Resveratrol (RES) and genistein (GEN) are the dietary natural products known to possess chemopreventive property and also the ability to repair DNA damage induced by mutagens/carcinogens. It is believed that the therapeutic activity of these compounds could be primarily due to their interaction with nucleic acids but detailed reports are not available. We here explore the interaction of these drugs with nucleic acids considering DNA and RNA as a potential therapeutic target. The interaction of RES and GEN has been analysed in buffered solution with DNA [saline sodium citrate (SSC)] and RNA [tris ethylene diammine tetra acetic acid (TE)] using UV-absorption and Fourier transform infrared (FTIR) spectroscopy. The UV analysis revealed lesser binding affinity with nucleic acids at lower concentration of RES (P/D = 5.00 and 10.00), while at higher drug concentration (P/D = 0.75, 1.00 and 2.50) hyperchromic effect with shift in the ${\lambda}_{max}$ is noted for DNA and RNA. A major RES-nucleic acids complexes was observed through base pairs and phosphate backbone groups with K = $35.782\;M^{-1}$ and K = $34.25\;M^{-1}$ for DNA-RES and RNA-RES complexes respectively. At various concentrations of GEN (P/D = 0.25, 0.50, 0.75, 1.00 and 2.50) hyperchromicity with shift in the ${\lambda}_{max}$ from 260 $\rightarrow$ 263 om and 260 $\rightarrow$ 270 nm is observed for DNA-GEN and RNA-GEN complexes respectively. The binding constant (from UV analysis) for GEN-nucleic acids complexes could not be obtained due to GEN absorbance overlap with that of nucleic acids at 260 nm. Nevertheless a detailed analysis with regard to the interaction of these drugs (RES/GEN) with DNA and RNA could feasibly be understood by FTIR spectroscopy. The NH band of free DNA and RNA which appeared at $3550-3100\;cm^{-1}$ and $3650-2700\;cm^{-1}$ shifted to $3450-2950\;cm^{-1}$ and $3550-3000\;cm^{-1}$ in DNA-RES and RNA-RES complexes respectively. Similarly shifts corresponding to $3650-3100\;cm^{-1}$ and $3420-3000\;cm^{-1}$ have been observed in DNA-GEN and RNA-GEN complexes respectively. The observed reduction in NH band of free nucleic acids upon complexation of these drugs is an indication of the involvement of the hydroxyl (OH) and imino (NH) group during the interaction of the drugs and nucleic acids (DNA/RNA) through H-bonded formation. The interaction of RES and GEN with bases appears in the order of G $\geq$ T > C > A and A > C $\geq$ T > G. Further interaction of these natural compounds with DNA and RNA is also supported by changes in the vibrational frequency (shift/intensity) in symmetrical and asymmetrical stretching of aromatic rings of drugs in the complex spectra. No appreciable shift is observed in the DNA and RNA marker bands, indicating that the B-DNA form and A-family conformation of RNA are not altered during their interaction with RES and GEN.
The ginsenoside-hydrolyzing β-glucosidase gene (bgy2) was cloned from Lactobacillus brevis. We expressed this gene in Escherichia coli BL21(DE3), isolated the resulting protein, and then utilized the enzyme for the biotransformation of ginsenosides. The bgy2 gene contains 2,223 bp, and encodes a protein of 741 amino acids that is a member of glycosyl hydrolase family 3. β-Glucosidase (Bgy2) cleaved the outer glucose moieties of ginsenosides at the C-20 position, and the inner glucose at the C-3 position. Under optimal conditions (pH 7.0, 30℃), we used 0.1 mg/ml Bgy2 in 20 mM sodium phosphate buffer (PBS) for enzymatic studies. In these conditions, 1.0 mg/ml ginsenoside Rb1 and ginsenoside F2 were converted into 0.59 mg/ml ginsenoside Rd and 0.72mg/ml compound K, with molar conversion productivities of 69% and 91%, respectively. In pharmaceutical and commercial industries, this recombinant Bgy2 would be suitable for producting ginsenoside Rd and compound K.
