Efficient Synthesis of (H2O)OHCbi

Ten milligrams of (CN)₂Cbi or dicyanocobinamide (Sigma-Aldrich), whose UV–visible spectrum was identical to the reported spectrum,^{35 (link),36 (link)} was dissolved in 3 mL of anaerobic 0.1 M NaOH, and small aliquots of sodium borohydride pellets were added every 5 min, until a molar excess of about 10-fold of borohydride over (CN)₂Cbi was reached. The reaction proceeded for about 1.5 h until the mixture was tranferred into a beaker and quenched by the addition of acidified water. The final pH was adjusted to 3, and the mixture was purified over a 1 g Sep Pak C₁₈ reverse phase column (Waters). The column was preconditioned prior to sample loading by washing once with 100% methanol and five times thereafter with 0.1% (v/v) TFA (trifluoroacetic acid). After the sample had passed through the column, salts were removed by washing with 0.1% TFA for five column volumes. This was followed by elution with 100% methanol, and the collected orange fractions were evaporated overnight at room temperature. The collected product (6 mg) was identified as (H₂O)OHCbi by its UV–visible absorption spectrum, which was identical to the spectrum reported in the literature.^{35 (link),36 (link)}