• Title/Summary/Keyword: ethanol precipitation

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Functional Characterization of an Exopolysaccharide Produced by Bacillus sonorensis MJM60135 Isolated from Ganjang

  • Palaniyandi, Sasikumar Arunachalam;Damodharan, Karthiyaini;Suh, Joo-Won;Yang, Seung Hwan
    • Journal of Microbiology and Biotechnology
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    • v.28 no.5
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    • pp.663-670
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    • 2018
  • The present study focused on the production, characterization, and in vitro prebiotic evaluation of an exopolysaccharides (EPS) from Bacillus sonorensis MJM60135 isolated from ganjang (fermented soy sauce). Strain MJM60135 showed the highest production ($8.4{\pm}0.8g/l$) of EPSs compared with other isolates that were screened for EPS production based on ropy culture morphology. Furthermore, MJM60135 was cultured in 5 L of medium and the EPS was extracted by ethanol precipitation. The emulsification activity of the EPS was higher in toluene than in o-xylene. Fourier transform infrared spectroscopy analysis showed the presence of hydroxyl and carboxyl groups and glycosidic linkages. The isolated EPS contained mannose and glucose, as observed by thin-layer chromatography analysis of the EPS hydrolysate. Lactic acid bacteria (LAB) and pathogenic E. coli K99 and Salmonella enterica serovar Typhimurium were tested for their growth utilizing the EPS from B. sonorensis MJM60135 as the sole carbon source for its possible use as a prebiotic. All the tested LAB exhibited growth in the EPS-supplied medium compared with glucose as carbon source, whereas the pathogenic strains did not grow in the EPS-supplied medium. These findings indicate that the EPS from B. sonorensis MJM60135 has potential application in the bioremediation of hydrocarbons and could also be used as a prebiotic.

Production of Inulase Using Jerusalem Artichoke Tuber Extract (돼지감자 추출물을 이용한 이눌라아제 생산)

  • Choi, Weon-Sang;Choe, Yong-Kyung;Kim, Su-Il;Byun, Si-Myung
    • Applied Biological Chemistry
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    • v.27 no.4
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    • pp.238-244
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    • 1984
  • To produce inulase from Kluyveromyces fragilis No.351 using Jerusalem artichoke tuber extract, optimization of the condition was conducted. As results, the optimal concentration of artichoke tuber extract was 3.5 % and bactocasitone showed better production than yeast extract. The optimal temperature and pH were $30^{\circ}C$ and 5.5, respectively. The addition of $NH_4H_2PO_4$ increased the enzyme production. Inulase synthesis was growth-associated and the enzyme production increased as concentration of dissolved oxygen increased. A higher quantity of industrial grade inulase was prepared by the combination of ultrafiltration and ethanol precipitation with 72% recovery.

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Study of Macrophage Activation and Structural Characteristics of Purified Polysaccharides from the Fruiting Body of Hericium erinaceus

  • Lee, Jong-Seok;Min, Kyoung-Min;Cho, Jae-Youl;Hong, Eock-Kee
    • Journal of Microbiology and Biotechnology
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    • v.19 no.9
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    • pp.951-959
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    • 2009
  • Most, if not all, Basidiomycetes mushrooms have biologically active polysaccharides showing potent antitumor activity with immunomodulating properties. These polysaccharides have various chemical compositions and belong primarily to the $\beta$-glucan group. In this study, the crude water-soluble polysaccharide HEF-P, which was obtained from the fruiting body of Hericium erinaceus by hot water extraction and ethanol precipitation, was fractionated by DEAE-cellulose and Sepharose CL-6B column chromatographies. This process resulted in four polysaccharide fractions, named HEF-NP Fr I, HEF-NP Fr II, HEF-AP Fr I, and HEF-AP Fr II. Of these fractions, HEF-AP Fr II was able to upregulate the functional events mediated by activated macrophages, such as production of nitric oxide and expression ofcytokines (IL-1${\beta}$ and TNF-${\alpha}$). The molecular mass of HEF-AP Fr II was estimated by gel filtration to be 13 kDa. Its structural characteristics were investigated by a combination of chemical and instrumental analyses, including methylation, reductive cleavage, acetylation, Fourier transform infrared spectroscopy (FT-IR), and gas chromatography-mass spectrometry (GC-MS). Results indicate that HEF-AP Fr II is a low-molecular-mass polysaccharide with a laminarin-like triple helix conformation of a ${\beta}$-1,3-branched-${\beta}$-1,6-glucan.

