Production and Functional Properties of Galdieria Sulphuraria Glycogen, a Novel Highly

Production and functional properties of Galdieria sulphuraria glycogen, a novel highly branched nanoparticle

Marta Martinez-Garcia, Akrivi Kormpa, Anastasia Kristijarti, Marc J.E.C. van der Maarel

Many microorganisms store excess energy when experiencing nitrogen limitation in the form of glycogen, a polymer of alpha,1-> 4-linked anhydroglucose residues with an occasional alpha,1-> 6 branch linkage. Glycogen is an interesting molecule as it is water soluble, forms nanoparticles and can easily be modified to provide specific functional properties. However, large-scale production of glycogen is limited and complicated as microbes tend to store it only under specific conditions and enzymatic conversion of a suitable sugar source is a multi enzyme process. We found that the extremophilic red microalga Galdieria sulphuraria, growing in acidic hot-springs (pH around 2 and approx. 50oC), produces large amounts (up to 50% of the dry biomass) of glycogen when growing in the dark on oxygen and glycerol, a waste product of the biodiesel industry. The structure, physico-chemical and functional properties of the G. sulphuraria (GS) glycogen and the octenylated derivative were studies in detail. The GS glycogen was extracted from Galdieria cells grown on 1-5% glycerol by ice-cold ethanol precipitation after the protein was removed from the cell extract by heating and TCA precipitation. H-NMR (500 MHz) showed that the GS glycogen has a high degree of alpha, 1-> 6 branching of 18-23%, being 2 to 3 times higher than has been found for bacterial glycogens. The side chain distribution was analysed by treating the GS glycogens with isoamylase/pullulanase and subsequently running the debranched samples on a Dionex HPLC system. The GS glycogen had only small side chains, with the longest having a DP of 11. The weight average molecular weight of the GS glycogen was about 2.5 x 105 kDa, as measured by size exclusion chromatography combined with a multi-angle laser light scattering and refractrometric index detectors. This is at least one order of magnitude lower than microbial glycogens. An octenylated derivative of the GS glycogen was produced and the emulsifying properties were analysed in detail, showing that it was as good as other emulsifiers produced from (waxy) starch or phytoglycogen. This research shows that GS glycogen can be produced from glycerol-grown microalgal cells. It is a highly branched carbohydrate nanoparticle that can be used as an alternative to (modified) starch or phytoglycogen in food products or specific medical applications.

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