Automated HPLC analyses of drugs of abuse via direct injection of biological fluids followed by simultaneous solid‐phase extraction and derivatization with fluorescence detection

Abstract

An automated system is described for the simultaneous extraction and derivatization of nucleophilic compounds from various biological media. The method includes the use of a solid-phase reagent containing a 9-fluorenylacetate activated ester. The reagent is based on a controlled pore, polystyrene divinylbenzene support prepared through a silica template procedure. An X-Y-Z robotic arm equipped with a needle is used in conjunction with a syringe pump for aspirating and dispensing samples and standards into the HPLC system. A precolumn cartridge containing the solid-phase reagent is put on-line in place of the fixed-volume injection loop. Injections of biological fluids such as urine or plasma with minimal sample treatment and handling are made directly into this reactor. The analytes are derivatized as they are extracted, allowing virtually unlimited sample volumes to be injected. The polymeric cartridge can be used for up to 100 injections without accruing unacceptable reductions in sensitivity. A detection limit of 500 p.p.t. (parts per trillion) of amphetamine in urine was achieved with this system.

Mixed‐bed polymeric o‐nitrobenzophenone reagents for the on‐line derivatization of amines in high performance liquid chromatography

“…The mechanism of TEA catalysis has been dctermined to be a combination of base catalysis and elimination of ion pairing effects between the reagent intermediate and substrate. It has previously been established with this reagent that, under optimized conditions, a kinetic plot of percent derivatization vs. time asymptoted to a constant value after reaching 50% derivatization (Bourque and Krull, 1991a). At this point, the hydrogen ion generated during the final stage of the addition-elimination reaction was protonating unreacted amines, so that addition of TEA scavenged this proton, allowing the reaction to proceed.…”

“…Polymeric activated ester reagents for the derivatization of amine compounds to amides or carbamates have recently been developed for improved detection in high performance liquid chromatography (HPLC) (Bourque and Krull, 1991a;Chou et al, 1989;Gao and Krull, 1989a,b;Gao et al, 1989;Gao and Krull, 1990;Gao, et al, 1991). Because primary and secondary amines often show poor chromatographic performance and detectability, chemical derivatization is employed to enhance both chromatographic behaviour and sensitivity.…”

Determination of amphetamine in dog plasma by gas chromatography with mass selective detection

“…Various analytical methods have been employed for the monitoring of amphetamine in biological fluids. The commonly employed methods are high-performance liquid chromatography with fluorescence detection (Bowyer et al, 1995;Farrell and Jeffries, 1983;Foster et al, 1998;Bourque et al, 1994), capillary electrophoresis (Kuroda et al, 1998), immunoassay (Caplan et el., 1987;Cody and Schwarzhoff 1993), GC with nitrogen-phosphorus detection (Cheung et al, 1997;Koide et al, 1998), flame-ionization detection (Kintz et al, 1989), electron-capture detection (Roy et el., 1984;Paetsch et al, 1992), and MS (Valentine et al, 1995;Battu et al, 1998;Marquet et al, 1997;Tsai et al, 1998). The matrices commonly assayed for amphetamine are human whole blood (Sato and Mitsui, 1997;Gjerde et al, 1993), plasma (Pizarro et al, 1999), urine (Fisher and Bourque, 1993), cerebrospinal fluid (Narasimhachari et al, 1979), and hair (Kintz et al, 1989).…”

Studies on styrene concentration in drinking water and hot beverages in some settings

“…Modern high-performance liquid chromatography (HPLC) with UV detection (Bourque et al, 1994;Fujii et al, 1999;Inoue et al, 1991), gas chromatography (GC) with flame ionization detection (Chakroum et al, 2008;Kataoka et al, 1991), and high-performance chromatography with mass spectrometry (Pacenti et al, 2008;Marais and Laurens, 2005) were performed for evaluating styrene. The level of styrene present in food and drinks depend on many factors such as heat, pH, fats, and the time of storage.…”