Validation Team

ANALYTICAL METHOD VALIDATION PROTOCOL

\ Company Name

PROTOCOLNO:-

Effective Date :

Revision No. : 00 COMPANY NAME & ADDRESS ANALYTICAL METHOD VALIDATION PROTCOL NO: EFFECTIVE DATE : Page No.:- 2 of 17

PRODUCT NAME :

Validation Team:

DEPARTMENT VALIDATION TEAM QUALITY ASSUARANCE QUALITY CONTROL

APPROVALS:

DEPARTMENT SIGN & DATE QUALITY ASSURANCE MANAGER QUALITY CONTROL MANAGER

REMARKS:

Prepared By: Checked By: Approved By: Date: Date: Date: COMPANY NAME & ADDRESS ANALYTICAL METHOD VALIDATION PROTCOL NO: EFFECTIVE DATE : Page No.:- 3 of 17

1.0 GENERAL: 1.1 INTRODUCTION: The Analytical method validation will be performed. The complete documentation for the validation comprises several independent documents, references to relevant documents will be given as part of this protocol, (see below). The results of the validation activities will be summarized in the validation report. ANALYTICAL METHOD VALIDATIONS:

The objective of validation of an analytical procedure is to demonstrate that it is suitable for its intended purpose.

Analytical methods used for the identification, ore assaying the purity and or the impurity profile For drug substances, packing components and drug products and will be validated for its parameters.

(I) Specificity:

Specificity is the ability to access unequivocally the analyte in the presence of components which may be expected to be present; typically these might include impurities, degradents, matrix, etc.

Lack of specificity of an individual procedure may be compensated by other supporting analytical procedure.

This definition has been following implications.

A) Identification: to ensure the identity of an analyte.

B) Purity Test: to ensure that all the analytical procedure performed allow an accurate statement of the content of impurities of an analyte i.e. related substances, heavy metals, residual solvents contents etc. C) Assay (Content or Potency): to provide an exact results which allows an accurate statement on the content or potency of the analyte in the sample.

(II) Accuracy:

The accuracy of an analytical procedure expresses the closeness of agreement between the value which is accepted either as a conventional true value or an accepted reference value and the value found.

This is sometimes termed as trueness. (III) Precision:

The precision of an analytical procedure expresses the closeness of agreement (degree of scatter) between a series of measurements obtained from multiple sampling of the same homogenous sample under the prescribed condition. Precision may be considered at three levels: Repeatability, intermediate precision, and reproducibility.

Precision should be investigated using homogenous authentic samples. However, if it is not possible to obtain a homogenous sample, it may be investigated using artificially prepared samples or a sample solution.

The precision of an analytical procedure is usually expressed as the variance, standard deviation or coefficient of variation of a series of measurements.

Repeatability: repeatability expresses the precision under the same operating conditions over a short interval of time, Repeatability is also termed intra-assay precision.

Intermediate precision: Intermediate precision expresses within-laboratories variations: different days, different analysts, different equipment, etc.

Reproducibility: Reproducibility expresses the precision between laboratories (collaborative studies, usually applied to standardization methodology.

(IV) Detection of Limit:

The detection limit of an individual analytical procedure is the lowest amount of analyte in a sample which can be detected but not necessarily quantitative as an exact value.

(V) Quantitation Limit:

The quantitation limit of an individual analytical procedure is the lowest amount of analyte in a sample which can be quantitatively determined with suitable precision and accuracy. The quantitation limit is a parameter of quantitative assay for levels of compounds in sample matrices, and is used particularly for the determination of impurities and/or degradation products.

(VI) Linearity The linearity of an analytical procedure is its ability (within a given range) to obtain test results which are directly proportional to the concentration (amount) of analyte in the sample.

(VII) Range The range of an analytical procedure is the interval between the upper and lower concentration (amounts) of analyte in the sample (including these concentrations) for which it has been demonstrated that the analytical procedure has a suitable level of precision , accuracy and linearity.

(VIII)Robustness The robustness of an analytical is a measure if its capacity to remain unaffected by small, but deliberate variations in method parameters and provides an indication of its reliability during normal usage.

1.2 List of Documents for Validation:  Validation protocol,  Details of sampling for the validation batches, test parameters with reference to test methods & Acceptance criteria. (acceptable Limit)  Methods for recording / evaluating results including statistical analysis.

 Reference to relevant documents.

2.0 PERSSONEL RESPONSIBILITIES.

SR ACTIVITY RESPONSIBILITY REMARKS

1 Preparation of validation protocol 2 Approval of Validation protocol 3 Testing of validation samples & Preparation of validation report 4 Approval of validation report.

