Estimation of postmortem interval by hypoxanthine and potassium evaluation in vitreous humor with a sequential injection system |
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Authors: | Marieta LC Passos Ana I Pereira Agostinho JC Santos M Lúcia MFS Saraiva José LFC Lima |
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Institution: | a REQUIMTE, Serviço de Química-Física, Faculdade de Farmácia, Universidade do Porto, Rua Aníbal Cunha, 164, 4099-030 Porto, Portugal b Serviço de Medicina Legal, Faculdade de Medicina, Universidade do Porto, Portugal c Serviço de Patologia Forense, Instituto Nacional de Medicina Legal, I.P. - Delegação Norte, Portugal |
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Abstract: | The estimation of the time since death known as postmortem interval (PMI) is a main issue in the field of forensic science and legal medicine. In this work it is proposed a sequential injection system for the determination of hypoxanthine and potassium in the same sample of vitreous humor since the concentrations of both parameters change with PMI and the vitreous humor has been regarded as the ideal extracellular fluid for these kinds of determinations. By measuring both parameters the accuracy of estimation of PMI can be increased, and the effects of factors which influence the values in postmortem chemistry minimized.Hypoxanthine determination is based on its oxidation to uric acid (290 nm), catalyzed by immobilized xanthine oxidase, and the quantification of potassium levels in vitreous humor was performed using a tubular potassium ion-selective electrode. With a unique analytical cycle both analytes were evaluated being potassium levels determined during the degradation of hypoxanthine in the enzymatic reactor.Working concentration ranges between 6.04-40.00 μmol L−1 and 7.00 × 10−5 to 1.00 × 10−1 mmol L−1 were obtained, for hypoxanthine and potassium, respectively.The method proved to be reproducible with R.S.D. <5% for hypoxanthine and <3% for potassium. Sampling rate was approximately 30 per hour for the sequential determination of both parameters being 15 and 60 determinations per hour if hypoxanthine or potassium, where evaluated independently. Statistical evaluation at the 95% confidence level showed good agreement between the results obtained, for the vitreous humor samples, with both the SIA system and the comparison batch procedures. Moreover the methodology has low environmental impact in agreement with the demands of green analytical chemistry as only 2.7 mL of chemical waste is produced during both determinations. |
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Keywords: | Sequential injection analysis Hypoxanthine Xanthine oxidase Potassium tubular electrode Vitreous humor Postmortem interval |
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