1

Verification of Verification of CompendialCompendial MethodsMethods

ภญภญ..ดรดร..สุภาณีสุภาณี ดวงธีรปรีชาดวงธีรปรีชาสํานักยาและวัตถเุสพติดสํานักยาและวัตถเุสพติด กรมวิทยาศาสตรการแพทยกรมวิทยาศาสตรการแพทย

7 7 มิถุนายนมิถุนายน 25525566

2

Scopes :Scopes :OverviewOverview- Asean Analytical Validation Guidelines

Verification of Verification of CompendialCompendial MethodsMethods- EDQM- AOAC- USP- ACTD- BDN ?

3

From ASEAN Analytical Validation GuidelinesFrom ASEAN Analytical Validation Guidelines

+-+-Linearity

+-+-Repeatability

+-+-Interm.Precision

AssayTesting for ImpuritiesIdentificationType of Analytical Procedure

+-+-Range

--+-LOQ

-+--LOD

++++Specificity

Precision

+-+-Accuracy

-dissolution-content/potency

Limit testQuantitativePerformance Characteristic

4

Specificity:Specificity:

The ability to measure accurately and specificity the analyte in the presence of components that may be expected to be present in the matrix

The degree of interference :- Active ingredients- Excipients- Impurities- Degradation Products- etc

5

Specificity:Specificity:

Identification TestsIdentification Tests- Demonstrate ability to select between compounds of

closely related structure- confirm positive and negative results

AssayAssay- Demonstrate that the results are unaffected by spiked

impurities or excipients

ImpuritiesImpurities- Spike the drug product/substance with impurities and demonstrate appropriate accuracy and precision

6

Specificity:Specificity:

Impurities are availableImpurities are available- Demonstrate that the assay is unaffected by the present of spiked material (impurities and/or excipients)

Impurities are not availableImpurities are not available- Compare test results to a second well-characterized procedure

- For assay : compare the two result- For Impurity Tests : compare purity profiles

-- Peak Purity Test (e.g. PDA, MS)

7

Linearity:Linearity:The ability of the method to elicit test results that

are directly proportational to concentration within a

given range

Range:Range:Interval between upper and lower levels of analyte

demonstrated by the method

Precision and accuracy expressed in the same units as the test results

8

Evaluation of LinearityEvaluation of Linearity- Established across the range of the method

- Dilutions- Separate weighings

- Evaluate by appropriate statistical methods (e.g.regression)

- include correlation coefficient, y-intercept, slope, plot

- Minimum 5 concentrations

9

Determination of appropriate rangeDetermination of appropriate range

Minimum specified range :Minimum specified range :- assay : 80-120%

- Impurity test : from QL to 120% spec

- Content unformity : 70-130% test conc

- Dissolution testing : +/- 20% over specified range

10

Precision:Precision:

The measure of the degree of agreement among test results when the method is applied repeatedly to multiple samplings of a homogeneous sample

Expressed as %RSD for a statiscally significant number of samples

11

Precision:Precision:should be performed at 3 levels

- Repeatability - Intermediate Precision- Reproducibility

12

Precision:Precision:RepeatabilityRepeatability

- Same operating conditions, short time interval - Min of 9 determinations covering specified range of procedure (3 levels, 3 reps each)

- Min of 6 determinations at 100% test conc

Intermediate PrecisionIntermediate Precision- Within-lab variations- Different days, analysts, equipment

ReproducibilityReproducibility- Precision between labs- Collaborative studies

13

Accuracy:Accuracy:

The closeness of test results to the true value obtained by the method

- established across the range

14

Accuracy:Accuracy:

Drug SubstanceDrug Substance- Analysis of reference material- Compare results to a second, well-characterized method

Drug ProductDrug Product- Analysis of synthetic mixtures spiked with known quantities of components

- Compare results to a second, well-characterized method

Determined concurrently with precision, linearlity and specificity

15

Accuracy:Accuracy:

Impurities (Impurities (QuantitationQuantitation))- Analysis of sample (Drug substances/Drug product)

spiked with known amounts of impurities

Recommended DataRecommended Data- Min of 9 determinations over a min of 3 conc levels

covering the specified range (e.g. 3 conc/3 replicates each)

