Voltammetric studies on the simple ion transfer (IT) behaviors of an important water-soluble B-vitamin, folic acid (FA), at the liquid–liquid (L–L) interface were firstly performed and then applied as a novel detection method for FA under physiological conditions. Meso-water–1,6-dichlorohexane (W–DCH) and meso-water–organogel interface arrays were built by using a hybrid mesoporous silica membrane (HMSM) with a unique structure of pores-in-pores and employed as the new platforms for the IT voltammetric study. In view of the unique structure of the HMSM, the impact of the ionic surfactant cetyltrimethylammonium bromide (CTAB), self-assembled within the silica nanochannels of the HMSM, was investigated. In particular, its effect on the IT voltammetric behavior and detection of FA at meso-L–L interface arrays was systematically examined by cyclic voltammetry (CV), differential pulse voltammetry (DPV) and differential pulse stripping voltammetry (DPSV). It was found that all the voltammetric responses of CV, DPV, and DPSV and the corresponding detection limit of FA at such meso-L–L interface arrays are closely related to the CTAB in the HMSM. Significantly, the calculated detection limit of FA could be improved to 80 nM after the combination of the DPSV technique with the additional preconcentration of FA in the silica-CTAB nanochannels, achieved through an anion-exchange process between FA⁻ and the bromide of CTAB in HMSM. This provides a new and attractive strategy for the detection of those biological anions.