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Glycerol is a byproduct of biodiesel production and is generated in large amounts, which has resulted in an increased interest in its valorization. In addition to its use as an energy source directly, the chemical modification of glycerol may result in value‐added derivatives. Herein, acid phosphatases employed in the synthetic mode were evaluated for the enzymatic phosphorylation of glycerol. Nonspecific acid phosphatases could tolerate glycerol concentrations up to 80 wt % and pyrophosphate concentrations up to 20 wt % and led to product titers up to 167 g L −1  in a kinetic approach. In the complementary thermodynamic approach, phytases were able to condense glycerol and inorganic monophosphate directly. This unexpected behavior enabled the simple and cost‐effective production of  rac ‐glycerol‐1‐phosphate from crude glycerol obtained from a biodiesel plant. A preparative‐scale synthesis on a 100 mL‐scale resulted in the production of 16.6 g of  rac ‐glycerol‐1‐phosphate with a reasonable purity (≈75 %).

Preparative‐Scale Enzymatic Synthesis of rac ‐Glycerol‐1‐phosphate from Crude Glycerol Using Acid Phosphatases and Phosphate