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原則和建議。以專屬性為例，為了確保原料藥或制劑的質量，需要考察分析過程對化合物中雜質的全面綜合分析能力。另外，文件提供的數據應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據及每個驗證項目的計算公式應一同提交并進行適當的論述(即得出相應結論)。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該適用并可接受。申請者的職責是制定最適合申報產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到預期目標。由于生物制品和生物科技產品本身的復雜性，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需要經過完全鑒定并標定純度。所需的純度取決于其應用類型(預期用途)。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. 事實上，設計試驗的時候，一些適當的驗證項目可以同時考慮，以便對分析方法的能力提供合理的，全面的了解(依據)，例如：專屬性、線性、范圍、準確度、精密度。provide aknowledge of 為提供依據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、雜質和含量測試方法的驗證都應考察方法的專屬性。證明專屬性的方法取決于分析方法的預

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.合適的鑒別方法應該能夠區(qū)分可能存在的結構相近的化合物?？梢酝阎獏⒖嘉镔|進行比較，從含有被分析物的樣品得到的正的結果和不含被分析物的樣品得到的負的結果來確定。此外，鑒別測試也可以用結構相近或相關的物質測試得不到正的反應來證實。在考慮可

15、能會造城干擾的前提下，應根據合理科學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，并恰當的注明每一個成分。其它的分離技術也應如此。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.色譜分離法應在一定程度上考察關鍵性的分離。對關鍵性的分離，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. 當采用非專屬性的方法測定含量時，應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，可結合使用合適的雜質檢測方法。supporting analytical procedures：輔助性分析方法；overall specificity：整體專屬性；combination：結合，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檢測。

19、is similar for：適用于可以得到雜質的情況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 levels of impurities and/or exc

20、ipients 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，專屬性應該包括提供被分析物在雜質和/或賦形劑存在時能被區(qū)分的證明；實際操作時，可通過向純物質(原料藥或制劑)中加入一定量的雜質和/或賦形劑的檢測結果和未添加雜質和/或賦形劑的純物質的檢測結果進行對比以此證明這些雜質和/或賦形劑的存在不會對含量檢測結果造成影響

21、。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 matrix. 對雜質檢測，可以向原料藥或制劑中加入一定量的雜質

22、，證明各雜質能夠分離且能與樣品中的其它組分分離。無法得到雜質的情況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-characterized procedure e.g.: pharmacopoeial method or other validate

23、d analytical procedure (independent procedure). As appropriate, this should include samples stored under relevant stress conditions: light, heat, humidity, acid/base hydrolysis and oxidation. 如果無法得到雜質或降解產物的對照品，檢測方法的專屬性可以通過將含有一定量的雜質或降解產物的樣品的檢測結果與另一種成熟的檢測方法如藥典方法或經驗證的其它方法(獨立的方法)的檢測結果進行比較來證明。必要時，應該包括放置在

24、強降解試驗條件，即強光，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果- For the impurity tests, the impurity profiles should be compared. - 對雜質檢測，需要對比雜質概況Peak purity tests may be useful to show that the analyte chromatographic peak is not att

25、ributable 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，質譜)。2. LINEARITY 線性A 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 d

26、ilution 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. 檢測方法的線性關系應該在范圍內(見章節(jié)3)進行評價。線性研究可通過所建議的分析方法，直接對原料藥(用標準儲備液稀釋)和/或分別稱取制劑組分的混合物測試來進行。后者應在方

27、法的范圍內進行研究。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 methods, for example, by calculation of a regression line by the method

28、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

29、 of linearity. 線性應關系應以信號對被測物濃度或含量作圖，根據圖形是否呈線性來評估。如果呈線性關系，測試結果應用適當的統(tǒng)計學方法進行評估，例如用最小二乘法進行線性回歸計算。在某些情況下，為了使含量與樣品濃度呈線性關系，在回歸分析前需要對測試數據進行數學轉化。由線性回歸評估所得的數據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, slope of the regression line and residual sum of squares should be s

30、ubmitted. 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. 相關系數，y軸上的截距，回歸曲線的斜率以及剩余方差應包含在遞交材料里。還應包括數據圖表。另外，實際數據點與回歸曲線的偏差也有助于對線性進行評價。Some analytical procedures, such as immu

31、noassays, 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，在任何轉換后，均不能證明呈線性。在這種情況下，分析的相應值應用被分析物的濃度(數量)的適當函數來表示。For the establishment of lin

32、earity, 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。若用其他方法應證明其合理性。3. RANGE 范圍The specified range is normally derived from linearity studies and depends on the intended application of the procedure. It is established by confirming that the analy

33、tical 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，它依賴于分析方法的應用目的。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)定的范圍內或在范圍末端，該分析方法均能獲得良好的線性，

34、精密度及準確度。The following minimum specified ranges should be considered: 以下是應考慮的最小規(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：一般應在測試濃度的80%120%；- for content uniformity, covering a minimum of 70 to 1

