药典分析方法确认与验证.doc

usp_1225 药典方法的验证 2011.02.26验证: Validation 针对新方法(没有法定方法, 需要用户自己建立的) 按照ICH Q2 或USP的方式均可; 主要是需要满足使用目的. VALIDATION OF COMPENDIAL PROCEDURES药典规程的验证Test procedures for assessment of the quality levels of pharmaceutical articles are subject to various requirements. According to Section 501 of the Federal Food, Drug, and Cosmetic Act, assays and specifications in monographs of the United States Pharmacopeia and the National Formulary constitute legal standards. The Current Good Manufacturing Practice regulations 21 CFR 211.194(a) require that test methods, which are used for assessing compliance of pharmaceutical articles with established specifications, must meet proper standards of accuracy and reliability. Also, according to these regulations 21 CFR 211.194(a)(2), users of analytical methods described in USP-NF are not required to validate the accuracy and reliability of these methods, but merely verify their suitability under actual conditions of use. Recognizing the legal status of USP and NF standards, it is essential, therefore, that proposal for adoption of new or revised compendial analytical procedures be supported by sufficient laboratory data to document their validity. 评估药品质量水平的实验方法受到多种要求的影响。要依据美国食品、药品、化妆品法案501款以及美国药典和国家处方集的各论中的含量测定和质量标准来构建法定标准。现行药品生产质量管理规范【21 CFR 211.194(a)】要求，用于评价药物与既有质量标准之间的符合性的分析规程必须在准确度和可靠性方面达到适当的标准。并且根据【21 CFR 211.194(a)(2)】的这些规定，不要求USP-NF（美国药典/国家处方集）中描述的分析方法/规程的使用者去验证这些规程的准确度和可靠性，而仅需确认其在正确实际使用条件下的适用性。认识到USP和NF标准的法定地位，因此，提议采纳新的或更改过的药典分析规程时，必须有充分的实验室数据作为支持，以记录其有效性。The text of this information chapter harmonizes, to the extent possible, with the Tripartite International Conference on Harmonization (ICH) documents Validation of Analytical Procedures and the Methodology extension text, which are concerned with analytical procedures included as part of registration applications submitted within the EC, Japan, and the USA. 本信息章节的内容尽可能地与三方国际协调会议（ICH）文件分析规程的验证和方法学的延伸内容保持一致，那是与包含欧盟、日本和美国注册申请内的分析方法相关的。SUBMISSIONS TO THE COMPENDIA向药典提交的文件Submissions to the compendia for new or revised analytical procedures should contain sufficient information to enable members of the USP Council of Experts and its Expert Committees to evaluate the relative merit of proposed procedures. In most cases, evaluations involve assessment of the clarity and completeness of the description of the analytical procedures, determination of the need for the procedures, and documentation that they have been appropriately validated. Information may vary depending upon the type of method involved. However, in most cases a submission will consist of the following sections. 向药典提交关于新的或更改过的分析规程的文件应该包括充足的信息，以使USP专家理事会和专家委员会能够评估拟议规程的相对优越性。在大多数情况下，这些评估涉及对分析规程描述的清晰度和完整度的评估，对规程的需求的确定，以及它们已经进行了适当验证的记录文件。这些信息可以根据所涉及规程的种类而变化。但是，在大多数情况下，提交的文件将有下面的章节组成。Rationale- This section should identify the need for the procedure and describe the capability of the specific procedure proposed and why it is preferred over other types of determinations. For revised procedures, a comparison should be provided of limitations of the current compendial procedure and advantages offered by the proposed procedure. 理论依据- 此部分应该辨明对于该规程的需求，并描述具体拟议中规程的能力，以及为什么优于其他种类检测。对于更改的方法/规程，应该提供对当前药典规程之缺陷与拟议中规程之优势的比较。Proposed Analytical Procedure- This section should contain a complete description of the analytical procedure sufficiently detailed to enable persons “skilled in the art” to replicate it. The write-up should include all important operational parameters and specific instructions such as preparation of reagents, performance of system suitability tests, description of blanks used, precautions, and explicit formulas for calculation of test results.