Systematic comparison of eight methods for preparation of high purity sulfated fucans extracted from the brown alga Pelvetia canaliculata.

Sulfated fucans from brown algae are a heterogeneous group of biologically active molecules. To learn more on their structure and to analyze and exploit their biological activities, there is a growing need to develop reliable and cost effective protocols for their preparation. In the present study, a brown alga Pelvetia canaliculata (Linnaeus) was used as a rich source of sulfated fucans. Sulfated fucan preparation methods included neutral and acidic extractions followed by purification with activated charcoal (AC), polyvinylpolypyrrolidone (PVPP), or cetylpyridinium chloride (CPC). Final products were compared in terms of yield, purity, monosaccharide composition and molecular weight. Acidic extractions provided higher yields compared to neutral ones, whereas the AC purification provided sulfated fucan products with the highest purity. Mass spectrometry analyses were done on oligosaccharides produced by the fucanase MfFcnA from the marine bacterium Mariniflexille fucanivorans. This has provided unique insight into enzyme specificity and the structural characteristics of sulfated fucans.

In-depth structural characterization of oligosaccharides released by GH107 endofucanase MfFcnA reveals enzyme subsite specificity and sulfated fucan substructural features.

The extracellular matrix of brown algae represents an abundant source of fucose-containing sulfated polysaccharides (FCSPs). FCSPs include sulfated fucans, essentially composed of fucose, and highly heterogeneous fucoidans, comprising various monosaccharides. Despite a range of potentially valuable biological activities, the structures of FCSPs are only partially characterized and enzymatic tools leading to their deconstruction are rare. Previously, the enzyme MfFcnA was isolated from the marine bacterium Mariniflexile fucanivorans and biochemically characterized as an endo-α-1 â†’ 4-l-fucanase, the first member of glycoside hydrolase family 107. Here, MfFcnA was used as an enzymatic tool to deconstruct the structure of the sulfated fucans from Pelvetia canaliculata (Fucales brown alga). Oligofucans released by MfFcnA at different time points were characterized using mass spectrometry coupled with liquid chromatography and tandem mass spectrometry through Charge Transfer Dissociation. This approach highlights a large diversity in the structures released. In particular, the analyses show the presence of species with less than three sulfates per two fucose residues. They also reveal species with monosaccharides other than fucose and the occurrence of laterally branched residues. Precisely, the lateral branching is either in the form of a hexose accompanied by a trisulfated fucose nearby, or of a side chain of fucoses with a pentose as the branching point on the polymer. Overall, the results indicate that the structure of sulfated fucans from P. canaliculata is more complex than expected. They also reveal the interesting capacity of MfFcnA to accommodate different substrates, leading to structurally diverse oligofucan products that potentially could be screened for bioactivities.