Selective Extraction and Isolation of Vitamin B12 Using Homogeneous Liquid–Liquid Extraction with Perfluoro Surfactant

Cyanocobalamin (vitamin B12) was selectively extracted from a biological sample by the homogeneous liquid–liquid extraction based on a pH-dependent phase separation with perfluorooctanoate ion (PFOA−). When the experimental conditions were [PFOA−]T = 2.86 × 10−3 mol L−1, [HCl]T = 0.66 mol L−1, and [Acetone]T = 2.86 vol %, 8.46 × 10−7 mol L−1 (1.14 ppm) of vitamin B12 was selectively extracted at 95.2% (sample solution; 35 mL → a liquid sedimented phase; 30 μL). This liquid sedimented phase consisted of large amounts of HPFOA. When microliter levels of THF and water were added to this sedimented phase, vitamin B12 is completely and rapidly back-extracted from the sedimented phase to the water phase. Vitamin B12 was then extracted into the water phase at 96.2% and all HPFOA remained in the sedimented phase. The rest of the sedimented phase after the back-extraction was dried and completely recovered as a powder of HPFOA. Consequently, this method was able to recycle HPFOA as a separation medium. Vitamin B12 in a water phase was also crystallized by the addition of excess THF. Moreover, this method was applied to the extraction from rare oysters. As a result, a few micrograms of vitamin B12 were recovered.