anal meth dev

7/28/2019 Anal Meth Dev

1/43

| Slide 1 of 43 April 2007

Training Workshop onPharmaceutical Development withfocus on Paediatric Formulations

Protea Hotel

Victoria Junction, Waterfront

Cape Town, South Africa

Date: 16 to 20 April 2007

Pharmaceutical Development


2/43


Pharmaceutical Development

Analytical Method Development

Presenter: Jnos Pogny, pharmacist, PhD

[email protected]

WHO expert
mailto:[email protected]:[email protected]


3/43


Analytical Method Development

Outline and Objectives of presentationIntroduction, guidelines

Dossier requirements Assay Related substances Other issues

Main points again


4/43

Training Workshop onPharmaceutical Development

with focus on Paediatric

Formulations

Introduction, guidelines


5/43


Interchangeability (IC)

INTERCHANGEABILITY (IC) OF MULTISOURCEFPPs = (ESSENTIAL SIMILARITY WITHINNOVATOR FPP ) =

PHARMACEUTICAL EQUIVALENCE (PE ) +BIOEQUIVALENCE (BE)

IC = PE + BE


6/43


Pharmaceutical equivalence

FPPs meet same or comparable standards (e.g.,marketing authorization, analytical methods ) Same API (chemical and physical equivalence) Same dosage form and route of administration Same strength Comparable labeling

Pharmaceutical development equivalenceStability equivalenceWHO-GMP (manufacturing equivalence)


7/43


Prequalification requirements

Analytical method validation is required by WHO for theprequalification of product dossiers. Non-compendial ARV

APIs and FPPs were/are tested with methods developedby the manufacturer.

Analytical methods should be used within GMP and GLPenvironments, and must be developed using the protocolsand acceptance criteria set out in the ICH guidelinesQ2(R1)


8/43


Guidelines used in PQP

WHO-GMP 4.11 It is of critical importance that particular attention is paid to the validation of analytical testmethods , automated systems and cleaning procedures.

Appendix 4. Analytical method validation (in WHO Expert

Committee on Specifications for Pharmaceutical Preparations. 40thReport. Geneva, WHO, 2006 (WHO Technical Report Series, No.937). http://whqlibdoc.who.int/trs/WHO_TRS_937_eng.pdf

Validation of analytical procedures: text and methodology

Q2(R1) ICH Harmonized Tripartite Guidelines, (2005)http://www.ich.org/LOB/media/MEDIA417.pdf
http://whqlibdoc.who.int/trs/WHO_TRS_937_eng.pdfhttp://www.ich.org/LOB/media/MEDIA417.pdfhttp://www.ich.org/LOB/media/MEDIA417.pdfhttp://www.ich.org/LOB/media/MEDIA417.pdfhttp://www.ich.org/LOB/media/MEDIA417.pdfhttp://whqlibdoc.who.int/trs/WHO_TRS_937_eng.pdfhttp://whqlibdoc.who.int/trs/WHO_TRS_937_eng.pdfhttp://whqlibdoc.who.int/trs/WHO_TRS_937_eng.pdf


9/43


General requirements

Qualified and calibrated instruments

Documented methods

Reliable reference standards

Qualified analysts

Sample selection and integrity

Change control


10/43


Measure of variation (spread of data)

95.46%68.26 %


11/43


Mean (average) chart

Normal

variation due tocommoncauses

USL Upper specification limit

LSL Lower specification limit

Abnormal variation of process

special causes

Abnormal variation of process special causes

average = mean


12/43


Capable process Almost all the measurements of astable process fall inside thespecification limits

USL LSL 6 8 10 12

Cp 1.00 1.33 1.66 2.00

OoS results: .27%. 6 ppm

64 ppm 2 ppb

.htm16 /pmc1 /handbook/pmc/section898http://www.itl.nist.gov/div
http://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htmhttp://www.itl.nist.gov/div898/handbook/pmc/section1/pmc16.htm


13/43


NEVIRAPINE Reference Standard

AtR Injection

124666.1601

123116.1672

124326.1663

125306.1724

124576.1655

124796.1686

124466.166Mean

740.004STD

0.59%0.06%RSD

System suitability

requirement:RSD is NMT 0.85%


14/43

Training Workshop onPharmaceutical Development

with focus on Paediatric

Formulations

Dossier requirements


15/43


Use of analytical methods - generics

MethodsPharmaceuticalClinicalAt initial phase of pharmaceutical development

-To understand the profile of related substances and to studystability and start measuring theimpact of key product and manufacturing process

parameters on consistent FPPquality

To develop a stable and reproducibleformulation for the manufactureof bioequivalence, dissolution,stability and pilot-scalevalidation batches

