In today's world, there is an increasing trend towards the use of renewable, cheap and readily available biomass in the production of a wide variety of fine and bulk chemicals in different biorefineries. Biorefineries utilize the activities of microbial cells and their enzymes to convert biomass into target products. Many of these processes require enzymes which are operationally stable at high temperature thus allowing e.g. easy mixing, better substrate solubility, high mass transfer rate, and lowered risk of contamination. Thermophiles have often been proposed as sources of industrially relevant thermostable enzymes. Here we discuss existing and potential applications of thermophiles and thermostable enzymes with focus on conversion of carbohydrate containing raw materials. Their importance in biorefineries is explained using examples of lignocellulose and starch conversions to desired products. Strategies that enhance thermostablity of enzymes both in vivo and in vitro are also assessed. Moreover, this review deals with efforts made on developing vectors for expressing recombinant enzymes in thermophilic hosts.

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Galectins are defined by a conserved β-galactoside binding site that has been linked to many of their important functions in e.g. cell adhesion, signaling, and intracellular trafficking. Weak adjacent sites may enhance or decrease affinity for natural β-galactoside-containing glycoconjugates, but little is known about the biological role of this modulation of affinity (fine specificity). We have now produced 10 mutants of human galectin-3, with changes in these adjacent sites that have altered carbohydrate-binding fine specificity but that retain the basic β-galactoside binding activity as shown by glycan-array binding and a solution-based fluorescence anisotropy assay. Each mutant was also tested in two biological assays to provide a correlation between fine specificity and function. Galectin-3 R186S, which has selectively lost affinity for LacNAc, a disaccharide moiety commonly found on glycoprotein glycans, has lost the ability to activate neutrophil leukocytes and intracellular targeting into vesicles. K176L has increased affinity for β-galactosides substituted with GlcNAcβ1–3, as found in poly-N-acetyllactosaminoglycans, and increased potency to activate neutrophil leukocytes even though it has lost other aspects of galectin-3 fine specificity. G182A has altered carbohydrate-binding fine specificity and altered intracellular targeting into vesicles, a possible link to the intracellular galectin-3-mediated anti-apoptotic effect known to be lost by this mutant. Finally, the mutants have helped to define the differences in fine specificity shown by Xenopus, mouse, and human galectin-3 and, as such, the evidence for adaptive change during evolution.

Antioxidants are widely used by humans, both as dietary supplements and as additives to different types of products. The desired properties of an antioxidant often include a balance between the antioxidizing capacity, stability, and solubility. This review focuses on flavonoids, which are naturally occurring antioxidants, and different common substituent groups on flavonoids and how these affect the properties of the molecules in vitro. Hydroxyl groups on flavonoids are both important for the antioxidizing capacity and key points for further modification resulting in O-methylation, -glycosylation, -sulfation, or -acylation. The effects of O-glycosylation and acylation are discussed as these types of substitutions have been most explored in vitro concerning antioxidizing properties as well as stability and solubility. Possibilities to control the properties by enzymatic acylation and glycosylation are also reviewed, showing that depending on the choice of enzyme and substrate, regioselective results can be obtained, introducing possibilities for more targeted production of antioxidants with predesigned properties.

Onion waste is a renewable raw material, rich in different molecular species of the antioxidant quercetin. To utilize this resource, an environmentally sustainable procedure has been developed, using pressurized hot water to extract the quercetin species, followed by biocatalytic conversion of the quercetin glycosides to quercetin and carbohydrates. Two different recombinantly expressed thermostable b-glucosidases, Thermotoga neapolitana b-glucosidase A and B, were utilized as catalysts. These enzymes maintain activity at temperatures around 90 uC, and are therefore ideal to use in combination with hot water extraction. Our results, based on experimental design, showed that they converted quercetin glycosides to active quercetin in less than 10 min reaction time in water at 90 uC, pH 5.0. Experimental design showed that the optimal extraction conditions included three 5 min extraction cycles with water at 120 uC and 50 bars, giving a total extraction time of 15 min. Several different types of quercetin and isorhamnetin glycosides as well as kaempferol were detected in onion waste using LC-MS/MS analysis. After converting the different glycosidic compounds to their respective aglycones, the quercetin content was 10 to 50 mg g 21 dry weight of onion waste (RSD 8%). In summary, our research demonstrates that subcritical water extraction followed by b-glucosidase-catalyzed hydrolysis is a rapid method to determine the content of quercetin and isorhamnetin in onion samples, and is environmentally sustainable as it only uses water as solvent and enzymes as catalysts.