- Report as % recovery of known, added amount, or difference between the mean and true value, with confidence intervals

16

Detection Limit (LOD):Detection Limit (LOD):

Lowest concentration of analyte in a sample that can be detected (not necessarily quantitated)

Limit test : above or below a certain level

Expressed as concentration (%, ppb)

Almost necessary not determine actual detection limit

17

Detection Limit (LOD):Detection Limit (LOD):

Visual (Non-instrumental methods)

Signal to noise ratio (3:1 or 2:1 generally accepted)

Detection limit may be based on the standard

deviation of the response and slope:

DL = (3.3)STD/SDL = (3.3)STD/S

STD = standard deviation of the response

S = slope of the calibration curve

18

QuantitationQuantitation Limit (LOQ):Limit (LOQ):

Lowest concentration of analyte in a sample that can be determined with acceptable precision andaccuracy under stated operational conditions

Expressed as concentration of analyte

- accuracy

- precision

19

QuantitationQuantitation Limit (LOQ):Limit (LOQ):

Visual (Non-instrumental methods)

Signal to noise ratio (10:1 is typical)

Quantitation limit may be based on the standard

deviation of the response and slope:

QL = (10)STD/SQL = (10)STD/S

STD = standard deviation of the response

S = slope of the calibration curve

Both DL and QL are validated by analyzing a suitable

number of samples. Method should be documented.

20

Verification of Verification of CompendialCompendial MethodsMethods

EDQMEDQM

AOACAOAC

USPUSP

ACTDACTD

21

From EDQM From EDQM ::1. Transfer of a Method

A verification of suitability under conditions of use (=method transfer check) has to be done

1.1 Pharmacopoeial (compendial) method1.1.1 Active substance1.1.1 Active substance

- Identification:Identification:no formal validation required

- Testing for Impurities:Testing for Impurities:no formal validation required

- Assay:no formal validation required

22

From EDQM From EDQM ::1.1 Pharmacopoeial (compendial) method1.1.2 Medicinal product1.1.2 Medicinal product

- Identification:Identification:- no formal validation required

- Testing for Impurities:Testing for Impurities:- specificity: no interference from excipients- reporting threshold (at least the quant. limit)

- Assay:Assay:- specificity- accuracy: mainly recovery, min 1 det.- precision (repeatability): around the target test

conc. (min 2 independent det.)- linearity at three measuring points in the range

around the target value.

23

From AOAC From AOAC ::

How to meet ISO 17025 requirements for Method Verification

Activities needed for method verification are a subset of those for validation.

24

From AOAC From AOAC ::

Performance Characteristics Included in a Validation

NoNoYesNoYesNoLinearity/Range

NoNoYesNoYesNoRuggedness

NoNoYes/NoNoYesNoLOQ

NoNoYes/NoYesYesNoLOD

YesYesYesYesYesYesSpecificity

NoYesYesNoYesNoPrecision

NoYesYesNoYesNoAccuracy

Quanlitative 6Analyte at high concLimit test 5

Analyte at high concquantitative 4

Analyte at low concLimit test 3

Analyte at low concquantitative 2

Identifica-tion 1

Performance Characteristic

25

From AOAC From AOAC ::How to meet ISO 17025 requirements for Method Verification

Category1 Category1 ::Confirmation of identity – A method that

ensures a material is what it purports to be or confirms the detection of the target analyte

26

The activity need only deal with the unique aspect’s of the lab’s samples or instrumentation

Yes – if differences between instruments could affect specificity

Same as those required for validation

Yes – if the lab’s sample differ from those in the standard method

NANo – If the lab’s samples are identical to those in the standard method and if and differences in instrumentation do not impact specificity

Specificity

Verification ActivitiesVerificationPerformance Characteristic

Category1 Category1 : : IdentificationIdentification

27

From AOAC From AOAC ::How to meet ISO 17025 requirements for Method Verification

Category2 Category2 ::Analyte at Low Concentration, Quantitative

28

If the concentration range for which the method is validated is narrow (<1 order of magnitude), analyze one reference material/standard/spike at one concentration level. Otherwise, demonstrate accuracy at each concentration level (low, middle, high) by anlyzing one reference material/standard/spike at each level.

