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Studies were carried out to identify and quantify the porcine follicular fluid (PFF) component(s) responsible for inhibition of murine oocyte maturation. A low molecular weight fraction of PFF (less than 1000) was prepared by dialysis and used in all experiments. This PFF fraction contained an inhibitor(s) of mouse oocyte maturation that absorbed maximally in the ultraviolet (UV) range at 250-260 nm. When the PFF fraction was charcoal-extracted, significant loss of absorbance at 250, 260, and 280 nm resulted, which corresponded to loss of inhibitory activity. Four major components of PFF were separated by ion-exchange chromatography and characterized according to their UV spectral characteristics and inhibitory activity. When individual fractions demonstrating identical spectra were pooled and analyzed by high-performance liquid chromatography, the first pooled fraction (A) was found to be impure, but adenine comprised 80% of the UV-absorbing material. Fractions B, C, and D were characterized as pure uracil, hypoxanthine, and 7-methylinosine, respectively. The concentrations of these compounds in PFF were estimated to be 0.06 mM adenine, 0.44 mM uracil, 1.41 mM hypoxanthine, and 0.19 mM 7-methylinosine. Comparison of the potencies of commercial preparations of these compounds established that hypoxanthine is the major inhibitory component of the low molecular weight PFF fraction. Moreover, a commercial preparation of hypoxanthine mimicked the action of PFF on mouse oocyte maturation in that it produced a transient inhibition of oocyte maturation that was significantly potentiated by follicle-stimulating hormone and dibutyryl cyclic adenosine monophosphate. When the inhibitory efficacies of purine and pyrimidine bases and nucleosides were compared, their relative potencies in decreasing order were purine bases greater than purine nucleosides greater than pyrimidine bases = pyrimidine nucleosides. We conclude that hypoxanthine is the predominant low molecular weight component of PFF that inhibits mouse oocyte maturation but that other purines/pyrimidines may also play a role in vivo in maintaining meiotic arrest.

Hypoxanthine is the principal inhibitor of murine oocyte maturation in a low molecular weight fraction of porcine follicular fluid.