ICH分析方法驗證指南--方法論

1、ICH Q2 (R1)INTRODUCTION 簡介This document is complementary to the parent document, which presents a discussion of the characteristics that should be considered during the validation of analytical procedures. Its purpose is to provide some guidance and recommendations on how to consider the various val

2、idation characteristics for each analytical procedure. In some cases (for example, demonstration of specificity), the overall capabilities of a number of analytical procedures in combination may be investigated in order to ensure the quality of the drug substance or drug product. In addition, the do

3、cument provides an indication of the data, which should be presented in a registration application. 本文作為前文的補(bǔ)充，旨在討論在分析方法驗證過程中在每一個具體的項目需要考慮哪些內(nèi)容。本文的目的是就不同類型的驗證該涵蓋哪些項目提供一個指導(dǎo)原則和建議。以專屬性為例，為了確保原料藥或制劑的質(zhì)量，需要考察分析過程對化合物中雜質(zhì)的全面綜合分析能力。另外，文件提供的數(shù)據(jù)應(yīng)該包含在注冊申請材料中。All relevant data collected during validation and formul

4、ae used for calculating validation characteristics should be submitted and discussed as appropriate. 驗證報告中的所有數(shù)據(jù)及每個驗證項目的計算公式應(yīng)一同提交并進(jìn)行適當(dāng)?shù)恼撌?即得出相應(yīng)結(jié)論)。Approaches other than those set forth in this guideline may be applicable and acceptable. It is the responsibility of the applicant to choose the validati

5、on procedure and protocol most suitable for their product. However it is important to remember that the main objective of validation of an analytical procedure is to demonstrate that the procedure is suitable for its intended purpose. Due to their complex nature, analytical procedures for biological

6、 and biotechnological products in some cases may be approached differently than in this document.本文闡述的原則之外的原則應(yīng)該適用并可接受。申請者的職責(zé)是制定最適合申報產(chǎn)品的驗證項目及驗證方案。最重要的是記住分析方法驗證的主要目的是證明該分析程序能達(dá)到預(yù)期目標(biāo)。由于生物制品和生物科技產(chǎn)品本身的復(fù)雜性，有時候，其分析方法的驗證也可能與本文提到的方法不同。Well-characterized reference materials, with documented purity, should be u

7、sed throughout the validation study. The degree of purity necessary depends on the intended use. 驗證研究用到的參比物質(zhì)需要經(jīng)過完全鑒定并標(biāo)定純度。所需的純度取決于其應(yīng)用類型(預(yù)期用途)。In accordance with the parent document, and for the sake of clarity, this document considers the various validation characteristics in distinct sections. The

8、arrangement of these sections reflects the process by which an analytical procedure may be developed and evaluated.與前文相同，該文件考慮了各個獨(dú)立章節(jié)的不同驗證項目。這些章節(jié)的安排也反映了分析方法的建立和評估的過程。In practice, it is usually possible to design the experimental work such that the appropriate validation characteristics can be consid

9、ered simultaneously to provide a sound, overall knowledge of the capabilities of the analytical procedure, for instance: specificity, linearity, range, accuracy and precision. 事實(shí)上，設(shè)計試驗的時候，一些適當(dāng)?shù)尿炞C項目可以同時考慮，以便對分析方法的能力提供合理的，全面的了解(依據(jù))，例如：專屬性、線性、范圍、準(zhǔn)確度、精密度。provide aknowledge of 為提供依據(jù)1. SPECIFICITY 專屬性An i

10、nvestigation of specificity should be conducted during the validation of identification tests, the determination of impurities and the assay. The procedures used to demonstrate specificity will depend on the intended objective of the analytical procedure.鑒別測試、雜質(zhì)和含量測試方法的驗證都應(yīng)考察方法的專屬性。證明專屬性的方法取決于分析方法的預(yù)

11、期目的。It is not always possible to demonstrate that an analytical procedure is specific for a particular analyte (complete discrimination). In this case a combination of two or more analytical procedures is recommended to achieve the necessary level of discrimination.一般來說，某一種分析方法不太可能完全證明其對某一特定被分析物具有專屬

