout the validation study. The degree of purity necessary depends on theintended use.In accordance with the parent document, and for the sake of clarity, this documentconsiders the various validation characteristics in distinct sections. The arrangementof these sections reflects the process by which an analytical procedure may bedeveloped and evaluated.In practice, it is usually possible to design the experimental work such that theappropriate validation characteristics can be considered simultaneously to provide asound, overall knowledge of the capabilities of the analytical procedure, for instance:specificity, linearity, range, accuracy and precision.1. SPECIFICITYAn investigation of specificity should be conducted during the validation ofidentification tests, the determination of impurities and the assay. The proceduresused to demonstrate specificity will depend on the intended objective of the analyticalprocedure.It is not always possible to demonstrate that an analytical procedure is specific for aparticular analyte (complete discrimination). In this case a combination of two orValidation of Analytical Procedures: Methodology more analytical procedures is recommended to achieve the necessary level of discrimination.1.1. IdentificationSuitable identification tests should be able to discriminate between compounds of closely related structures which are likely to be present. The discrimination of a procedure may be confirmed by obtaining positive results (perhaps by comparison with a known reference material) from samples containing the analyte, coupled with negative results from samples which do not contain the analyte. In addition, the identification test may be applied to materials structurally similar to or closely related to the analyte to confirm that a positive response is not obtained. The choice of such potentially interfering materials should be based on sound scientific judgement with a consideration of the interferences that could occur.1.2. Assay and Impurity Test(s)For chromatographic procedures, representative chromatograms should be used to demonstrate specificity and individual components should be appropriately labelled. Similar considerations should be given to other separation techniques.Critical separations in chromatography should be investigated at an appropriate level. For critical separations, specificity can be demonstrated by the resolution of the two components which elute closest to each other.In cases where a non-specific assay is used, other supporting analytical procedures should be used to demonstrate overall specificity. For example, where a titration is adopted to assay the drug substance for release, the combination of the assay and a suitable test for impurities can be used.The approach is similar for both assay and impurity tests:1.2.1 Impurities are availableFor the assay , this should involve demonstration of the discrimination of the analyte in the presence of impurities and/or excipients; practically, this can be done by spiking pure substances (drug substance or drug product) with appropriate levels of impurities and/or excipients and demonstrating that the assay result is unaffected by the presence of these materials (by comparison with the assay result obtained on unspiked samples).For the impurity test, the discrimination may be established by spiking drug substance or drug product with appropriate levels of impurities and demonstrating the separation of these impurities individually and/or from other components in the sample matrix.1.2.2 Impurities are not availableIf impurity or degradation product standards are unavailable, specificity may be demonstrated by comparing the test results of samples containing impurities or degradation products to a second well-characterized procedure e.g.: pharmacopoeial method or other validated analytical procedure (independent procedure).As appropriate, this should include samples stored under relevant stress conditions: light, heat, humidity, acid/base hydrolysis and oxidation.- for the assay, the two results should be compared;Validation of Analytical Procedures: Methodology- for the impurity tests, the impurity profiles should be compared.Peak purity tests may be useful to show that the analyte chromatographic peak is not attributable to more than one component (e.g., diode array, mass spectrometry).2. LINEARITYA linear relationship should be evaluated across the range (see section 3) of the analytical procedure. It may be demonstrated directly on the drug substance (by dilution of a standard stock solution) and/or separate weighings of synthetic mixtures of the drug product components, using the proposed procedure. The latter aspect can be studied during investigation of the range.Linearity should be evaluated by visual inspection of a plot of signals as a function of analyte concentration or content. If there is a linear relationship, test results should be evaluated by appropriate statistical methods, for example, by calculation of a regression line by the method of least squares. In some cases, to obtain linearity between assays and sample concentrations, the test data may need to be subjected to a mathematical transformation prior to the regression analysis. Data from the regression line itself may be helpful to provide mathematical estimates of the degree of linearity.The correlation coefficient, y-intercept, slope of the regression line and residual sum of squares should be submitted. A plot of the data should be included. In addition, an analysis of the deviation of the actual data points from the regression line may also be helpful for evaluating linearity.Some analytical procedures, such as immunoassays, do not demonstrate linearity after any transformation. In this case, the analytical response should be described by an appropriate function of the concentration (amount) of an analyte in a sample.For the establishment of linearity, a minimum of 5 concentrations is recommended. Other approaches should be justified.3. RANGEThe specified range is normally derived from linearity studies and depends on the intended application of the procedure. It is established by confirming that the analytical procedure provides an acceptable degree of linearity, accuracy and precision when applied to samples containing amounts of analyte within or at the extremes of the specified range of the analytical procedure.The following minimum specified ranges should be considered:- for the assay of a drug substance or a finished (drug) product: normally from 80 to 120 percent of the test concentration;- for content uniformity, covering a minimum of 70 to 130 percent of the test concentration, unless a wider more appropriate range, based on the nature of the dosage form (e.g., metered dose inhalers), is justified;- for dissolution testing: +/-20 % over the specified range;e.g., if the specifications for a controlled released product cover a region from 20%, after 1 hour, up to 90%, after 24 hours, the validated range would be 0-110% of the label claim.Validation of Analytical Procedures: Methodology - for the determination of an impurity: from the reporting level of an impurity1to120% of the specification;-for impurities known to be unusually potent or to produce toxic or unexpected pharmacological effects, the detection/quantitation limit should be commensurate with the level at which the impurities must be controlled;Note: for validation of impurity test procedures carried out during development, it may be necessary to consider the range around a suggested (probable) limit.- if assay and purity are performed together as one test and only a 100% standard is used, linearity should cover the range from the reporting level of the impurities1 to 120% of the assay specification.4. ACCURACYAccuracy should be established across the specified range of the analytical procedure.4.1. Assay4.1.1 Drug SubstanceSeveral methods of determining accuracy are available:a) application of an analytical procedure to an analyte of known purity (e.g. referencematerial);b) comparison of the results of the proposed analytical procedure with those of asecond well-characterized procedure, the accuracy of which is stated and/or defined (independent procedure, see 1.2.);c) accuracy may be inferred once precision, linearity and specificity have beenestablished.4.1.2 Drug ProductSeveral methods for determining accuracy are available:a) application of the analytical procedure to synthetic mixtures of the drug productcomponents to which known quantities of the drug substance to be analysed have been added;b) in cases where it is impossible to obtain samples of all drug product components ,it may be acceptable either to add known quantities of the analyte to the drug product or to compare the results obtained from a second, well characterized procedure, the accuracy of which is stated and/or defined (independent procedure, see 1.2.);c) accuracy may be inferred once precision, linearity and specificity have beenestablished.1 see chapters “Reporting Impurity Content of Batches” of the corresponding ICH-Guidelines: “Impurities in New Drug Substances” and “Impurities in New Drug Products”Validation of Analytical Procedures: Methodology4.2. Impurities (Quantitation)Accuracy should be assessed on samples (drug substance/drug product) spiked with known amounts of impurities.In cases where it is impossible to obtain samples of certain impurities and/or degradation products, it is considered acceptable to compare results obtained by an independent procedure (see 1.2.). The response factor of the drug substance can be used.It should be clear how the individual or total impurities are to be determined e.g., weight/weight or area percent, in all cases with respect to the major analyte.4.3. Recommended DataAccuracy should be assessed using a minimum of 9 determinations over a minimum of 3 concentration levels covering the specified range (e.g., 3 concentrations/3 replicates each of the total analytical procedure).Accuracy should be reported as percent recovery by the assay of known added amount of analyte in the sample or as the difference between the mean and the accepted true value together with the confidence intervals.5. PRECISIONValidation of tests for assay and for quantitative determination of impurities includes an investigation of precision.5.1. RepeatabilityRepeatability should be assessed using:a) a minimum of 9 determinations covering the specified range for the procedure(e.g., 3 concentrations/3 replicates each);orb) a minimum of 6 determinations at 100% of the test concentration.5.2. Intermediate PrecisionThe extent to which intermediate precision should be established depends on the circumstances under which the procedure is intended to be used. The applicant should establish the effects of random events on the precision of the analytical procedure. Typical variations to be studied include days, analysts, equipment, etc. It is not considered necessary to study these effects individually. The use of an experimental design (matrix) is encouraged.5.3. ReproducibilityReproducibility is assessed by means of an inter-laboratory trial. Reproducibility should be considered in case of the standardization of an analytical procedure, for instance, for inclusion of procedures in pharmacopoeias. These data are not part of the marketing authorization dossier.5.4. Recommended DataThe standard deviation, relative standard deviation (coefficient of variation) and confidence interval should be reported for each type of precision investigated.Validation of Analytical Procedures: Methodology 6. DETECTION LIMITSeveral approaches for determining the detection limit are possible, depending on whether the procedure is a non-instrumental or instrumental. Approaches other than those listed below may be acceptable.6.1. Based on Visual EvaluationVisual evaluation may be used for non-instrumental methods but may also be used with instrumental methods.The detection limit is determined by the analysis of samples with known concentrations of analyte and by establishing the minimum level at which the analyte can be reliably detected.6.2. Based on Signal-to-NoiseThis approach can only be applied to analytical procedures which exhibit baseline noise.Determination of the signal-to-noise ratio is performed by comparing measured signals from samples with known low concentrations of analyte with those of blank samples and establishing the minimum concentration at which the analyte can be reliably detected. A signal-to-noise ratio between 3 or 2:1 is generally considered acceptable for estimating the detection limit.6.3 Based on the Standard Deviation of the Response and the SlopeThe detection limit (DL) may be expressed as:DL = 3.3Swhere = the standard deviation of the responseS = the slope of the calibration curveThe slope S may be estimated from the calibration curve of the analyte. The estimate of may be carried out in a variety of ways, for example:6.3.1 Based on the Standard Deviation of the BlankMeasurement of the magnitude of analytical background response is performed by analyzing an appropriate number of blank samples and calculating the standard deviation of these responses.6.3.2 Based on the Calibration CurveA specific calibration curve should be studied using samples containing an analyte in the range of DL. The residual standard deviation of a regression line or the standard deviation of y-intercepts of regression lines may be used as the standard deviation.6.4 Recommended DataThe detection limit and the method used for determining the detection limit should be presented. If DL is determined based on visual evaluation or based on signal to noise ratio, the presentation of the relevant chromatograms is considered acceptable for justification.。

