• Journal of Microbiology and Biotechnology
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• v.31 no.10
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• pp.1455-1464
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• 2021

Trehalose is a non-reducing disaccharide in increasing demand for applications in food, nutraceutical, and pharmaceutical industries. Single-step trehalose production by trehalose synthase (TreS) using maltose as a starting material is a promising alternative process for industrial application due to its simplicity and cost advantage. Pseudomonas monteilii TBRC 1196 was identified using the developed screening method as a potent strain for TreS production. The TreS gene from P. monteilii TBRC 1196 was first cloned and expressed in Escherichia coli. Purified recombinant trehalose synthase (PmTreS) had a molecular weight of 76 kDa and showed optimal pH and temperature at 9.0 and 40℃, respectively. The enzyme exhibited >90% residual activity under mesophilic condition under a broad pH range of 7-10 for 6 h. Maximum trehalose yield by PmTreS was 68.1% with low yield of glucose (4%) as a byproduct under optimal conditions, equivalent to productivity of 4.5 g/l/h using enzyme loading of 2 mg/g substrate and high concentration maltose solution (100 g/l) in a lab-scale bioreactor. The enzyme represents a potent biocatalyst for energy-saving trehalose production with potential for inhibiting microbial contamination by alkaline condition.

• Journal of Microbiology and Biotechnology
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• v.29 no.5
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• pp.721-730
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• 2019

The probiotic Lactobacillus acidophilus KBL409 was encapsulated with alginate (Al) and alginate-chitosan (Al/Chi) through extrusion method. The sizes and zeta potentials of microspheres were measured to confirm encapsulation. To evaluate the protective effect of microspheres against gastrointestinal fluids, all the samples were exposed to simulated gastric fluids (SGFs, pH 1.5) at $37^{\circ}C$ for 1 or 2 h, followed by incubation with simulated intestinal fluids (SIFs, pH 6.5) for 2 h. The mucoadhesive ability of microspheres was evaluated using the intestinal epithelial cell line HT29-MTX. To extend the shelf-life of probiotics, lyoprotectants such as disaccharide and polysaccharide were mixed with free or encapsulated cells during the freeze-drying process. The size of the microspheres demonstrated a narrow distribution, while the zeta potentials of Al and Al/Chi-microspheres were $-17.9{\pm}2.3$ and $20.4{\pm}2.6mV$, respectively. Among all the samples, Al/Chi-encapsulated cells showed the highest survival rate even after exposure to SGF and SIF. The mucoadhesive abilities of Al and Al/Chi-microspheres were higher than 94%, whereas the free L. acidophilus showed 88.1% mucoadhesion. Ten percent of sucrose showed over 80% survival rate in free or encapsulated cells. Therefore, L. acidophilus encapsulated with Al and Al/Chi-microspheres showed higher survival rates after exposure to the gastrointestinal tract and better mucoadhesive abilities than the free cells. Also, sucrose showed the highest protective effect of L. acidophilus during the freeze-drying process.

• Bulletin of the Korean Chemical Society
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• v.14 no.2
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• pp.200-206
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• 1993

Structures of two polymorphs of ${\alpha},{\alpha}$-trehalose octaacetate monohydrate, $C_{28}H_{38}O_{19}\;{\cdot}\;H_2O$, have been studied by X-ray diffraction method. ${\alpha},{\alpha}$-trehalose (${\alpha}$-D-glucopyranosyl ${\alpha}$-D-glucopyranoside) is a nonreducing disaccharide. The polymorph I belongs to the monoclinic $P2_1$, and has unit cell parameters of a=10.725(l), b=15.110(4), c=11.199(5) ${\AA}$, ${\beta}=108.16(2)^{\circ}$ and Z=2. The polymorph II is orthorhombic $P2_12_12_1$, with a=13.684(4), b=15.802(4), c=17.990(9) ${\AA}$ and Z=4. The final R and R$_w$ values for monoclinic polymorph I are 0.043 and 0.048 and for orthorhombic polymorph II are 0.116 and 0.118, respectively. Those R values of polymorph II are high because the large thermal motions of acetyl groups and the poor quality of the crystal. The molecular conformations in the two polymorphs are similar. Both D-glucopyranosyl rings have chair $^4C_1$ conformations and atoms of glycosidic chain ${\alpha}(1{\rightarrow}1)$ linkage are coplanar. The primary acetate groups of the pyranose residues assume both gauche-trans conformations. The molecules of two polymorphs have pseudo-C$_2$ symmetry at glycosidic O(1) atom. The bond lengths and angles are normal compared with those in other acetylated sugar compounds. The molecules in the monoclinic crystal are held by the hydrogen bonds with the water molecules and by van der Waals forces.

