The main aim of the present study was to investigate the plasma phospholipids (PL) fatty acids status and its association with plasma Hcy in patients with type 2 diabetes mellitus (T2DM). One hundred and four T2DM (aged 57.3±13.4 y) and 150 healthy subjects (aged 48.4±8.7 y) were recruited. Plasma Hcy and PL fatty acids were determined by standard methods. Plasma Hcy concentration in T2DM was significantly higher than that in healthy subjects (p<0.001). The prevalence of hyperhomocysteinemia was significantly higher in T2DM (36.54%) than that in healthy subjects (17.32%) (p=0.012). Plasma PL 20:4n-6 (r=0.303, p=0.012), 22:5n-3 (r=0.312, p=0.01), total PUFA (r=0.303, p=0.012), n-6 PUFA (r=0.261, p=0.032) were significantly positively associated with plasma Hcy concentration in T2DM. While, plasma PL n-3:n-6 PUFA (r=-0.400, p=0.046) was negatively associated with plasma Hcy in T2DM. In healthy subjects, plasma PL 22:6n-3 (r=-0.201, p=0.042) was negatively associated with plasma Hcy. In addition, plasma PL 22:6n-3 (r=0.193, p=0.044) and 22:5n-6 (r=0.234, p=0.038) were significantly negatively associated with plasma vitamin B-12 in healthy subjects. Our results suggested that increased plasma Hcy levels in T2DM associated with low n-3:n-6 ratio intake. We suggest that T2DM increase their long chain n-3 PUFA intake from fish or fish oil while decrease n-6 PUFA intake.