Bioanalytical Method ValidationGuidance for Indust

Bioanalytical Method ValidationGuidance for Indust

Guidance for Industry Bioanalytical Method ValidationU.S. Department of Health and Human ServicesFood and Drug AdministrationCenter for Drug Evaluation and Research (CDER)Center for Veterinary Medicine (CVM)May 2001BPGuidance for Industry Bioanalytical Method ValidationAdditional copies are available from:Drug Information Branch (HFD-210)Center for Drug Evaluation and Research (CDER)5600 Fishers Lane, Rockville, MD 20857 (Tel) 301-827-4573Internet at /cder/guidance/index.htmorCommunications Staff (HFV-12)Center for Veterinary Medicine (CVM)7500 Standish Place, Rockville, MD 20855 (Tel) 301–594-1755Internet at /cvmU.S. Department of Health and Human ServicesFood and Drug AdministrationCenter for Drug Evaluation and Research (CDER)Center for Veterinary Medicine (CVM)May 2001BPTable of ContentsI.INTRODUCTION (1)II.BACKGROUND (1)A.F ULL V ALIDATION (2)B.P ARTIAL V ALIDATION (2)C.C ROSS-V ALIDATION (3)III.REFERENCE STANDARD (4)IV.METHOD DEVELOPMENT: CHEMICAL ASSAY (4)A.S ELECTIVITY (4)B.A CCURACY, P RECISION, AND R ECOVERY (5)C.C ALIBRATION/S TANDARD C URVE (5)D.S TABILITY (6)E.P RINCIPLES OF B IOANALYTICAL M ETHOD V ALIDATION AND E STABLISHMENT (8)F.S PECIFIC R ECOMMENDATIONS FOR M ETHOD V ALIDATION (10)V.METHOD DEVELOPMENT: MICROBIOLOGICAL AND LIGAND-BINDING ASSAYS (11)A.S ELECTIVITY I SSUES (11)B.Q UANTIFICATION I SSUES (12)VI.APPLICATION OF VALIDATED METHOD TO ROUTINE DRUG ANALYSIS (13)A CCEPTANCE C RITERIA FOR THE R UN (15)VII.DOCUMENTATION (16)A.S UMMARY I NFORMATION (16)B.D OCUMENTATION FOR M ETHOD E STABLISHMENT (17)C.A PPLICATION TO R OUTINE D RUG A NALYSIS (17)D.O THER I NFORMATION (19)GLOSSARY (20)GUIDANCE FOR INDUSTRY1Bioanalytical Method ValidationI.INTRODUCTIONThis guidance provides assistance to sponsors of investigational new drug applications (INDs), new drug applications (NDAs), abbreviated new drug applications (ANDAs), and supplements in developing bioanalytical method validation information used in human clinical pharmacology, bioavailability (BA), and bioequivalence (BE) studies requiring pharmacokinetic (PK) evaluation. This guidance also applies to bioanalytical methods used for non-human pharmacology/toxicology studies and preclinical studies. For studies related to the veterinary drug approval process, this guidance applies only to blood and urine BA, BE, and PK studies.The information in this guidance generally applies to bioanalytical procedures such as gas chromatography (GC), high-pressure liquid chromatography (LC), combined GC and LC mass spectrometric (MS) procedures such as LC-MS, LC-MS-MS, GC-MS, and GC-MS-MS performed for the quantitative determination of drugs and/or metabolites in biological matricessuch as blood, serum, plasma, or urine. This guidance also applies to other bioanalytical methods, such as immunological and microbiological procedures, and to other biological matrices, such as tissue and skin samples.This guidance provides general recommendations for bioanalytical method validation. The recommendations can be adjusted or modified depending on the specific type of analytical method used. II.BACKGROUND1 This guidance has been prepared by the Biopharmaceutics Coordinating Committee in the Center for Drug Evaluation and Research (CDER) in cooperation with the Center for Veterinary Medicine (CVM) at the Food and Drug Administration.This guidance has been developed based on the deliberations of two workshops: (1) Analytical Methods Validation: Bioavailability, Bioequivalence, and Pharmacokinetic Studies (held on December 3B5, 19902 ) and (2) Bioanalytical Methods Validation C A Revisit With a Decade of Progress (held on January 12B14, 20003).Selective and sensitive analytical methods for the quantitative evaluation of drugs and their metabolites (analytes) are critical for the successful conduct of preclinical and/or biopharmaceutics and clinical pharmacology studies. Bioanalytical method validation includes all of the procedures that demonstrate that a particular method used for quantitative measurement of analytes in a given biological matrix, such as blood, plasma, serum, or urine, is reliable and reproducible for the intended use. The fundamental parameters for this validation include (1) accuracy, (2) precision, (3) selectivity, (4) sensitivity, (5) reproducibility, and (6) stability. Validation involves documenting, through the use of specific laboratory investigations, that the performance characteristics of the method are suitable and reliable for the intended analytical applications. The acceptability of analytical data corresponds directly to the criteria used to validate the method.Published methods of analysis are often modified to suit the requirements of the laboratory performing the assay. These modifications should be validated to ensure suitable performance of the analytical method. When changes are made to a previously validated method, the analyst should exercise judgment as to how much additional validation is needed. During the course of a typical drug development program, a defined bioanalytical method undergoes many modifications. The evolutionary changes to support specific studies and different levels of validation demonstrate the validity of an assay’s performance. Different types and levels of validation are defined and characterized as follows:A.Full Validation•Full validation is important when developing and implementing a bioanalytical method for the first time.•Full validation is important for a new drug entity.• A full validation of the revised assay is important if metabolites are added to an existing assay for quantification.B.Partial ValidationPartial validations are modifications of already validated bioanalytical methods. Partial validation can range from as little as one intra-assay accuracy and precision determination to a nearly full2 Workshop Report: Shah, V.P. et al., Pharmaceutical Research: 1992; 9:588-592.3 Workshop Report: Shah, V.P. et al., Pharmaceutical Research: 2000; 17:in press.validation. Typical bioanalytical method changes that fall into this category include, but are not limited to:•Bioanalytical method transfers between laboratories or analysts•Change in analytical methodology (e.g., change in detection systems)•Change in anticoagulant in harvesting biological fluid•Change in matrix within species (e.g., human plasma to human urine)•Change in sample processing procedures•Change in species within matrix (e.g., rat plasma to mouse plasma)•Change in relevant concentration range•Changes in instruments and/or software platforms•Limited sample volume (e.g., pediatric study)•Rare matrices•Selectivity demonstration of an analyte in the presence of concomitant medications•Selectivity demonstration of an analyte in the presence of specific metabolitesC.Cross-ValidationCross-validation is a comparison of validation parameters when two or more bioanalytical methods are used to generate data within the same study or across different studies. An example of cross-validation would be a situation where an original validated bioanalytical method serves as thereference and the revised bioanalytical method is the comparator. The comparisons should be done both ways.When sample analyses within a single study are conducted at more than one site or more than one laboratory, cross-validation with spiked matrix standards and subject samples should be conducted at each site or laboratory to establish interlaboratory reliability. Cross-validation should also be considered when data generated using different analytical techniques (e.g., LC-MS-MS vs.ELISA4) in different studies are included in a regulatory submission.All modifications should be assessed to determine the recommended degree of validation. The analytical laboratory conducting pharmacology/toxicology and other preclinical studies for regulatory submissions should adhere to FDA=s Good Laboratory Practices (GLPs)5 (21 CFR part 58) and to sound principles of quality assurance throughout the testing process. The bioanalytical method for human BA, BE, PK, and drug interaction studies must meet the criteria in 21 CFR 320.29. The analytical laboratory should have a written set of standard operating procedures (SOPs) to ensure a complete system of quality control and assurance. The SOPs should cover all aspects of analysis from the time the sample is collected and reaches the laboratory until the results of the analysis are reported. The SOPs also should include record keeping, security and chain of sample custody4 Enzyme linked immune sorbent assay5 For the Center for Veterinary Medicine, all bioequivalence studies are subject to Good Laboratory Practices.(accountability systems that ensure integrity of test articles), sample preparation, and analytical tools such as methods, reagents, equipment, instrumentation, and procedures for quality control and verification of results.The process by which a specific bioanalytical method is developed, validated, and used in routine sample analysis can be divided into (1) reference standard preparation, (2) bioanalytical method development and establishment of assay procedure, and (3) application of validated bioanalytical method to routine drug analysis and acceptance criteria for the analytical run and/or batch. These three processes are described in the following sections of this guidance.III.REFERENCE STANDARDAnalysis of drugs and their metabolites in a biological matrix is carried out using samples spiked with calibration (reference) standards and using quality control (QC) samples. The purity of the reference standard used to prepare spiked samples can affect study data. For this reason, an authenticated analytical reference standard of known identity and purity should be used to prepare solutions of known concentrations. If possible, the reference standard should be identical to the analyte. When this is not possible, an established chemical form (free base or acid, salt or ester) of known purity can be used. Three types of reference standards are usually used: (1) certified reference standards (e.g., USP compendial standards); (2) commercially supplied reference standards obtained from a reputable commercial source; and/or (3) other materials of documented purity custom-synthesized by an analytical laboratory or other noncommercial establishment. The source and lot number, expiration date, certificates of analyses when available, and/or internally or externally generated evidence of identity and purity should be furnished for each reference standard.IV.METHOD DEVELOPMENT: CHEMICAL ASSAYThe method development and establishment phase defines the chemical assay. The fundamental parameters for a bioanalytical method validation are accuracy, precision, selectivity, sensitivity, reproducibility, and stability. Measurements for each analyte in the biological matrix should be validated. In addition, the stability of the analyte in spiked samples should be determined. Typical method development and establishment for a bioanalytical method include determination of (1) selectivity, (2) accuracy, precision, recovery, (3) calibration curve, and (4) stability of analyte in spiked samples.A.SelectivitySelectivity is the ability of an analytical method to differentiate and quantify the analyte in thepresence of other components in the sample. For selectivity, analyses of blank samples of theappropriate biological matrix (plasma, urine, or other matrix) should be obtained from at leastsix sources. Each blank sample should be tested for interference, and selectivity should be ensured at the lower limit of quantification (LLOQ).Potential interfering substances in a biological matrix include endogenous matrix components, metabolites, decomposition products, and in the actual study, concomitant medication and other exogenous xenobiotics. If the method is intended to quantify more than one analyte, each analyte should be tested to ensure that there is no interference.B.Accuracy, Precision, and RecoveryThe accuracy of an analytical method describes the closeness of mean test results obtained by the method to the true value (concentration) of the analyte. Accuracy is determined by replicate analysis of samples containing known amounts of the analyte. Accuracy should be measured using a minimum of five determinations per concentration. A minimum of three concentrations in the range of expected concentrations is recommended. The mean value should be within 15% of the actual value except at LLOQ, where it should not deviate by more than 20%. The deviation of the mean from the true value serves as the measure of accuracy.The precision of an analytical method describes the closeness of individual measures of an analyte when the procedure is applied repeatedly to multiple aliquots of a single homogeneous volume of biological matrix. Precision should be measured using a minimum of five determinations per concentration. A minimum of three concentrations in the range of expected concentrations is recommended. The precision determined at each concentration level should not exceed 15% of the coefficient of variation (CV) except for the LLOQ, where it should not exceed 20% of the CV. Precision is further subdivided into within-run, intra-batch precision or repeatability, which assesses precision during a single analytical run, and between-run, inter-batch precision or repeatability, which measures precision with time, and may involve different analysts, equipment, reagents, and laboratories.The recovery of an analyte in an assay is the detector response obtained from an amount of the analyte added to and extracted from the biological matrix, compared to the detector response obtained for the true concentration of the pure authentic standard. Recovery pertains to the extraction efficiency of an analytical method within the limits of variability. Recovery of the analyte need not be 100%, but the extent of recovery of an analyte and of the internal standard should be consistent, precise, and reproducible. Recovery experiments should be performed by comparing the analytical results for extracted samples at three concentrations (low, medium, and high) with unextracted standards that represent 100% recovery.C.Calibration/Standard CurveA calibration (standard) curve is the relationship between instrument