Sorbitol metabolism in spermatozoa

Abstract

Sorbitol was identified as a chemical constituent of the seminal plasma in the ram, rabbit, bull, boar, stallion, cock and man. Its concentration was determined by an enzymic-spectrophotometric method based on the use of purified sorbitol dehydrogenase from liver. Ram spermatozoa oxidize sorbitol to fructose, and they are also capable of reducing fructose, and to a smaller extent glucose, to sorbitol. The oxidative conversion of sorbitol to fructose makes it probable that sorbitol contributes to the lactic acid which accumulates in semen incubated i n vitro . Triphenyl tetrazonium chloride was used as an electron acceptor instead of molecular oxygen, to study the oxidative behavior of ram spermatozoa towards sorbitol, fructose, and glucose. 'Cold shock’ treatment, which rapidly immobilizes spermatozoa, produced little or no effect on the rate of triphenyl tetrazonium chloride reduction, either in the absence or in the presence of sorbitol, fructose, or glucose. Prolonged aerobic incubation of a washed sperm suspension which depletes the spermatozoa of ‘endogenous’ substrate, did not abolish the ability of the‘exhausted’ sperm cells to reduce triphenyl tetrazonium chloride in the presence of sorbitol, fructose, or glucose. Sorbitol dehydrogenase, the enzyme responsible for the reversible oxidoreduction of sorbitol to fructose in ram spermatozoa, was prepared in a soluble and highly active form, by grinding sperm with lavigated aluminium oxide and extraction with phosphate buffer. The soluble enzyme preparation requires diphosphopyridine nucleotide ( DPN ) as coenzyme, and catalyzes the reaction. sorbitol + DPN+ ⇌ fructose + DPNH + H + . The oxidizing activity of sperm sorbitol dehydrogenase is highest towards D-sorbitol and L-iditol, and much lower towards D-xylitol; D-ribitol, D-mannitol, and D-arabitol are only poorly oxidized. In the reverse reaction, which involves the reduction of sugars to polyols, only fructose, and to a smaller extent, sorbose, are reduced. The Michaelis constants ( K m ) obtained were 9.8 x 10 -3 for serbitol, and 9.0 x 10 -3 M for iditol. In intact spermatozoa the steady state of the reversible enzymic conversion of sorbitol to fructose depends upon the actual ratio between oxidized and reduced DPN . It is suggested that upon the respective levels of fructose and sorbitol in seminal plasma may depend the ratio between the oxidized and reduced form of DPN within the sperm cells themselves.