推荐的分析方法/规程- 此部分包含对该分析规程的完整描述，应足够具体以便能让业内技术熟练的人重复它。文章应该包括所有重要的操作参数和具体的指令，例如：试剂制备、系统适用性测试表现、所使用空白对照的描述、预防措施、用于计算测试结果的明确公式。Data Element- This section should provide thorough and complete documentation of the validation of the analytical procedure. It should include summaries of experimental data and calculations substantiating each of the applicable analytical performance characteristics. These characteristics are described in the following section. 数据要素- 此部分应该提供完全彻底的分析规程验证记录文件。其应该包括对于证明每一个实用功能特性的实验数据和计算的概况、总结。这些特性在下面的部分描述。VALIDATION验证Validation of an analytical procedure is the process by which it is established, by laboratory studies, that the performance characteristics of the procedure meet the requirements for the intended analytical applications. Typical analytical performance characteristics that should be considered in the validation of the types of procedures described in this document are listed in Table 1. Because opinions may differ with respect to terminology and use, each of the performance characteristics is defined in the next section of this chapter, along with a delineation of a typical method or methods by which it may be measured. 分析规程的验证是，通过实验室研究，确定该规程的工作特性达到了预定分析用途要求的过程。在此文件中所描述的规程种类的验证中，应当考虑的常见分析工作特性在表1中列出。因为对于术语和使用的意见可能不同，在此通则的下个部分定义了每个工作性能，以及可以对其进行测量的常用的一个或几个方法的描绘。Table 1. Typical Analytical Characteristics Used in Method Validation表1. 在方法验证中使用的常用分析特性Accuracy准确度Precision精密度Specificity专属性Detection Limit检测限度Quantitation Limit定量限度Linearity线性Range范围Robustness耐用性In the case of compendial procedures, revalidation may be necessary in the following cases: a submission to the USP of a revised analytical procedure; or the use of an established general procedure with a new product or raw material (see below in Data Elements Required for Validation).在药典方法的情况下，在下面的情况有必要进行再验证：向USP提交修改的分析规程；或将已确立的通用规程用于新产品或原料（见下面验证必需的数据要素）The ICH documents give guidance on the necessity for revalidation in the following circumstances: changes in the synthesis of the drug substance; changes in the composition of the drug product; and changes in the analytical procedure.ICH文件对于下列情况下再验证的必要性做出了指导：药物合成过程有变更；药品组成的有变更；以及分析规程中有变更。Analytical Performance Characteristics分析性能特征ACCURACY准确度Definition- The accuracy of an analytical procedure is the closeness of test results obtained by that procedure to the true value. The accuracy of an analytical procedure should be established across its range. 定义- 分析规程的准确度是由该规程得到的测试结果与真实值的接近程度。分析规程的准确度应通过它的适用范围来建立。Determination- In the case of the assay of a drug substance, accuracy may be determined by application of the analytical procedure to an analyte to known purity (e.g., a Reference Standard) or by comparison of the results of the procedure with those of a second, well-characterized procedure, the accuracy of which has been stated or defined. 测定- 当对一种药物进行分析时，准确度可以通过该分析规程来分析一个已知纯度的物质（例如，某个标准物质）来进行测定，或者通过比较运用这个分析方法所得的结果与另一个已经鉴定的、其准确度已被说明或被解释过的方法所得的结果来进行测定。In the case of the assay of a drug in a formulated product, accuracy may be determined by application of the analytical procedure to synthetic mixtures of the drug product components to which known amounts of analyte have been added within the range of the procedure. If it is not possible to obtain samples of all drug product components, it may be acceptable either to add known quantities of the analyte to the drug product (i.e., “to spike”) or to compare results with those of a second, well-characterized procedure, the accuracy of which has been stated or defined. 