12、性。這種情況下，建議采用兩種或兩種以上的分析方法以確保完全鑒別水平。1.1. Identification 鑒別Suitable 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 compariso

13、n 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

14、response is not obtained. The choice of such potentially interfering materials should be based on sound scientific judgment with a consideration of the interferences that could occur.合適的鑒別方法應(yīng)該能夠區(qū)分可能存在的結(jié)構(gòu)相近的化合物?？梢酝阎獏⒖嘉镔|(zhì)進(jìn)行比較，從含有被分析物的樣品得到的正的結(jié)果和不含被分析物的樣品得到的負(fù)的結(jié)果來確定。此外，鑒別測試也可以用結(jié)構(gòu)相近或相關(guān)的物質(zhì)測試得不到正的反應(yīng)來證實(shí)。在考慮可

15、能會造城干擾的前提下，應(yīng)根據(jù)合理科學(xué)的判斷來選擇可能存在的干擾物。likely to be present：可能存在的；discrimination：比較；1.2. Assay and Impurity Test(s) 含量和雜質(zhì)測定For chromatographic procedures, representative chromatograms should be used to demonstrate specificity and individual components should be appropriately labelled. Similar consideratio

16、ns should be given to other separation techniques.在色譜法測定中，需要用有代表性的圖譜證明專屬性，并恰當(dāng)?shù)淖⒚髅恳粋€成分。其它的分離技術(shù)也應(yīng)如此。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 clos

17、est to each other.色譜分離法應(yīng)在一定程度上考察關(guān)鍵性的分離。對關(guān)鍵性的分離，可用兩個洗脫程度最接近的化合物的分離度來證明其專屬性。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 comb

18、ination of the assay and a suitable test for impurities can be used. 當(dāng)采用非專屬性的方法測定含量時，應(yīng)采用輔助性分析方法來證明整個方法具有專屬性。例如用滴定法測定放行原料藥的含量，可結(jié)合使用合適的雜質(zhì)檢測方法。supporting analytical procedures：輔助性分析方法；overall specificity：整體專屬性；combination：結(jié)合，聯(lián)合；The approach is similar for both assay and impurity tests: 下述方法均適用于含量和雜質(zhì)檢測。

19、is similar for：適用于1.2.1 Impurities are available 可以得到雜質(zhì)的情況For 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

20、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). 對含量檢測，專屬性應(yīng)該包括提供被分析物在雜質(zhì)和/或賦形劑存在時能被區(qū)分的證明；實(shí)際操作時，可通過向純物質(zhì)(原料藥或制劑)中加入一定量的雜質(zhì)和/或賦形劑的檢測結(jié)果和未添加雜質(zhì)和/或賦形劑的純物質(zhì)的檢測結(jié)果進(jìn)行對

21、比以此證明這些雜質(zhì)和/或賦形劑的存在不會對含量檢測結(jié)果造成影響。discrimination：區(qū)分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 m

22、atrix. 對雜質(zhì)檢測，可以向原料藥或制劑中加入一定量的雜質(zhì)，證明各雜質(zhì)能夠分離且能與樣品中的其它組分分離。1.2.2 Impurities are not available 無法得到雜質(zhì)的情況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-cha

23、racterized 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. 如果無法得到雜質(zhì)或降解產(chǎn)物的對照品，檢測方法的專屬性可以通過將含有一定量的雜質(zhì)或降解

24、產(chǎn)物的樣品的檢測結(jié)果與另一種成熟的檢測方法如藥典方法或經(jīng)驗證的其它方法(獨(dú)立的方法)的檢測結(jié)果進(jìn)行比較來證明。必要時，應(yīng)該包括放置在強(qiáng)降解試驗條件，即強(qiáng)光，高溫，高濕，酸/堿水解及氧化條件下的樣品測試。As appropriate：必要時；- For the assay, the two results should be compared; - 對含量檢測，需要對比兩種方法的檢測結(jié)果- For the impurity tests, the impurity profiles should be compared. - 對雜質(zhì)檢測，需要對比雜質(zhì)概況Peak purity tests may