35、30 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：應至少在測試濃度的70%130%之內，超出此范圍，應有正當理由，主要是根據劑型的特點(如定量吸入劑)；- for dissolution testing: +/-20 % over the specified range; - 對溶出度測試，應為規(guī)定范圍的

36、+/-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小時后達到20%，24小時后達到90%，它的驗證范圍應為標示量的0110%。- for the determination of an impurity: from the reportin

37、g level of an impurity1 to 120% of the specification; - 對雜質測定，應為雜質的報告水平至標準規(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 commensurate with the level at which the impurities must be controlled

38、; - 對已知有異常功效的，有毒的或者有意外藥理作用的雜質，其檢測限度和定量限度應與該雜質必須被控制的水平相當。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：在研制階段進行雜質檢測方法驗證時，有必要根據建議(可能)的限度水平來考慮范圍；- if assay and purity are performed togeth

39、er 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 corresponding ICH-Guidelines: “Impurities in New Drug Substances” and “Impur

40、ities in New Drug Products” - 如果一個試驗同時進行含量和純度檢測，且僅使用100%的標準品，線性范圍應覆蓋雜質的報告水平(見相應ICH指南“新原料藥中的雜質”和“新制劑中的雜質”中“批雜質含量的報告”章節(jié))至含量指標120%。4. ACCURACY 準確度Accuracy should be established across the specified range of the analytical procedure. 應在分析方法規(guī)定的范圍內建立(考察方法的)準確度。4.1. Assay 含量原料藥Several methods of determinin

41、g accuracy are available: 以下幾種方法可用于測定準確度：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)；b) comparison of the results of the proposed analytical procedure with those of a second well-characterized procedure, the accuracy of

42、 which is stated and/or defined (independent procedure, see 1.2.);用建議采用的分析方法的結果與另一種完全驗證過的方法的結果作對比，對比的方法的準確度是規(guī)定的(一定的)和/或已定義的(獨立的方法，見1.2節(jié))c) accuracy may be inferred once precision, linearity and specificity have been established. 準確度可以在精密度，線性及專屬性建立之后推論得到；制劑Several methods for determining accuracy are

43、 available: 以下幾種方法可用于測定準確度：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 been 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 dru

44、g 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.); 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

45、者與另一種經過完整驗證過的準確度是規(guī)定的(一定的)和/或已定義的方法的結果作對比，(獨立的方法，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. 準確度可以在精密度，線性及專屬性建立之后推論得到；4.2. Impurities (Quantitation) 雜質(定量)Accuracy should be assessed on samples (drug substance/drug product) spiked with kn

46、own amounts of impurities. 準確度可以通過向樣品(原料藥/制劑)中加入已知量雜質的方法來評價。In cases where it is impossible to obtain samples of certain impurities and/or degradation products, it is considered acceptable to compare results obtained by an independent procedure (see 1.2.). The response factor of the drug substance c

47、an be used. 如果無法得到雜質和或降解產物的樣品，可以通過與其它獨立方法(見1.2節(jié))的檢測結果進行對比評估其準確度?？梢允褂迷纤幍捻憫蜃?。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. 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量分數或面積百分比。4.3. Recommended Da

48、ta 可接受數據(數據要求)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 the total analytical procedure). 準確度的評價需要在方法的線性范圍內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進樣三次)。Accuracy should be re

49、ported 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. 準確度應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 q

50、uantitative determination of impurities includes an investigation of precision. 含量和雜質的定量分析需要考察方法的精密度。5.1. Repeatability 重復性Repeatability should be assessed using: 重復性可以通過一下方法進行考察：a) a minimum of 9 determinations covering the specified range for the procedure (e.g., 3 concentrations/3 replicates each

51、); 在方法的線性范圍內至少檢測九次(三種濃度每種濃度重復進樣三次)；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 precision should be established depends on the circumstances under which the procedure is intended to be us

52、ed. 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 enc

53、ouraged. 中間精密度的考察程度應根據分析方法的操作環(huán)境而定。申請者應確定(弄清楚)隨機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。建議使用試驗設計(矩陣法)。5.3. Reproducibility 重現(xiàn)性Reproducibility is assessed by means of (通過) an inter-laboratory trial. Reproducibility should be considered in case of the standardization of an analytical procedur

54、e, for instance, for inclusion of procedures in pharmacopoeias. These data are not part of the marketing authorization dossier. 重現(xiàn)性可通過實驗室之間的試驗進行評估。如果方法需要標準化，例如藥典方法，則應考慮重現(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 數據要求The standard deviation, relative standard deviation

55、 (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報告標準偏差，相對標準偏差(變異系數)和置信區(qū)間。6. DETECTION LIMIT 檢測限Several approaches for determining the detection limit are possible, depending on whether the procedure is a non-instrumental or i

56、nstrumental. Approaches other than those listed below may be acceptable. 根據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(根據直觀評價)Visual evaluation may 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

57、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. 檢測限的確定是通過一系列已知濃度的分析物樣品進行分析，并以能準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 applie

58、d 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 Res