response and known concentrations of the analyte. A calibration curve should be generated for each analyte in thesample. A sufficient number of standards should be used to adequately define the relationship between concentration and response. A calibration curve should be prepared in the same biological matrix as the samples in the intended study by spiking the matrix with known concentrations of the analyte. The number of standards used in constructing a calibration curve will be a function of the anticipated range of analytical values and the nature of theanalyte/response relationship. Concentrations of standards should be chosen on the basis of the concentration range expected in a particular study. A calibration curve should consist of a blank sample (matrix sample processed without internal standard), a zero sample (matrix sample processed with internal standard), and six to eight non-zero samples covering the expected range, including LLOQ.1.Lower Limit of Quantification (LLOQ)The lowest standard on the calibration curve should be accepted as the limit ofquantification if the following conditions are met:C The analyte response at the LLOQ should be at least 5 times the responsecompared to blank response.C Analyte peak (response) should be identifiable, discrete, and reproducible witha precision of 20% and accuracy of 80-120%.2.Calibration Curve/Standard Curve/Concentration-ResponseThe simplest model that adequately describes the concentration-response relationshipshould be used. Selection of weighting and use of a complex regression equation should be justified. The following conditions should be met in developing a calibration curve:C#20% deviation of the LLOQ from nominal concentrationC#15% deviation of standards other than LLOQ from nominal concentrationAt least four out of six non-zero standards should meet the above criteria, including the LLOQ and the calibration standard at the highest concentration. Excluding thestandards should not change the model used.D.StabilityDrug stability in a biological fluid is a function of the storage conditions, the chemical properties of the drug, the matrix, and the container system. The stability of an analyte in a particular matrix and container system is relevant only to that matrix and container system and should not be extrapolated to other matrices and container systems. Stability procedures should evaluate the stability of the analytes during sample collection and handling, after long-term (frozen at theintended storage temperature) and short-term (bench top, room temperature) storage, and after going through freeze and thaw cycles and the analytical process. Conditions used in stability experiments should reflect situations likely to be encountered during actual sample handling and analysis. The procedure should also include an evaluation of analyte stability in stock solution.All stability determinations should use a set of samples prepared from a freshly made stock solution of the analyte in the appropriate analyte-free, interference-free biological matrix. Stock solutions of the analyte for stability evaluation should be prepared in an appropriate solvent at known concentrations.1.Freeze and Thaw StabilityAnalyte stability should be determined after three freeze and thaw cycles. At least three aliquots at each of the low and high concentrations should be stored at the intendedstorage temperature for 24 hours and thawed unassisted at room temperature. Whencompletely thawed, the samples should be refrozen for 12 to 24 hours under the sameconditions. The freeze–thaw cycle should be repeated two more times, then analyzedon the third cycle. If an analyte is unstable at the intended storage temperature, thestability sample should be frozen at -700C during the three freeze and thaw cycles.2.Short-Term Temperature StabilityThree aliquots of each of the low and high concentrations should be thawed at roomtemperature and kept at this temperature from 4 to 24 hours (based on the expectedduration that samples will be maintained at room temperature in the intended study) and analyzed.3.Long-Term StabilityThe storage time in a long-term stability evaluation should exceed the time between the date of first sample collection and the date of last sample analysis. Long-term stabilityshould be determined by storing at least three aliquots of each of the low and highconcentrations under the same conditions as the study samples. The volume of samples should be sufficient for analysis on three separate occasions. The concentrations of allthe stability samples should be compared to the mean of back-calculated values for the standards at the appropriate concentrations from the first day of long-term stabilitytesting.4.Stock Solution StabilityThe stability of stock solutions of drug and the internal standard should be evaluated at room temperature for at least 6 hours. If the stock solutions are refrigerated or frozenfor the relevant period, the stability should be documented. After completion of thedesired storage time, the stability should be tested by comparing the instrumentresponse with that of freshly prepared solutions.5.Post-Preparative StabilityThe stability of processed samples, including the resident time in the autosampler, should be determined. The stability of the drug and the internal standard should be assessedover the anticipated run time for the batch size in validation samples by determiningconcentrations on the basis of original calibration standards.Although the traditional approach of comparing analytical results for stored samples with those for freshly prepared samples has been referred to in this guidance, other statistical approaches based on confidence limits for evaluation of an analyte=s stability in abiological matrix can be used. SOPs should clearly describe the statistical method andrules used. Additional validation may include investigation of samples from dosedsubjects.E.Principles of Bioanalytical Method Validation and Establishment•The fundamental parameters to ensure the acceptability of the performance of a bioanalytical method validation are accuracy, precision, selectivity, sensitivity,reproducibility, and stability.• A specific, detailed description of the bioanalytical method should be written. This can be in the form of a protocol, study plan, report, and/or SOP.•Each step in the method should be investigated to determine the extent to which environmental, matrix, material, or procedural variables can affect the estimation of analyte in the matrix from the time of collection of the material up to and including the time ofanalysis.•It may be important to consider the variability of the matrix due to the physiological nature of the sample. In the case of LC-MS-MS-based procedures, appropriate steps should be taken to ensure the lack of matrix effects throughout the application of the method,especially if the nature of the matrix changes from the matrix used during method validation.• A bioanalytical method should be validated for the intended use or application. All experiments used to make claims or draw conclusions about the validity of the methodshould be presented in a report (method validation report).•Whenever possible, the same biological matrix as the matrix in the intended samples should be used for validation purposes. (For tissues of limited availability, such as bone marrow, physiologically appropriate proxy matrices can be substituted.)•The stability of the analyte (drug and/or metabolite) in the matrix during the collection process and the sample storage period should be assessed, preferably prior to sampleanalysis.•For compounds with potentially labile metabolites, the stability of analyte in matrix from dosed subjects (or species) should be confirmed.•The accuracy, precision, reproducibility, response function, and selectivity of the method for endogenous substances, metabolites, and known degradation products should beestablished for the biological matrix. For selectivity, there should be evidence that thesubstance being quantified is the intended analyte.•The concentration range over which the analyte will be determined should be defined in the bioanalytical method, based on evaluation of actual standard samples over the range,including their statistical variation. This defines the standard curve.• A sufficient number of standards should be used to adequately define the relationship between concentration and response. The relationship between response and concentration should be demonstrated to be continuous and reproducible. The number of standards used should be a function of the dynamic range and nature of the concentration-responserelationship. In many cases, six to eight concentrations (excluding blank values) can define the standard curve. More standard concentrations may be recommended for nonlinear than for linear relationships.•The ability to dilute samples originally above the upper limit of the standard curve should be demonstrated by accuracy and precision parameters in the validation.•In consideration of high throughput analyses, including but not limited to multiplexing, multicolumn, and parallel systems, sufficient QC samples should be used to ensure control of the assay. The number of QC samples to ensure proper control of the assay should be determined based on the run size. The placement of QC samples should be judiciously considered in the run.•For a bioanalytical method to be considered valid, specific acceptance criteria should be set in advance and achieved for accuracy and precision for the validation of QC samples over the range of the standards.F.Specific Recommendations for Method Validation•The matrix-based standard curve should consist of a minimum of six standard points, excluding blanks, using single or replicate samples. The standard curve should cover the entire range of expected concentrations.•Standard curve fitting is determined by applying the simplest model that adequately describes the concentration-response relationship using appropriate weighting and statistical tests for goodness of fit.•LLOQ is the lowest concentration of the standard curve that can be measured with acceptable accuracy and precision. The LLOQ should be established using at least five samples independent of standards and determining the coefficient of variation and/orappropriate confidence interval. The LLOQ should serve as the lowest concentration on the standard curve and should not be confused with the limit of detection and/or the low QC sample. The highest standard will define the upper limit of quantification (ULOQ) of an analytical method.•For validation of the bioanalytical method, accuracy and precision should be determined using a minimum of five determinations per concentration level (excluding blank samples).The mean value should be within ±15% of the theoretical value, except at LLOQ, where it should not deviate by more than ±20%. The precision around the mean value should not exceed 15% of the CV, except for LLOQ, where it should not exceed 20% of the CV.Other methods of assessing accuracy and precision that meet these limits may be equally acceptable.•The accuracy and precision with which known concentrations of analyte in biological matrix can be determined should be demonstrated. This can be accomplished by analysis ofreplicate sets of analyte samples of known concentrations C QC samples C from anequivalent biological matrix. At a minimum, three concentrations representing the entire range of the standard curve should be studied: one within 3x the lower limit of quantification (LLOQ) (low QC sample), one near the center (middle QC), and one near the upperboundary of the standard curve (high QC).•Reported method validation data and the determination of accuracy and precision should include all outliers; however, calculations of accuracy and precision excluding values that are statistically determined as outliers can also be reported.•The stability of the analyte in biological matrix at intended storage temperatures should be established. The influence of freeze-thaw cycles (a minimum of three cycles at twoconcentrations in triplicate) should be studied.•The stability of the analyte in matrix at ambient temperature should be evaluated over a time period equal to the typical sample preparation, sample handling, and analytical run times.•Reinjection reproducibility should be evaluated to determine if an analytical run could be reanalyzed in the case of instrument failure.•The specificity of the assay methodology should be established using a minimum of six independent sources of the same matrix. For hyphenated mass spectrometry-basedmethods, however, testing six independent matrices for interference may not be important.In the case of LC-MS and LC-MS-MS-based procedures, matrix effects should beinvestigated to ensure that precision, selectivity, and sensitivity will not be compromised.Method selectivity should be evaluated during method development and throughout methodvalidation and can continue throughout application of the method to actual study samples.•Acceptance/rejection criteria for spiked, matrix-based calibration standards and validation QC samples should be based on the nominal (theoretical) concentration of analytes.Specific criteria can be set up in advance and achieved for accuracy and precision over therange of the standards, if so desired.V.METHOD DEVELOPMENT: MICROBIOLOGICAL AND LIGAND-BINDING ASSAYSMany of the bioanalytical validation parameters and principles discussed above are also applicable to microbiological and ligand-binding assays. However, these assays possess some unique characteristics that should be considered during method validation.A.Selectivity IssuesAs with chromatographic methods, microbiological and ligand-binding assays should be shown to be selective for the analyte. The following recommendations for dealing with two selectivity issues should be considered:1.Interference From Substances Physiochemically Similar to the Analyte•Cross-reactivity of metabolites, concomitant medications, or endogenouscompounds should be evaluated individually and in combination with the analyteof interest.•When possible, the immunoassay should be compared with a validated reference method (such as LC-MS) using incurred samples and predetermined criteria foragreement of accuracy of immunoassay and reference method.。