当分析一种成品药时，准确度可以通过分析规程来分析成药各组分的合成物质来测定，而添加进去的分析物质是在方法的范围之内的、是已知数量的。如果不能得到成药的所有组分，也可以将已知数量的待分析物加入到该成药中（例如，“增敏”），或者将结果与用第二种、成熟的、已知准确度的规程得到的结果进行比较，In the case of quantitative analysis of impurities, accuracy should be assessed on samples (of drug substance or drug product) spiked with known amount of impurities. Where it is not possible to obtain samples of certain impurities or degradation products, results should be compared with those obtained by an independent procedure. In the absence of other information, it may necessary to calculate the amount of an impurity based on comparison of its response to that of the drug substance; the ratio of the response of equal amounts of the impurity and the drug substance (relative response factor) should be used if known. 对于杂质的定量分析，应使用以已知数量杂质增敏的样品来评估准确度。当不能获得特定杂质或降解产物的样品时，应将结果与另一独立方法获得的结果进行比较。在没有其他信息的情况下，可能必需通过将某种杂质的响应值与药物的响应值进行比较来计算杂质的数量；同等数量的杂质与药物的响应值的比值（相对响应因子），如果已知，则应使用。Accuracy is calculated as the percentage of recovery by the assay of the known added amount of analyte in the sample, or as the difference between the mean and the accepted true value, together with confidence interval. 通过测定被加入到样品中的已知数量的被分析物来计算准确度，得到回收百分比，或得到平均值与可接受的真实值之间的差异，同置信区间一起。The ICH documents recommended that accuracy should be assessed using a minimum of nine determinations over a minimum of three concentration levels, covering the specified range (i.e., three concentrations and three replicates of each concentration).ICH文件建议精密度的评估应当使用覆盖规定范围的至少三个浓度水平进行至少九次测试（例如，三个浓度并且每个浓度三次重复进样）。Assessment of accuracy can be accomplished in a variety of ways, including evaluating the recovery of the analyte (percent recovery) across the range of the assay, or evaluating the linearity of the relationship between estimated and actual concentrations. The statistically preferred criterion is that the confidence interval for the slope be contained in an interval around 1.0, or alternatively, that the slope be close to 1.0. In either case, the interval or the definition of closeness should be specified in the validation protocol. The acceptance criterion will depend on the assay and its variability and on the product. Setting an acceptance criterion based on the lack of statistical significance of the test of the null hypothesis that the slop is 1.0 is not an acceptable approach. 准确度的评估可以通过多种不同的方式完成，包括评价在含量测定的整个范围内被分析物的回收率，或评价估计浓度与实际浓度之间关系的线性。具统计学意义的标准有二，一是斜率的置信区间被限定在约1.0的区间，二是此斜率接近1.0。在任意一种情况下，此区间或接近程度的定义应该在验证方案中明确规定。接受标准将取决于含量和其差异性，以及取决于该产品。通过测试证明该斜率为1.0的零假设没有统计学意义，这样的方法不能用于设定接受标准。PRECISION精密度Definition- The precision of an analytical procedure is the degree of agreement among individual test results when the procedure is applied separately to multiple samplings of a homogeneous sample. The precision of an analytical procedure is usually expressed as the standard deviation or relative standard deviation (coefficient of variation) of a series of measurements. Precision may be a measure of either the degree of reproducibility or of repeatability of the analytical procedure under normal operating conditions. In this context, reproducibility refers to the use of the analytical procedure in different laboratories, as in a collaborative study. Intermediate precision (as known as ruggedness) express within-laboratory variation, as on different days, or with different analysts or equipment within the same laboratory. Repeatability refers to the use of the analytical procedure within a laboratory over a short period of time using the same analyst with the same equipment. 