25、be useful to show that the analyte chromatographic peak is not attributable to more than one component (e.g., diode array, mass spectrometry).峰純度測試是非常有用的，它能顯示被測物的色譜峰是一個成分還是多個成分(如二極管陣列，質(zhì)譜)。2. LINEARITY 線性A linear relationship should be evaluated across the range (see section 3) of the analytical proc

26、edure. 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. 檢測方法的線性關(guān)系應(yīng)該在范圍內(nèi)(見章

27、節(jié)3)進(jìn)行評價。線性研究可通過所建議的分析方法，直接對原料藥(用標(biāo)準(zhǔn)儲備液稀釋)和/或分別稱取制劑組分的混合物測試來進(jìn)行。后者應(yīng)在方法的范圍內(nèi)進(jìn)行研究。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 method

28、s, 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 its

29、elf may be helpful to provide mathematical estimates of the degree of linearity. 線性應(yīng)關(guān)系應(yīng)以信號對被測物濃度或含量作圖，根據(jù)圖形是否呈線性來評估。如果呈線性關(guān)系，測試結(jié)果應(yīng)用適當(dāng)?shù)慕y(tǒng)計學(xué)方法進(jìn)行評估，例如用最小二乘法進(jìn)行線性回歸計算。在某些情況下，為了使含量與樣品濃度呈線性關(guān)系，在回歸分析前需要對測試數(shù)據(jù)進(jìn)行數(shù)學(xué)轉(zhuǎn)化。由線性回歸評估所得的數(shù)據(jù)本身又有助于精確的評價線性的程度。In some cases：在某些情況下，有時候；The correlation coefficient, y-intercept, sl

30、ope 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. 相關(guān)系數(shù)，y軸上的截距，回歸曲線的斜率以及剩余方差應(yīng)包含在遞交材料里。還應(yīng)包括數(shù)據(jù)圖表。另外

31、，實(shí)際數(shù)據(jù)點(diǎn)與回歸曲線的偏差也有助于對線性進(jìn)行評價。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. 一些分析方法，如免疫測定法，在任何轉(zhuǎn)換后，均不能證明呈

32、線性。在這種情況下，分析的相應(yīng)值應(yīng)用被分析物的濃度(數(shù)量)的適當(dāng)函數(shù)來表示。For the establishment of linearity, a minimum of 5 concentrations is recommended. Other approaches should be justified. 為建立線性，建議至少用5個濃度。若用其他方法應(yīng)證明其合理性。3. RANGE 范圍The specified range is normally derived from linearity studies and depends on the intended applicatio

33、n 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. 特定的范圍一般是從線性研究中得到的，它

34、依賴于分析方法的應(yīng)用目的。確定范圍的方法是：樣品中含有被分析物的量在分析方法規(guī)定的范圍內(nèi)或在范圍末端，該分析方法均能獲得良好的線性，精密度及準(zhǔn)確度。The following minimum specified ranges should be considered: 以下是應(yīng)考慮的最小規(guī)定范圍：- for the assay of a drug substance or a finished (drug) product: normally from 80 to 120 percent of the test concentration; - 對原料藥或成品藥(制劑)的含量測定：一般應(yīng)在測試

35、濃度的80%120%；- 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; - 對含量均勻度檢測：應(yīng)至少在測試濃度的70%130%之內(nèi)，超出此范圍，應(yīng)有正當(dāng)理由，主要是根據(jù)劑型的特點(diǎn)(如定量吸入劑)；- for disso

36、lution testing: +/-20 % over the specified range; - 對溶出度測試，應(yīng)為規(guī)定范圍的+/-20%； 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. 例如：如果是控釋劑，規(guī)定1小時后達(dá)到20%，24小時后達(dá)到90%，它的驗證范圍應(yīng)為標(biāo)

