betadex and methylformamide

The cryopreservation of dromedary camel (Camelus dromedarius) sperm has proved challenging with little success reported. The routine application of artificial insemination with frozen semen would assist the flow of valuable genetic material nationally and internationally. The current study sought to examine the effects of cholesterol (cholesterol-loaded cyclodextrin [CLC]) preloading on camel sperm cryosurvival. Ejaculates (n = 3 males; 3 ejaculates per male) were collected using an artificial vagina during the breeding season and extended in HEPES-buffered Tyrode's albumin lactate pyruvate (TALP) and allowed to liquefy in the presence of papain (0.1 mg/mL) before removal of the seminal plasma by centrifugation. Sperm pellets were resuspended (120 million/mL) in fresh TALP and incubated (15 minutes; 37 °C) with 0, 1.5, or 4.5 mg CLC/mL. Sperm suspensions were then centrifuged and reconstituted in INRA-96 containing 20% (v:v) egg yolk and 2.5% (v:v) methylformamide, loaded in 0.5-mL plastic straws, sealed, and cooled for 20 minutes at 4 °C. Straws were frozen over liquid nitrogen (4 cm above liquid; 15 minutes), plunged, and stored. Sperm motility, forward progressive status, and acrosomal integrity were recorded at 0 and 3 hours after thawing and compared with these same parameters before freezing. Aliquots also were stained with chlortetracycline hydrochloride to assess spontaneous sperm capacitation status before freezing and post-thaw. Pretreatment with CLC (1.5 and 4.5 mg/mL) enhanced cryosurvival. Post-thaw sperm motility was highest (P < 0.05) in 1.5 mg CLC/mL immediately after thawing (44%) and after 3 hours incubation at room temperature (34%). Highest post-thaw sperm progressive status was also achieved in the presence of 1.5 CLC. Greater proportions of spermatozoa retained acrosomal membrane integrity when cryopreserved in the presence of CLC, but there was no difference between 1.5 and 4.5 CLC. Although thawed spermatozoa underwent spontaneous capacitation during in vitro incubation, cryopreservation and CLC treatment exerted no effect. In summary, dromedary camel sperm benefit from exposure to CLC before cryopreservation; this may facilitate the routine collection and storage of sperm from this species.

Topics: Acrosome; Animals; beta-Cyclodextrins; Camelus; Cholesterol; Cryopreservation; Cryoprotective Agents; Formamides; Hot Temperature; Male; Semen; Semen Preservation; Sperm Capacitation; Sperm Motility; Spermatozoa

The use of nonaqueous background electrolytes in capillary electrophoresis (CE) is a promising new trend which should widen the scope of this technique. We demonstrate the chiral separation of dansyl-amino acids (Dns-AAs) in N-methylformamide (NMF) using beta-cyclodextrin (beta-CD) as chiral selector. The solubility of beta-CD is much better in NMF than in water, allowing high concentration of the chiral selector and successful enantioseparation despite the weak host-guest interaction between the Dns-AAs and beta-CD. The association constants for the complexation between Dns-AAs and beta-CD could be calculated from the electrophoretic mobilities, with attention paid to the change in viscosity of the electrolyte upon addition of beta-CD. The association constants ranged between 2 and 13 M(-1).

Topics: Amino Acids; beta-Cyclodextrins; Cyclodextrins; Dansyl Compounds; Electrophoresis, Capillary; Formamides; Molecular Structure