NDLI: Simple Reversed-Phase HPLC Method with Spectrophotometric Detection for Measuring Acetaminophen-Protein Adducts in Rat Liver Samples

Content Provider	PubMed Central
Author	Acharya, Miteshkumar Lau-cam, Cesar A.
Copyright Year	2012
Abstract	A simple reversed-phase HPLC method for measuring hepatic levels of acetaminophen- (APAP-) protein adduct following an overdose of APAP was developed. An aliquot of liver homogenate in phosphate-buffered saline pH 7.4 (PBS) was placed on a Nanosep centrifugal device, which was centrifuged to obtain a protein residue. This residue was incubated with a solution of p-aminobenzoic acid (PABA), the internal standard, and bacterial protease in PBS, transferred to a Nanosep centrifugal device, and centrifuged. A 100 μ L portion of the filtrate was analyzed on a YMC-Pack ODS-AMQ C18 column, using 100 mM potassium dihydrogen phosphate-methanol-acetic acid (100 : 0.6 : 0.1) as the mobile phase, a flow rate of 1 mL/min, and photometric detection at 254 nm. PABA and APAP-cystein-S-yl (APAP-Cys) eluted at ~14.7 min and 22.7 min, respectively. Method linearity, based on on-column concentrations of APAP-Cys, was observed over the range 0.078–40 μ g. Recoveries of APAP-Cys from spiked blank liver homogenates ranged from ~83% to 91%. Limits of detection and of quantification of APAP-Cys, based on column concentrations, were 0.06 μ g and 0.14 μ g, respectively. RSD values for interday and intraday analyses of a blank liver homogenate spiked with APAP-Cyst at three levels were, in all cases, ≤1.0% and <1.5%, respectively. The proposed method was found appropriate for comparing the antidotal properties of N-acetylcysteine and taurine in a rat model of APAP poisoning.
Related Links	http://dx.doi.org/10.1100/2012/145651
Starting Page	145651
File Format	PDF
ISSN	1537744X
e-ISSN	1537744X
Journal	The Scientific World Journal
Volume Number	2012
Language	English
Publisher	The Scientific World Journal
Publisher Date	2012-01-01
Access Restriction	Open
Rights Holder	The Scientific World Journal
Subject Keyword	Biochemistry, Genetics and Molecular Biology(all) Environmental Science(all) Medicine(all) Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Medicine Biochemistry, Genetics and Molecular Biology Environmental Science

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