A total of 82 postpartum buffalo cows were used in the present work. These animals were assigned into 5 random groups. The 1" one (n=15) was inseminated with frozen buffalo semen during their natural heat and used as a control group. Ovulation in the females of the 2nd group (n=16); 3rd group (n=17); 4'h group (n=19) and 5'h group (n=16) were synchronized by i.m injections of GnRH and PGF2α. They inseminated at 0 - 8, 9 - 16, 17 - 24 and 25 - 32 hours respectively. Milk progesterone profile was determined in all groups studied, starting from 0-time of insemination up to 28'h days post insemination. The highest pregnancy rates of 66.67 and 55.56% were recorded for the 1'' and 4'h groups respectively. Meanwhile, lower rates were observed for animal in the 2nd, 3rd and 5'h groups respectively. The rest of buffalo-cows did not observed in heat for 3 months post-partum and were considered sub-estrous as indicated by rectal palpation of ovarian follicle. Moreover, irrespective of animals groups and time of insemination the widening of the cervix at the time of insemination might affect the pregnancy rate. It was 56.52O/0, 42.85% and 6.45% in buffaloes with medium, low and high difficulty to pass the inseminating gun through cervix respectively. Regarding the level of progesterone hormone in milk, it was very low during estrous phase and very high on 21" to 28'h days post insemination in non-returned animals.