Diet and lifestyle play an important role in etiology of various tumors. Serum concentration of folate and vitamin B12may be associated with carcinogenesis since they are involved in DNA methylation and nucleotide synthesis. However, the role of these micronutrients on development of cervical cancer is still controversial. Thus, the aim of this study was to analyze the association of lower status of folate and vitamin B12 with the risk of pre-neoplastic cervical lesions. Our sample group was divided in Control group (n=120) - women with normal cytology, and Case groups (n=57) - women presenting Atypical Squamous Cells of Undetermined Significance (ASC-US, n=21), Low Grade Squamous Intraepithelial Lesion (LSIL; n=16), and High-Grade lesions (n=20). We obtained cervical samples for cytology analysis and HPV detection, and blood samples for evaluation of serum concentration of folate and vitamin B12. No difference of serum folate was observed among Cases and Control groups. On the other hand, women with High-Grade lesions presented significant lower median concentration of vitamin B12 if compared to another groups. Then, we observed increased risk of High-Grade lesions among participants with low vitamin B12 levels was observed in relation to women that presented high levels of the micronutrient and from Control group [OR (95% CI): 2.09 (0.65-6.76), p=0.216], ASC-US [OR (95% CI): 3.15 (0.82-12.08), p=0.095], and LSIL [OR (95% CI): 3.10 (0.76-12.70), p=0.116]. Low concentration of vitamin B12 was associated with an increased risk of High-Grade cervical lesions. Besides, we did not observe any difference of serum folate among women with normal cytology and women with pre-neoplastic cervical lesions.