Separation and Quantification of Microalgal Carbohydrates

Structural carbohydrates can constitute a large fraction of the dry weight of algal biomass and thus accurate identification and quantification is important for summative mass closure. Two limitations to the accurate characterization of microalgal carbohydrates are the lack of a robust analytical procedure to hydrolyze polymeric carbohydrates to their respective monomers and the subsequent identification and quantification of those monosaccharides. We address the second limitation, chromatographic separation of monosaccharides, here by identifying optimum conditions for the resolution of a synthetic mixture of 13 microalgae-specific monosaccharides, comprised of 8 neutral, 2 amino sugars, 2 uronic acids and 1 alditol (myo-inositol as an internal standard). The synthetic 13-carbohydrate mix showed incomplete resolution across 11 traditional high performance liquid chromatography (HPLC) methods, but showed improved resolution and accurate quantification using anion exchange chromatography (HPAEC) as well as alditol acetate derivatization followed by gas chromatography (for the neutral- and amino-sugars only). We demonstrate the application of monosaccharide quantification using optimized chromatography conditions after sulfuric acid analytical hydrolysis for three model algae strains and compare the quantification and complexity of monosaccharides in analytical hydrolysates relative to a typical terrestrial feedstock, sugarcane bagasse.