Application of MALDI-MS for characterization of fucoidan hydrolysates and screening of endo-fucoidanase activity.

Brown macroalgae synthesize large amounts of fucoidans, sulfated fucose-containing polysaccharides, in the ocean. Fucoidans are of importance for their recently discovered contribution to marine carbon dioxide sequestration and due to their potential applications in biotechnology and biomedicine. However, fucoidans have high intra- and intermolecular diversity that challenges assignment of structure to biological function and the development of applications. Fucoidan-active enzymes may be used to simplify this diversity by producing defined oligosaccharides more applicable for structural refinement, characterization, and structure to function assignment for example via bioassays. In this study, we combined MALDI mass spectrometry with biocatalysis to show that the endo-fucoidanases P5AFcnA and Wv323 can produce defined oligosaccharide structures directly from unrefined macroalgal biomass. P5AFcnA released oligosaccharides from seven commercial fucoidan extracts in addition to unrefined biomass of three macroalgae species indicating a broadly applicable approach reproducible across 10 species. Both MALDI-TOF/TOF and AP-MALDI-Orbitrap systems were used, demonstrating that the approach is not instrument-specific and exploiting their combined high-throughput and high-resolution capabilities. Overall, the combination of MALDI-MS and endo-fucoidanase assays offers high-throughput evaluation of fucoidan samples and also enables extraction of defined oligosaccharides of known structure from unrefined seaweed biomass.

Fucoid brown algae inject fucoidan carbon into the ocean.

Brown algae annually convert gigatons of carbon dioxide into carbohydrates, including the complex extracellular matrix polysaccharide fucoidan. Due to its persistence in the environment, fucoidan is potentially a pathway for marine carbon sequestration. Rates of fucoidan secretion by brown algae remain unknown due to the challenge of identifying and quantifying complex polysaccharides in seawater. We adapted the techniques of anion exchange chromatography, enzyme-linked immunosorbent assay, and biocatalytic enzyme-based assay for detection and quantification of fucoidan. We found the brown alga Fucus vesiculosus at the Baltic Sea coast of south-west Finland to secrete 0.3% of their biomass as fucoidan per day. Dissolved fucoidan concentrations in seawater adjacent to algae reached up to 0.48 mg L-1. Fucoidan accumulated during incubations of F. vesiculosus, significantly more in light than in darkness. Maximum estimation by acid hydrolysis indicated fucoidan secretion at a rate of 28 to 40 mg C kg-1 h-1, accounting for 44 to 50% of all exuded dissolved organic carbon. Composed only of carbon, oxygen, hydrogen, and sulfur, fucoidan secretion does not consume nutrients enabling carbon sequestration independent of algal growth. Extrapolated over a year, the algae sequester more carbon into secreted fucoidan than their biomass. The global utility of fucoidan secretion is an alternative pathway for carbon dioxide removal by brown algae without the need to harvest or bury algal biomass.

Structures and functions of algal glycans shape their capacity to sequester carbon in the ocean.

Algae synthesise structurally complex glycans to build a protective barrier, the extracellular matrix. One function of matrix glycans is to slow down microorganisms that try to enzymatically enter living algae and degrade and convert their organic carbon back to carbon dioxide. We propose that matrix glycans lock up carbon in the ocean by controlling degradation of organic carbon by bacteria and other microbes not only while algae are alive, but also after death. Data revised in this review shows accumulation of algal glycans in the ocean underscoring the challenge bacteria and other microbes face to breach the glycan barrier with carbohydrate active enzymes. Briefly we also update on methods required to certify the uncertain magnitude and unknown molecular causes of glycan-controlled carbon sequestration in a changing ocean.