美国FDA分析方法验证指引中英文对照

美国FDA分析方法验证指引中英文对照

美国FDA分析方法验证指南中英文对照美国FDA分析方法验证指南中英文对照八、、I.INTRODUCTIONThis guida nee provides recomme ndati ons to applica nts on submitt ing an alytical procedures, validati on data, and samples to support the docume ntati on of the identity, strength, quality, purity, and potency of drug substances and drug products.1.绪论本指南旨在为申请者提供建议,以帮助其提交分析方法,方法验证资料和样品用于支持原料药和制剂的认定,剂量,质量,纯度和效力方面的文件。

This guida nce is in ten ded to assist applica nts in assembli ng in formati on, submitt ing samples, and prese nti ng data to support an alytical methodologies. The recomme ndati ons apply to drug substa nces and drug products covered in new drug applicati ons (NDAs), abbreviated new drug applicati ons (ANDAs), biologics license applications (BLAs), product license applications (PLAs), and supplements to these即plicatio ns.本指南旨在帮助申请者收集资料,递交样品并资料以支持分析方法。

How to do - ICH Q7a(2010)-English & Chinese

How to do - ICH Q7a(2010)-English & Chinese

5.
Process Equipment工艺设备 ................................................................................................................................................................................ 16 5.1 5.2 5.3 5.4 Design and Construction设计与建造 ....................................................................................................................................................... 16 Equipment Maintenance and Cleaning设备保养和清洁 ......................................................................................................................... 17 Calibration校准 ......................................................................................................................................................................................... 17 Computerized Systems计算机化系统 ...................................................................................................................................................... 18

