An Automatic Flow Procedure for the Determination of 3-Hydroxybutyrate in Animal Serum and Plasma

An automatic flow procedure based on the multicommutation concept, comprising three-way solenoid valves, for the spectrophotometric determination of 3-hydroxybutyrate in animal serum and plasma is proposed. The 3-hydroxybutyrate was enzymatically converted to acetoacetate with the reduction of NAD+ to NADH monitored at 340 nm. It was possible to carry out up to 600 determinations without a significant decrease in the analytical signal, with 5 mg of 3-hydroxybutyrate dehydrogenase immobilized on porous silica beads and packed in a column. The system enabled 60 determinations/h of 3-hydroxybutyrate in the range of 10−150 mg L-1, with a consumption of 0.9 mg of NAD+ and 200 μL of sample per determination. A detection limit of 2 mg L-1 for both animal serum and plasma and coefficients of variation of 1.4% and 1.2% (n = 17), respectively, were determined. Animal serum and plasma samples were analyzed without previous treatment, the results of which agreed with those obtained using the conventional method (UV kit, Sigma). Keywords: Flow analysis; multicommutation; 3-hydroxybutyrate; spectrophotometry; enzyme reactor; plasma and serum