The production of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS) are known to be modulated by a variety of factors. Recent study showed that endotoxin-induced NO synthesis and iNOS expression were greatly enhanced by elevation of extracellular glucose concentration in murine macrophages. Although this was suggested to be due to the activation of protein kinase C (PKC) via sorbitol pathway, there was lack of evidence for this speculation. This study was performed to delineate the underlying intracellular mechanisms of glucose-enhancing effect on endotoxin-induced NO production in Raw264.7 macrophages. The levels of NO release induced by lipopolysaccharide (LPS) significantly increased by the treatment of glucose in a concentration dependent manner and also, this effect was observed in LPS-preprimed cells. Concurrent incubation of cells with PKC inhibitors, H-7 or chelerythrine, and LPS resulted in the diminution of NO production regardless of glucose concentration but this was not in the case of LPS-prepriming, that is, chelerythrine showed a minimal effect on the glucose- enhancing effect. PMA, a PKC activator, did not show any significant effect on glucose-associated NO production. Modulation of sorbitol pathway with zopolrestat, an aldose reductase inhibitor, did not affect LPS-induced NO production and iNOS expression under high glucose condition. And also, sodium pyruvate, which is expected to normalize cytosolic $NADH/NAD^+$ ratio, did not show any significant effect at concentrations of up to 10 mM. Glucosamine marginally increased the endotoxin-induced nitrite release in both control and high glucose treated group. 6-diazo-5-oxonorleucine (L-DON) and azaserine, glutamine: fructose- 6-phosphate amidotransferase (GFAT) inhibitors, significantly diminished the augmentation effect of high glucose on endotoxin-induced NO production. On the other hand, negative modulation of GFAT inhibitors was not reversed by the treatment of glucosamine, suggesting the minimal involvement, if any, of glucosamine pathway in glucose-enhancing effect. In summary, these results strongly suggest that the hexosamine biosynthesis pathway and the activation of PKC via sorbitol pathway do not contribute to the augmenting effect of high glucose on endotoxin induced NO production in macrophage-like Raw264.7 cells.
Kwun, Hyeon-Sook;Nam, Seoul-Hee;Park, Min-Kyoung;Cho, Mi SooK;Cheon, Sae-Hee
Journal of the Korea Academia-Industrial cooperation Society
/
v.15
no.9
/
pp.5644-5651
/
2014
Propolis is an extremely safe natural antimicrobial substance that has been reported to have powerful antibacterial efficacy. The aim of this study was to evaluate the inhibitory effects of propolis against Candida albicans (C. albicans). Propolis was collected from the honey bee Apis mellifera. The strain of C. albicans was cultivated overnight in liquid media incubated at $37^{\circ}C$. The antimicrobial activity was investigated using phosphate buffered saline (PBS), 3% sodium hypochlorite (NaOCl), 0.1% chorhexidine (CHX), and propolis extracts ($5{\mu}l/ml$, $10{\mu}l/ml$). C. albicans were sensitive to 3% NaOCl, 0.1% CHX, and propolis ($5{\mu}l/ml$, $10{\mu}l/ml$) with zones of inhibition of 15, 14.5, 16, and 17 mm, respectively. The CFU of PBS, 3% NaOCl, 0.1% CHX, $5{\mu}l/ml$ and $10{\mu}l/ml$ of propolis led a 1, 7, 7, 5 and 7-log reduction. Among the groups tested, C. albicans was most sensitive to $10{\mu}l/ml$ of propolis, which showed the largest inhibition zones. Therefore, propolis can be a new antimicrobial therapy for oral mucosa disease in traditional medicine.
As functionality investigation of Korean traditional soybean fermentation foods, an antihypertensive peptide was separated during Chunggugjang fermentation by Bacillus subtilis CH-1023 and investigated inhibitory effect against angiotensin converting enzyme. After incubation at $20^{\circ}C,\;30^{\circ}C,\;40^{\circ}C,\;50^{\circ}C,\;60^{\circ}C$ for the $0{\sim}72$ hrs, protein content, protease activity and angiotensin converting enzyme inhibitory rate were determined. The protein content and protease activity were increased and reached maximum at 60 hrs fermentation with $40^{\circ}C$ and decreased after the 60 hrs fermentation. The optimum condition for antihypertensive peptide from Chunggugjang was appeared for 60 hrs at $40^{\circ}C$. Crude extract of Chunggugjang was partially purified by Amicon YM-3 membrane filtration and Sephadex G-10, G-25 gel filtration. The purified peptide showed inhibitory rate of 94.3% with 0.5 mg peptide content. The most prominent amino acid composition of the peptide from Chunggugjang was alanine, followed by phenylalanine, histidine.