37、示量的0110%。- for the determination of an impurity: from the reporting level of an impurity1 to 120% of the specification; - 對雜質(zhì)測定，應(yīng)為雜質(zhì)的報告水平至標(biāo)準(zhǔn)規(guī)定的120%；- for impurities known to be unusually potent or to produce toxic or unexpected pharmacological effects, the detection/quantitation limit should be comm

38、ensurate with the level at which the impurities must be controlled; - 對已知有異常功效的，有毒的或者有意外藥理作用的雜質(zhì)，其檢測限度和定量限度應(yīng)與該雜質(zhì)必須被控制的水平相當(dāng)。Note: for validation of impurity test procedures carried out during development, it may be necessary to consider the range around a suggested (probable) limit. 注意：在研制階段進(jìn)行雜質(zhì)檢測方法驗證

39、時，有必要根據(jù)建議(可能)的限度水平來考慮范圍；- 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. 1 see chapters “Reporting Impurity Content of Batches” of the correspond

40、ing ICH-Guidelines: “Impurities in New Drug Substances” and “Impurities in New Drug Products” - 如果一個試驗同時進(jìn)行含量和純度檢測，且僅使用100%的標(biāo)準(zhǔn)品，線性范圍應(yīng)覆蓋雜質(zhì)的報告水平(見相應(yīng)ICH指南“新原料藥中的雜質(zhì)”和“新制劑中的雜質(zhì)”中“批雜質(zhì)含量的報告”章節(jié))至含量指標(biāo)120%。4. ACCURACY 準(zhǔn)確度Accuracy should be established across the specified range of the analytical procedure. 應(yīng)在分析

41、方法規(guī)定的范圍內(nèi)建立(考察方法的)準(zhǔn)確度。4.1. Assay 含量4.1.1 Drug Substance 原料藥Several methods of determining accuracy are available: 以下幾種方法可用于測定準(zhǔn)確度：a) application of an analytical procedure to an analyte of known purity (e.g. reference material); 用該分析方法測定已知純度的被分析物(例如參照物質(zhì))；b) comparison of the results of the proposed an

42、alytical procedure with those of a second well-characterized procedure, the accuracy of which is stated and/or defined (independent procedure, see 1.2.);用建議采用的分析方法的結(jié)果與另一種完全驗證過的方法的結(jié)果作對比，對比的方法的準(zhǔn)確度是規(guī)定的(一定的)和/或已定義的(獨(dú)立的方法，見1.2節(jié))c) accuracy may be inferred once precision, linearity and specificity have be

43、en established. 準(zhǔn)確度可以在精密度，線性及專屬性建立之后推論得到；4.1.2 Drug Product 制劑Several methods for determining accuracy are available: 以下幾種方法可用于測定準(zhǔn)確度：a) application of the analytical procedure to synthetic mixtures of the drug product components to which known quantities of the drug substance to be analysed have bee

44、n added; 用該分析方法測定按處方量制成的混合物，其中加入了已知量的待測原料藥。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

45、accuracy of which is stated and/or defined (independent procedure, see 1.2.); 如果不能得到制劑的所有成分，向制劑中加入已知量的被測物或者與另一種經(jīng)過完整驗證過的準(zhǔn)確度是規(guī)定的(一定的)和/或已定義的方法的結(jié)果作對比，(獨(dú)立的方法，見1.2節(jié))也是可以接受的；c) accuracy may be inferred once precision, linearity and specificity have been established. 準(zhǔn)確度可以在精密度，線性及專屬性建立之后推論得到；4.2. Impuritie

46、s (Quantitation) 雜質(zhì)(定量)Accuracy should be assessed on samples (drug substance/drug product) spiked with known amounts of impurities. 準(zhǔn)確度可以通過向樣品(原料藥/制劑)中加入已知量雜質(zhì)的方法來評價。In cases where it is impossible to obtain samples of certain impurities and/or degradation products, it is considered acceptable to co