Method Validation for Related Substances

Method Validation for  Related Substances

3METHOD V ALIDATION FORHPLC ANALYSIS OF RELATED SUBSTANCES IN PHARMACEUTICAL DRUG PRODUCTSY.C.L EE,P H.D.Patheon YM,Inc.3.1INTRODUCTIONIn this chapter we outline the general requirements for analytical method valida-tion for HPLC analysis of related substances in pharmaceutical products.Most of the discussion is based on method validation for pharmaceutical products of synthetic origin.Even though most of the requirements are similar for other types of pharmaceutical drug products(e.g.,biopharmaceutical drug products),detailed discussion of method validation for other types of pharmaceutical drug products is outside the scope of this chapter.The discussion focuses on current regulatory requirements in the pharmaceutical industry.Since the expectations for method validation are different at different stages of the product development process, the information given in this chapter is most suitable forfinal method valida-tion according to the ICH requirements to prepare for regulatory submissions (e.g.,NDA).Even though the method validation is related to HPLC analysis, most of the principles are also applicable to other analytical techniques(e.g., TLC,UV).Analytical Method Validation and Instrument Performance Verification,Edited by Chung Chow Chan,Herman Lam,Y.C.Lee,and Xue-Ming ZhangISBN0-471-25953-5Copyright 2004John Wiley&Sons,Inc.2728METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCES3.2BACKGROUND INFORMATION3.2.1DefinitionsDefinitions for some of the commonly used terms in this chapter are given below.žDrug substance(active pharmaceutical ingredient):a pharmaceutical active ingredient.žRelated substances:impurities derived from the drug substance and there-fore not including impurities from excipients.Related substances include degradation products,synthetic impurities of drug substance,and manufac-turing process impurities from the drug product.žAuthentic sample:a purified and characterized sample of a related substance.Unlike reference standards,authentic samples may not be of high purity.However,the purity of an authentic sample has to be determined before use.Authentic samples are used in method development to identify related substances in the analysis.In addition,they are used extensively to prepare the spiked samples in method validation.žSpiked sample:a sample added with a known amount of related substances, prepared from authentic samples during method development or validation.žControl sample:a representative batch of drug substance(or drug product).Typically,control samples are tested in all analyses to ensure consistency in method performance across different runs.Sometimes,they are used as part of the system suitability test to establish the run-to-run precision(e.g., intermediate precision,reproducibility).žResponse factor:the response of drug substance or related substances per unit weight.Typically,the response factor of drug substance(or related substance)can be calculated by the following equation:Response factor=response(in response units) concentration(in mg/mL)žRelative response factor:the ratio of the response factor of individual related substance to that of a drug substance to correct for differences in the response of related substances and that of the drug substance.It can be determined using the following equation:Relative response factor=response factor of individual related substance response factor of drug substanceIf a linearity curve(Figure3.1)is constructed for both the related substance and the drug substance by plotting the response versus the concentration, the relative response factor can also be determined byRelative response factor=slope related substance slope drug substanceBACKGROUND INFORMATION 29P e a k a r e a Concentration (mg/mL)Figure 3.1.Relative response factor.3.2.2Different Types of Related Substance AnalysisArea Percent.In this approach,the level of an individual related substance is calculated by the following equation:%related substance =area related substance total area×100%where the area related substance is the peak area of the individual related substance and the total area is the peak area (i.e.,response)of the drug substance plus the peak areas of all related substances.This is one of the simplest approaches for related substance analysis because there is no need for a reference standard.This is particularly important during the early phase of the project when a highly purified reference standard is not available.It is the preferred approach as long as the method performance meets the criteria described below.Linearity over a Wide Range of Concentration .Since the areas of the related sub-stances (typically,less than 1%)and drug substance (typically,more than 95%)are summed,it is important that the method is linear from the concentration of related substances (e.g.,1%)to that of the drug substance (e.g.,95%).However,in some cases,the peak shape of the drug substance may not be totally sym-metrical at such a high concentration.Therefore,the response may not be linear in such a wide concentration range,and the use of area percentage may not be appropriate.If the response of the analyte is nonlinear at higher concentrations,the related substances would be overestimated.Although this is conservative from a safety perspective,it is inaccurate and therefore unacceptable.Sample Concentration (Method Sensitivity).To maintain linearity at the con-centration range of the drug substance,scientists may try to lower the sample concentration to improve peak shape for the drug substance.However,if the sample concentration is too low,it will affect the method sensitivity,and the ability to detect low levels of related substances may not be adequate.30METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCESResponse Factor .The response factors of the related substances should be sim-ilar to that of the drug substance (i.e.,relative response factors close to unity).Otherwise,a response factor correction must be used in the calculation.High–Low.This approach can be used to overcome the limitation of linear range in the area percent method discussed above.In this approach,samples are prepared at a concentration (i.e.,high concentration)similar to that of the area percent method (Figure 3.2).In addition,the high concentration sample solutions are diluted further,to low concentrations (Figure 3.3).Samples from both high-and low-concentration solutions are injected for analysis.In the injections of the high concentration,the responses of all related substances are determined as these small peaks are detectable.The high sample concentration is used to allow all related substances to be detected and quantitated.In the injection of Peak area for related substancesTime (min)A b s o r b a n c e (m A U )510152025010203040506070Figure 3.2.chromatogram from high concentration.Time (min)A b s o r b a n c e (m A U )010203040506070Figure 3.3.chromatogram from low concentration.BACKGROUND INFORMATION 31low-concentration sample,the response of the drug substance is determined.Low concentration is used to ensure that the response of the drug substance is within the linearity range.After dilution,response of the drug substance in the low-concentration sample is similar to that of related substance in the high-concentration sample.Therefore,only a small linearity range is required for this method.In addition,since high sample concentration is used for the determination of related substances,high method sensitivity can be achieved.The limitation of the high–low approach is that each sample is injected at least twice (i.e.,high and low concentrations)and the total analysis time will be doubled.In addition,an additional step is required to dilute the high concentration to a low concentration,and dilution error can occur during the second dilution.External Standard.In this approach,related substance levels are determined by calculation using a standard curve.The concentration of related substance is determined by the response (i.e.,peak area of individual related substance)and the calibration curve.A reference standard of the drug substance is typically used in the calibration.Therefore,a response factor correction may be required if the response of related substance is very different from that of the drug substance.A single-point standard curve (Figure 3.4)is appropriate when there is no significant y -intercept.Otherwise,a multipoint calibration curve (Figure 3.5)has to be used.Different types of calibration are discussed in Section 3.2.3.The external standard approach offers several advantages over the area per-cent method,as discussed below.Reduced Linear Range .Unlike the area percent and high–low methods,which use the response of the drug substance in sample injections for calculation,an external standard method uses a standard curve.Typically,the concentration range of the calibration curve is similar to that of related substances in the sample (e.g.,1to 5%of the nominal sample concentration).Therefore,this method requires a small linear range.P e a k a r e a Concentration (% related substance)Reference standard calibration curveArea found Figure 3.4.Single-point calibration.32METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCESConcentration (% related substance)% related substance foundat different levelsP e a k a r e a Area found Figure 3.5.Multi-point calibration.Improved Method Sensitivity .In this approach,only the responses of individual related substances are used in the calculation.Since the area of drug substance peak in the sample injections is not necessary for the calculation,high sample concentrations can be used without worrying about the off-scale response of the drug substance.This approach is particularly useful when the scientists want to improve the method sensitivity by increasing the sample concentration.Reference Standard .One of the limitations of the external standard method is that a well-characterized reference standard is essential.In addition,each anal-ysis requires accurate weighings of small quantities (e.g.,10mg)of reference standard.Therefore,weighing error can affect method precision and accuracy.3.2.3Suitability of Related Substance AnalysisAs discussed in Section 3.2.2,linear range is a critical factor for determining the suitable type of related substance analysis.The following are different situations to illustrate the rationales.Typically,the low end of a linearity curve is about 50%of the ICH reporting limit (e.g.,50%of 0.1%=0.05%).This is to ensure that the method will be able to calculate results accurately below the ICH reporting limit.The high end of the linearity curve is the nominal concentration (i.e.,100%).This is the target sample concentration for the drug substance.Case 1.Linearity demonstrated from 50%of the ICH reporting limit to a nominal concentration of drug substance in the sample solution.In addition,no signif-icant y -intercept is observed (Figure 3.6).In this case,area percent calculation is suitable because the linearity range covers the responses of related substances and that of the drug substance in the sample solution.Therefore,these responses can be used directly to calculate the area percentage of each related substance.Case 2.Linearity demonstrated from 50%of the ICH reporting limit to 150%of the shelf life specification of related substance.No significant y -intercept is observed (Figure 3.7).In this case,a high–low calculation is more suitable,asBACKGROUND INFORMATION33P e a k a r e a Concentration (% related substance)ICH reporting limitFigure 3.6.Linearity:case 1.Concentration (% related substance)reporting limitP e a k a r e a Figure 3.7.Linearity:case 2.the response is linear only up to the shelf life specification level.Drug substance concentration in sample solution (high concentration)should be diluted to the linear range to obtain the low-concentration solution.Therefore,the response of drug substance in low concentration will be within the linearity range and suitable for calculation.Alternatively,a single-point external standard calibration of concentration within the linearity range can also be used.Case 3.Linearity demonstrated from 50%of the ICH reporting limit to 150%of the shelf life specification of a related substance,and a significant y -intercept is observed (Figure 3.8).Due to the significant y -intercept,a single-point cali-bration (e.g.,high–low or one-point external standard calibration)is not suitable.In this case,multiple-point external standard calibration is the most appropriate.See Section 3.3.3for more discussion of the significant y -intercept.3.2.4Preparation before Method ValidationCritical Related Substances.Critical related substances are those that may exist at significant levels in the drug product.Authentic samples of these critical related34METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCESConcentration (% related substance)ICH reporting limit P e a k a r e a Figure 3.8.Linearity:case 3.substances should be available for method validation.According to the ICH guidelines,all related substances at a level exceeding the identification threshold have to be identified.These related substances should be considered critical and included in the method validation.To determine the critical related substances,one can review the related sub-stance profile when the drug substance (or drug product)is subject to stress testing.The most significant related substances in stress testing should be con-sidered critical.In addition,significant related substances (i.e.,greater than ICH identification threshold)observed in stability studies during product development should also be included in the method validation.The related substance method has to be validated with respect to each critical related substance;therefore,the workload associated with method validation will increase drastically if the number of critical related substances is large.Lower and Upper Concentration Range for Method Validation.The concentra-tion range of related substances is typically related to the targeted quantitation limit (QL)at the low end and the proposed shelf life specification at the high end.Therefore,it is important to have a good estimate of these limits;otherwise,inappropriate concentrations may be used in method validation.Even though ICH proposes a method validation range from the ICH reporting limit to 120%of specification,one would want to extend the range to 50%of the ICH report-ing limit to 150%of specification to ensure that the method is suitable for most intended uses.The ICH reporting limit is given in Table 3.1.In general,the quantitation limit should be lower than the corresponding ICH reporting limit.This is to ensure that the method is