The study aimed to compare the acid-base balance and steroid concentrations between follicular fluids (FF) of preovulatory follicles derived from a spontaneous (SE), synchronized estrus (IE) and follicular cysts (CYS) and between FF and blood in dairy cows. Forty two dairy cows were included in this study. The animals were allocated into three groups, SE (n = 23), IE (n = 11) using GnRH at Day 0 and Day 9, PGF2α at Day 7 and the animals with CYS (n = 10). The follicular fluids (FF) were aspirated from the cyst / preovulatory ( ≥ 15mm) after SE and after second GnRH dose in IE by transvaginal ultrasound-guided ovum-pick-up technique. Blood samples (BL) were collected in herparinized vacutaner tubes. The oxygen tension (pO2) in FF of IE was higher (P  0.05) than in SE and CYS groups. There were negative correlations (P  0.001, r = -0.89) between FF and blood pO2. The carbon dioxide tension (pCO2) and lactate in FF of CYS was higher (P  0.05) than in SE and IE groups. There were negative correlations (P  0.01, r = -0.73) between blood and FF pO2. Estradiol-17β concentration in preovulatory follicles and plasma of the SE was higher (P < 0.001) than in IE and cyst groups. Progesterone concentration in preovulatory follicles and plasma of the SE and IE was lower (P < 0.01) than in cyst group. Plasma androstendione concentration in SE and IE was higher (P < 0.05) than that in cyst group. In conclusion, acid base parameters, E2 and P4 levels in the follicular fluid of both synchronized estrus and cyst were deviated greatly from the physiological naviou (disturbances of intrafollicular/intracystical environment) which may affect the quality of both the oocyte and the granulosa cells.