1-Ethyl-β-N-acetylglucosaminide increases hyaluronan production in human keratinocytes by being converted to N-acetylglucosamine via β-N-acetylglucosaminidase-dependent manner

Regulation of hyaluronan (HA) is important for the maintenance of epidermal homeostasis. Here we examined the mechanism by which 1-ethyl-β-N-acetylglucosaminide (β-NAG2), a newly developed N-acetylglucosamine (NAG) derivative, increases HA production in cultured human epidermal keratinocytes. When keratinocytes were treated with β-NAG2, mRNA expression of HA synthase 3, which is responsible for HA production in human keratinocytes, was not influenced, but the intracellular level of UDP-NAG, a substrate used for HA synthesis, was increased. By using a synthetic substrate for β-N-acetylglucosaminidase (β-NAGase), keratinocytes were found to possess β-NAGase activity, and treatment of o-(2-acetamido-2-deoxy-D-glucopyranosylidene) amino N-phenyl carbamate (PUGNAc), an inhibitor of β-NAGase, abolished the release of NAG from β-NAG2 in keratinocytes. Furthermore, PUGNAc attenuated the β-NAG2-induced intracellular UDP-NAG and HA production in keratinocytes. These results suggest that β-NAG2 is converted to NAG by endogenous β-NAGase in keratinocytes, and the resulting NAG is further metabolized to UDP-NAG and utilized for HA production.

Simple enumeration of Escherichia coli concentrations in river water samples by measuring β-D-glucuronidase activities in a microplate reader

Monitoring of Escherichia coli concentrations in river water (RW) is essential to identify fecal pollution of the river. The objective of this study was to assess the suitability of a novel simple and high throughput method developed in our laboratory to enumerate E. coli concentrations in RW samples. The method is based on the use of the synthetic substrate specific for the β-D-glucuronidase (GUS) produced by E. coli. GUS activities and E. coli concentrations were monitored at eight selected sites in rivers running through Sapporo, Japan. Because the fluorescence intensities of the synthetic substrate in the RW samples increased linearly over a 4-h incubation period, we could estimate the GUS activities of the RW samples. The GUS activities were highly correlated with E. coli concentrations at >100 most probable numbers 100 mL−1 with a correlation coefficient of 0.87. The GUS activities of the RW samples collected from all sampling sites fitted well to a single correlation equation, which indicates that it was applicable to the estimation of E. coli concentrations regardless of the sampling sites. This method is simple, rapid, reliable, inexpensive, and high throughput, and is therefore useful for monitoring E. coli in RW.

Effect of the type and method of additives on the productivity of mushrooms grown in industrial conditions

The paper summarizes and analyzes the results of practical studies on the cultivation of Agaricus bisporus champignon mushrooms on a synthetic substrate with the introduction of organic additives into it – food and processing wastes: brewing waste, soybean and sunflower meal semolina. A yield analysis was carried out; indicators characterizing the quality of Agaricus bisporus mushrooms were studied; dependence of yield and quality of mushrooms on flush was demonstrated. It was noted that the preparation time of the substrate affects the number of flushes, respectively, and, correspondingly, on the yield of mushrooms: when growing mushrooms on a synthetic substrate prepared in the spring-summer period, two flushes are noted and, as a result, a higher yield is noted compared to mushrooms grown on a substrate prepared in the autumn-winter period. The study of the quality of fruit bodies showed that the use of additives increased the content of protein, fiber, fat and ash elements (potassium, phosphorus, calcium, sodium and magnesium). The results showed that the brewing waste is the best organic additive, which contributes not only to the growth of yield, but also to the improvement of the quality of cultured mushrooms. The study demonstrated that the quality of mushrooms obtained in the first flush exceeds the qualitative indicators of mushrooms grown in the second flush. The authors concluded that it is advisable to use organic waste (brewing waste, sunflower meal) as a substrate element when growing mushrooms in industrial conditions.

Proteinaceous Pancreatic Lipase Inhibitor from the Seed of Litchi chinensis

A study of the pancreatic lipase inhibitory activity of a protein from the seed of Litchi chinensis was carried out. Protein was isolated by 70 % ammonium sulphate precipitation followed by dialysis. Lipase inhibitory activity of the protein was evaluated using both synthetic (p-nitrophenyl palmitate) and natural (olive oil) substrates. Protein at the final concentration of 100 µg/mL was able to inhibit 68.2 % pancreatic lipase on synthetic substrate and 60.0 % on natural substrate. Proteinaceous nature of the inhibitor was determined using trypsinization assay. Pancreatic lipase inhibitory protein was sensitive to 0.05 % trypsin treatment with the loss of 61.9 % activity. IC50 of this proteinaceous pancreatic lipase inhibitor was 73.1 µg/mL using synthetic substrate. This inhibitory protein was sensitive to pH, with the highest inhibitory activity at pH=8.0 and the lowest at pH=3.0. Protein was further analyzed using 10 % non-reducing sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and, interestingly, it showed the presence of a single band of (61±2) kDa when stained with Coomassie brilliant blue. The isolated protein was finally crystallized to see its homogeneity by batch crystallization method. Crystals were well formed with distinct edges. The isolated protein showed good pancreatic lipase inhibitory activity.

A comparative study of microporous polyolefin silica-based paper and cellulose paper substrates utilizing paper spray-mass spectrometry in drug analysis

This comparative study proposes an efficient technique by employing a synthetic substrate, which is commercially available, for direct, sensitive, efficient and fast analysis in Paper Spray Mass Spectrometry (PS-MS).

Inhibitors of human collagenase, MMP1

Different common drugs (Meloxicam, Tenoxicam and Piroxicam, and sodium alendronate) were tested both experimental and theoretically as inhibitors of interstitial human collagenase, also known as matrix metalloproteinase 1 (MMP-1). The in vitro collagenase activity, alone and in the presence of inhibitors, was quantified by the reaction with a fluorescent synthetic substrate and measuring the change of emission. Collagenase-inhibitor interaction was studied theoretically by computational calculations. Three among the four tested substances showed moderate inhibiting activity against the human collagenase.