In this study, development of folic acid-loaded liposomes using a basic amino acid, histidine as a solubilizing agent for folic acid was presented, which tackled the poor solubility of this vitamin. The effect of the liposomal membrane modifiers, cholesterol and SPAN 20 on the characteristics of the final formulations was examined. Liposomes prepared from a commercially available purified soybean lecithin (Phospholipon 90G) by proliposome method were between 503 and 877 nm in average diameter, where cholesterol induced enlargement and SPAN.20 reduction of vesicles. High encapsulation efficiency of 84% and drug loading of 0.123 mg g−1 were achieved, irrespective to the composition. According to AFM images, folic acid-loaded liposomes of a fraction with a nano size were flattened compared to globular empty liposomes. FTIR analysis revealed possible interactions between phospholipids and histidine, while DSC study suggested interactions between folic acid and lipids during heating. Release study done by a Franz diffusion cell showed prolonged release of folic acid from liposomes and the release rate was determined by folic acid solubility. Display omitted •Liposomes were used as a model system for encapsulation of folic acid.•A basic amino acid-histidine was used as a solubilizing agent for folic acid.•Interactions between folic acid and phospholipids were determined by DSC analysis.•Interactions between histidine and phospholipids were analyzed by ATR-IR.•Cholesterol and SPAN 20 did not contribute to the effect of prolonged release.