A crucial area in developing new drugs is drug metabolism & pharmacokinetics is the study of a drug’s absorption, distribution, metabolism, and excretion.
Drug metabolism and pharmacokinetics are dealt by bioanalytical assay development method, which quantifies the drug in biological matrices such as plasma, urine, and tissue samples to determine its pharmacokinetics (PK) and pharmacodynamics (PD).
In determining the concentration, the samples of the relevant substances are collected and tested by bioanalytical methods. In addition to the active medication, compounds of interest can also contain metabolites (for small molecules) or in vivo breakdown products (for proteins). The drug concentration in the serum, plasma, or blood is measured and utilized as a proxy for the drug concentration at the site of action because blood sampling is comparatively noninvasive. These steps are necessary for the process of new drug discovery and development.
Analyses in tissues and other fluids such as urine, cerebrospinal fluid, and bile are also performed to understand better the action, toxicity, or elimination of the drug.
In addition to PK, large-molecule development necessitates immunogenicity testing, which is now included in bioanalysis because it employs the same methods as PK analysis and measures a large molecule’s ability to provoke an immunological response in a patient.
Due to the analytical methods’ similarities to PK, molecular biomarker assays are also a part of bioanalysis.
Setting Up A Bioanalytical Assay Validation
1. Critical Reagents and Reference Standards
All reference standards and crucial reagents, including antibodies, tagged analytes, and matrices, should be appropriately characterized and documented (e.g., to ascertain their identification, purity, and stability) by the sponsor, who should also store them under distinct guidelines.
2. The calibrating curve
The sponsor should select the quantitation range of the assay and the concentrations of the calibration standards s during method development based on the concentration range anticipated in a specific study.
3. Samples for Quality Control
The accuracy, precision, and stability of a sample and the assay stability are evaluated using quality controls (QCs).
Sponsors must create QCs using the same matrix as the research samples tested using the approved technique.
For precision and accuracy analyses during method development, freshly created QCs are advised because stability data are typically not yet available.
4. Selected and Particularity
To reduce or prevent interference, the sponsor should ensure the material being tested is the target analyte throughout technique development.
Analysis of blank samples of the appropriate biological matrix (such as plasma) from various sources is a standard procedure to demonstrate the method’s selectivity.
The LLOQ should be established during the method development process since it describes the method sensitivity. The technique should be created and tested to ensure that it can fulfill the needs of the study samples that are intended.
6. Recoverability, accuracy, and precision
Analyzing repeated QCs at various concentrations across the assay range is necessary to assess the accuracy and precision across the quantitation range during method development to establish whether the method is prepared for assay validation.
The sponsor should ascertain the chemical stability of the analyte in a specific matrix during the method development process, taking into account the impacts of sample collection, handling, and analyte storage.
The sponsor should evaluate the analyte’s long-term stability, autosampler, benchtop, processed or extracted samples, freeze-thaw, stock solution, and processed or extracted samples.
8. The Impact of Dilution
QC samples above the ULOQ with a similar matrix to bring to within quantitation range should be diluted during assay validation if the method assesses diluted samples, and the accuracy and precision of these diluted QCs should be proven.
The dilutions utilized during the validation should correspond to those anticipated for the study. LBAs should be used to demonstrate the prozone impact.
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