Fabrication of vitamin B12-loaded multivesicular liposomes via microfluidic double-emulsification: Extended pharmacokinetics and bioavailability

Abstract

Vitamin B12 (VB12) is essential for treating pediatric methylmalonic acidemia (MMA) by activating methylmalonyl-CoA mutase and reducing toxic metabolite levels. However, its clinical utility is constrained by frequent intramuscular injections (leading to poor compliance) and the low, highly variable bioavailability associated with oral administration. Thus, a long-acting, sustained-release VB12 system is urgently needed. Herein, VB12-loaded multivesicular liposomes (VB12-MVLs) were reported for the first time, prepared via an improved microfluidic-double emulsification method. To overcome the loading limitation imposed by VB12's water solubility, dimethyl-β-cyclodextrin (DM-β-CD) was introduced to construct VB12-β-MVLs. The resulting VB12-MVLs exhibited a characteristic multi-chambered “honeycomb” structure with uniform size (D50 ∼ 14.30 μm) and high encapsulation efficiency (92.43%). DM-β-CD significantly increased the drug concentration from 10.96 to 17.60 mg mL−1. In vivo studies showed that the lipid phase remained at the subcutaneous injection site for >4 days, while the aqueous phase sustained release for up to 7 days. Pharmacokinetic analysis revealed that compared with free VB12 (mean residence time, MRT = 1.50 h), VB12-MVLs extended MRT to 80.69 h, and VB12-β-MVLs further extended it to 121.42 h, with a relative bioavailability of 191.89%. Preliminary safety evaluations indicated no hemolysis, minimal tissue irritation, and no significant changes in hematological or serum biochemical indices. Collectively, the VB12-MVLs platform—particularly VB12-β-MVLs formulation—represents a promising long-acting formulation for MMA, with the potential to reduce dosing frequency and improve the quality of life of pediatric patients while maintaining therapeutic efficacy.