The objective of this study is to investigate and compare the prevalence, knowledge, and attitudes of Chinese university students with respect to nutritional supplements. We conducted a cross-sectional study in several universities around China from January to December 2017, and enrolled a total of 8752 students. Of these, 4252 were medical students and 4500 were non-medical students. The use of nutritional supplements was reported by 58.9% in universities students, with a higher rate for medical students as compared to non-medical students. It was found 24.2% of participants had taken supplements in the past year. Medical students had a higher level of knowledge on nutritional supplements than non-medical students (p < 0.001). The most commonly used nutritional supplements were vitamin C, calcium, and vitamin B. Gender (p < 0.001), household income (p < 0.001), and health status (p < 0.001) were related to the nutritional supplement use after adjustment for related factors. In conclusion, in China, nutritional supplement use was found to be more common in medical students than those studying other disciplines, and was associated with sex, income, and health status. The attitude towards nutritional supplements by medical students was positive. Students' knowledge levels about nutritional supplements need to be improved.