Objective: To reveal the relationship between physical fitness and body fat percentage (BFP) among female college students. Methods: During March 2017, BFP and fat free mass were measured among 633 randomly selected healthy female college students. All subjects were divided into five groups based on BFP: low weight, normal weight, high body fat, grade I obesity and grade II/III obesity. Grip strength, leg strength, back strength, vital capacity, maximal oxygen uptake, reaction time, sitandreach, back scratch and standing on one foot with closed eyes were tested. Results: There were significant differences between all groups in grip strength, back strength, vital capacity, the absolute and relative value of VO2max, back scratch on both sides and standing on one foot with closed eyes(P<0.05). Grade I obesity group showed significant higher vital capacity, but lower relative value of VO2max, back scratch in both sides compared with normal group. Significant differences were observed in relative value of maximum oxygen uptake and back scratch in both sides between high body fat group and grade I obesity(P<0.05). Moreover, grade II/III obesity group showed significantly higher grip strength, back strength, vital capacity and absolute VO2max, but lower relative VO2max, back scratch in both sides and standing on one foot with closed eyes compared with normal and high body fat groups(P<0.05). There were significant differences in all the indicators except back strength and the relative VO2max between grade II/III obesity and grade I obesity group(P<0.05). Conclusion The results showed that physical fitness of female college students with normal body fat percentage is significantly better than that of obese peers. Body fat percentage confers damage for cardiopulmonary endurance fitness and upper limb flexibility fitness when higher than 30% and for balance fitness when higher than 35%.