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It has long been known that seminal plasma functions to transport and sustain motility of spermatozoa (19). Detrimental (7, 9, 10,13) or beneficial (3) effects of seminal plasma on sperm motility and viabil~y have been described. Likewise, seminal piasma contains factors that may influence male fertility. Comparisons of seminal plasma compos~ion between males of different fertility have been made, (17, 18), as well as the isolation of factors from seminal plasma that facilitate or inhibit sperm capacitation or fertilization, (8, 16). Since the adsorption of seminal plasma components could modify the spermatozoon surface, part~ion in aqueous two.phase systems is a useful technique to analyse seminal plasma interactions. This technique is based on the different cell surface affinity for inmiscible aqueous solutions of polymers, such as dextran and polyethylene glycol (PEG) (2). The upper phase is rich in PEG and is relatively more hydrophobic than the lower phase, which is rich in dextran. When cells are added to the system they partition between the interphase and the PEG-rich upper phase. The extent of partition is dependent on the cell surface properties. W~h populations of homogeneous cells repeated phase part~ioning (countercurrent distribution or CCD) is generally used to improved the resolution of the separation process. CCD is a chromatography process with one stationary (lower) phase and one mobile (upper) phase. The cell sample is partitioned in one system and the two phases are then, in a systematic way, brought into contact with fresh opposite phases. This separation method has already been used for studying mammalian sperm characteristics (5, 6, 14). However, the long period of time necessary for phase separation may increase cell death during the run. Shorter separation procedures can be carried out by using new equipment in which centrifugation speeds up the phase separation process (CCCD, for centrifugal countercurrent distribution) (1). Thus, we have shown that CCCD in aqueous two-phase systems is a resolutive technique to reveal spermatozoa heterogeneity (20, 22, 23) and that the acquis~ion of adsorbed seminal plasma proteins by the sperm cell surface modifies the partition behaviour of bull spermatozoa reproducing that of live cells (enhanced affin~y for the upper PEG-rich phase) (11, 21). The aim of this work was to characterize the effect of seminal plasma proteins on the distribution behaviour of fresh ram spermatozoa, as well as the effect of some seminal plasma fractions on ram sperm populations submitted to a freezing-thawing process.

Seminal plasma effect on ram spermatozoa studied by partitioning in an aqueous two-phase system.