WHO Method Validation

8/12/2019 WHO Method Validation

1/34


2/34

The prequalification programme --Assessor's training | 19-20 January 20112 |

Outline

HPLC methodology

- Content of HPLC test procedure

- System Suitability Testing (SST)

- Relative Response actor (RR)

!alidation of HPLC method

case study


3/34


Information Sources

"# C"$R revie%er guideline for validation ofchromatographic methods (&'')

H* TRS '+, #ppendi .#nalytical /ethod !alidation

0112

3CH 40(R&) 0115

Compendial 6eneral Chapters

/ethods and !alidation presentation7Lynda Paleshnui8


4/34


High Performance Li9uid Chromatography (HPLC)

HPLC is a separation techni9ue based on a solid stationary phase and

a li9uid mobile phase: Separations are achieved by partition;

adsorption; or ion-echange processes; depending upon the type of

stationary phase used:

Chiral

3on--echange

3on--paire eclusion

The reversed-phase HPLC %ith ?! detectionis most commonly usedform of HPLC; is selected to illustrate the parameters of HPLC

method and validation:


5/34


A flow scheme for HPLC


6/34

The prequalification programme --Assessor's training | 19-20 January 2011! |

Content of HPLC test procedure

#ny analytical procedure submitted should be described in

sufficient detail; includes@

Preparation of mobile phase

Chromatographic condition@

7 Column@ type (e:g:; C&A or CA); dimension (length; innerdiameter); particle si>e (&1Bm; 5 Bm)

7 "etector@ %avelength

7 3nection volume

7column T

7 flo% rate;


7/34

The prequalification programme --Assessor's training | 19-20 January 2011" |

Content of HPLC test procedure

$lution procedure@ isocratic or gradient elution

Preparation of standards and samples

*peration procedure@ se9uence of inections

System suitability testing (SS) and criteria

Calculations

4*S 0:+:R:0analytical procedures and validation summaries


8/34

The prequalification programme --Assessor's training | 19-20 January 2011# |

Compendial methods

hen claim a compendial method; there should be no change in@

The type of column i:e the stationary phases

"etector %avelength

Components in /obile phase

System suitability testing and criteria

#dustments to ratio of components in mobile phase; flo% rate; columntemp; dimension of column; particle si>e (reduction only); may be

necessary to achieve the system suitability criteria: The allo%ablevariations for each parameter; see 3nt:Ph &:&: or ?SP generalchapter D20&E:


9/34


System suitability testin! "SST#

Precision@7 #ssay@ RS" F&G (#P3) or F 0G (PP); n 5

7 3mpurities@ in general; RS" F 5G at the limit level; up to &1G or

higher at L*4; n 2

Resolution (R)@ E0


10/34



Tailing factor


11/34



=umber of theoretical plates (=)@ column efficiency 0111

6radient elution is one %ay to increase the =


12/34



# SST should contain@

or #ssay@

precision I one or more other parameter

or impurity test@

resolution I precisionI one or more other parameter


13/34


$elati%e $esponse &actor "$$

'uantitation of Impurities#gainst impurity RSJs@ %hen reference standard available

#gainst #P3 itself

Relative response factor should be considered


14/34


15/34



$ifampicine)

y K*+,*+- I :'2+

$ifampicine 'uinone)

y K -.,+/0 I &:&5

RRK -.,+/0< *+,*+-

K1:A


16/34



To revie%@

a) RR calculation; and

b) if RR is properly used in the final calculation for G

impurity

3f RR %ithin 1:A-&:0; correction may not be necessay

Correction factorK &


17/34


$e%ie1 points for HPLC method

is the analytical proceduredescribed in detail includin! all the parameters 2 is SS1ell defined to ensure the consistency of system performance2

he preparation of solutions)

7 assay) concentration of reference standard should be close to the samplesolution

