[38] Aldose reductase from seminal vesicle and placenta of ruminants

This chapter describes the assay method, purification procedure, and properties of aldose reductase from seminal vesicle and placenta of ruminants. Glucose is reduced to sorbitol in the presence of ruminant placental or seminal vesicle aldose reductase and reduced pyridine nucleotide at pH 7.5. The reaction rate is, however, higher using glyceraldehydes or lactaldehyde as substrate. Although the equilibrium of the reaction strongly favors the reduction of the aldehyde, the oxidation reaction takes place when the reaction is carried out at pH 10 using the corresponding alcohols as substrate. For practical purposes, DL-glyceraldehyde is used as substrate in the standard assay system, and the reaction is measured spectrophotometrically by following the decrease in absorption at 340 nm caused by the formation of NADP+. All operations of purification procedure are carried out at 0–4°. Fractionations with ammonium sulfate are made by slow addition, with stirring of the calculated amount of solid ammonium sulfate. The resulting suspensions are kept for 30 min before centrifugation. In the purified form both the placental and seminal vesicle enzymes are NADP specific. The most reactive substrates are lactaldehyde and glyceraldehyde. Both enzymes are sensitive to heavy metal ions. Sulfate ions cause a marked stimulation of the enzyme when glyceraldehyde is used as substrafe, while inhibition by sulfate is observed when the substrate is propanediol. Other properties are also discussed.