accurate and precise enough to report results at the level of the ICH reporting limit.Method Procedure.Since the method procedure is undergoing constant modifi-cations during method development,it is very important to define the procedure before method validation.This will ensure that the same method procedure will be used in all method validation experiments.METHOD V ALIDATION EXPERIMENTS35 Table3.1.Various ICH Thresholds Regarding Degra-dation Products in New Drug Products as Stated inthe Current ICH Guidelines Q3B(R)Maximum Daily Dose a Threshold bThresholds for Reporting≤1g0.1%>1g0.05%Thresholds for Identification<1mg 1.0%or5µg TDI c whicheveris lower1–10mg0.5%or20µg TDI,whicheveris lower>10mg–2g0.2%or2mg TDI,whicheveris lower>2g0.1%Thresholds for Qualification<10mg 1.0%or50µg TDI,whicheveris lower10–100mg0.5%or200µg TDI,whichever is lower>100mg–2g0.2%or2mg TDI,whicheveris lower>2g0.1%a The amount of drug substance administered per day.b Threshold is based on percent of the drug substance.c Total daily intake.Critical Experimental Parameters for Robustness.Critical experimental param-eters should be identified during method development,and they will be investi-gated in the robustness experiments.System Suitability Tests.The appropriate system suitability tests should be defi-ned before method validation(e.g.,precision,resolution of critical related sub-stances,tailing,detector sensitivity).These system suitability tests should be performed in each method validation experiments.System suitability results from the method validation experiment can be used to determine the appropriate system suitability acceptance criteria.3.3METHOD V ALIDATION EXPERIMENTSIn this section we outline the requirements for method validation according to current ICH guidelines.36METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCES3.3.1SpecificityICH definition:Specificity is the ability to assess unequivocally the analyte in the presence of components that may be expected to be present.Most related substance methods will be used in a stability study,and therefore they have to be stability indicating.Stability indicating means that the method has sufficient specificity to resolve all related substances and the drug substance from each other.Typically,for the related substance method for a drug product, degradation products are the most critical related substances.Therefore,as a minimum requirement,the method should have sufficient specificity to resolve the degradation products and the drug substance.In addition,all degradation products should be resolved from potential interference with the excipients. Samples for SpecificityžBlank solution to show no interference with any HPLC system artifact peak.žPlacebo to demonstrate the lack of interference from excipients.žDrug substance to show that all significant related substances are resolved from the drug substance.žAuthentic samples of critical related substances to show that all known related substances are resolved from each other.žTypically,a stressed sample of about10to20%degradation is used to demonstrate the resolution among degradation products.A10to20%de-graded sample is used because it has a sufficiently high concentration level of critical related substance.Therefore,these related substances can be detected easily.In addition,10to20%degradation is not too excessive,and the related substance profile should be close to that of a typical stability sample.žStressed placebo to show that the degradation products from the excipients will not interfere with the degradation products of the drug substance. Different Approaches1.When authentic samples of related substance are available.Analyzestressed drug product,placebo,drug substance,stressed placebo,and solutions spiked with authentic samples of related substances.The HPLC chromatograms are used to show the resolution among related substances, drug substance,and other potential interferences.In addition,check the peak homogeneity of the significant degradation products and drug substance by a photodiode array detector(PDA)or mass spectrometer.This verifies that no significant related substance coelute with each other.2.When authentic samples of impurities are not available.A stressed drugproduct can be analyzed to show separation of the most significant related substances.In addition,the peak homogeneity of the stressed sample should be investigated by PDA or mass spectrometry.Alternatively,one may use an orthogonal procedure to verify the method specificity.The orthogonalMETHOD V ALIDATION EXPERIMENTS37method can be a different technique(e.g.,capillary electrophoresis,thin-layer chromatography)or different type of HPLC analysis(e.g.,reversed phase versus normal phase).For example,compare the related substance profile in the original HPLC method and that of the orthogonal method.To demonstrate method specificity,the significant related substances should be consistent in these methods.3.3.2Quantitation Limit(and/or Detection Limit)ICH definition:The quantitation limit of an individual analytical procedure is the lowest amount of analyte in a sample that can be determined quantitatively with suitable precision and accuracy.The detection limit of an individual analytical procedure is the lowest amount of analyte in a sample that can be detected but not necessarily quantitated as an exact value.Two types of approaches can be used to determine the quantitation limit or detection limit,as described below.Signal-to-Noise Approach.Quantitation limit(QL;Figure3.9)is defined as the concentration of related substance in the sample that will give a signal-to-noise (S/N)ratio of10:1.Detection limit(DL)corresponds to the concentration that will give a signal-to-noise ratio of3:1.The quantitation limit of a method is affected by both the detector sensitivity and the accuracy of sample preparation at such a low concentration.In practice,the quantitation limit should be lower than the corresponding ICH reporting limit(Table3.1).To investigate the effect of both factors(i.e.,sample preparation and detector sensitivity),solutions of different concentrations near the ICH reporting limits are prepared by spiking known amounts of related substances into excipients. Each solution is prepared according to the procedure and analyzed repeatedly to determine the S/N ratio.The average S/N ratio from all analyses at each concen-tration level is used to calculate the QL or DL.The following equation can be used to estimate the QL at each concentration level.Since different concentration levels give different QLs,typically the worst-case QL will be reported as the QL of the method.QL at each concentration=10×concentration(in%related substance) S/N(average at each concentration)Noise (N) Figure3.9.Quantitation limit.38METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCES Alternatively,the spike solution can be diluted serially to lower concentrations. The S/N ratio at each concentration level is determined.The concentration level (in percent related substance)that gives an S/N value of about10will be reported as the QL.Standard Deviation Approach.The following equations can be used to deter-mine quantitation limit and detection limit by standard deviation of the response at low concentrations:QL=10×SD SDL=3.3×SD Swhere SD is the standard deviation of the response near QL and S is the slope of the linearity curve near QL.There are two ways to determine SD:ing experiments similar to those given for the signal-to-noise approach,determine the standard deviation of the responses by repeat analysis of a solution near the targeted QL.2.Construct a calibration curve near the targeted QL:a.Determine the residual standard deviation of the regression line of cal-ibration,orb.Determine the standard deviation of the y-intercept.Other Considerations for QL.To account for instrument-to-instrument variation, one may need to verify the QL in multiple runs using different instruments. The desired QL should be less than the ICH reporting limit(e.g.,50%of ICH reporting limit).QL should be appropriate;too high indicates that the method is not sensitive enough to report results at the ICH reporting limit.Too low indicates that insignificant degradation products,even though much lower than the ICH reporting limit,may need to be reported.To ensure that the HPLC system in each analysis is sufficiently sensitive to report results at the ICH reporting limit,one may use a detector sensitivity solution as part of the system suitability test.Since the ICH reporting limit corresponds to QL(i.e.,S/N=10),one-third of the ICH reporting limit should correspond to DL(i.e.,S/N=3).Therefore,as part of the system suitability test, a detector sensitivity solution of a concentration of about one-third of the ICH reporting limit level would be injected.The response of the detector sensitivity solution should meet the detection limit and should be visually distinguishable from baseline.Alternatively,one may evaluate the S/N ratio of the standard solution during method development or validation.Part of the routine system suitability test is toMETHOD V ALIDATION EXPERIMENTS39 determine the S/N of the standard solution before each analysis.Therefore,the S/N of each analysis needs to be greater than the established limit.3.3.3LinearityICH definition:The linearity of an analytical procedure is its ability(within a given range)to obtain test results that are directly proportional to the concentra-tion(amount)of analyte in the sample.General RequirementsRange.Ideally,linearity should be established from50%of the ICH reporting limit to the nominal concentration of drug substance in the sample solution(for area percent method).If the linearity does not support such a wide range of concentration,determine the linearity from50%of the ICH reporting level to 150%of the proposed shelf life specifications of the related substance(for the high–low and external standard methods)as a minimum.This will ensure a linear response for related substances at all concentration levels to be detected during stability.Experimental Requirements.Solutions of known concentrations are used to deter-mine the linearity.A plot of peak area versus concentration(in percent related substance)is used to demonstrate the linearity.Authentic samples of related sub-stances with known purity are used to prepare these solutions.In most cases,for the linearity of a drug product,spiking the related substance authentic sample into excipients is not necessary,as the matrix effect should be investigated in method accuracy.Acceptance Criteria.Visual inspection is the most sensitive method for detecting nonlinearity.Therefore,the plot has to be linear by visual inspection.In addition, according to ICH guidelines,the following results should be reported:slope, correlation coefficient,y-intercept,and residual sum of squares.y-Intercept.There are several approaches to evaluating the significance of the y-intercept.žIntercept/slope ratio.The intercept/slope ratio is used to convert the y-intercept from the response unit(peak area)to the unit of percent related substance.The intercept/slope ratio should be compared to the proposed specifications to determine its significance.For example,if the shelf life specification is2.0%,an intercept/slope ratio of0.2%may be considered significant,as0.2%represents10%relative to the specification.žStatistical approach.The linearity results can be subjected to statistical analysis(e.g.,use of statistical analysis in an Excel spreadsheet).The p-value of the y-intercept can be used to determine if the intercept is sta-tistically significant.In general,when the p-value is less than0.05,the40METHOD V ALIDATION FOR HPLC ANALYSIS OF RELATED SUBSTANCES y-intercept is considered statistically significant.The p-value,which com-pares the y-intercept with the variation of responses,indicates the probability that the y-intercept to be not equal to zero.For example,when the p-value is less than0.05,this indicates that it is95%confident that the y-intercept is not equal to zero.In other words,it is95%certain that the y-intercept is significant.Typically,a positive y-intercept indicates the existence of interference with the response or the saturation of responses at high concentrations.A negative y-intercept indicates the possibility of method sensitivity problem(i.e.,a low response cannot be detected)or analytes get retained in the glassware or HPLC system(i.e.,a compatibility issue between sample solvent and mobile phase). Different Approaches for Linearity Determination.Thefirst approach is to weigh different amounts of authentic sample directly to prepare linearity solu-tions of different concentrations.Since solutions of different concentration are prepared separately from different weights,if the related substances reach their solubility limit,they will not be completely dissolved and will be shown as a nonlinear response in the plot.However,this is not suitable to prepare solutions of very low concentration,as the weighing error will be relatively high at such a low concentration.In general,this approach will be affected significantly by weighing error in the preparation.Another approach is to prepare a stock solution of high concentration,then perform serial dilution from the stock solution to obtain solutions of lower con-centrations for linearity determination.This is a more popular approach,as serial dilution can be used to prepare solutions of very low concentrations.Since the low concentrations are prepared by serial dilution,this approach does not need to weigh a very small quantity of related substance.In addition,since all solutions are diluted from the same stock solution,weighing error in preparing the stock solution will not affect the linearity determination.Relative Response Factor.The relative response factor(RRF)can be used to correct for differences in relative response between the related substances and the drug substance.In the area percent and high–low method,the related sub-stances are calculated against the response of the drug substance.In the external standard calculation,the standard curve of drug substance is generally used in the calculation.Since the related substances are calibrated by the response of the drug substance,it is necessary to determine the relative response of the related substance to that of the drug substance.After the linearity of the related sub-stances and the drug substance are determined,one can calculate the relative response factor by comparing the slope of the related substance to that of the drug substance.If the relative response factor is significantly different from unity, a correction factor may need to be used in the calculation.Otherwise,the reported results will be grossly over-or underestimated(Figure3.10).。