To determine bioavailability inhealthy volunteers

At advanced phase of pharmaceutical developmentTo be robust , transferable , accurate ,

and precise for specificationsetting, stability assessment,and QC release of prequalified

batches

To optimize, scale-up, and transfer a

stable and controlledmanufacturing process for the prequalification product

To prove

bioequivalence after criticalvariations to the

prequalified dossier


16/43


Analytical procedure characteristics

Type of characteristic Identification ImpuritiesQuantitative Limit Assay

Accuracy - + - +

Precision - + - +

Specificity + + + +Detection limit - - + -

Quantitation limit - + - -

Linearity - + - +

Range - + - +

Robustness + + + +


17/43


Accuracy - ISO 5725 1-6

Accuracy

Trueness Precision(Random errors)

Systematic errorsIntra-assayvariability

Intra-laboratoryvariability

Inter-laboratoryvariability

Repeatability Intermediateprecison Reproducibility

Source: ISO. 1994. ISO 5725 1-6: Accuracy (Trueness and Precision) of Measurement Methods and Results. ISO, Geneva, Switzerland.


18/43


Accuracy and precision

Inaccurate &

imprecise

Inaccurate butprecise

Accurate butimprecise Accurate and precisePrecise Accurate

Inaccurate and imprecise


19/43


Percent accuracy (hypothetical figures)

The data show that the recovery of analyte in spiked samples met the evaluationcriterion for accuracy (100 2.0% across 50 130% of target concentrations).

RSD%

Recovery%

Nevirapine, mgLA, %Sample RecoveredAdded

0.6499.20.4950.499501

0.3199.80.7010.703702

0.2799.90.7950.796803

1.88100.41.0051.0011004

0.3899.81.2091.2111205

1.1299.81.2961.2991306

0.7799.80.9170.918Mean


20/43


Percent accuracy (hypothetical figures)

96979899

100101102103104

0 1 2 3 4 5 6 7

Number of samples

% R

e c o v e r y

Red line: LA Green lines: USL and LSL


21/43


Precision ( of any process )

The precision (VARIABILITY ) of ananalytical procedure is usuallyexpressed as the standard deviation(S), variance (S 2), or coefficient of

variation (= relative standarddeviation, RSD%.) of a series of measurements.

The confidence interval (CI) should

be reported for each type of precisioninvestigated.

Measured mean Real mean

PRECISION


22/43


Repeatability ( of any process )

Repeatability expresses theprecision (spread of the data,variability ) under the sameoperating conditions over a shortinterval of time. Repeatability is

also termed intra-assay precision.

REPEATABILITY

Measured mean


23/43


Repeatability (hypothetical figures)

The repeatability precisionobtained by one analyst inone laboratory was 1.25%RSD for the analyte and,

therefore, meets theevaluation criterion of RSD2%.

Imp1Peak areaInjection

0.301579351

0.301578332

0.299574973

0.3005761740.301577785

0.298572316

0.30057649Mean

0.0013257STD

0.4%0.4%RSD

0.001427095% CI


24/43


Intermediate Precision and Reproducibility ( of any process )

Intermediate precision expresseswithin-laboratories variations. #1, #2and #3 : different days, differentanalysts, different (manufacturing ) equipment , etc.

Reproducibility expresses theprecision between laboratories #1, #2and #3 (collaborative studies, usuallyapplied to standardization of methodology). ( Transfer of

technology )

Measured means

Intermediate precision orReproducibility


25/43


Intermediate precision (ruggedness)

Assay (mg/5ml)Sample52.651.71

52.151.92

52.353.03

52.952.54

53.252.35

53.152.76

52.752.4Mean

0.440.49STD

0.8%0.9%RSD

0.460.5195% CI

Combined values52.5Mean

0.48STD

0.9%RSD

0.3195% CI


26/43


Specificity (selectivity)

Specificity is the ability to assess unequivocally theanalyte in the presence of components , which may beexpected to be present. Typically these might includeimpurities, degradants and excipients .

An example of specificity criterion for an assay method isthat the analyte peak will have baseline chromatographicresolution of at least 2.0 minutes from all other samplecomponents

Stability indicating analytical methods should always bespecific.


27/43


28/43


Specificity (hypothetical figures and data)

HPLC chromatograms of (a) API referencestandard, (b) FPP and (c) placebo


29/43


SPECIFICITY degradants

Purity thresholdPurity angleA (%) *Stress0.2800.040

100.0Initial 0.3800.1051.6300.7250.2800.045

99.3Acid 0.4100.1201.6101.0400.2700.060

99.8Peroxide 0.3600.1101.2500.690

NANA100.0All others

There were no peaks in the placebo chromatogram at the retention times of nevirapine (N),methylparaben (MP) and propylparaben (PP) peaks.

*Sum of N, MP and PP peak areas. The three ingredients can be assessed in the presence of (non-expected) degradants. The peaks are homogeneous and pure. The method is selective, specificand stability-indicating.


