Accumulation and metabolism of selenium in the rare yeast Kazachstania unispora during the selenium enrichment process.

Selenium (Se)-enriched yeast is a good nutritional source for human being. Kazachstania unispora (K. unispora) has shown the positive physiological functionality for human health, whose potential for Se enrichment, however, remains elusive. This study demonstrated the ability of K. unispora to convert inorganic Se to organic Se, and then comprehensively investigated the accumulation and metabolism of Se in K. unispora. The results indicated that K. unispora can effectively accumulate organic Se, of which 95% of absorbed Se was converted to organic forms. Among these organic Se, 46.17% of them was bound to protein and 16.78% was combined with polysaccharides. In addition, some of the organic Se was metabolized to selenomethionine (30.26%) and selenocystine (3.02%), during which four low-molecular weight selenometabolites were identified in K. unispora. These findings expand the scope of Se-enriched yeast species, and provide useful knowledge for further investigation of Se enrichment mechanism in K. unispora.

Tibetan kefir grains fermentation alters physicochemical properties and improves antioxidant activities of Lycium barbarum pulp polysaccharides.

There is a lack of research on how Tibetan kefir grains fermentation alters the physicochemical properties and biological activity of Lycium barbarum pulp polysaccharides, despite some reports that fermentation can affect the structure and activity of plant polysaccharides. This study demonstrated that, through fermentation, the molecular weight of polysaccharides decreased from 25.33 to 15.11 kg/mol while the contents of total sugar and uronic acid increased by 19.11% and 40.38%, respectively. Furthermore, after fermentation, the polysaccharides exhibited an uneven and rough surface along with a reduced number of branched chains and triple helix structures. Tibetan kefir grains fermentation enhanced the antioxidant activity of polysaccharides, which may be attributed to an increase in arabinose, galactose, and uronic acid content and a decrease in polysaccharide molecular weight. This research offers an alternative viewpoint on the potential application of Tibetan kefir grains-fermented Lycium barbarum pulp polysaccharides in functional foods.

Immobilisation of Candida rugosa lipase on a highly hydrophobic support: A stable immobilised lipase suitable for non-aqueous synthesis.

Lipase from Candida rugosa (CrL) was immobilised on highly hydrophobic, octadecyl methacrylate resin (Lifetech™ ECR8806M) via interfacial adsorption. The aim was to produce a stable biocatalyst suitable for use in a range of lipid-modifying reactions. Immobilisation was carried out in 10 mM phosphate buffer (pH 6.0) over 24 h at 21 °C. High protein binding of 58.7 ±â€¯4.9 mg/g dry support accounted for ∼53 % of the applied protein. The activity recovery against tributyrin was 74.0 ±â€¯1.1 %. The specific activity of immobilised CrL against tributyrin was considerably higher than that of Novozym® 435, at 1.79 ±â€¯0.05 and 1.08 ±â€¯0.04 U/mg bound protein, respectively. Incubation with high concentrations (10 % w/v) of both Triton X-100 and SDS resulted in only a small reduction in immobilised lipase activity. Solvent-free synthesis of glycerides by the FFA-saturated immobilised CrL was successful over 6 reaction cycles, with no apparent loss of activity.

A simplified method for active-site titration of lipases immobilised on hydrophobic supports.

The aim of this work was to develop a simple and accurate protocol to measure the functional active site concentration of lipases immobilised on highly hydrophobic supports. We used the potent lipase inhibitor methyl 4-methylumbelliferyl hexylphosphonate to titrate the active sites of Candida rugosa lipase (CrL) bound to three highly hydrophobic supports: octadecyl methacrylate (C18), divinylbenzene crosslinked methacrylate (DVB) and styrene. The method uses correction curves to take into account the binding of the fluorophore (4-methylumbelliferone, 4-MU) by the support materials. We showed that the uptake of the detection agent by the three supports is not linear relative to the weight of the resin, and that the uptake occurs in an equilibrium that is independent of the total fluorophore concentration. Furthermore, the percentage of bound fluorophore varied among the supports, with 50 mg of C18 and styrene resins binding approximately 64 and 94%, respectively. When the uptake of 4-MU was calculated and corrected for, the total 4-MU released via inhibition (i.e. the concentration of functional lipase active sites) could be determined via a linear relationship between immobilised lipase weight and total inhibition. It was found that the functional active site concentration of immobilised CrL varied greatly among different hydrophobic supports, with 56% for C18, compared with 14% for DVB. The described method is a simple and robust approach to measuring functional active site concentration in immobilised lipase samples.

The use of immobilised digestive lipase from Chinook salmon (Oncorhynchus tshawytscha) to generate flavour compounds in milk.

The aim of this research was to determine the potential of immobilised digestive lipase from Chinook salmon (Oncorhynchus tshawytscha) to generate flavour compounds in milk. The lipase was immobilised on hydrophobic resin (Toyopearl® Butyl) and used to hydrolyse milk lipids in a batch reactor. The lipase was stable when immobilised and there was no significant resin fouling or enzyme inhibition between cycles. Eight cycles were achieved before the hydrolysis rate dropped significantly because of physical losses of the immobilised lipase. The immobilised lipase showed the highest specificity towards short-chain fatty acids butanoic and hexanoic acids, the main dairy product flavour and odour compounds. Based on the performance of the reactor, and the ability of the lipase to alter free fatty acid composition and sensory characteristics of milk, the immobilised salmon lipase has potential applications in developing dairy products with unique flavours.

Purification and properties of digestive lipases from Chinook salmon (Oncorhynchus tshawytscha) and New Zealand hoki (Macruronus novaezelandiae).

Lipases were purified from delipidated pyloric ceca powder of two New Zealand-sourced fish, Chinook salmon (Oncorhynchus tshawytscha) and hoki (Macruronus novaezelandiae), by fractional precipitation with polyethylene glycol 1000, followed by affinity chromatography using cholate-Affi-Gel 102, and gel filtration on Sephacryl S-300 HR. For the first time, in-polyacrylamide gel activity of purified fish lipases against 4-methylumbelliferyl butyrate has been demonstrated. Calcium ions and sodium cholate were absolutely necessary both for lipase stability in the gel and for optimum activity against caprate and palmitate esters of p-nitrophenol. A single protein band was present in native polyacrylamide gels for both salmon and hoki final enzyme preparations. Under denaturing conditions, electrophoretic analysis revealed two bands of 79.6 and 54.9 kDa for salmon lipase. It is proposed that these bands correspond to an uncleaved and a final form of the enzyme. One band of 44.6 kDa was seen for hoki lipase. pI values of 5.8±0.1 and 5.7±0.1 were obtained for the two salmon lipase forms. The hoki lipase had a pI of 5.8±0.1. Both lipases had the highest activity at 35°C, were thermally labile, had a pH optimum of 8-8.5, and were more acid stable compared to other fish lipases studied to date. Both enzymes were inhibited by the organophosphate paraoxon. Chinook salmon and hoki lipases showed good stability in several water-immiscible solvents. The enzymes had very similar amino acid composition to mammalian carboxyl ester lipases and one other fish digestive lipase. The salmon enzyme was an overall better catalyst based on its higher turnover number (3.7±0.3 vs. 0.71±0.05 s(-1) for the hoki enzyme) and lower activation energy (2.0±0.4 vs. 7.6±0.8 kcal/mol for the hoki enzyme) for the hydrolysis of p-nitrophenyl caprate. The salmon and hoki enzymes are homologous with mammalian carboxyl ester lipases.