Yes 1. Accuracy

Verification ActivitiesVerificationPerformance Characteristic

Category2:Analyte at Low Concentration, Quantitative

29

Perform the repeatability test once. If the method covers a concentration range>1 order of magnitude, then the repeatability test must include low, middle and high concentrations.

Yes 2. Precision

Verification ActivitiesVerificationPerformance Characteristic

Category2:Analyte at Low Concentration, Quantitative

30

No/Yes3. Specificity

Run a sample close to LODYes4. LOD

Run a sample close to LOQYes 5. LOQ

Verification ActivitiesVerificationPerformance Characteristic

Category2:Analyte at Low Concentration, Quantitative

31

From AOAC From AOAC ::How to meet ISO 17025 requirements for Method Verification

Category3 Category3 ::Analyte is present above or below a

specified, low concentration (Limit Test)

32

No/Yes1. Specificity

Run a sample close to LODYes2. LOD

Run a sample close to LOQYes 3. LOQ

Verification ActivitiesVerificationPerformance Characteristic

Category3Category3 :: Limit TestLimit Test

33

From AOAC From AOAC ::How to meet ISO 17025 requirements for Method Verification

Category4 Category4 ::Quantifying an analyte at high concentration

34

If the concentration range for which the method is validated is narrow (<1 order of magnitude), analyze one reference material/standard/ spike at one concentration level. Otherwise, demonstrate accuracy at each concentration level (low, middle, high) by anlyzing one reference material/standard/spike at each level.

Yes 1. Accuracy

Verification ActivitiesVerificationPerformance Characteristic

Category4 : Assay

35

Perform the repeatability test once. If the method covers a concentration range>1 order of magnitude, then the repeatability test must include low, middle and high concentrations.

Yes 2. Precision

No/Yes3. Specificity

Verification ActivitiesVerificationPerformance Characteristic

Category4 : Assay

36

From USP From USP ::

Verification of Verification of CompendialCompendialProcedures <1226>Procedures <1226>

37

USP: Validation of USP: Validation of CompendialCompendial Procedures <1225>Procedures <1225>

-

-

-

-

-

+-

Category III(dissolution)

--++Linearity

--++Precision

Category IV(Identification)

Category II(Impurities)Category I

(Assay)

--++Range

--+-LOQ

-+--LOD

++++Specificity

--++Accuracy

Limit testQuantitative

Performance Characteristic

38

Verification of Verification of CompendialCompendial Procedures<1226>Procedures<1226>

Verification of Verification of CompendialCompendial Procedures. The purpose Procedures. The purpose of this new general information chapter is to provide of this new general information chapter is to provide guidelines for guidelines for verifying the suitability of a verifying the suitability of a compendialcompendial procedure under actual conditions of procedure under actual conditions of useuse, as stated in 21 CFR 211.194(a)(2). Because it is not , as stated in 21 CFR 211.194(a)(2). Because it is not necessary for users of necessary for users of compendialcompendial procedures to procedures to perform a complete validation, but rather, to confirm that perform a complete validation, but rather, to confirm that the procedure works for their the procedure works for their drug substancedrug substance, , excipientexcipient, , or or dosage formdosage form, verification can be accomplished by , verification can be accomplished by assessing a subset of validation characteristics. This assessing a subset of validation characteristics. This new chapter is considered an extension of the general new chapter is considered an extension of the general information chapterinformation chapter Validation of Validation of CompendialCompendial Methods Methods <1225>, and both chapters use similar terminology. <1225>, and both chapters use similar terminology. Interested parties are encouraged to submit comments Interested parties are encouraged to submit comments by July 15, 2005. by July 15, 2005.

