Measurement of ethanol in microdialysis samples by means of enzymatic assay using alcohol dehydrogenase and NAD

• Main Author: Ghoncheh, Shahin
• Format: Others
• Language: English
• Published: Högskolan i Borås, Institutionen Ingenjörshögskolan 2008
• Subjects: alcohol dehydrogenase; ethanol; four-parameter logistic model; microdialysis; blood flow changes; Engineering and Technology; Teknik och teknologier
• Online Access: http://urn.kb.se/resolve?urn=urn:nbn:se:hb:diva-18812

The enzymatic method for ethanol measurement can detect very low concentration of ethanol at samples, consequently it can’t be applied for samples with high concentration and implies as very sensitive method at limited range of detection.The alcohol dehydrogenase method is based on oxidation of alcohol in the presence of ADH as enzyme and NAD+ as coenzyme and formation of acetaldehyde and NADH that can be monitored by spectrophotometric measurement at 334,340 or 365 nm wavelengths.Ethanol +NAD+ ADH↔ Acetaldehyde +NADH+H+For optimum conditions of measurements all the parameters that affect the enzymatic reaction including temperature, pH, trapping agent for product and proper mixing need to be optimized.In order to calculate the unknown concentration of ethanol in a sample based on this method,it is crucial to find right mathematical model to calculate the unknown concentrations of ethanol in the sample using a mathematical equation that generalizes relationships among the reactants in the reaction including the reaction products. In most enzymatic reactions many parameters are involved meaning that the reaction seldom follows simple linear relation between concentration and signal. Four-parameter logistic model is well suited for modeling sigmoid relationships frequently found in biology.The aim of this project is determination of ethanol at microdialysis samples and the fundamental reason for developing the present measurement method was to study changes in blood flow in living tissues using wash out of the very dissolvable ethanol as a flow marker using the Microdialysis technique.Result from this measurement technique for microdialysis samples shows that ethanol can be detected at range of 0,5-16mmol/L and whole detected concentration for different samples during one microdialysis test follows the inverse relation of blood flow changes in tissue.Also the reported result from Urea test as general method for studying blood flow changes and ethanol test for microdialysis sample has been compared and leads to this conclusion that ethanol techniques is as reliable tool for studying blood flow changes. === Uppsatsnivå: D