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Production of superoxide anion and hydrogen peroxide by capacitating buffalo ( Bubalus bubalis ) spermatozoa

Animal Reproduction Science, ISSN: 0378-4320, Vol: 103, Issue: 3, Page: 260-270
2008
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Article Description

In the present study attempts were made to detect and quantify the generation of superoxide anion (O 2 − ) and hydrogen peroxide (H 2 O 2 ) by capacitating buffalo spermatozoa. Ejaculated buffalo spermatozoa were suspended in sp-TALP medium at 50 × 10 6 mL −1 and incubated at 38.5 °C with 5% CO 2 in air in the absence or presence of heparin (a capacitation inducer) or reduced nicotinamide adenine dinucleotide phosphate (NADPH) or diphenyleneiodonium (DPI, a flavoprotein inhibitor) for 6 h. Production rate of O 2 − and H 2 O 2 by spermatozoa at different hours of capacitation were measured by cytochrome c reduction and phenol red oxidation assays, respectively. Spermatozoa generated both O 2 − and H 2 O 2 spontaneously and following stimulation with heparin and a significant increase of O 2 − production was observed in the presence of NADPH. However, DPI inhibited this NADPH-induced O 2 − production and suggested for existence of putative NADPH-oxidase that constitute a specific O 2 − generating systems in buffalo spermatozoa. Results of our study indicated that buffalo spermatozoa generate O 2 − and H 2 O 2 and production of these free radicals is induced during capacitation.

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