Lipase activity in stallion seminal plasma and the effect of lipase on stallion spermatozoa during storage at 5 °C

D. A. Carver, B. A. Ball

Research output: Contribution to journalArticlepeer-review

25 Scopus citations

Abstract

Previous studies have demonstrated a detrimental effect of seminal plasma on the maintenance of motility of cooled equine spermatozoa; however, the mechanism for the adverse effect of seminal plasma during cooled storage remains undetermined. In goats, a glycoprotein component of bulbourethral gland secretion contains lipase activity that is detrimental to sperm motility when stored in skim milk-based extenders. The objective of the current study was to determine the amount of lipase activity in stallion seminal plasma and to determine the effect of added lipase on spermatozoal motility during cooled semen storage. In the first experiment, seminal plasma (1.0 ml) was assayed for lipase activity based upon hydrolysis of triglycerides (olive oil substrate) into free fatty acids and subsequent titration of pH change (Sigma Diagnostic Lipase Kit). Lipase activity in stallion seminal plasma was 0.36 ± 0.02 Sigma units/ml, (mean + S.E.M.; n = 16 ejaculates from six stallions). In the second experiment, equine semen (three ejaculates from each of four stallions) was divided into five treatment aliquots. In Treatment 1, semen was extended 1:3 with nonfat dried skim milk extender (NFDSM). In treatment groups 2 through 5, spermatozoa were washed by centrifugation (300 × g for 15 min) and resuspended in NFDSM to a final concentration of 25 × 106 spermatozoa/ml. Porcine pancreatic lipase (pPL) was added to Treatment 3(10pPL units/ml), Treatment 4(100 pPL units/ml) and Treatment 5(100 pPL units/ ml,heat inactivated at 100 °C for 5 min), while Treatment 2 had no pancreatic lipase added and served as the control. Samples were cooled slowly to 5 °C, and stored at 5 °C until evaluation. Sperm motility was evaluated at time 0, 24, 48 and 72 h by computerized semen analysis, and data were analyzed viarepeated measures ANOVA. The addition of 100 units/ml but not 10 units/ml of pPL decreased (P < 0.01) total and progressive motility of stored sperm. Heat-inactivated pPL (Treatment 5) did not significantly decrease motility of spermatozoa during storage. Because the lipase activity assayed (Sigma units) and the lipase activity added to cooled semen (pPL units) were not equivalent, pPL was assayed in the Sigma Diagnostic Lipase assay. The relationship between Sigma Units (Y) and pPL units (X) appeared to be a log-linear relationship with log(Y) = -0.912 + 0.007X; R2 = 0.90. Mean lipase activity assayed in stallion seminal plasma was equivalent to approximately 64 pPL units/ml. These data suggest that endogenous lipase activity in stallion seminal plasma may be a factor in the adverse effects of seminal plasma on cooled spermatozoa in some stallions.

Original languageEnglish
Pages (from-to)1587-1595
Number of pages9
JournalTheriogenology
Volume58
Issue number8
DOIs
StatePublished - Nov 2002

Bibliographical note

Funding Information:
This research was supported by the John P. Hughes Endowment and by the Center for Equine Health with the funds provided by the Oak Tree Racing Association, the State of California pari-mutuel fund, and contributions by private donors. The authors thank Anthony Vo for technical assistance.

Keywords

  • Lipase
  • Seminal plasma
  • Spermatozoal motility
  • Stallion

ASJC Scopus subject areas

  • Small Animals
  • Food Animals
  • Animal Science and Zoology
  • Equine

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