Validation

Defination

Establishing documentary evidence which provides a high degree of assurance that specification process will consistently produce a product meeting its predetermined specifications and quality attributes.

Methods need to be validated or revalidated

• before their introduction into routine use

• whenever the conditions change for which the method has been validated, e.g., instrument with different characteristics

• whenever the method is changed, and the change is outside

Validation parameters evaluated(Q2R1)

•Specificity

•Linearity

•Accuracy and range

•Precision

Repeatability

Intermediate precision

•Robustness

Specificity

• Specificity Is ability to measure analyte

Of interest accurately & specifically in presence of other componants that could be expected to be present in sample matrix.

Linearity• Minimum of 5 concentrations is recommended.

• Range studied 0 to 150% of nominal (7 solutions, including a ‘blank’ solution)

• Linearity experiments conducted in the presence and absence of excipients

• Determine equation of the line, correlation coefficient, 95% confidence interval of the slope and the intercept. Verify that the 95% confidence interval of the intercept includes zero, or that the result is insignificant in the context of the experiment

• Check that the response for both experiments is the same – calculate the slopes as a % of each other

• Typically results of 98 to 102% would be considered acceptable

Range

• The following minimum specified ranges should be considered:

For the assay of a drug substance or a finished (drug) product: normally from 80 to 120 percent of the test concentration;

• For content uniformity, covering a minimum of 70 to 130 percent of the test concentration, unless wider more appropriate range, based on the nature of the dosage form (e.g., metered dose inhalers), is justified;-

• For dissolution testing: +/-20 % over the specified range;

Precision - Repeatability• One analyst conducting multiple replicates of the

same sample, on the same day using the same equipment, and applying the method as written.

• Determine the mean, the range, the standard deviation and the relative standard deviation.

• Repeatability should be assessed using: a) a minimum of 9 determinations covering the

specified range for the procedure (e.g. 3 concentrations/3 replicates each)

Precision – Intermediate precision

• A second operator conducts the ‘repeatability experiment’ on a different day using different equipment where possible.

• Determine the individual data and overall mean, range, standard deviation and relative standard deviation.

• Consider testing statistically that there is no significant difference between operators before combining the data set.

System Suitability

• System suitability is checking of systemto ensure system performance before or durring the analysis of unknown

• Plate count (N), Tailing Factor (T) , Resolution (R) & Reproducibilty (RSD) are measured for rplicate injections of sample

• If RSD specification is less than 2%,Five replicate injections should be used.

• If RSD specification is more than 2%,Six replicate injections should be used.

• System suitability should be done Before,Durring,after the analysis.

Accuracy

• Accuracy is a measure of Exactness of analytical method

• It is measured as % analyte recovered by assay ,by spiking samples

• Data from minimum Three conentrations & three replicate of each concentration to be reported.

Limit of Detection

• The limit of Detection (LOD) is lowest conc of analyte in the sample that can be detected & Not quantified.

• It can be measured by A signal-to-noise ratio .

• A signal-to-noise ratio should be between 3 or 2:1 is generally considered acceptable for estimating the detection limit.

Limit of Quantitation

• The limit of Quantitation (LOQ) is lowest conc of analyte in the sample that can be determined (quantitated).

• It can be measured by A signal-to-noise ratio .

• A signal-to-noise ratio should be between 10:1 is generally considered acceptable for estimating the detection limit.

RobustnessA number of attributes were evaluated prior to

validation using an experimental design approach:

• Injector temperature• Detector temperature• Initial temperature• Ramp rate• Hold time• Split flow• Column head pressure

A validation report should be prepared that includes:

•objective and scope of the method (applicability, type)

•type of compounds and matrix

•detailed chemicals, reagents, reference standards and control sample •preparations

•procedures for quality checks of standards and chemicals used

•safety considerations

•method parameters

•critical parameters indicated from robustness testing

•listing of equipment and its functional and performance requirements, e.g. cell

A validation report should be prepared that includes:

• dimensions, baseline noise, column temperature range • detailed conditions on how the experiments were conducted,

including sample preparation • statistical procedures and representative calculations • procedures for quality control in the routine (e.g., system suitability

tests) • representative plots, e.g. chromatograms, spectra and calibration

curves • method acceptance limit performance data • the expected uncertainty of measurement results • criteria for revalidation • person who developed and initially validated the method • summary and conclusions