QUANTITATIVE ANALYSIS OF PARACETAMOL IN HUMAN POSTMORTEM BLOOD & URINE VIA HPLC

Abstract

Paracetamol becomes as one of the most commonest drug that be a respondent for a large number of suicidal and poisonings due to its cheap, free availability, and can easily purchase without medical prescriptions. There were several cases received to the forensic toxicology laboratory, Government Analyst’s Department for qualitative and quantitative analysis of paracetamol in blood or urine due to bad circumstances. In this study a sensitive, rapid and selective methodology was developed for the qualitative and quantitative analysis of paracetamol levels in blood and urine samples by High Performance Liquid Chromatography (HPLC). A simple, selective single step liquid-liquid extraction method was developed to extract paracetamol from blood and urine. In this process, sample protein is successfully precipitated by dilution with a mixture of ethanol: ethyl acetate. Then it is centrifuged and clean extract is completely dried at 80◦C under N2 gas. HPLC analysis is carried out by reconstituted dried sample with methanol. The separation is achieved on C8 column (15cm x 4.6mm, 5µm), using an isocratic elution with acetonitrile/ methanol/ 0.1M potassium dihydrogen-orthophosphate/ water (20:10:5:65 v/v/v/v) mixture as the mobile phase. The detection was done by Photodiode Array Detector. For the quantification of paracetamol, Phenacetin was used as an internal standard. The HPLC method was validated for all the required validation parameters such as linearity, extraction efficiency, accuracy and precision. Calibration curve was obtained ranges of linearity, were set between 1-70 mg/L (correlation coefficient ≥ 0.99). The method limit of detection and limit of quantification were 0.25 mg/L and 1.5 mg/L, respectively. Blood and urine paracetamol recovery percentages were varied in between 78.2% - 78.9% and 83.5% - 85.4% respectively. The results obtained from postmortem samples, paracetamol was identified in the all test samples. The paracetamol amounts are found in the cases are ranged from 1.9 mg/L to 331 mg/L. Out of test samples, one sample is contained paracetamol in toxic level. These findings help to the judicial medical officers to make their judgments.