7 impurities) concentration of the reference standards should be close tothe limit

he 1ay of 3uantitation of impurities

In case API is used as the reference( $$& should be used or 4ustification ofe5clusion should be pro%ided,

o chec6 the determination of $$&( chec6 the correction of calculation ofimpurities

confirm7complete the 'OS -,*,$,-


18/34


Validation 8 compendial methods

Assay 8 API

=o validation generally re9uired: $ception@ specificity for maor impuritiesnot in the monograph:

Assay 8 &PP

Specificity; accuracy and precision (repeatability),

Purity 8 API and &PP

ull validation for specified impurities that are not included in themonograph (specificity; linearity; accuracy; repeatability; intermediateprecision; L*"


19/34


20/34


Specificity

lan8 solution to sho% no interference

Placebo to demonstrate the lac8 of interference from ecipients

Spi8ed samples to sho% that all 8no%n related substances are

resolved from each other

Stressed sample of about &1 to 01G degradation is used to

demonstrate the resolution among degradation products

7 Chec8 pea8 purity of drug substance by photodiode array detector (P"#)@ eg

purity angle is lo%er than the purity threshold:

Representative chromatograms should be provided %ith time scale

and attenuation indicated


21/34


Linearity 7 $an!e

The %or8ing sample concentration and samples tested foraccuracy should be in the linear range (concentrations

!s: Pea8 areas)

Minimum : concentrations

"ilute of stoc8 solution or separate %eighings


22/34


23/34


Linearity 7 $an!e

Correlation coefficient "r#

API) B ;,//0

Impurities) B ;,//

y9Intercept and slope should be indicated to!ether 1ith

plot of the data


24/34


25/34


Accuracy

3mpurities@ #P3


26/34


Precision

System precision@7 by multiple inections (n 5) of a homogeneous sample

(standard solution):

7 RS" F &G is recommended for assayM

7 RS" F 5G is recommended for related substances (referencestandards at the limit)

7 3ndicates the performance of the HPLC system

7 #s a system suitability test


27/34


Precision

$epeatability "method precision#7 Multiple measurements of a sample by the same analyst

7 A minimum of . determinations at the test concentration ".

times of a sin!le batch#( or

7 * le%els "0;


28/34


Precision

Intermediate precision "part of ru!!edness#7 est a sample on multiple days( analysts( e3uipments

7 $epeat the method precision by different analyst in

different e3uipment usin! different lot of column on

different days

7 $S? should be the same re3uirement as method precision

$eproducibility "inter9laboratory trial#

7 Not re3uested in the submission


29/34


LO?7LO'

signal to noise ratio@ L*" +@& ; L*4 &1@&7 /ay vary %ith lamp aging; model


30/34


LO?7LO'

LO?) belo1 the reportin! threshold

LO') at or belo1 the specified limit

=ot re9uired for assay


31/34


$obustness

The methodOs capability to remain unaffected by small butdeliberate variationsin method parameters

7 3nfluence of variations of pH in a mobile phase

7 3nfluence of variations in mobile phase composition

7 "ifferent columns (different lots and


32/34


33/34


Conclusion

HPLC methods play a critical role in analysis ofpharmaceutical product

!alidation of HPLC should demonstrate that the method

is suitable for its intended use

Revie% the information in dossier against 4*S 0:+:R:0

"ata for acceptance; release; stability %ill only be

trust%orthy if the methods used are reliable


34/34


Case Study

Case Study &--HPLC/ethod:doc

4*S 0:+:R:doc

Case Study 0 -- !alidation of HPLC/ethod:doc
http://case%20study%201--hplc%20method.doc/http://case%20study%201--hplc%20method.doc/http://qos%202.3.r.doc/http://case%20study%202%20--%20validation%20of%20hplc%20method.doc/http://case%20study%202%20--%20validation%20of%20hplc%20method.doc/http://case%20study%202%20--%20validation%20of%20hplc%20method.doc/http://case%20study%202%20--%20validation%20of%20hplc%20method.doc/http://qos%202.3.r.doc/http://case%20study%201--hplc%20method.doc/http://case%20study%201--hplc%20method.doc/

Documents

WHO Method Validation