The enzyme xylose isomerase (E.C. 5.3.1.5, XI) is responsible for the conversion of an aldose to ketose, especially xylose to xylulose. Owing to the ability of XI to isomerize glucose to fructose, this enzyme is used in the food industry to prepare high-fructose corn syrup. Therefore, we studied the characteristics of XI from Anoxybacillus kamchatkensis G10, a thermophilic bacterium. First, the gene coding for XI (xylA) was inserted into the pET-21a(+) expression vector and the construct was transformed into the Escherichia coli competent cell BL21 (DE3). The expression of recombinant XI was induced in the absence of isopropyl-thio-${\beta}$-galactopyranoside and purified using Ni-NTA affinity chromatography. The optimum temperature of recombinant XI was $80^{\circ}C$ and measurement of the heat stability indicated that 55% of residual activity was maintained after 2 h incubation at $60^{\circ}C$. The optimum pH was found to be 7.5 in sodium phosphate buffer. Magnesium, manganese, and cobalt ions were found to increase the enzyme activity; manganese was the most effective. Additionally, recombinant XI was resistant to the presence of $Ca^{2+}$ and $Zn^{2+}$ ions. The kinetic properties, $K_m$ and $V_{max}$, were calculated as 81.44 mM and $2.237{\mu}mol/min/mg$, respectively. Through redundancy analysis, XI of A. kamchatkensis G10 was classified into a family containing type II XIs produced by the genera Geobacillus, Bacillus, and Thermotoga. These results suggested that the thermostable nature of XI of A. kamchatkensis G10 may be advantageous in industrial applications and food processing.
Pharmacokinetics of prednisolone and prednisone undergoing reversible interconversion were analyzed from the model including this metabolic process. Blood samples were drawn serially upto 12 hours after I,V. bolus injection of 1 mg/kg prednisolone sodium phosphate and prednisone into 8 dogs as a crossover manner. Plasma concentrations of those two steroids were simultaneously measured with the method of HPLC. After injection, plasma concentrations of administered prednisolone and prednisone were declined with a biexponential pattern and their metabolic partner was rapidly formed. Plasma concentrations of those metaboite were decayed in parallel with their parent steroids throught the elimination phase. Apparent clearances of prednisolone and prednisone were $11.1{\pm}2.0\;ml/min/kg$ and $45.9{\pm}6.4\;ml/min/kg$, and they were underestimated by 29.4% and 33.6% compared to their real clearances$(15.7{\pm}4.4\;and\;69.2{\pm}17.7\;ml/min/kg)$ estimated using reversible interconversion model. Apparent volume of distribution of prednisolone$(1.32{\pm}0.43\;L/kg)$ and prednisone$(4.81{\pm}2.75\;L/kg)$ were overestimated by 53.5 and 52.7% and were compared to the real volumes $(0.86{\pm}0.30\;and\;3.15{\pm}2.13\;L/kg)$. Mean residence time of prednisolone$(2.0{\pm}0.61\;h)$ and prednisone$(1.74{\pm}0.74\;h)$ were much longer than the real sojourn time$(0.93{\pm}0.26\;and\;0.88{\pm}0.54\;h)$. Essential clearances In the reversible interconversion were greater as following orders: $Cl_{21}$(44.3 ml/min/kg) > $Cl_{20}$(24.2 ml/min/kg) > $Cl_{12}$ (7.9 ml/min/kg) > $Cl_{10}$(7.8 ml/min/kg). Estimated mean values of RF, EE, $%X^1_{ss}$ and $RHO^2_1$ were $0.31{\pm}0.10$, $1.49{\pm}0.23$, $69.3{\pm}16.7%$ and $0.65{\pm}0.10$, respectively. These results suggested that true pharmacokinetic parameters estimated from the model including reversible interconversion were significantly different from the apparent parameters estimated from the conventional mamillary model, and disposition of these two steroids seemed to be well explained by the model including reversible interconversion.
