This project was designed to study some biochemical constituents of ovarian follicular fluid (FF) and peripheral blood in buffaloes. For this purpose, ovaries were collected from adult buffaloes immediately after slaughter, FF was aspirated and stored at -4°C. Blood samples were also collected from these buffaloes before slaughter, plasma was separated and stored for further analysis. Samples were classified into two groups according to the follicle diameter i.e. small (3-10 mm) and large (11-20 mm). The FF and plasma samples were analyzed for various biochemical constituents, including glucose, cholesterol, total proteins, albumin, globulin, sodium, potassium, calcium, magnesium, zinc, triiodothyronine and thyroxine contents, using commercial kits.The results showed that small follicles had significantly lower (P<0.05) glucose contents than large follicles, while blood plasma had significantly higher (P<0.05) glucose contents than fluid from both classes of follicles. The differences in concentrations of cholesterol, total proteins, albumin and globulin between small and large follicles were non-significant. The concentrations of these compounds were higher in the blood than in FF, except albumin, which was higher in FF than in the blood. Contents of electrolytes and trace elements did not vary between the two follicle classes. However, the plasma levels of these electrolytes and trace elements, except potassium and zinc, were significantly higher (p<0.05) than their levels in FF. The level of potassium was significantly higher (P<0.05) in FF than in the plasma, while serum zinc level did not differ from FF. The differences in concentrations of triiodothyronine and thyroxine in fluid from small and large sized follicles were non-significant. Likewise, the levels of these hormones were non-significantly higher in blood plasma than in FF. It was concluded that FF levels of glucose differed between small and large follicle groups, while blood levels of most of constituents were higher than their levels in FF.