Objective: We analyzed the correlation between mutation in intron 4 and exon 7 of endothelial nitric oxide synthase (eNOS) and avascular necrosis of femoral head (ANFH). Method: A total of 260 ANFH cases without history of hip joint injuries were diagnosed and subject to staging according to Ficat standard, with 262 health subjects as control. Venous blood was collected to extract genome DNA, which was then amplified by PCR. The polymorphism of 27 bp repeat sequence in intron 4 and G894T polymorphism in exon 7 of eNOS gene was detected. Results: The b/b, b/a and a/a genotype frequency of intron 4 was 77.7%, 19.2% and 3.1% in ANFH group, respectively, and that in the control group was 58.0%, 32.8% and 9.2%, respectively. The b allele frequency in ANFH group was obviously higher than that in the control (P<0.0001). The frequency of 894 G/G wild type, G/T heterozygote and T/T homozygote in eNOS exon 7 was analyzed by PCR-RLFP: 65.4%, 26.5% and 8.1% in ANFH group, and 46.2%, 37.8% and 16% in normal control, respectively. The frequency of TT genotype in ANFH was obviously higher than that in the control group (P<0.001). Conclusion: Polymorphism of eNOS was correlated with ANFH.