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A holistic perspective on analytical method validation under International Council for Harmonisation Q2(R1)
*Corresponding author: Ankita Raikwar, Department of Pharmaceutical Sciences, Jaipur National University, Jaipur, Rajasthan, India. raikwarankita147@gmail.com
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Received: ,
Accepted: ,
How to cite this article: Panda A, Raikwar A, Panda S. A holistic perspective on analytical method validation under International Council for Harmonisation Q2(R1). Am J Biopharm Pharm Sci. 2025;5:1. doi: 10.25259/AJBPS_6_2025
Abstract
Validation is a fundamental requirement for ensuring compliance with Good Laboratory Practices and Current Good Manufacturing Practices. Each and every pharmaceutical sector ensures testing of excipients. IPQC material like excipients, drugs packaging material, and raw materials efficiently evaluated for their efficacy study. Hence, validation of analytical methods is a necessary prerequisite for testing of pharmaceuticals. Analyzing active pharmaceutical ingredients, pharmaceuticals excipients, and finished dosage forms includes the development of analytical evaluation procedure, which is essential to provide a highly accurate, exact, and intended outcome; hence, to obtain such a precise outcome, the analytical technique needs to be verified and validated according to the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines. The ICH guideline Q2(R2), endorsed by the European Medicines Agency, offers a harmonized and updated framework for analytical procedure validation across different regulatory regions including the USA (Food and Drug Administration), Japan (Pharmaceuticals and Medical Devices Agency), and the European Union and provides a standardized framework for validating analytical methods, addressing parameters such as accuracy, precision, specificity, detection limit, quantitation limit, linearity, and range. This comprehensive review outlines the principles and key requirements of ICH Q2(R1), emphasizing its role in regulatory compliance and quality assurance. It elaborates on the fundamental aspects of method validation, including systematic approaches to evaluate method performance, criteria for acceptance, and practical implementation strategies.
Keywords
Analytical method development and validation
Current Good Manufacturing Practices
Good Laboratory Practices
Q2
Q2(R1)
INTRODUCTION
An analytical procedure, as defined by the European Medicines Agency (EMA)/International Council for Harmonisation ICH Q2(R2), refers to a set of documented instructions that describe the steps to be followed for the analysis of a specific substance or product.[1] Quantification and qualification assessments of chemicals involve the analysis of chemicals to determine the identity, strength, quality, and purity of pharmaceutical substances. The various chemical combination compounds or samples are first segregated and then are analyzed for its quality [Table 1].[2] Quantification process involves the determination of amount of specific chemical components by analytical procedure. Analytical methodology to be followed for a procedure depends on a number of factors, including the sample matrix, analyte concentration, physical and chemical characteristics sample to be analyzed, the cost and speed of the analysis, and quantitative or qualitative assay. Qualitative analytical methodology includes numerical data and determination of amount of ingredient in sample. On the basis of developed methodology according to the ICH, different processes are being utilized for method development and validation ICH Q2(R2).[1,2]
| Type of validation | Purpose/when required |
|---|---|
| New analytical procedure | For a newly developed method |
| Verification of compendial procedure | To confirm suitability of pharmacopeial methods |
| Method transfer | When method is transferred to another laboratory |
| Post-change validation | After significant changes to an existing method |
| Ongoing performance verification | To confirm continued suitability during routine use |
Planning the selection of the appropriate analytical procedure based on intended use
Collecting all relevant information, including physicochemical properties and previous method data
Performing both qualitative and quantitative analytical procedure development under controlled laboratory conditions
Establishing standardized procedures for the testing of pharmaceutical samples.
A well-developed analytical procedure, in alignment with ICH Q2(R2) guidelines, facilitates straightforward validation by adhering to established scientific principles and validation parameters. The development of analytical method and its validation is crucial from the beginning of drug discovery and hence accounting for research and development of medicine.[1,3]
ANALYTICAL METHOD DEVELOPMENT
The process of defining and applying an appropriate analytical testing strategy for a specific intended use is referred to as analytical procedure validation, as outlined in the EMA-ICH guidelines Q2(R2). Stability studies have been an important part of research as it confirms quality of drug during the manufacturing of drug. Ultimately developed process can boost evaluation of performance of drug to ensure safety parameters and the study of physicochemical characteristics of pharmaceuticals.[1,4]
Analytical procedure development is an iterative and systematic process that occurs concurrently with the development of the pharmaceutical dosage form. The goal is to establish a procedure that is scientifically sound, cost-effective, time-efficient, and capable of producing reliable results that support the performance and quality of the drug product. Thus, robust and appropriate analytical procedure development is essential for pharmaceutical research and development. Understanding the physicochemical properties of the active pharmaceutical ingredient (API) is critically important during the early phases of drug development. These properties guide the selection of analytical techniques and influence subsequent steps, including safety evaluation for pre-clinical studies, pre-formulation investigations, and stability assessments. As the development process advances, analytical procedures are refined and expanded based on the evolving understanding of the API and formulation characteristics. In line with ICH Q2(R2) and EMA regulatory expectations, the selected procedures must be suitably validated, scientifically justified, and sufficiently robust to ensure reproducibility, specificity, and reliability.