30/43


Limit of Quantitation (LOQ)

Limit of Detection (LOD)

Signal to Noise Ratio (SNR)

noise

Peak ALOD

Peak B

LOQ

Baseline

LOD, LOQ and SNR


31/43


LOD and LOQ (hypothetical figures)Impurity 2Impurity 1

Injection LOQLODLOQLOD

78923497723541761

77914258809936082

82923275795041963

80503464816643034

83684008784739325

82844702841552386

8113386779524242Mean

238551402548STD

2.9%14.3%5.1%12.9%RSD0.2140.1070.1710.086Conc. (g/ml)

0.0390.0190.0330.017Conc. (%w/w)


32/43


LOD and LOQ

The limit of detection ( LOD) is defined as the lowest concentration of ananalyte in a sample that can be detected, not quantified. It is expressed as aconcentration at a specified signal : noise ratio (SNR) , usually between 3and 2 : 1 .

In this study, the LOD was determined to be 0.086 g/m l (Impurity 1) with asignal : noise ratio of 3.6 : 1

The limit of quantitation ( LOQ ) is defined as the lowest concentration of ananalyte in a sample that can be determined with acceptable precision andaccuracy under the stated operational conditions of the method. The ICHhas recommended a signal : noise ratio (SNR) of 10:1 .

The LOQ was 0.171 g/m l (Impurity 1) with a signal:noise ratio of 11.3 . TheRSD for six injections of the LOQ solution was 2%.


33/43


Linearity

Linearity expresses differencesin precision at different points of a given range.The linearity of an analyticalprocedure is its ability (within agiven range) to obtain test

results, which are directlyproportional to theconcentration (amount) of analyte in the sample.

Measured Realmean mean Precision


34/43


Linearity and range

Acceptance criterion: correlation coefficient should not be less than 0.9990

y = 36.124x - 7.2984

R 2 = 0.9998

0

1000

2000

3000

4000

5000

0 20 40 60 80 100 120 140

Concentration, %

A s s a y m e a n


35/43


Linearity and range

Concentration range 1.0 1.3 mg/ml ( 10 130% of the theoreticalconcentration in the test preparation, n=3 )

Regression equation was found by plotting the means of peak area(y) against the analyte concentration (x) expressed in %:y = 36.124x - 7.2984 ( R 2 = 0.9998 ).

The regression coefficient demonstrates an excellent relationshipbetween peak area and concentration of analyte.

The analyte response is linear across 10-130% of the target

nevirapine concentration.


36/43


Range (minimum requirements)

Assay of an API or a FPP : 20% of the test concentration.

Content uniformity : 30% of the test concentration (unless a wider more appropriate range, based on the nature of the dosage form (e.g., metereddose inhalers), is justified).

Dissolution testing : 20 % over the specified range.

Impurity : from the reporting level of an impurity to 120% of thespecification. (Unusually potent or toxic impurities, LOD and LOQ should becommensurate with ICH requirement.)

If assay and purity are performed together as one test and only a

100% standard is used, linearity should cover the range from thereporting level of the impurities to 120% of the assay specification


37/43


Stability of analytical solution

Stability (of the analytical solution)expresses variation of themeasured mean as a function of time.

#1 First measurements

#2, #3, #4 , n Series of measurements of the same sample

within a relatively short period of time.

Stability Measured means


38/43


Stability of test analytical solution

An analytical solution preparedfrom Nevirapine 50mg/5ml OralSuspension was spiked withImpurity-1 at specification leveland stored in a capped volumetricflask on a laboratory bench atuncontrolled room temperatureunder normal lighting conditionsfor 25 hours.

Conclusion : the stability of theanalytical solution of Impurity-1 isnot a source of variation.

Impurity-1Time in

hours DifferenceArea

720790

0.7%715741

0.5%717402

0.2%719603

-0.4%723524

0.7%715735

-0.3%7232210

-0.3%7231015-0.3%7231220

-0.8%7267025


39/43


Sensitivity and robustness


40/43


41/43


Methods for cleaning validation

Method for assay and related substances used in stabilitystudies of API and FPP Specificity (in samples taken from a cleaning assessment) Linearity of response (from 50% of the cleaning limit to 10x this

concentration; R 2 0.9900; ) Precision

Repeatability (RSD 5%) , intermediate precision [ruggedness (USP)], and reproducibility

Limits of detection and quantitation Accuracy or recovery from rinsate ( 80%), swabs ( 90%), and

process surface ( 70%) Range (lowest level is at least 2x higher than LOQ)


42/43


Main Points Again

Analytical procedures play a critical role inpharmaceutical equivalence and risk assessment /management : establishment of product-specific acceptance criteria , and stability of APIs and FPPs.

Validation should demonstrate that the analyticalprocedure is suitable for its intented purpose.

HPLC systems and method validation deserves specialattention during the assessment of dossiers for prequalification .


43/43


THANK YOU

anal meth dev

Documents