High-performance liquid chromatography with electrochemical detection applied to the analysis of 3,4-dihydroxymethamphetamine in human plasma and urine.
Metabolic activation in the disposition of 3,4-methylenedioxymethamphetamine (MDMA, "ecstasy") has been implicated in some of its pharmacological and toxicological effects, with the major metabolite 3,4-dihydroxymethamphetamine (HHMA) as a putative toxicant through the formation of thioether adducts. We describe the first validated method for HHMA determination based on acid hydrolysis of plasma and urine samples, further extraction by a solid-phase strong cation-exchange resin (SCX, benzenesulfonic acid), and analysis of extracts by high-performance liquid chromatography with electrochemical detection. The chromatographic separation was performed in an n-butyl-silane (C4) column and the mobile phase was a mixture of 0.1 M sodium acetate containing 0.1 M 1-octanesulphonic acid and 4 mM EDTA (pH 3.1) and acetonitrile (82:18, v/v). Compounds were monitored with an electrochemical cell (working potentials 1 and 2, +0.05 and +0.35 V, respectively, gain 60 microA). A mobile phase conditioning cell with a potential set at +0.40 V was connected between the pumping system and the injector. Calibration curves were linear within the working concentration ranges of 50-1000 microg/L for urine and plasma. Limits of detection and quantification were 10.5 and 31.8 microg/L for urine and 9.2 and 28.2 microg/L for plasma. Recoveries for HHMA and DHBA (3,4-dihydroxybenzylamine, internal standard) were close to 50% for both biological matrices. Intermediate precision and inter-day accuracy were within 3.9-6.5% and 7.4-15.3% for urine and 5.0-10.8% and 9.2-13.4% for plasma.
Metabolism of racemic 3,4-methylenedioxyethylamphetamine in humans. Isolation, identification, quantification, and synthesis of urinary metabolites.
Studies on the isolation, identification, quantification, and synthesis of the urinary metabolites of racemic 3,4-methylenedioxyethylamphetamine (MDE) in humans are presented. After oral administration of 140 mg of racemic MDE to healthy volunteers, the following phase I metabolites could be isolated and identified by GC/MS: unchanged racemic MDE (I), racemic 3,4-dihydroxyethylamphetamine (II), racemic 4-hydroxy-3-methoxyethylamphetamine (IIIa), racemic 3,4-methylenedioxyamphetamine (IV), racemic 3,4-dihydroxyamphetamine (V), racemic 4-hydroxy-3-methoxyamphetamine (VIa), methylenedioxyphenylacetone (IXa), 3,4-methylenedioxyhippuric acid (X), and hydroxymethoxyhippuric acid (XII). The probable intermediate metabolite 3,4-dihydroxyhippuric acid (XI) could not be detected. Therefore, two overlapping phase I metabolic pathways for racemic MDE in humans could be postulated. The first and predominant pathway leads, via ring degradation by O-dealkylation, to the corresponding 3,4-dihydroxy metabolites, which are subsequently methylated at the hydroxyl group at position 3 of the aromatic ring. The second pathway leads, via side chain degradation by N-dealkylation, to the corresponding primary amines (IV, V, and VI). Oxidative N-deamination forms the substituted phenylacetones, which are degraded to the corresponding benzoic acids. This is followed by conjugation with glycine to form substituted hippurates. The structures of all of these metabolites were confirmed by chemical syntheses, which are described in this paper. All of the metabolites containing hydroxy groups are partly excreted in a conjugated form, because the amounts of these metabolites were much higher in urine extracts after enzymatic cleavage of conjugates. Quantification of the urinary excretion by HPLC revealed that 19% of the MDE dose was eliminated as I, 31.6% as IIIa, and 2.8% as IV within 32 hr.
Chip scale packaging using chip-on-flex technology
An increasing number of electronic designers, fabricators and users see Chip Scale Packaging (CSP) as a way to obtain the benefits foreseen in multichip packaging and Chip-on-Board (COB) without the problems and limitations currently associated with each. The Chip-on-Flex (COF) multichip packaging technology has been demonstrated to be applicable to single chip packages that meet chip scale packaging goals. This paper looks at the Chip-on-Flex Chip Scale technology and addresses issues including the process, structure, assembly, yields and reliability.