EQUIP LIST:

SR EQUIPMENT/ INSTRUMENT I.D.NO. STATUS REMARK NAME

GLASS WARE

SR GLASS WARE NAME NOMINAL STATUS REMARK CAPACITY

METHOD OF ANALYSIS: Method :- STP. No. :- Ref. :- Product Name :- Parameter Results Acceptance Criteria. RSD is Less than 1 % Precision 1. (Repeatability)

2.Reproducibility RSD is Less than 1 % Accuracy

2 Linearity R is NLT 0.98 Detection limit Method should be able to detect

minimum analyte concentration Method should be able to detect

Specificity impurity/ degradable product &

distinguish between std.

( Undegradable product)

Method :- STP. No. :- Ref. :- Product Name :- Label :- Precision ( Repeatability )

Taken 6 Sample individually and followed method Calculation :-

Sample Reading Assay in gm Assay in % Xi (Xi-X) (Xi-X)2

T6 Σ Xi Σ (Xi-X)2 X =

S = Σ (Xi-X) 2 (n-1)

S = ______5

RSD = 100 S X

NOTE: Minimum six determination at 100 % of test concentration

Accuracy RECOVERY BY SPIKING STD. TO TEST PREPARATION

Taken Std.-Reference standard ______gm Std. Potency: - ______

Test Smp. T1______ml / gm T2______ml / gm T3______ml / gm

Recovery Smp. R______ml of test preparation ,______gm -______Std. Spiked, mixed.

(i.e. gm Std./ ml/ gm ) & followed method .

Reading Mean

Std. S

Test Smp. T1

Recovery Smp. R

Reading Mean

Std. S

Test Smp. T2

Recovery Smp. R

Reading Mean

Std. S

Test Smp. T3

Recovery Smp. R

Calculation :-

Calculate % recovery using following formula

% Recovery = gm in Recovery sample –gm in Test Sample X 100 gm Std. Spiked

Note: Three concentration and three replicates of each concentration.

e.g. 100 + 10 % i.e. 90 % , 100 % , 110 %

Linearity

Take Sample ______ml / gm. ______ml / gm.______ml / gm.______ml / gm. ______ml / gm. and followed method.

Calculation:

zSample Taken in ml Concentration in % % Recovered From 80 %

90 % 100 % 110 % 120 %

Note: Minimum of five concentrations * Assay of drug substance: - from 80 % to 120 % of the test concentration.

Linearity

150 118.04 % 100 98.87 108.46 78.21 88.54 50

0 0.0096 0.0108 0.012 0.0132 0.0144 y = 9.962x + 68.508 R2 = 0.9996 % of Conceration (Limit :- NLT 0.98)

Method :- STP. No. :- Ref. :- Product Name :- Label :- Precision ( Reproducibility )

Test by different analyst on different days. Sample A1:______Analyst 1 on:______

Sample A2:______Analyst 2 on:______

Sample A3:______Analyst 3 on:______

Calculation :-

Sample A1

Sr Reading Assay in gm Assay in % Xi (Xi-X) (Xi-X)2 1 2 3 4 5 6 Σ Xi Σ (Xi-X)2 X =

Sample A2

Sample A3

S = Σ (Xi-X) 2 (n-1)

S = ______5

RSD = 100 S X

NOTE: Minimum six determination at 100 % of test concentration

Method :- STP. No. :- Ref. :- Product Name :- Label :- Specificity:

Include comparison of test results of samples containing impurities or degradation products ( 15,30,45, minutes of degradation product)

Factor for degradation;  Light  Heat  Humidity  Acid base Hydrolysate  Oxidation.

Sample A1: 15 minutes of degradation

Sample A4: Undegradable sample

Sample RESULTS A1 A2 A3 A4

NOTE: Recommended data : *In case of chromatographic procedure – chromatogram should be presented. * Peak purity test- Chromatographic peak should be presented.

Method :- STP. No. :-

Ref. :- Product Name :- Label :- Detection of limit:

Approach is used for detection limit: - Visual evaluation.

Take samples with known concentration of analyte and established the minimum level at which the analyte can be reliably detected.

Sample A1: ______

Sample A2: ______

Sample A3: ______

Sample A4: ______

Sample Known Concentration RESULTS A1 A2 A3 A4

Known concentration of analyte is decided from on the basis of required detection level i.e. above and below the required detection level.

Prepared By: Checked By: Approved By: Date: Date: Date: