Purification of glycosaminoglycans from bovine follicular fluid

The follicular glycosaminoglycans, dermatan sulfate and heparan sulfate, are not available in sufficient quantities for detailed analytical studies or to use as reagents in cell cultures. The purpose of this study was to purify follicular glycosaminoglycans simultaneously from bovine follicular fluid. The use of a chloroform:methanol extraction of 10 ml of fluid resulted in recoveries of at least 90% of the glycosaminoglycans, otherwise an insoluble product resulted. Heparan sulfate and dermatan sulfate eluted at .11 and .46 M NaCl, respectively, when subjected to HPLC with an anion exchange column. Fluid from small follicles allowed to "age" in the ovaries exhibited marked reductions in the concentrations of glycosaminoglycans. Aging of the fluid from large follicles resulted in greater glycosaminoglycan concentrations and appearance of glycosaminoglycans reflecting other charge distributions. Known amounts of purified follicular fluid heparan sulfate and dermatan sulfate were applied to a gel filtration HPLC column, and corresponding integrated peak areas were plotted in relation to the glycosaminoglycan concentrations. Slopes of the lines for commercial chondroitin sulfate and follicular dermatan sulfate were not significantly different, but the slope of the line for follicular heparan sulfate was 13.5-fold greater than the slope for commercial heparin.