A well-developed analytical procedure ensures the achievement of key objectives throughout every phase of drug development. It facilitates the use of optimized laboratory equipment and systems, contributing to the generation of high-quality, accurate, and reproducible testing results. Analytical procedures may include a range of techniques such as electrochemical methods, chromatography, and spectroscopic analysis, selected based on the nature of the analyte and intended purpose. A scientifically validated procedure serves as a reliable tool for verifying compliance with predefined specifications during manufacturing, release testing, and ongoing quality control. It also supports extended research activities related to the chemical profile, safety assessment, and pharmacological action of drug substances and products. Stability studies, as a component of the overall validation and lifecycle management process, are routinely performed to confirm that the drug product maintains its identity, purity, and physical characteristics from raw material sourcing through to final formulation. These studies align with EMA and ICH expectations to ensure product quality over its entire shelf life.[5]
Reason for analytical method development
Required for product that degrades easily and indicates lesser stability.
To analyze and evaluate characteristics of API, i.e., uniformity of drug, crystalline property of chemical, API bioavailability in body, study of bioavailability.
To study impurities for the identification of drug profile and safety.
To understand physiochemical properties and biological activities of finished product and key elements for the calculation of the correct dosage form.[1,6,7]
VALIDATION
The validation is documented evidence that provides high degree of assurance to a desired result with predetermined compliance.[6] A validated procedure ensures reliability and consistency in planned outcome. Further, it focuses on the compliance of product and analysis of final product, which is significant in pharmaceutical industry.[8]
Reason for validation
Mandatory for the enrollment of pharmaceuticals
Ensures at most precision of outcomes
Cost-effective
To ensure accreditation of the research center by the International Organisation for Standardisation guidelines
To acknowledge drugs by worldwide organization.
Types of validation
Process validation
Analytical method validation
Cleaning validation
Process validation
The manufacturing operational procedure must be flexible, but certain complications arise during manufacturing. The achievement of the good pharmaceutical qualities should be ensured with the prevention of unnecessary properties and, hence, achieve process validation.[10]
Analytical method validation
Analytical method validation is a fundamental necessity for pharmaceutical evaluation.[3] Method validation principally involves method of analysis with appropriate expected outcome explanation which are equipped, and ensures acceptable measurement as per ICH and Good Manufacturing Practices regulation to produce quality pharmaceuticals. To determine the outcomes, the sample must be tested multiple times.[2] Analytical validation method includes analysis of excipients, pharmaceuticals, and analytical conditions, which ensures validation of a product-specific analytical technique.[11]
Objective
When formulation or concentration of formulation requires substitution
Certain chances of regulatory non-compliance is decreased
Critical parameter analysis
Minimization of interference on accuracy and precision
Required for product authorization and marketing licenses for new, but non-pharmacopeia pharmaceuticals.
Cleaning validation
In order to ensure pharmaceuticals are free from contamination cleaning validation process is performed.[1] The unwanted contamination should be less as that of regulatory requirement. In the drug factory, cleaning validation is a crucial fundamental process, which is performed by different techniques i.e. the swab test. It is essential to utilize appropriate sampling technique and equipment that are free from chemical and microbial pollutants. Contaminants present if any must be in detection limit.[1,12]
Objectives
To ensure drug safety and purity
Patients’ requirement and satisfaction is ensured
Contamination and cross- contamination due to microbes, must be in acceptable criteria according to the ICH guidelines
Consistency of product and API is ensured during formulation.
The equipment and facilities should be cleaned efficiently. It is necessary to remove the product residues from the equipment, since they are a source of contamination. It is possible to guarantee the excipients’ and the API’s microbiological and chemical stability.[11,12]
ANALYTICAL METHOD VALIDATION CHARACTERISTICS
In accordance with the ICH guidelines, the validation of analytical method includes the following parameters:
Specificity
Accuracy
-
Precision.
Repeatability
Intermediate precision
Reproducibility.
Limit of detection
Limit of quantification
Linearity
Range
Robustness.
The required conditions for revalidation
Remodeling in manufacturing procedure.
Remodeling in the ingredients in the final product of drug
Remodeling analytical method procedure.[13]
Parameters of validation of analytical method
Specificity
Evaluation for the presence of impurities that might result in chemical degradation and has the highest possibility for its presence in the raw substance. It includes:
Identification: Identification and reorganization of ingredients.
Purity tests: Since it is nearly impossible to eliminate certain impurities, impurity limits are established. Impurities may exist in the form of heavy metals, related compounds, residual solvent content, etc., and hence can be tested by limit test, atomic absorption spectroscopy, and flame photometer.