• The Plant Pathology Journal
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• v.37 no.6
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• pp.566-579
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• 2021

Ralstonia syzygii subsp. indonesiensis (Rsi, former name: Ralstonia solanacearum phylotype IV) PW1001, a causal agent of potato wilt disease, induces hypersensitive response (HR) on its non-host eggplant (Solanum melongena cv. Senryo-nigou). The disaccharide trehalose is involved in abiotic and biotic stress tolerance in many organisms. We found that trehalose is required for eliciting HR on eggplant by plant pathogen Rsi PW1001. In R. solanacearum, it is known that the OtsA/OtsB pathway is the dominant trehalose synthesis pathway, and otsA and otsB encode trehalose-6-phosphate (T6P) synthase and T6P phosphatase, respectively. We generated otsA and otsB mutant strains and found that these mutant strains reduced the bacterial trehalose concentration and HR induction on eggplant leaves compared to wild-type. Trehalose functions intracellularly in Rsi PW1001 because addition of exogenous trehalose did not affect the HR level and ion leakage. Requirement of trehalose in HR induction is not common in R. solanacearum species complex because mutation of otsA in Ralstonia pseudosolanacearum (former name: Ralstonia solanacearum phylotype I) RS1002 did not affect HR on the leaves of its non-host tobacco and wild eggplant Solanum torvum. Further, we also found that each otsA and otsB mutant had reduced ability to grow in a medium containing NaCl and sucrose, indicating that trehalose also has an important role in osmotic stress tolerance.

• Journal of Microbiology and Biotechnology
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• v.32 no.7
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• pp.911-917
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• 2022

As valuable antibiotics, microbial natural products have been in use for decades in various fields. Among them are polyene compounds including nystatin, amphotericin, and nystatin-like Pseudonocardia polyenes (NPPs). Polyene macrolides are known to possess various biological effects, such as antifungal and antiviral activities. NPP A1, which is produced by Pseudonocardia autotrophica, contains a unique disaccharide moiety in the tetraene macrolide backbone. NPP B1, with a heptane structure and improved antifungal activity, was then developed via genetic manipulation of the NPP A1 biosynthetic gene cluster (BGC). Here, we generated a Streptomyces artificial chromosomal DNA library to isolate a large-sized NPP B1 BGC. The NPP B1 BGC was successfully isolated from P. autotrophica chromosome through the construction and screening of a bacterial artificial chromosome (BAC) library, even though the isolated 140-kb BAC clone (named pNPPB1s) lacked approximately 8 kb of the right-end portion of the NPP B1 BGC. The additional introduction of the pNPPB1s as well as co-expression of the 32-kb portion including the missing 8 kb led to a 7.3-fold increase in the production level of NPP B1 in P. autotrophica. The qRT-PCR confirmed that the transcription level of NPP B1 BGC was significantly increased in the P. autotrophica strain containing two copies of the NPP B1 BGCs. Interestingly, the NPP B1 exhibited a previously unidentified SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) inhibition activity in vitro. These results suggest that the Streptomyces BAC cloning of a large-sized, natural product BGC is a valuable approach for titer improvement and biological activity screening of natural products in actinomycetes.