47、mpare results obtained by an independent procedure (see 1.2.). The response factor of the drug substance can be used. 如果無法得到雜質(zhì)和或降解產(chǎn)物的樣品，可以通過與其它獨(dú)立方法(見1.2節(jié))的檢測結(jié)果進(jìn)行對比評估其準(zhǔn)確度。可以使用原料藥的響應(yīng)因子。It should be clear how the individual or total impurities are to be determined e.g., weight/weight or area percent, i

48、n all cases with respect to the major analyte. 需要說明單雜和總雜是如何測定的，如相對于主要被分析物所占的質(zhì)量分?jǐn)?shù)或面積百分比。4.3. Recommended Data 可接受數(shù)據(jù)(數(shù)據(jù)要求)Accuracy 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 t

49、he total analytical procedure). 準(zhǔn)確度的評價需要在方法的線性范圍內(nèi)的三種濃度至少測定九次(按完整分析步驟對三種濃度每種濃度重復(fù)進(jìn)樣三次)。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. 準(zhǔn)

50、確度應(yīng)以向樣品中加入已知量的被測物所得的百分回收率或者平均值和可接受真實(shí)值之間的差值及置信區(qū)間來報告。5. PRECISION 精密度Validation of tests for assay and for quantitative determination of impurities includes an investigation of precision. 含量和雜質(zhì)的定量分析需要考察方法的精密度。5.1. Repeatability 重復(fù)性Repeatability should be assessed using: 重復(fù)性可以通過以下方法進(jìn)行考察：a) a minimum of

51、 9 determinations covering the specified range for the procedure (e.g., 3 concentrations/3 replicates each); 在方法的線性范圍內(nèi)至少檢測九次(三種濃度每種濃度重復(fù)進(jìn)樣三次)；b) or a minimum of 6 determinations at 100% of the test concentration. 以100%測試濃度至少檢測六次。5.2. Intermediate Precision 中間精密度The extent (程度) to which intermediate p

52、recision 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 c

53、onsidered necessary to study these effects individually. The use of an experimental design (matrix) is encouraged. 中間精密度的考察程度應(yīng)根據(jù)分析方法的操作環(huán)境而定。申請者應(yīng)確定(弄清楚)隨機(jī)時間對分析方法的精密度的影響。需要研究的典型變化有：日期，分析者，儀器等。沒有必要逐項考察這些因素。建議使用試驗設(shè)計(矩陣法)。5.3. Reproducibility 重現(xiàn)性Reproducibility is assessed by means of (通過) an inter-labor

54、atory 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. 重現(xiàn)性可通過實(shí)驗室之間的試驗進(jìn)行評估。如果方法需要標(biāo)準(zhǔn)化，例如藥典方法，則應(yīng)考慮重現(xiàn)性。這些資料不是上市申請文檔的一部分(申報注冊不

55、需要考察藥典收錄方法的重現(xiàn)性，這是藥典委需要考慮的問題)。5.4. Recommended Data 數(shù)據(jù)要求The standard deviation, relative standard deviation (coefficient of variation) and confidence interval should be reported for each type of precision investigated. 每一種精密度研究中都應(yīng)報告標(biāo)準(zhǔn)偏差，相對標(biāo)準(zhǔn)偏差(變異系數(shù))和置信區(qū)間。6. DETECTION LIMIT 檢測限Several approaches for d

56、etermining 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. 根據(jù)檢測方法是用儀器分析還是非儀器分析，可用幾種方法來確定檢測限。除了下面所列的方法外，其它的分析方法也可能被接收。6.1. Based on Visual Evaluation 視覺判定(根據(jù)直觀評價)Visual evaluation m

57、ay be used for non-instrumental methods but may also be used with instrumental methods. 視覺判定可用于非儀器分析方法，也可用于儀器分析方法。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. 檢測限

58、的確定是通過一系列已知濃度的分析物樣品進(jìn)行分析，并以能準(zhǔn)確測得被分析物的最小水平來建立。6.2. Based on Signal-to-Noise This 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 cons