Page 26 - Modern Derivatization Methods for Separation Sciences

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1.1.1—
Introduction

Drug analysis is mainly divided into three fields; materials, formulations and bioanalysis of specimens
obtained in vitro and in vivo. The former two are controlled by regulations such as UPS in the USA, and
the physico-chemical properties of drugs are summarized in a series of books entitled 'Analytical
Profiles of Drug Substances', published periodically by Academic Press Inc. from 1972. In this section,
methods for derivatization for drugs, in order to monitor their levels in biological samples, are
described.

Drug level monitoring in biological fluids such as blood and urine, and in tissues is essential to
elucidate its disposition in the body regarding pharmacological and toxicological properties. From the
early 70's, chromatographic methods, especially using HPLC, have played important roles in trace level
drug monitoring. In this field, sensitivity and selectivity of the target drug in a complex matrix, are the
most important parameters.

UV detection methods in HPLC, which is the most widely used one, sometimes lack sensitivity or
selectivity for trace level drug analysis. Chemical derivatization can modify drugs to give efficient
absorption in UV or visible wavelength and luminescent properties such as fluorescence and chemi-or
bio-luminescence, or electrochemical activity can attain highly sensitive and selective determination of
drugs using HPLC.

A vast number of drugs exhibit the property of chirality and some of them are used therapeutically

as the racemates. When racemic drugs are administered, individual enantiomers often have different
activities, toxicities, and pharmacokinetic properties. For enantioseparation, HPLC is one of the most
powerful techniques. There are two principles in chiral separation using HPLC; direct separation using
a chiral stationary phase column, and diastereomer formation with a suitable chiral reagent. The
diastereomeric derivatization method has the advantage for trace analysis of enantiomers in biological
matrix because of the utilization of the reagent with high sensitivity in detection.

Papers published recently on the derivatization of drugs using HPLC are mostly classified into the two
categories mentioned above. In the following section, our recent publications on the above categories
are mainly described.

1.1.2—
Application of Derivatization of Drugs

1.1.2.1—
Determination of Pravastatin Sodium in Plasma by HPLC with Laser-induced Fluorescence
Detection after Immobilized Antibody Extraction

Pravastatin sodium, Fig. 1.1.1, is an inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A (HMGCoA)
reductase, a key enzyme in cholesterol biosynthesis [1]. Since plasma concentration in rats and humans
is quite low due to specific uptake [2], the development of a highly sensitive and specific assay in
biological matrices has been required to study of the pharmacokinetics of

http://emedia.netlibrary.com/nlreader/nlreader.dll?bookid=17968&filename=Page_2.html 30/09/2003