• Korean Journal of Community Nutrition
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• v.28 no.3
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• pp.181-191
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• 2023

Objectives: The incidence of inflammatory bowel disease (IBD) is increasing globally, and excessive added sugar consumption has been identified as one of the contributing factors. In the context of IBD, it is essential to explore functional sweeteners that can improve metabolic health and minimize the risk of IBD-related symptoms. This review article aims to shed light on the effects of natural mono- and di-saccharides as alternative sweeteners, specifically focusing on potential benefits for IBD. Methods: A comprehensive literature review was performed using PubMed and Google Scholar databases with articles published after the year 2000. The search terms 'IBD', 'added sugar', 'sweeteners', 'mono-saccharide', and 'di-saccharide' were combined to retrieve relevant articles. A total of 21 manuscripts, aligning with the objectives of the study, were selected. Papers focusing on artificial or high-intensity sweeteners were excluded to ensure relevant literature selection. Results: Multiple studies have emphasized the association between the high consumption of added sugars such as simple sugars and the increased risk of developing IBD. This is suggested to be attributed to the induction of pro-inflammatory cytokine productions and dysbiosis of the gut microbiota. Consequently, there is a growing demand for safe and functional sweeteners, in particular mono- and di-saccharides, that can serve as alternatives for IBD patients. Those functional sweeteners regulate inflammation, oxidative stress, and Intestinal barrier protection, and restore microbiome profiles in various IBD models including cells, animals, and humans. Conclusions: Understanding these mechanisms resolves the link between how sugar consumption and IBD, and highlights the beneficial effects of natural alternative sweeteners on IBD when they were administered by itself or as a replacement for simple sugar. Further, exploration of this relationship leads us to recognize the necessity of natural alternative sweeteners in dietary planning. This knowledge could potentially lead to more effective dietary strategies for individuals with IBD.

• Journal of Embryo Transfer
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• v.17 no.2
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• pp.137-143
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• 2002

This study was conducted to evaluate the effect of sugar kinds and combination of various sugars, and straw size and pre-freezing time on motility of post-thaw spermatozoa in canine. In general, the extender was supplemented with fructose or glucose for semen freezing. The extender used in .this study was Tris-citric acid extender (Tris-buffer) supplemented with 20％ Egg-yolk, 8％ glycerol, 1％ Equex STM paste, and 70 mM sugars such as monosaccharide (fructose and xylose) and disaccharide(trehalose). To evaluate of sugar combination, the sugars supplemented in Tris-buffer. were combined such as control (fructose, xylose, trehalose), two combinations (Fru＋Tre, Fru＋Xy1, Tre＋Xy1) and three combinations (Fru＋Tre＋Xy1). The motility after CASA analysis in Fru＋Tre＋Xy1 was higher than those in fructose, trehalose, xylose, Fru＋Tre, Fru＋Xy1, Tre＋Xy1 (69 vs. 58, 61, 50, 65, 20, 54). However, the progressive motility after CASA analysis in Fru＋Tre group was higher than those in fructose, trehalose, xylose, Fru＋Xy1, Tre＋Xy1, Fru＋Tre＋Xy1 (59％ vs. 47, 55, 42, 13, 49, 44％). The survival rate of post-thaw spermatozoa in 0.25 $m\ell$ straw for 10 min pre-freezing was significantly higher than those in 0.25 $m\ell$ straw for 10 min, 0.25 and 0.5 $m\ell$ for 5 min (80＋0.0 vs. 65＋7, 68＋16, 58＋8％; P＜0.05). The results indicated that the progressive motility of post-thaw spermatozoa in 70 mM Fru ＋Tre (two combination) following CASA analysis and 0.25 ml straw for 10 min pre-freezing time could be better for freezing of semen in canine.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.11
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• pp.1699-1704
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• 2007