Bacillus polyfermenticus SCD which is commonly called as Bisroot strain is being used for functional foods through the treatment of long-term intestinal disorders, since the live strains in the form of active endospores can successfully reach the target intestine in humans. The cells of B. polyfermenticus SCD were treated for 4h in artificial gastric juice (pH 2.0,3.0) and bile acid. Final viability of the strain in artificial gastric Juice (pH 2.0, 3.0) is reached to 62.8% and 81.2% respectively B. polyfermenticus SCD is resistant to antibiotics such as streptomycin, rifampicin, nystatin and ampicilin. B. polyfermenticus SCD is well known supplies the nutrients by synthesizing vitamin $B_1$, $B_2$, C and K. B. polyfermenticus SCD produces various digestive enzymes and the enzymes enable to completely digest diets in our body. Above all, $\alpha$-amylase and pretense activities are very higher than B. subtilis KCTC 1020, about two fold and twenty five fold respectively. B. polyfermenticus SCD is very stable during long-term storage period in phosphate buffers of wide-range pH, solutions of various concentrations of sodium chloride, 5% glucose solution and water.
One bacterium, which showed strong antagonistic activity against H. pylori KCCM 41756, was isolated from kimchi. The strain NO1 was designated as Lactobacillus plantarum NO1 based on Gram staining, biochemical properties, and 16S rRNA gene sequencing. The culture medium $(2{\sim}4{\mu}g/ml)$ of Lb. plantarum NO1 reduced $(40{\sim}60%)$ the urease activity of H. pylori KCCM 41756. Lb. plantarum NO1 inhibited the binding of H. pylori to human gastric cancer cell line, AGS cells, by more than 33%. Lb. plantarum NO1 exhibited high viability (maintained initial viable cell count of $10^9CFU/ml$) in 0.05 M sodium phosphate buffer (pH 3.0) for 2 h, in artificial gastricjuice for 2 h and in 0.3%, 0.5% oxgall for 24 h. Hemolysis phenomena did not observed when Lb. plantarum NO1 was incubated in the blood agar media. We concluded that Lb. plantarum NO1 can be a good candidate as a probiotic, harboring anti-H. pylori activity.
There are two problems to be solved by our efforts in the enamel frit. One is how we can cover the enamel frit thin with complete milk white as possible, and the other is how it can be, made resistant for chemicals than before one. The frit which can solved the two problems just mentioned above is titanium enamel frit. This frit has been developed in America after War Ⅱ, and now the research for concerning antimony frit into titanium frit is under development entirely. In order to develope the enamel industry in Korea, it is urgent problem to convert antimony frit into titanium frit. By the way the titanium frit is emulsified titanium oxide crystal which made through reheating the supersaturated solution of titanium oxide in the basis of glass. Unfortunately, there are many obscure points in active fact or which influence on its composition and characteristics yet. However, this task was tried for the first in Korea. As first step, the test was carried on the reference books, and we can be possible convert antimony frit into titanium frit as a result of this experiment. As a conclusion, for the purpose of developing the enamel industry in Korea, we studied that the research for converting antimony enamel frit which has been used popularly into titanium enamel frit which is more economic and resistant for chemicals. As a result of experiments, the following points concerning with titanium frit have become clearly. 1. It is better when the composition of titanium enamel frit has as following table.Man Duck San Silica 24 An Yang Feldspar 20 Borax 28 Sodium Nitrate 4 Cryolite 7 Calcium Carbonate 3.6∼1 Titanium Oxide 10 Calcium phosphate 0 ∼3.2 Calcium Fluoride 0∼1.8 Antimony Oxide 0∼0.5 2. The amount of $TiO_2$, to be added is $10%\;to\;12{%,\;CaF_2\;is\;under\;1.8%,\;P_2O_5\;is\;under\;1.6%,\;Sb_2O_3\;is\;under\;0.5%$. 3. In the titanium frit, the limit of iron oxide amount to be included is under 0. 5%. 4. Comparing the titanium enamel frit with antimony enamel frit not only the titanium frit can be savely 20.6% in the price of raw materials, but one time of glazing and heating process is omitted in each case, and it is known the titanium frit is more resistant for chemicals than antimony frit.
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