The aim of the study was to evaluate the bioavailability and clinical benefits of oral new formulation (HB ) of hydroxocobalamin (Hdrx) with Hibiscus sabdariffa (HS). First, in an observational study, a cohort of 30 vitamin B -deficient patients (vit B < 200 pg/mL) with neurological symptoms received oral fixed dose of Hdrx containing 15 mg Hdrx daily for 10 days followed by 15 mg monthly. Clinical benefits were evaluated on haematological and biochemical parameters, and neurological improvement at days 10 and 90 compared to day 0. To understand the mechanism, intestinal mucosa from mice were mounted in vitro in Ussing chambers to measure Hdrx Fluxes. In the clinical study, serum vitamin B level increased from 55.1 ± 36.9 to 1330 ± 335.5 pg/mL at day 10 and 431.0 ± 24.27 pg/mL at day 90, without overt adverse effects. In mice ileum, (i) intestinal bioavailability of Hdrx increased in dose-dependent manner with HB . The apparent permeability of Hdrx was P = 34.9 ± 4.6 × 10 cm/s in the presence of 3 mg/mL (HB B) compared to the control P = 6.2 ± 0.7 × 10 cm/s. (ii) Total transepithelial electrical conductance (G ) increased in dose-dependent manner with HB , G = 161.5 ± 10.8 mS/cm² with HB B (Hdrx 1 mg + HS 3 mg) compared to the control Hdrx, G = 28.7 ± 4.0 mS/cm². In conclusion, the clinical study suggests that injections are not required when Hdrx is given orally. Intestinal bioavailability of Hdrx increased in vitro when it was used concomitantly with HS.