iDOVE包的文档 持久性疫苗有效性评估与SARS-CoV-2感染关联的非参数最大似然方法说明书

iDOVE包的文档  持久性疫苗有效性评估与SARS-CoV-2感染关联的非参数最大似然方法说明书

Package‘iDOVE’December14,2023Type PackageTitle Durability of Vaccine Efficacy Against SARS-CoV-2InfectionVersion1.5Date2023-12-13Author Yu Gu[aut],Shannon T.Holloway[aut,cre],Dan-Yu Lin[aut]Maintainer Shannon T.Holloway<****************************> Description Implements a nonparametric maximum likelihood method for assessing potentially time-varying vaccine efficacy(VE)against SARS-CoV-2infection under staggered enrollment and time-varying community transmission,allowing crossover of placebo volunteers to the vaccine arm.Lin,D.Y.,Gu,Y.,Zeng,D.,Janes,H.E.,and Gilbert,P.B.(2021)<doi:10.1093/cid/ciab630>.License GPL-2Encoding UTF-8Suggests rmarkdown,knitrVignetteBuilder utils,knitrImports Rcpp(>=1.0.4.6),methods,stats,graphics,RcppArmadilloLinkingTo Rcpp,RcppArmadilloRoxygenNote7.2.3Collate'VEplot.R''VEcal.R''postProcess.R''EMmeth.R''RcppExports.R''verifyInputs.R''idove.R''idoveData.R''intCens.R''plot.iDOVE.R''print.iDOVE.R'NeedsCompilation yesDepends R(>=3.5.0)Repository CRANDate/Publication2023-12-1323:40:02UTC1R topics documented:idove (2)idoveData (5)intCens (5)plot (6)print (7)Index9 idove Durability of Vaccine Efficacy Against Asymptomatic SARS-CoV-2In-fectionDescriptionAssesses potentially time-varying vaccine efficacy(VE)against SARS-CoV-2infection under stag-gered enrollment and time-varying community transmission,allowing crossover of placebo volun-teers to the vaccine arm.The infection time data are interval-censored,and the log hazard ratio is assumed to be a piece-wise linear function of time.Usageidove(formula,data,constantVE=FALSE,plots=TRUE,changePts=NULL,timePts=NULL,tol=1e-04,maxit=2000)Argumentsformula A formula object,with all of the time variables on the left hand side of a’~’operator and the covariates on the right.The time variables must be specifiedthrough the intCens()function.See?intCens and Details for further information.data A data.frame object.The data.frame in which to interpret the variable names in formula.Must contain the entry time,the left interval time,the right intervaltime,the vaccination time,and any covariates.See Details.constantVE A logical object.If FALSE(default),VE is assumed to be potentially waning after the last change point;otherwise it is assumed to be constant after the lastchange point.plots A logical object.If TRUE(default),plots of the estimated VE in reducing attack rate,the estimated VE in reducing the hazard rate,and their95%confidenceintervals will be automatically generated.If FALSE,plots will not be generated.changePts An integer vector object or NULL.The potential change points(in days)of the piece-wise log-linear hazard ratio.See Details for further information.If NULL,the Akaike information criterion(AIC)will be used to select one change pointfrom{28,35,42,49,56}(weeks4,5,6,7,8).timePts An integer vector object or NULL.The endpoints(in days)of the time periods for which the VE in reducing the attack rate are to be estimated.The estimatedVE in reducing the hazard rate at these endpoints are also returned.If NULL,adefault sequence t1,2t1,3t1,...will be used,where t1is thefirst change point.The sequence ends at the maximum of the left and and right ends of the timeintervals from all participants.This input is ignored when constantVE=TRUE.tol A numeric scalar object.The convergence threshold for the EM algorithm.The default value is0.0001.maxit A positive integer object.The maximum number of iterations for the EM algo-rithm.The default value is2000.DetailsThe information required for an analysis isEntry Time:The time when the participant enters the trial in whole units days.Left Interval Time:The last examination time when the test is negative in whole units days.Right Interval Time:Thefirst examination time when the test is positive in whole units days.If the participant does not test positive during the trial,use NA or Inf.Vaccination Time:The time when vaccination takes place in whole units days.If the participant is not vaccinated during the trial,use NA or Inf.Covariates:Baseline covariates(e.g.,priority group,age,ethnicity).The covariates can include categorical variables,for which all other categories are compared to the first category.A model without covariates is also allowed.Note that all of the time variables are measured from the start of the clinical trial and are specified in whole units of days.Though they need not be provided as integer,all non-NA andfinite values must be able to be cast as integers without loss of information.For each individual,the entry_time and left_time should satisfy entry_time≤left_time.For each individual that tests positive,entry_time ≤left_time≤right_time.For each individual that is vaccinated,entry_time≤vaccination_time.The general structure of the formula input isintCens(entry_time,left_time,right_time,vaccination_time)~covariatesThe left-hand side contains all of the time information.It must be specified through function’int-Cens()’.Specifically,intCens(entry_time,left_time,right_time,vaccination_time)If entry_time>left_time,or left_time>right_time,the case will be removed from the analysis anda message will be generated.The special case of right-censored data is implemented by dove2()in the DOVE package available through CRAN.ValueAn S3object of class iDOVE containing a list with elementscall The unevaluated call.changePts The changePts of the analysis.covariates A matrix containing the estimated(log)hazard ratio of each covariate,togetherwith the estimated standard error,the95%confidence intervals,and the two-sided p-value for testing no covariates effect.NA if only an intercept is given asthe right hand side in input formula.vaccine A list containing one or three elements,depending on the value of constantVE.IfconstantVE=TRUE,the only element is named’VE’and is a vector containingthe estimate of constant VE,its standard error estimate,and the95%confidenceinterval.If constantVE=FALSE,three matrices are returned.Thefirst matrixnamed’VE_a’contains the estimates of the VE in reducing the attack rate at alltime points given in timePts,together with the95%confidence intervals.Thesecond matrix named’VE_h’contains the estimates of the VE in reducing thehazard rate at timePts.The third matrix named’VE_period’contains the esti-mates of VE in reducing the attack rate over successive time periods accordingto timePts,together with the95%confidence intervals.ReferencesLin,D-Y,Gu,Y.,Zeng,D.,Janes,H.E.,and Gilbert,P.B.(2021).Evaluating Vaccine EfficacyAgainst SARS-CoV-2Infection.Clinical Infectious Diseases,ciab630,https:///10.1093/cid/ciab630 Examplesdata(idoveData)set.seed(1234)smp<-sample(1L:nrow(x=idoveData),size=250L)#NOTE:This sample size is chosen for example only--larger data sets#should be used.#See the vignette for a full analysis of the idoveData dataset#Fit the model with default settingsidove(formula=intCens(entry.time,left.time,right.time,vaccine.time)~1,data=idoveData[smp,])#Specify Week4as the change point#Assume a potentially waning VE after4weeks#Estimate VE_a over0-4,4-16,16-28,28-40weeksidove(formula=intCens(entry.time,left.time,right.time,vaccine.time)~1,data=idoveData[smp,],changePts=4*7,timePts=c(4,16,28,40)*7)#Specify multiple change points at Weeks4and8idoveData5 #Assume a constant VE after8weeksidove(formula=intCens(entry.time,left.time,right.time,vaccine.time)~1, data=idoveData[smp,],changePts=c(4,8)*7,constantVE=TRUE)idoveData Toy Dataset For IllustrationDescriptionThis data set is provided for the purposes of illustrating the use of the software.It was simulated under a blinded,priority-tier dependent crossover design.Usagedata(idoveData)FormatidoveData is a data.frame containing40,000participants.The data.frame contains6columns, entry.time The entry time in days.left.time The left end of the time interval in days.right.time The right end of the time interval in days.vaccine.time The time of vaccination in days.priority A composite baseline risk score taking values1-5.sex A binary indicator of sex(male/female).intCens Specify Time VariablesDescriptionThis function is used in the model statement of idove()to specify the entry time,left interval time, right interval time,and vaccination time.UsageintCens(entry_time,left_time,right_time,vaccination_time)6plotArgumentsentry_time The variable for the time when the participant enters the trial.Entry times must be integer(or be able to be cast as integer without loss of information),non-negative,and complete.left_time The variable for the last examination time when the test is negative.Left in-terval times must be integer(or be able to be cast as integer without loss ofinformation),non-negative,and complete.right_time The variable for thefirst examination time when the test is positive.Right in-terval times must be integer(or be able to be cast as integer without loss of in-formation),non-negative,and NA or Inf should be used if the participant nevertested positive during the trial.vaccination_timeThe variable for the time when vaccination takes place.Vaccination times mustbe integer(or be able to be cast as integer without loss of information),non-negative,and NA or Inf should be used if the participant was not vaccinatedduring the trial.DetailsTimes must obey the following relationships:(i)For all participants,entry_time<=left_time;(ii) For all participants that tested positive during the trial,entry_time<=left_time<=right_time;and (iii)For all participants that received vaccination,entry_time<=vaccination_time.If a case is found to violate one or more of these relationships,its entry_time is set to NA.ValueThis function is intended to be used only in the model statement of idove().The result,a matrix,is used internally.plot Plot Estimated Vaccine EfficacyDescriptionGenerates plots of the estimated vaccine efficacy in reducing attack rate,the estimated vaccine efficacy in reducing the hazard rate,and their95%confidence intervals.Usage##S3method for class iDOVEplot(x,...)Argumentsx An iDOVE object.The value object returned by idove()....ignoredValueNo return value,called to produce graphical elements.Examplesdata(idoveData)set.seed(1234)smp<-sample(1L:nrow(x=idoveData),size=250L)#NOTE:This sample size is chosen for example only--larger data sets#should be used.#See the vignette for a full analysis of the idoveData dataset#Fit the model with default settingsresult<-idove(formula=intCens(entry.time,left.time,right.time,vaccine.time)~1, data=idoveData[smp,])plot(x=result)print Print the Primary Results of an idove()AnalysisDescriptionPrint the primary results of an idove()analysis.Usage##S3method for class iDOVEprint(x,...)Argumentsx An iDOVE object.The value object returned by a call to idove()...ignoredValueNo return value,called to display key results.Examplesdata(idoveData)set.seed(1234)smp<-sample(1L:nrow(x=idoveData),size=250L)#NOTE:This sample size is chosen for example only--larger data sets#should be used.#See the vignette for a full analysis of the idoveData dataset#Fit the model with default settingsresult<-idove(formula=intCens(entry.time,left.time,right.time,vaccine.time)~1, data=idoveData[smp,])print(x=result)Index∗datasetsidoveData,5idove,2idoveData,5intCens,5plot,6print,79。

