Levomefolic acid is an orally available, bioactive form of folic acid that crosses the blood-brain barrier[1]. Levomefolic acid, a ligand with low binding energy, can form three hydrogen bonds with Arg169, Asp305 and Tyr276 of COVID-19 PLpro protein and form π-π stacking interaction with Tyr271[2]. Levomefolic acid has been widely used as a model compound and an internal reference to develop novel LC-MS/MS methods for the identification of related compounds[3].
In vitro, Levomefolic acid treatment at 10µg/ml for 24 hours significantly inhibited Caco-2 cell proliferation and induced cell cycle arrest[4]. Treatment of BEAS-2B cells with 0.012g/L Levomefolic acid for 24h protected BEAS-2B cells from oxidative stress and inflammatory injury induced by glucose (50mM) and restored cell viability[5]. Treatment with 50µM Levomefolic acid for 48h significantly induced DNA damage in MRC5 and IMR90 cells, resulting in decreased MTR gene expression and increased MTHFD1, EGFR, XRCC1, and DNA Polβ gene expression[6].
In vivo, intraperitoneal injection of Levomefolic acid (3µg/kg) at 30 minutes before ischemia and again 3 hours after reperfusion can improve renal function in Sprague-Dawley rats, alleviate oxidative stress, restore glutathione levels in the kidneys, and reduce lipid peroxidation[7]. Within 5 weeks after weaning, oral administration of 4ml of drinking water containing a dose of 47μg/ml of Levomefolic acid daily significantly increased the body weight of female mice and promoted the maturation of oocytes[8].
References:
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[2] Hosseini M, Chen W, Xiao D, et al. Computational molecular docking and virtual screening revealed promising SARS-CoV-2 drugs[J]. Precision clinical medicine, 2021, 4(1): 1-16.
[3] Golja M V, Trontelj J, Geršak K, et al. Simultaneous quantification of intracellular concentrations of clinically important metabolites of folate-homocysteine cycle by LC-MS/MS[J]. Analytical biochemistry, 2020, 605: 113830.
[4] Akoglu B, Milovic V, Caspary W F, et al. Hyperproliferation of homocysteinetreated colon cancer cells is reversed by folate and 5-methyltetrahydrofolate[J]. European journal of Nutrition, 2004, 43(2): 93-99.
[5] Pathikkal A, Puthusseri B, Divya P, et al. Folate derivatives, 5-methyltetrahydrofolate and 10-formyltetrahydrofolate, protect BEAS-2B cells from high glucose–induced oxidative stress and inflammation[J]. In Vitro Cellular & Developmental Biology-Animal, 2022, 58(5): 419-428.
[6] Karkera C, Senejani A G. Evaluating toxicity and level of DNA damage in human fetal lung cells upon exposure to 5-methyltetrahydrofolate (bioactive folate)[J]. Nutrition and Health, 2024: 02601060241302895.
[7] Wijerathne C U B, Au-Yeung K K W, Siow Y L, et al. 5-methyltetrahydrofolate attenuates oxidative stress and improves kidney function in acute kidney injury through activation of Nrf2 and antioxidant defense[J]. Antioxidants, 2022, 11(6): 1046.
[8] Nasr‐Esfahani F, Jafarpour F, Varnosfaderani S R, et al. Differential Effects of Folic Acid and 5‐Methyltetrahydrofolate on Mice Oocytes and Embryo Development In Vitro and In Vivo: Implications for Reproductive Health[J]. The FASEB Journal, 2025, 39(20): e71090.