This experiment was carried out to evaluate the effects of adding encapsulated-glucose, glucose, sorbic acid or prefermented juice of epiphytic lactic acid bacteria (FJLB) on the fermentation quality and residual mono- and disaccharide composition of Italian ryegrass (Lolium multiflorum Lam) silages. The additive treatments were as follows: (1) control (no addition), (2) encapsulated-glucose addition at 0.5% for glucose, (3) glucose addition at 1%, (4) sorbic acid addition at 0.1%, (5) FJLB addition at a theoretical application rate of $2.67{\times}10^5$ CFU (colony forming unit) $g^{-1}$, on a fresh weight basis of Italian ryegrass. Although control and encapsulated-glucose treatments had higher contents of butyric acid (33.45, 21.50 g $kg^{-1}$ DM) and ammonia-N/Total nitrogen (114.91, 87.01 g $kg^{-1}$) as compared with the other treated silages, the fermentation in all silages was clearly dominated by lactic acid. This was well indicated by the low pH (4.38-3.59), and high lactic acid/acetic acid (4.39-22.97) and lactic acid content (46.85-121.76 g $kg^{-1}$ DM). Encapsulated-0.5% glucose and glucose addition increased lactic acid/acetic acid, and significantly (p<0.05) decreased ammonia-N/total nitrogen, and the contents of butyric acid and total volatile fatty acids (VFAs) as compared with the control. However, there were higher butyric acid and lower residual mono-and di-saccharides on the two treatments as compared with sorbic acid and FJLB addition, and their utilization efficiency of water soluble carbohydrates (WSC) was lower than that of both sorbic acid and FJLB additions. Sorbic acid addition showed the lowest content of ethanol and ammonia-N/total nitrogen, and the highest content of residual fructose and total mono-and disaccharides as well as the higher lactic acid/acetic acid value. Sorbic acid addition decreased the loss of mono-and disaccharides, and inhibited the activity of clostridial and other undesirable bacteria, and greatly increased the utilization efficiency of fermentable substrates by epiphytic LAB. FJLB addition had the lowest pH value and the highest lactic acid content among all additive treatments, with the most intensive lactic acid fermentation occurring in FJLB treated silage. This resulted in the faster accumulation of lactic acid and faster pH reduction. Sorbic acid and FJLB addition depressed clostridia or other undesirable bacterial fermentation which decreased the WSC loss and saved the fermentable substrate for lactic acid fermentation.

• Journal of Preventive Medicine and Public Health
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• v.7 no.1
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• pp.107-113
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• 1974

Many factors exert an influence on enzyme activity and thus on the rate of reactions that they catalyse. The most important of these factors are pH, temperature, substrate concentration, and the concentration of some inhibitors present. A solution of the enzyme diastase, which breaks down molecules of the polysaccharide starch to the disaccharide maltose by hydrolysis, was provided. Activity of this enzyme was measured by the rate at which starch was removed from the reaction mixture. These experiments were designed to study this reaction rate under varying conditions and the following results were obtained. 1. The range of optimum pH for this enzyme at room temperature was 4.0-7.0 and the optimum pH was 5.0. 2. The range of optimum temperatures for this enzyme at pH 7.0 was $30^{\circ}C-50^{\circ}C$ and the optimum temperature was $40^{\circ}C$. 3. The relationship between the enzyme activity and substrate concentration could be expressed by the Michaelis-Menten equation. The limiting velocity of this enzyme at room temperature and pH 7.0 was $415{\mu}g$ starch removed/ml of reaction mixture/min and $K_m$, Michaelis constant, was $343{\mu}g/ml$. 4. Inhibitors NaCl and $HgCl_2$ blocked this enzyme activity completely at 1% and 0.01% respectively.

• Journal of Life Science
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• v.13 no.1
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• pp.110-117
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• 2003

A thermotolerant yeast strain, Saccharomyces cerevisiae KNU5377 (abbreviated as KNU5377, was exposed to inorganic acids including sulfuric, nitric and hydrochloric acid. As a stressor, each inorganic acid is very easily dissociated in water, resulting in lowering environmental pH. When compared with a reference S. cerevisiae ATCC24858, KNU5377 could overcome such a severe condition containing a final 0.4% concentration of sulfuric acid or nitric acid to grow at the overnight culture, but this reference could not. Additionally, this strain showed a surprisingly strong tolerance by surviving despite of exposure to the regime of 0.35% of hydrochloric acid for over 90 min and also to 0.6% of sulfuric acid for 30 min. On the contrary, both strains could not survive against a final 0.45% concentration of nitric acid. This strain KNU5377 could produce ethanol of 3% in 2 days by using the fermentation medium containing a final 0.3% concentration of sulfuric arid. Moreover, change into a final 0.2% concentration of sulfuric acid caused this strain to enhance fermentation productivity up to about 4.5% even at $40^{\circ}C$. In exposure to a final 0.2% of sulfuric acid for 60 min, trehalose was most accumulated within 30 min in KNU5377, and this suggested a cellular defense system led by this disaccharide was profitable for this strain to lead to no morphological changes.