Folate deficiency has been confirmed to be related to various diseases. Unfortunately, there are few reports on the folate status of Chinese adults. This study aims to evaluate the serum folate status of blood donors in south-central China. In this study, 248 blood donors were included. The information on subjects was collected by a brief questionnaire concerning alcohol consumption habits, smoking habits, fruit and vegetable consumption and physical activity. The serum folate concentration was measured by electrochemiluminescence immunoassay. The geometric mean serum folate concentration was 13.4 nmol l-1 (95% CI, 12.7–14.1). The prevalence of serum folate concentrations below 6.8 nmol l-1 was 5.2% (95% CI, 2.5–8.0). There were significant differences in serum folate concentrations with respect to sex (p-values < 0.05), age (p-values < 0.05), fruit and vegetable consumption (p-values < 0.05), and alcohol consumption habits (p-values < 0.05). The concentration of serum folate increased with age (p-values < 0.05) and fruit and vegetable consumption (p-values < 0.05). Individuals with an age of 30 years or younger were nearly 3.5 times as likely as those aged over 30 years to have an insu±cient level of serum folate (OR = 3.48; 95% CI: 1.01–11.99). An age of 30 years or younger was a risk factor for folate deficiency. Most blood donors had su±cient serum folate concentrations in south-central China. National surveys of folate status should be implemented in China.