Biomedical Engineering Reference
In-Depth Information
1
Introduction
New drug development is a lengthy and costly process. It typically lasts 15-17 years
and roughly 1 out of 5,000 compounds may come out as commercial drug. In order
to ensure the safety of the drug and the well-being of the public population, the new
drug development is conducted under a very constrained environment and closely
monitored by regulatory agencies. Except early drug screening and discovery which
is exempt from regulatory monitoring, most preclinical and clinical trials, i.e., animal
toxicology studies, in vitro laboratory experiments, and clinical trials, are conducted
according to FDA guidance, white paper recommendations and the SOPs [
1-
3
] .
Preclinical study is typically classified as regulated Good Laboratory Practice (GLP)
study. GLP specifically refers to management controls for laboratories and institu-
tions to ensure the uniformity, consistency, reliability, reproducibility, quality, and
integrity of pharmaceutical safety and efficacy tests. Although the scope and rigid-
ness of clinical study are similar as preclinical study, clinical study is normally
classified as regulated but non-GLP study due to lack of central study director, for
clinical study typically involves multiple testing facilities.
Throughout new drug development, multiple bioanalytical studies are conducted
in which the amount/concentration of drug and/or its metabolites in biological
matrix is measured and evaluated. For the same drug candidate, multiple method-
ologies may be developed to meet the specific goals at each stage. In early drug
discovery, the emphasis is speed and fast turnaround. Therefore, it is imperative to
use a nonspecific method suitable for wide range of analytes with diverse structures.
For later stage drug development such as GLP toxicology study and clinic study, the
reliability and reproducibility of the methodology are critical and essential.
Therefore, it is worthwhile to develop robust and rugged method.
The most frequently used techniques in bioanalytical studies are liquid chromatog-
raphy coupled with tandem mass spectrometry (LC-MS/MS) and noncompetitive
hybridization enzyme-linked immunosorbent assay (ELISA). Traditionally, hybrid-
ization ELISA is very useful technique for biopharmaceuticals and biomarkers such
as protein and oligonucleotides while the LC-MS/MS is more suitable for small mol-
ecules [
4,
5
]. This paradigm has changed in recent years. LC-MS/MS has become
more popular for the quantitation of macromolecules due to the advantage of high
selectivity, wide linearity range, and fast method development [
6,
7
] . The focus of this
chapter highlights the systematic approaches for method development, validation, and
sample analysis for regulated quantitative bioanalysis using LC-MS/MS technique.
2
Sample Preparation
The common biological matrixes for bioanalysis are various tissues, body fluid,
whole blood, plasma, serum, and urine. More recently, dried-blood spot sample as
alternative matrix has gained popularity in the industry [
8
] . Prior to LC-MS/MS
