Novel Two-Step Process Utilizing a Single Enzyme for the Production of High-Titer 3,6-Anhydro-l-galactose from Agarose Derived from Red Macroalgae.

3,6-Anhydro-l-galactose (l-AHG), a major component of agarose derived from red macroalgae, has excellent potential for industrial applications based on its physiological activities such as skin whitening, moisturizing, anticariogenicity, and anti-inflammation. However, l-AHG is not yet commercially available due to the complexity, inefficiency, and high cost of the current processes for producing l-AHG. Currently, l-AHG production depends on a multistep process requiring several enzymes. Here, we designed and tested a novel two-step process for obtaining high-titer l-AHG by using a single enzyme. First, to depolymerize agarose preferentially into agarobiose (AB) at a high titer, the agarose prehydrolysis using phosphoric acid as a catalyst was optimized at a 30.7% (w/v) agarose loading, which is the highest agarose or agar loading reported so far. Then AB produced by the prehydrolysis was hydrolyzed into l-AHG and d-galactose (d-Gal) by using a recently discovered enzyme, Bgl1B. We suggest that this simple and efficient process could be a feasible solution for the commercialization and mass production of l-AHG.