Assay (content or potency): Quantitative determination of API, required for potency of drug.[14]
Linearity
Linearity is analyzed by calibration curve that measures data of testing substance that is directly correlated with the amount of sample amount.[15] It is necessary to prepare a minimum of five concentrations for analysis [Table 2].[16]
| Characteristics | Test of identification | Kind of analytical procedure | ||
|---|---|---|---|---|
| Impurities test | Assay | |||
| Quantification | Limit | Test of dissolution and content/potency | ||
| Accuracy | X | Y | X | Y |
| Precision Repeatability |
X X |
Y Y |
X | Y |
| Intermediate precision | ||||
| Limit of detection | Y | Y | Y | Y |
| Limit of quantification | X | X | Y | X |
| Linearity | X | Y | X | X |
| Range | X | Y | X | Y |
| Robustness | X | Y | X | Y |
X: Not essential parameter to performed, Y: Essential parameter to performed
Range
Range is used to analyze API (Active Pharmaceutical Ingredients) concentration. It is used to analyze API concentration for its upper and lower limits. Between the interval of upper and lower limits, API must demonstrate good efficacy. The range needs to be chosen such that it can demonstrate linearity, accuracy, and precision at a respectable level.[17]
The criteria of ranges that should be followed for range analysis
Assay for finished goods must be range between 80% and 120% of label claims
The content homogeneity must be between 70% and 130% of the label claim.[18]
Accuracy
Accuracy is closeness of agreements of data as conventional true value or accepted reference value.[19]
To perform the accuracy at least nine conclusions with a minimum of three concentration levels, that ensures predefined range must be used for analysis. For the analytical technique, three replications from three concentrations must be carried out each for accuracy calculations. As a result of accuracy, either the percentage recovery or the difference between the mean and the acceptable true value along with the confidence intervals must be documented.[14,20]
Precision
Precision is performed in predetermined condition for homogeneous sample, which should be performed multiple time, the result obtained ust be in accordance to standard observation. The three types of precision which are repeatability, intermediate precision, and reproducibility are assessed for calculation:
Repeatability: Must be analyzed in a brief amount of time and carried out in a suitable working environment and parameter. For instance, a minimum of three replications from each of the three concentrations is carried out.
Intermediate precision: Tests are conducted under varying laboratory conditions, on different days, by different personnel, and by different machines, etc.
Reproducibility: Accomplished outside of laboratories. It may involve cooperative research projects between various laboratories.[2,21]
Limit of detection
It is lowest concentration of a substance that can be analyzed with a given analytical method.
Visual evaluation basis: While comparison to the instrumental method, the non-instrumental method makes greater utilization of visual assessment with establishment of threshold. The limit of detection is controlled by the samples that have a known concentration.
Signal-to-noise basis: Signal-to-noise assesses baseline noise. The analyte with the lowest sample concentration is compared to that signal. In addition, the blank sample is examined. It is possible to determine the lowest concentration that can be determined. Generally speaking, a signal-to-noise ratio of 3:1 or higher is sufficient.[1]
Standard deviation of blank basis: It analyzes the adequate quantity of blanks set up and examined, to estimate the standard deviation.[1]
Standard deviation of the blank basis: Calculation of analytical background response magnitude. An appropriate quantity of blanks is created and examined. Based on the responses, the standard deviation is estimated.[3,7]
Calibration curve: Standard deviation can be drawn from the residual standard deviation of a regression line or from the regression y-intercepts.[22]
Robustness
Robustness is application of little but deliberate changes, but analytical method must remain same. It is laborious work to conduct numerous system appropriateness tests. However, this procedure guarantees the test acceptability for usage at all times.[15,23]
CONCLUSION
Analytical method validation i.e. ICH Q2(R1) guidelines, is an indispensable component of pharmaceutical quality assurance. It ensures that analytical procedures are scientifically sound, reproducible, and capable of delivering accurate and reliable results throughout the drug development lifecycle. By adhering to standardized validation parameters such as accuracy, precision, specificity, linearity, and robustness, pharmaceutical industries maintains compliance with regulatory requirements while safeguarding product quality and patient safety. A well-planned validation strategy not only minimizes errors and variability but also supports regulatory approval and market sustainability. Ultimately, rigorous method validation strengthens confidence in analytical outcomes, enabling consistent delivery of safe, effective, and high-quality pharmaceutical products.
Ethical approval:
Institutional review board approval is not required.
Declaration of patient consent:
Patient’s consent not required as there are no patients in this study.
Conflicts of interest:
There are no conflicts of interest.
Use of artificial intelligence (AI)-assisted technology for manuscript preparation:
The authors confirm that there was no use of artificial intelligence (AI)-assisted technology for assisting in the writing or editing of the manuscript and no images were manipulated using AI.
Financial support and sponsorship: Nil.
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