Method Validation 2009

Method Validation 2009

Observed %Assay (on the dried basis)
测得含量
• At=Peak area of API in the test sample solution • As=Average peak area of API from the 5 standard injection • Ct=Concentration of the sample solution in ug/mL • Ca=Concentration of the API standard solution in ug/mL • P=Purity of API standard in % • E=Total volatiles (%)=water% + Total residual solvents% (from test sample COA)
*
Category II 杂质 Category I 主要成份 Yes Yes Yes No No Quantitative 定量 Yes Yes Yes No Yes Limit-Test 限度
*
Category III 功能特性
*
Category IV 成分ID No No Yes No
No Yes Yes
API ACCURACY API准确性
• • • • • • API: evaluated at 80%, 100%, 120%, three samples at each concentration (Use test sample) 用待测样品配制80%,100%,120%溶液,各三个 Inject each solution once 每个样品测一次 Observed %assay (on the dried basis) 测得含量 Theoretical %Assay (From COA) 理论含量 API %Recovery 计算API回收率 Acceptance criteria: Mean %Recovery of %Assay 98.0-102.0% at each level 接受标准:平均API回收率在各个浓度在98.0-102.0%之间

美国FDA分析方法验证指南中英文对照--6

美国FDA分析方法验证指南中英文对照--6

美国FDA分析⽅法验证指南中英⽂对照--6XI. METHODOLOGYSections II through IX provide general information on the submission of analytical procedures and methods validation information, including validation characteristics. Additional information on certain methodologies is provided below.XI.⽅法学II章到第IX章提供了分析⽅法和分析⽅法验证资料⽅⾯的基本信息,包括验证项⽬。

下⽂就⼀些具体的⽅法给出了说明:A. High-Pressure Liquid Chromatography (HPLC)The widespread use of HPLC analytical procedures and the multitude of commercial sources of columns and packings frequently have created problems in assessing comparability. Many of the following points may also apply to other chromatographic analytical procedures.⾊谱(HPLC)⾼效液相⾊谱A.⾼效液相HPLC分析⽅法的⼴泛应⽤及⾊谱柱和柱填充的众多来源都经常会给可⽐性评估带来很多问题。

如下这些要点中,很多都适⽤于其它⾊谱分析⽅法。

1. ColumnThe following characteristics are useful for defining a particular column and, if known, should be included in the analytical procedure description. If method development has indicated that columns from only one commercial source are suitable, this information should be included as part of the analytical procedure. If more than one column is suitable, a listing of columns found to be equivalent should be included.1.⾊谱柱在定义某⼀⾊谱柱时,如下这些性质是很有⽤的,也应当要包括在分析⽅法描述中。

201507FDA行业指南:分析方法验证(中英文)(上)

201507FDA行业指南:分析方法验证(中英文)(上)

201507FDA行业指南:分析方法验证(中英文)(上)Analytical Procedures and Methods Validation for Drugs and Biologics药品和生物制品分析方法验证Guidance for Industry行业指南U.S. Department of Health and Human ServicesFood and Drug AdministrationCenter for Drug Evaluation and Research (CDER)Center for Biologics Evaluation and Research (CBER)July 2015Pharmaceutical Quality/CMCAnalytical Procedures and Methods Validation for Drugs and BiologicsGuidance for IndustryAdditional copies are available from:Office of Communications, Division of Drug InformationCenter for Drug Evaluation and ResearchFood and Drug Administration10001 New Hampshire Ave., Hillandale Bldg., 4th FloorSilver Spring, MD 20993Phone: 855-543-3784 or 301-796-3400; Fax: 301-431-6353 Email:****************.gov/Drugs/GuidanceComplianceRegulatoryInformation/Guidan ces/default.htmand/orOffice of Communication, Outreach and DevelopmentCenter for Biologics Evaluation and ResearchFood and Drug Administration10903 New Hampshire Ave., Bldg. 71, Room 3128Silver Spring, MD 20993Phone: 800-835-4709 or 240-402-7800Email:************.gov/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInf ormation/Guidances/default.htmU.S. Department of Health and Human ServicesFood and Drug AdministrationCenter for Drug Evaluation and Research (CDER)Center for Biologics Evaluation and Research (CBER)July 2015Pharmaceutical Quality/CMCAnalytical Procedures and Methods Validation for Drugs and Biologics药物和生物制品分析方法验证Guidance for Industry[1]行业指南This guidance represents the current thinking of the Food and Drug Administration (FDA or Agency) on this topic. It does not create any rights for any person and is not binding on FDA or the public. You can use an alternative approach if it satisfies the requirements of the applicable statutes and regulations. To discuss an alternative approach, contact the FDA staff responsible for this guidance as listed on the title page.本指南代表了FDA对本专题的当前想法。

  1. 1、下载文档前请自行甄别文档内容的完整性,平台不提供额外的编辑、内容补充、找答案等附加服务。
  2. 2、"仅部分预览"的文档,不可在线预览部分如存在完整性等问题,可反馈申请退款(可完整预览的文档不适用该条件!)。
  3. 3、如文档侵犯您的权益,请联系客服反馈,我们会尽快为您处理(人工客服工作时间:9:00-18:30)。

1. Validation of Rating discriminatory power:−Cumulative Accuracy Profile (“CAP”) and itssummary index, the Accuracy Ratio (“AR”);−Receiver Operating Characteristic (“ROC”) and itssummary indices, the ROC measure and the PietraIndex;−Bayesian error rate (“BER”);−Conditional entropy, Kullback-Leibler distance, andConditional Information Entropy Ratio (“CIER”);−Information value (“IV”);−Kendall’s τ and Somers’ D (for shadow ratings);−Brier score (“BS”); (this is similar to RMSE which we will do in validation)−Divergence.Not applicable since we will follow Basel’s rankingand our validation is not on discriminatory power.2. Validation of PD calibration−Binomial test with assumption of independentdefault events;−Binomial test with assumption of non-zero defaultcorrelation;−Chi-square test.We will do this validation on the calibration, for the accuracy instead of conservative.3. Validation of LGD estimates−Comparisons between internal LGD estimatesand relevant external data sources This may not be applicable since currently in HK and PRC region, no bank has fully completed the IFRS9 modelling work, including HSBC. For bond, the external LGD has been used in IFRS9 model already.−Comparisons between realised LGD of newdefaulted facilities and their LGD estimates We will do this.4. Validation of EAD estimates−back-test their internal EAD estimates against therealised EAD of the new defaulted facilitiesWe will do this for some segments which Basel EAD will be modified in IFRS9−Where available, AIs should compare their internalestimates with external benchmarks.This may not be applicable since currently in HK and PRC region, no bank has fully completed the IFRS9 modelling work, including HSBC.−compare the estimated aggregate EAD amount forthe subject facility type with the realised aggregateEAD amount for that facility typeWe will do this for some segments which Basel EAD will be modified in IFRS95. Benchmarking−HKMA will expect AIs to obtain theirbenchmarks from third parties, provided that relevantexternal benchmarks for a specific portfolio areavailable. When external benchmarks are not used,despite being available, the HKMA will expect AIs toprovide valid justifications and demonstrate that theyhave other compensating measures(prehensive back-testing at a frequency higher thanrequired, such as quarterly,with sufficient defaultobservations to ensure the reliability of the back-testingresults) to ensure the accuracy of their rating systems.The HKMA will not accept cost implications as the solejustification for not using external benchmarks.For external benchmark, it may not be applicable since currently in HK and PRC region, no bank has fully completed the IFRS9 modelling work, including HSBC. And that’s why we suggest using a higher frequency data to do the back testing.−Where a relevant external benchmark is not available(e.g. PD of SME and retail exposures, LGD and EAD),an AI should develop an internal benchmark. Forexample, to benchmark against a model-based ratingsystem, an AI might employ internal rating reviewers tore-rate a sample of credits on an expert-judgementbasis.Internal benchmark will be useful for IRB since we could ask rating reviewers to do the judgmental rating for some sampling. However our IFRS9 methodology is based on Basel rating, this is not applicable for IFRS9 validation. And normally we do not build new models with other methodology since it is not comparable for the model performance under different methodologies.−the HKMA willnormally expect AIs to use in validating their ratingsystems and internal estimates:∙comparison of internal estimates with benchmarkswith respect to a common or similar set ofborrowers/facilities;∙comparison of internal ratings and migrationmatrices with the ratings and migration matrices ofthird parties such as rating agencies or data pools;∙comparison of internal ratings with external expertjudgements, for example, where a portfolio has notexperienced recent losses but historical experiencesuggests that therisk of loss is greater than zero;∙comparison of internal ratings or estimates withmarket-based proxies for credit quality, such asequity prices, bond spreads, or premiums for creditderivatives;∙analysis of the rating characteristics of similarlyrated exposures; and∙comparison of the average rating output for theportfolio as a whole with actual experience for theportfolio rather than focusing on estimates forindividualborrowers/facilities.As HKMA required, if we choose external benchmark, we need to assess the quality in adequately representing the riskcharacteristics of the portfolio under consideration, including definition of default, rating criteria, data quality, frequency of rating updates and assessment horizon.This will be challenging to us since it is not easy to get all these information for peers.Also currently in HK and PRC region, no bank has fully completed the IFRS9 modelling work, including HSBC.6. The other information that HKMA mentioned in CA-G-4:Validation methodology mentioned in Terminology:−“k-fold cross validation” means a kind of testemploying resamplingtechniques. The data set isdivided into k subsets. Each time, one of the ksubsets is used as the validation data set and theother k-1 subsets are put together to form thedevelopment data set. Byrepeating theprocedures k times, the targeted test statisticacross all k trials is then computed;−“bootstrapping” means a resampling technique withreplacement of the data sampled, aiming togenerate information on the distribution of theunderlying data set;−“in-sample validation” means validation of a ratingsystem employing observations that have beenused for developing the rating system;−“out-of-sample validation” means validation of arating system employing observations that have notbeen used for developing the rating system;−“out-of-time validation” means validation of a ratingsystem employing observations that are notcontemporary with the data used for developing therating system;Besides, the HKMA CA-G-4 also mentioned the validation methodology which AI should consider due to the data limitations is described as below:If out-of-sample and out-oftimevalidations cannot be conducted due to dataconstraints, AIs will be expected to employ statisticaltechniques such as k-fold cross validation orbootstrapping for this purpose.。

相关文档
最新文档