This study was performed to determine the relationship between the corpus luteum weight/diameter, follicular diameter and the levels of vitamin C, progesterone and oestradiol 17-β of the plasma, corpus luteum and follicular fluid of pregnant Holstein cows in a slaughterhouse. Progesterone and oestradiol 17-β concentrations were determined by the microtitration plate EIA method. Vitamin C levels were determined by spectrophotometer. Although there was a positive correlation between follicular fluid oestradiol 17-β levels and the follicle diameter in the follicular fluid, in contrast to oestradiol 17-β levels, small follicles have higher progesterone content than large follicles. This negative correlation was due to the fact that progesterone is the precursor of oestradiol 17-β. There was no correlation between corpus luteum progesterone levels and corpus luteum weight/diameter. However, a positive correlation between corpus luteum progesterone levels and the corpus luteum weight/diameter was observed; therefore, we suggest that this situation resulted from placenta-originated progesterone synthesis in pregnancy. Furthermore, levels of corpus luteum vitamin C were found to be higher and we concluded that this increase resulted from the effect of vitamin C on steroid and collagen biosynthesis. Nevertheless, negative correlations between corpus luteum vitamin C concentrations and corpus luteum weight/diameter were seen, and hence we suggest that this may be the result of the inhibitory effect of vitamin C on PGF2α synthesis.