A luminometric assay for determination of ethanol in microdialysates

In microdialysis, samples from the extracellular fluid are analysed in the outgoing dialysate. By adding ethanol to the ingoing perfusate and following its exchange in the dialysate, an outflow/inflow ratio is obtained. This ratio has been shown to correlate with the tissue blood flow. An automatic luminometric ethanol assay that quantifies the light produced from three coupled enzyme reactions is described. Alcohol dehydrogenase catalyses the oxidation of ethanol with formation of NADH, which is monitored using NADH : FMN oxido­reductase and bacterial luciferase. Each assay can be individually calibrated by measuring the increased rate of NADH formation through the addition of a known amount of ethanol. The linear range of the assay was 0.05 - 10 mmol l-1 in microdialysate samples. The within-run imprecision was 1.4% CV in 10 mmol l-1 samples, and ethanol recovery was almost 100%. The assay was applied to microdialysates that were generated from probes implanted in the vastus lateralis muscle in fasting ...