HPLC Method for Quantification of Oxidative Stress by Salicilate Hydroxylation in Human Plasma

• Published in: Journal of chromatographic science Vol. 48; no. 8; pp. 675 - 679
• Main Authors: Lazalde-Ramos, Blanca P., Lares-Asseff, Ismael, Villanueva-Fierro, Ignacio, Sosa-Macías, Martha, Yahuaca-Mendoza, Patricia, Zamora-Perez, Ana
• Format: Journal Article
• Language: English
• Published: Niles, IL Oxford University Press 01.09.2010; Preston Publications
• Subjects: Biological and medical sciences; Biomarkers - blood; Catechols - blood; Chromatography, High Pressure Liquid - methods; Drug Stability; Gentisates - blood; Humans; Hydroxybenzoates - analysis; Hydroxylation; Investigative techniques, diagnostic techniques (general aspects); Linear Models; Medical sciences; Miscellaneous. Technology; Oxidative Stress; Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques; Reproducibility of Results; Salicylates - chemistry; Salicylates - metabolism; Sensitivity and Specificity; Human; Validation; Salicylic acid; Biological fluid; Oxidative stress; Chemical analysis; Hydroxylation; HPLC chromatography; Biological marker; Blood; Electrochemical detector; Blood plasma; Non steroidal antiinflammatory agent; Reversed phase chromatography; Clinical biology; Salicylates; Quantitative analysis
• ISSN: 0021-9665; 1945-239X
• DOI: 10.1093/chromsci/48.8.675

The aim of the present study was to modify and validate a high-performance liquid chromatographic (HPLC) method for determining 2,3 and 2,5 dihydroxybenzoic acid (2,3-DHBA and 2,5-DHBA) from salicylic acid in human plasma. The mobile phase was a mixture of sodium acetate/citrate (pH 2.5) 30 mM-methanol (93:7, v/v). The injection volume was 10 µL. Retention time for 2,5- DHBA, and 2,3-DHBA was 4.5 ± 0.10 and 5.8 ± 0.15 min, respectively. The detection and quantification limits were 10 and 40 nM for 2,3-DHBA and 8 and 20 nM for 2,5-DHBA. Linearity was evaluated in the range of 40–1600 nM for both metabolites. Inter-and intra-analysis variation coefficient was below 10%. Good recoveries of more than 99% were obtained for both metabolites using this method.