定义- 分析规程的精密度是当该分析规程单独分析均质样品的多个样本时，若干检验结果的一致程度。分析规程的精密度通常以一系列测量数值的标准差或相对标准差（变异系数）来表示。精密度可以是分析规程在普通操作条件下可重现性或可重复性程度的度量单位。在这样的背景下，重现性指的是该分析规程在不同实验室的应用，例如在一个协作实验室里进行研究。中间精密度（也称为“耐用性”）体现了在实验室内的差异，如在相同的实验室，但在不同的日期，或使用不同的分析员或设备。可重复性指的是在同一个实验室内，一段较短的时间内，相同的分析员使用相同的设备、同一个分析方法的应用情况。Determination- The precision of an analytical procedure is determined by assaying a sufficient number of aliquots of a homogeneous sample to be able to calculate statistically valid estimates of standard deviation or relative standard deviation (coefficient of variation). Assays in this context are independent analyses of samples that have been carried through the complete analytical procedure from sample preparation to final test result. 测定-一个分析方法的精密度是通过对足够的同一样品的分析来计算有统计学意义的标准偏差或相对标准偏差来测定的。在这个背景下的分析是从样品准备到最终实验结果的完整分析程序的对样品的独立分析。The ICH documents recommended that repeatability should be assessed using a minimum of nine determinations covering the specified range for the procedure (i.e., three concentration and three replicates of each concentration or using a minimum of six determinations at 100% of the test concentration).ICH文件建议可重复性的评估应该使用最少九次检测，覆盖该分析规程所规定的范围（例如，三个浓度和每个浓度三次重复进样，或在100%测试浓度上进行最少六次测定）。SPECIFITY专属性Definition- The ICH documents define specificity as the ability to assess unequivocally the analyt in the presence of components that may be expected to be present, such as impurities, degradation products, and matrix components. Lack of specificity of an individual analytical procedure may be compensated by other supporting analytical procedures. NOTE- Other reputable international authorities (IUPAC, AOAC-I) have preferred the term “selectivity”, reserving “specificity” for those procedures that are completely selective. For the tests discussed below, the above definition has the following implications. 定义- ICH文件将专属性定义是当待分析物含有预期会有的其他组分（例如，杂质、降解产物、矩阵组分）时，准确可靠地评估待分析物的能力。某个分析规程缺乏专属性可以通过其他辅助性分析规程进行补偿。【注意：其他声誉卓著的国际权威机构（IUPAC、AOAC-I）已经提出术语“选择性”，而将“专属性”留给已经具有完全选择性的规程。】对于下面讨论的测试，上述定义具有以下的含义。Identification Tests: ensure the identity of the analyte.鉴别检测：确保待分析物的鉴别。Purity Tests: ensure that all the analytical procedures performed allow an accurate statement of the content of impurities of an analyte (e.g., related substances test, heavy metals limit, organic volatile impurities). 纯度检测：确保执行的所有分析规程能够令对于待分析物各杂质含量的准确陈述得以做出。Assays: provide an exact result, which allow an accurate statement on the content or potency of the analyte in a sample. 含量检测：提供准确的结果，令对样品中待分析物的含量或效力的准确陈述得以做出。Determination- In the case of qualitative analyses (identification tests), the ability to select between compounds of closely related structure that are likely to be present should be demonstrated. This should be confirmed by obtaining positive results (perhaps by comparison to a known reference material) from samples containing the analyte, coupled with negative results from samples that do not contain the analyte and by confirming that a positive response is not obtained from materials structurally similar to or closely related to the analyte. 测定- 对于定性分析（鉴别检验），应当论证其在可能存在的、结构密切相关的物质中进行选择的能力。从含有待分析物的样品中得到阳性结果（可能通过与已知标准物质的比较），而从不含待分析物的样品得到阴性结果，以对其选择能力加以确认，并还要确认阳性响应不是来自与待分析物结构相似或密切相关的物质。