Vitamin B12 and folate deficiency are not only linked to hematological, neurological and cardiovascular diseases, but also associate with insulin resistance. Metformin can decrease vitamin B12 and folate concentrations.To examine (i) effects of short-term metformin treatment on serum holotranscobalamin (holoTC) and folate and (ii) their association with insulin sensitivity in recent-onset type 2 diabetes.This cross-sectional analysis comprised patients (known disease duration <12 months) on metformin monotherapy (MET, n=123, 81 males, 53±12 years) or non-pharmacological treatment (NPT, n=126, 77 males, 54±11 years) of the German Diabetes Study.HoloTC (ELISA), cobalamin and folate (electrochemiluminescence); beta-cell function and whole-body insulin sensitivity, measured during fasting (HOMA-B, HOMA-IR) and intravenous glucose tolerance tests (IVGTT) combined with hyperinsulinemic-euglycemic clamp tests.HoloTC (105.4 [82.4, 128.3] vs. 97 [79.7, 121.9] pmol/L) and folate concentrations (13.4 [9.3, 19.3] vs. 12.7 [9.3, 22.0] nmol/L) were similar in both groups. Overall, holoTC neither associated with fasting nor glucose-stimulated beta-cell function and insulin-stimulated glucose disposal. Cobalamin measurements yielded similar results in representative subgroups. In NPT but not MET, folate levels were inversely correlated with HOMA-IR (r=-0.239, P=0.007). Folate levels neither related to insulin sensitivity nor insulin secretion in the whole cohort and in each group separately after adjustment for age, BMI and sex.Metformin does not affect circulating holoTC and folate concentrations in recent-onset type 2 diabetes, rendering monitoring of vitamin B12 and folate dispensable at least during the first 6 months after diagnosis or initiation of metformin.