A biocatalytic route for the synthesis of novel glycosyl acrylate monomers produced from hemicellulosic glycans.

We present here a series of thermoresponsive glycopolymers in the form of poly( N -isopropylacrylamide) -co- (2-[β-manno[oligo]syloxy] ethyl methacrylate)s. These copolymers were prepared from oligo-β-mannosyl ethyl methacrylates that were synthesized through enzymatic catalysis, and were subsequently investigated with respect to their aggregation and phase behavior in aqueous solution using a combination of 1 H NMR spectroscopy, dynamic light scattering, cryogenic transmission electron microscopy (TEM), and small-angle X-ray scattering (SAXS). The thermoresponsive glycopolymers were prepared by conventional free radical copolymerization of different mixtures of 2-(β-manno[oligo]syloxy)ethyl methacrylates (with either one or two saccharide units) and N -isopropylacrylamide (NIPAm). The results showed that below the lower critical solution temperature (LCST) of poly(NIPAm), the glycopolymers readily aggregate into nanoscale structures, partly due to the presence of the saccharide moieties. Above the LCST of poly(NIPAm), the glycopolymers rearrange into a heterogeneous mixture of fractal and disc/globular aggregates. Cryo-TEM and SAXS data demonstrated that the presence of the pendant β-mannosyl moieties in the glycopolymers induces a gradual conformational change over a wide temperature range. Even though the onset of this transition is not different from the LCST of poly(NIPAm), the gradual conformational change offers a variation of the temperature-dependent properties in comparison to poly(NIPAm), which displays a sharp coil-to-globule transition. Importantly, the compacted form of the glycopolymers shows a larger colloidal stability compared to the unmodified poly(NIPAm). In addition, the thermoresponsiveness can be conveniently tuned by varying the sugar unit-length and the oligo-β-mannosyl ethyl methacrylate content.

Spent sulfite liquor (SSL) from softwood processing is rich in hemicellulose (acetyl galactoglucomannan, AcGGM), lignin, and lignin-derived compounds. We investigated the effect of sequential AcGGM purification on the enzymatic bioconversion of AcGGM. SSL was processed through three consecutive purification steps (membrane filtration, precipitation, and adsorption) to obtain AcGGM with increasing purity. Significant reduction (~99%) in lignin content and modest loss (~18%) of polysaccharides was observed during purification from the least pure preparation (UFR), obtained by membrane filtration, compared to the purest preparation (AD), obtained by adsorption. AcGGM (~14.5 kDa) was the major polysaccharide in the preparations; its enzymatic hydrolysis was assessed by reducing sugar and high-performance anion-exchange chromatography analysis. The hydrolysis of the UFR preparation with Viscozyme L or Trichoderma reesei β-mannanase TrMan5A (1 mg/mL) resulted in less than ~50% bioconversion of AcGGM. The AcGGM in the AD preparation was hydrolyzed to a higher degree (~67% with TrMan5A and 80% with Viscozyme L) and showed the highest conversion rate. This indicates that SSL contains enzyme-inhibitory compounds (e.g., lignin and lignin-derived compounds such as lignosulfonates) which were successfully removed.

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Protein engineering to functionalize the self-assembling enamel matrix protein amelogenin with a cellulose binding domain (CBD) is used. The purpose is to examine the binding of the engineered protein, rh174CBD, to cellulose materials, and the possibility to immobilize self-assembled amelogenin nanospheres on cellulose. rh174CBD assembled to nanospheres ≈35 nm in hydrodynamic diameter, very similar in size to wild type amelogenin (rh174). Uniform particles are formed at pH 10 for both rh174 and rh174CBD, but only rh174CBD nanospheres showes significant binding to cellulose (Avicel). Cellulose binding of rh174CBD is promoted when the protein is self-assembled to nanospheres, compared to being in a monomeric form, suggesting a synergistic effect of the multiple CBDs on the nanospheres. The amount of bound rh174CBD nanospheres reached ≈15 mg/g Avicel, which corresponds to 4.2 to 6.3 × 10 mole/m . By mixing rh174 and rh174CBD, and then inducing self-assembly, composite nanospheres with a high degree of cellulose binding can be formed, despite a lower proportion of rh174CBD. This demonstrates that amelogenin variants like rh174 can be incorporated into the nanospheres, and still retain most of the binding to cellulose. Engineered amelogenin nanoparticles can thus be utilized to construct a range of new cellulose based hybrid materials, e.g. for wound treatment.