In the case of analytical procedures for impurities, specificity may be established by spiking the drug substance or product with appropriate levels of impurities and demonstrating that these impurities are determined with appropriate accuracy and precision. 对于检测杂质的分析规程，专属性可以通过以适当水平的杂质将原料药或成药增敏，并论证这些杂质的测定达到了适当的准确度和精密度。In the case of the assay, demonstration of specificity requires that it can be shown that the procedure is unaffected by the presence of impurities or excipients. In practice, this can be done by spiking the drug substance or product with appropriate levels of impurities or excipients and demonstrating that the assay result is unaffected by the presence of these extraneous materials. 对于含量检测，对专属性的论证要求能够显示出该分析规程不受各杂质或辅料的影响。在实际操作中，通过以适当水平的杂质或辅料将原料药或成药增敏，并证明含量检验结果不受这些外来物质的影响，来完成论证。If 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., Pharmacopeial or other validated procedure). These comparisons should include samples stored under relevant stress conditions (e.g., light, heat, humidity, acid/base hydrolysis, oxidation). In the case of assay, the results should be compared; in the case of chromatographic impurity test, the impurity profiles should be compared. 如果没有杂质或降解产物的标准品，可以通过将含有杂质或降解产物的样品的测试结果与第二种、成熟规程（例如，药典或其他验证过的规程）的结果进行比较，来论证专属性。这些比较应该包括在相关破坏性条件下（例如，光、热、湿度、酸/碱水解、氧化作用）存储的样品。对于含量测定，应比较其结果；对于色谱法杂质检测，应比较杂质概况。The ICH documents state that when chromatographic procedures are used, representative chromatograms should be presented to demonstrate the degree of selectivity, and peaks should be appropriately labeled. Peak purity tests (e.g., using diode array or mass spectrometry) may be useful to show that the analyte chromatographic peak is not attributable to more than one component. ICH文件声明，当使用色谱分析规程时，应提交具代表性的色谱图，以论证选择性的程度，而且应对色谱峰作适当的标识。也可以使用色谱峰纯度测试（例如，使用二极管阵列或质谱仪），来显示待分析物的色谱峰仅产生于一个组分。DETECTION LIMIT检测限度Definition- The detection limit is a characteristic of limit tests. It is the lowest amount of analyte in a sample that can be detected, but not necessarily quantitated, under the stated experimental conditions. Thus, limit tests merely substantiate that the amount of analyte is above or below a certain level. The detection limit is usually expressed as the concentration of analyte (e.g., percentage, parts per billion) in the sample. 定义- 检测限度是限度检测的特性。它是指在规定的试验条件下，样品中可被检测到的待分析物的最小数量，但是无需定量。因此，限度检测仅仅说明了待分析物的数量高于或低于某个特定水平。检测限度通常以在样品中的待分析物浓度（例如，百分比、十亿分率）表示。Determination- For non-instrumental procedures, the detection limit is generally 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. 测定- 对于非仪器分析规程，检测限度的测定方法通常为，对含有已知浓度待分析物的样品进行分析，并确立能够可靠地被检测出来的待分析物的最低水平。For instrumental procedures, the same approach may be used as for non-instrumental procedures. In the case of procedures submitted for consideration as official compendial procedures, it is almost never necessary to determine the actual detection limit. Rather, the detection limit is shown to be sufficiently low by the analysis of samples with known concentrations of analyte above and below the required detection level. For example, if it is required to detect an impurity at the level of 0.1%, it should be demonstrated that the procedure will reliably detect the impurity at that level.对于仪器分析规程，可以使用与非仪器分析规程相同的方法。对于提交用于官方药典规程的备选方法，其几乎从来不需要确定实际的检测限度。而是，通过分析含有高于和低于必需的检测水平的、已知待分析物浓度的样品，以显示检测限度足够低。例如，如果必需检测浓度在0.1%的杂质，则应当证明该分析规程将可靠地检测在这个水平的杂质。In the case of instrumental analytical procedures that exhibit background noise, the ICH documents describe a common approach, whic