39

Verification of Verification of CompendialCompendial Procedures<1226>Procedures<1226>

““verifying the suitability of a verifying the suitability of a compendialcompendial procedure under procedure under

actual conditions of useactual conditions of use””

But No GuidelinesBut No Guidelines

40

Data Elements for Verification of Data Elements for Verification of Drug SubstancesDrug Substances and and ExcipientsExcipients**

Technique Category (assay)

Category II(impurities)

Category III(dissolution)

Category IV(identification)

Quantitative Limit test

HPLC/GC PrecisionPrecision

SpecificityLOQ

SpecificityLOD - Specificity

Spectrophotometric/colorimetric

Precision PrecisionLOQ

SpecificityLOD - Specificity

Titrimetric Precision Precision - - -

TLC - SpecificityLOQ

SpecificityLOD - Specificity

Gel ElectroPhoresis - Specificity

LOQSpecificity

LOD - Specificity

* Not official* Not official

41

Data Elements for Verification of Data Elements for Verification of Dosage FormsDosage Forms

Technique Category I(assay)

Category II(impurities)

Category III(dissolution)

Category IV(identification)

Quantitative Limit test

HPLC/GCHPLC/GC

PrecisionPrecisionSpecificitySpecificityLinearityLinearityRangeRange

PrecisionPrecisionSpecificitySpecificity

LOQLOQ

SpecificitySpecificityLODLOD PrecisionPrecision SpecificitySpecificity

Spectrophotometric/colorimetric

PrecisionLinearityRange

PrecisionLOQ

SpecificityLOD Precision Specificity

TitrimetricPrecisionLinearityRange

Precision - - -

TLC - SpecificityLOQ

SpecificityLOD - Specificity

Gel Electrophoresis - Specificity

LOQSpecificity

LOD - Specificity

* * Not officialNot official

42

Category I:Category I: Analytical procedures for Analytical procedures for quantitationquantitation of of major components of drug substances or active major components of drug substances or active ingredients (including preservatives) in finished dosage ingredients (including preservatives) in finished dosage forms.forms.

Category II:Category II: Analytical procedures for determination of Analytical procedures for determination of impurities in drug substances or degradation compounds impurities in drug substances or degradation compounds in finished pharmaceutical products. These procedures in finished pharmaceutical products. These procedures include quantitative assays and limit tests. include quantitative assays and limit tests.

Category III:Category III: Analytical procedures for determination Analytical procedures for determination of performance characteristics (e.g., dissolution, drug of performance characteristics (e.g., dissolution, drug release). release).

Category IV:Category IV: Identification procedures. Identification procedures.

43

From ACTDFrom ACTD

P5 : Control of Finished Product

5.3 Validation of Analytical Procedure

Verification of Verification of compendialcompendial method applicability method applicability --

AccuracyAccuracyPrecisionPrecision

44

Comparison : AssayComparison : Assay

AccuracyAccuracy-

--Range

-LinearityLinearity

PrecisionPrecisionPrecision

SpecificitySpecificitySpecificity

AOACAOACEDQMEDQMUSPUSP

45

Recommendation (BDN):Recommendation (BDN):

For Assay:For Assay:

- Specificity

- Precision at 100 % (6 det.)

- Accuracy at 100% (3 det.)

- Linearity (5 points)

46

Comparison : Impurity (Quantitative)Comparison : Impurity (Quantitative)

Accuracy--

LOQ, LODLOQLOQ

Precision-Precision

SpecificitySpecificitySpecificity

AOACAOACEDQMEDQMUSPUSP

47

Recommendation (BDN):Recommendation (BDN):

For Impurity (Quantitative) :For Impurity (Quantitative) :

- Specificity

- LOQ

- Precision (6 det.)

- Accuracy (3 det.)

48

Comparison : Impurity (Limit tests)Comparison : Impurity (Limit tests)

LOQ, LODLOQ (LOD?)LOD

SpecificitySpecificitySpecificity

AOACAOACEDQMEDQMUSPUSP

49

Recommendation (BDN):Recommendation (BDN):

For Impurity (Limit tests) :For Impurity (Limit tests) :

- Specificity

- LOD

50

Comparison : IdentificationComparison : Identification

SpecificityNo formal validation required

Specificity

AOACAOACEDQMEDQMUSPUSP

51

Verification documentation :Verification documentation :

The verification exercise must be planned, approved, and documented and a final report kept on file to demonstrate the lab can perform the method adequately.

Verification documentation should be created that:

Described the procedure to be verified

Details the analytical performance characteristics to be evaluated

Established acceptance criteria used to determine that the procedure perform suitably

Justifies deviations from reference method

52

Thank You Thank You for for

Your AttentionYour Attention