Safety evaluation of calcium L-methylfolate

Abstract

Calcium L-methylfolate (L-5-MTHF-Ca; CAS Number 151533-22-1) is a source of folate and an alternative to folic acid for use in human food and food supplements. The safety of L-5-MTHF-Ca was evaluated by testing for genotoxicity, subchronic and prenatal developmental toxicity. In in vitro assays L-5-MTHF-Ca was not mutagenic and did not induce other chromosomal events. Additionally, L-5-MTHF-Ca was not genotoxic in the in vivo micronucleus test nor did it induce DNA damage in rat liver cells. In a subchronic toxicity study, rats administered up to 400 mg/kg bw/day of L-5-MTHF-Ca via oral gavage for 13 weeks had no treatment-related mortalities, and no treatment-related effects were identified on behaviour, body weight, food consumption, ophthalmology, haematology, or organ weights. No treatment-related macroscopic or histopathological findings were observed. Calcium and sodium levels increased with increasing dosage, however the slight increases were within historical control ranges and reversible after the recovery period. L-5-MTHF-Ca is neither teratogenic nor embryotoxic. Based on the results of the in vitro and in vivo studies, the safe use of L-5-MTHF-Ca as an ingredient in foods is supported. The no observed adverse effect level was the highest dose in the subchronic toxicity study, i.e. 400 mg/kg bw/day for male and female rats.

Keywords: 5-MTHF, 5-methyltetrahydrofolate; ANOVA, analysis of variance; BaP, benzo[a]pyrene; Calcium L-methylfolate; Developmental toxicity; EFSA, European Food Safety Authority; GD, gestation day; GLP, Good Laboratory Practice; GRAS, generally recognized as safe; Genotoxicity; HPLC, High Performance Liquid Chromatography; JECFA, Joint FAO/WHO Expert Committee on Food Additives; L-5-MTHF-Ca; L-5-MTHF-Ca, calcium L-methylfolate; MTT, 3-[45-dimethylthiazole-2-yl]-2,5-diphenylbromide; NNG, net grains/nucleus; NOAEL, No Observed Adverse Effect Level; OECD, Organisation for Economic Co-operation and Development; TFT, 5-trifluorothymidine; Toxicity; USP, United States Pharmacopeia; WE-I, Williams E medium-Incomplete; bw, body weight.