You are hereStrategies & Considerations for Bioanalytical Method Development and Validation Using LCMS / MS - A Review

Strategies & Considerations for Bioanalytical Method Development and Validation Using LCMS / MS - A Review

Bioanalytical Method Validation
Bioanalytical method validation (BMV) employed for the quantitative determination of drugs and their metabolites in biological fluids plays a significant role in the evaluation and interpretation of bioavailability, bioequivalence, pharmacokinetic, and toxicokinetic study data. These studies generally support regulatory filings (12). Validation is a tool of quality assurance which provides confirmation of the quality in equipment system, manufacturing processes, software and testing methods. Validation assures that products with pre-determined quality characteristics and attributes can be reproduced consistently within the establish limits (13). Bioanalytical validation is the process of demonstrating that analytical procedures are suitable for intended use.  In May 2001, FDA issued a guidance document about validating bioanalytical methodsbased on the deliberations of two workshops: Analytical Methods Validation: Bioavailability, Bioequivalence, and Pharmacokinetic Studies (held on December 3-5, 1990) and Bioanalytical Methods Validation (held on January 12-14, 2000) (14). These guidance take into account a variety of physicochemical, pharmacokinetic, and clinical characteristics for ensuring  the validation parameters of bioanalytical methods used for drug and its metabolites analysis (15).  Some validation parameters that, at least should be evaluated in a method validation process are : Selectiviy, calibration model, stability, accuracy, and limit of quantification(16).

Need of Validation
Bioanalytical method validation is required to establish, a documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its pre-determined specifications and quality attributes (13). It is essential to employ well-characterized and fully validated bioanalytical methods to yield reliable results that can be satisfactorily interpreted. It is recognized that bioanalytical methods and techniques are constantly undergoing changes and improvements; and in many instances, they are at the cutting edge of the technology. It is also important to emphasize that each bioanalytical technique has its own characteristics, which will vary from analyte to analyte—in these instances, specific validation criteria may need to be developed for each analyte. Moreover, the appropriateness of the technique may also be influenced by the ultimate objective of the study. When sample analysis for a given study is conducted at more than one site, it is necessary to validate the bioanalytical method(s) at each site and provide appropriate validation information for different sites to establish interlaboratory reliability (17).

Benefits of Bioanalytical Method Validation
The validation process provides the highest degree of assurance of the quality of product and process which ought to meet the predetermined quality specification. With this note method validation is also beneficial in many ways (13):

· Decreases the risk of preventing problems and thus assures the smooth running of the process
· Decreases the running cost
· Decreases the risk of regulatory non-compliance
· Fully validated process may require less in-process controls and end product testing (13)

Fundamental Elements of Method Validation
Some elements or points on which any method validation process stands (13)

· Validation plan and Standard operating procedures (SOP)
· Establishment of Acceptance criteria (i.e. testing parameters, limits of acceptability of the components)
· Demonstration of proper equipment system calibration and operation
· Demonstration of meeting of process with an established range of operation for multiple runs
· Demonstration of accuracy and precision for any analytical test methods (13)

Types of Method Validation
It is accepted that during the course of a typical drug development program, a defined bioanalytical method will undergo many modifications. These evolutionary changes (e.g. addition of metabolite, lowering of the lower limit of quantification) require different levels of validation to demonstrate continuity of the validity of the analytical process. Three different types/levels of method validation are defined and characterized as follow (18):

· Full Validation
· Partial Validation
· Cross Validation

a.) Full Validation

· Full validation is important when developing and implementing a bioanalytical method for the first time .
· Full validation is required for a new drug entity
· If metabolites are added to an existing assay for quantification, than full validation of the revised assay will be necessary for the analysis (19).

b.) Partial Validation
Partial Validations are modifications of validated bioanalytical methods that do not necessarily require full revalidations. Partial Validation can range from as little as one intra-assay accuracy and precision determination to a “nearly” Full Validation. Typical bioanalytical method changes which fall into this category include, but are not limited to (19):

· Bioanalytical method transfers between laboratories or analysts.
· Instrument and / or Software Platform changes.
· Change in species within matrix (e.g. rat plasma to mouse plasma).
· Change in matrix within a species (e.g., human plasma to human urine).
· Selectivity demonstration of an analyte in the presence of specific metabolites.
· Selectivity demonstration of an analyte in the presence of concomitant medications.
· Change in analytical methodology (e.g., change in detection systems).
· Change in sample processing procedure(s).
· Rare matrices.
· Change in anti-coagulant in harvesting biological fluid.
· Limited volume changes (e.g. planned pediatric study) (19).



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