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Poster Sessions AP13: Novel Techniques and Technologies. A simple and sensitive HPLC‐method for simultaneous monitoring of oxidative stress indices and monoamine metabolites
Author(s) -
Yao J. K.,
Cheng P.
Publication year - 2002
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.81.s1.13_1.x
Subject(s) - chemistry , chromatography , coulometry , high performance liquid chromatography , elution , sample preparation , monoamine neurotransmitter , analytical chemistry (journal) , electrode , biochemistry , receptor , serotonin , electrochemistry
Studies of the antioxidant defense system and the monoamine metabolic pathways are often complicated by cumbersome analytical methods, which require separate and multistep extraction and chemical reaction procedures. Thus, measurements of multiple parameters are limited in relatively small biological samples. High performance liquid chromatography (HPLC) coupled with a Coulometric Multi‐Electrode Array System (CMEAS) provides us a convenient and most sensitive tool to measure low molecular weight, redox‐active compounds in biological sample. The deproteinized sample was analyzed on a HPLC coupled with a 16‐channel CMEAS, which incremented from 60 to 960 mV in 60 mV steps. Each sample was run on a single column (Meta‐250, 4.6 × 250 mm) under a 150‐minute complex gradient that ranged from 0% B (A: 1.1% pentane sulfonic acid) to 20% B (B: 0.1 m lithium acetate in mixture of methanol, acetonenitrile and isopropanol), with a flow rate of 0.5 mL/min. We have developed an automated procedure to simultaneously measure various antioxidant, oxidative stress marker, and monoamine metabolites in a single column with binary gradient. No other chemical reactions are necessary. In order to reduce the running time and yet achieve a reproducible retention time by the autosampler injection, our gradient elution profile was modified to produce a shorter equilibration time and to compensate for the initial contamination of mobile phase B following the first injection. Without the use of two columns in series and peak suppresser/gradient mixer, we have simplified the previously published method to measure over 20 different antioxidants, oxidative stress markers and monoamine metabolites simultaneously in biological samples.