High-performance liquid chromatography assay for N-acetylcysteine in biological samples following derivatization with N-(1-pyrenyl)maleimide.
N-Acetylcysteine is a thiol antioxidant with expanding clinical importance. A sensitive, rapid method for determining reduced N-acetylcysteine (NAC) concentration in biological samples has been developed which uses a modified reversed-phase high-performance liquid chromatography (HPLC) technique in conjunction with the derivatizing agent N-(1-pyrenyl)maleimide (NPM). The NAC-NPM adduct was analyzed by HPLC with fluorescence detection. The calibration curve for NAC was linear over the range 8-2500 nM and the coefficient of variation obtained for the within-run precision and the between-run precision for 0.5 mM NAC was 1.5% and 2.7%, respectively. Relative recovery of NAC from biological materials ranged between 86% and 96% and the limit of quantitation from biological samples was 32 nM. These results suggest practical advantages relative to other widely-accepted methods of NAC measurement.
Ercal
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Department of Chemistry, University of Missouri, Rolla 65401, USA.
Oztezcan
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Hammond
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Matthews
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RH
Spitz
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DR
D000111:Acetylcysteine / Q000008:administration & dosage / Q000032:analysis* / Q000737:chemistry / Q000493:pharmacokinetics; D000818:Animals; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004355:Drug Stability; D016166:Free Radical Scavengers / Q000008:administration & dosage / Q000032:analysis* / Q000737:chemistry / Q000493:pharmacokinetics; D016014:Linear Models; D008297:Male; D008301:Maleimides / Q000737:chemistry*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013439:Sulfhydryl Reagents / Q000737:chemistry*; D014018:Tissue Distribution
D016166:Free Radical Scavengers; D008301:Maleimides; D013439:Sulfhydryl Reagents; C003944:N-(3-pyrene)maleimide; D000111:Acetylcysteine