Study of Macrophage Activation and Structural Characteristics of Purified Polysaccharide from the Fruiting Body of Cordyceps militaris

  • Lee, Jong-Seok;Kwon, Jeong-Seok;Won, Dong-Pil;Lee, Jung-Hyun;Lee, Keun-Eok;Lee, Shin-Young;Hong, Eock-Kee
    • Journal of Microbiology and Biotechnology
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    • v.20 no.7
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    • pp.1053-1060
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    • 2010
  • Cordyceps militaris, an entomopathogenic fungus belonging to the class Ascomycetes, has been reported to have beneficial biological activities such as hypoglycemic, anti-inflammatory, antitumor, antimetastatic, hypolipidemic, immunomodulatory, and antioxidant effects. In this study, the crude water-soluble polysaccharide CMP, which was obtained from the fruiting body of C. militaris by hot water extraction and ethanol precipitation, was fractionated by DEAE-cellulose and Sepharose CL-6B column chromatographies. This process resulted in three polysaccharide fractions, termed CMP Fr I, CMP Fr II, and CMP Fr III. Of these fractions, CMP Fr II, with an average molecular mass of 127 kDa, was able to upregulate effectively the phenotypic functions of macrophages such as NO production and cytokine expression. The chemical property of the stimulatory polysaccharide, CMP Fr II, was determined based on its monosaccharide composition, which consisted of glucose (56.4%), galactose (26.4%), and mannose (17.2%). Its structural characteristics were investigated by a combination of chemical and instrumental analyses, including methylation, reductive cleavage, acetylation, Fourier transform infrared spectroscopy (FTIR), and gas chromatography-mass spectrometry (GCMS). Results indicated that CMP Fr II consisted of the (1${\rightarrow}$4) or (1${\rightarrow}$2) linked glucopyranosyl or galactopyranosyl residue with a (1${\rightarrow}$2) or (1${\rightarrow}$6) linked mannopyranosyl, glucopyranosyl, or galactopyranosyl residue as a side chain. The configuration of the ${\beta}$-linkage and random coil conformation of CMP Fr II were confirmed using a Fungi-Fluor kit and Congo red reagent, respectively.

Effect of Template Existence on the Textural Properties of Iron-based Catalyst for Fischer Tropsch Reaction

  • Sirikulbodee, Papahtsara;Tungkamani, Sabaithip;Phongksorn, Monrudee;Ratana, Tanakorn;Sornchamni, Thana
    • International Journal of Internet, Broadcasting and Communication
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    • v.7 no.2
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    • pp.96-104
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    • 2015
  • Fischer Tropsch reaction is one of the interesting topic for renewable and clean energy. Polymerization of carbon monoxide or carbon dioxide with hydrogen over metal supported catalyst can produce long chain hydrocarbons. Synthetic liquid hydrocarbons are promising alternative to fossil fuels. This research work has been focused on the synthesis of Fe based catalyst for Fischer Tropsch reaction. Mesoporous silica (MS) support prepared by a precipitation method using two different washing solution, distilled water (DW) and acid in ethanol solution (ET), and different calcination temperature. Then, Fe/MS was prepared by an incipient wetness impregnation method. All of samples were systematically characterized using various physical and chemical techniques. TEM and XRD analysis were used to ensure that the cubic Ia3d mesostructure is stable after calcination. FTIR spectra are useful to ascertain the existence of template in the support. TPR studies were also used to understand the nature of Fe species and their reducibility. The results reveal that washing the support with distilled water and calcination at $550^{\circ}C$ can efficiently remove the triblock copolymer templates. The existence of template in the support affects the textural properties of all catalyst investigated.

An Anticoagulant Polysaccharide Isolated from Ganoderma lucidum (한국산 영지버섯에서 분리된 항응고성 다당)

  • 나경수;이별나;이현순;권미향
    • The Korean Journal of Food And Nutrition
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    • v.10 no.3
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    • pp.375-381
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    • 1997
  • The anticoagulant polysaccharide was screened from the ediable mushrooms. Among them, alkali extract of Ganoderma lucidum showed the highest activity in aPTT. A crude polysaccharide fraction (GL-I) was prepared from the 1N NaOH solution extract of Ganoderma lucidum followed by methanol-reflux, precipitation with ethanol, dialysis and lyophilization. GL-I inhibited the intrinsic pathway in blood coagulation pathway and exhibited concentration dependent anticoagulation effects. The anticoagulant activity of GL-I was decreased greatly by periodate oxidation, but was not changed by pronase digestion. These suggest that carbohydrate moiety may be related to the anticoagulant activity. GL-I consisted of glucose, galactose, fucose, xylose, mannose, arabinose in a molar ratio of 19.3:3.0:2.3:1.3:1.0:0.3. GL-I was partially purified on the DEAE-Toyopearl 650C(GL-IalongrightarrowGL-If) and on the Sephadex G-100(GL-Ic-ilongrightarrowGL-Ic-II).

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Cellulolytic Enzymes from Acrophialophora nainiana

  • Punnapayak, Hunsa
    • Proceedings of the Korean Society for Applied Microbiology Conference
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    • 2005.06a
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    • pp.245-247
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    • 2005
  • A cellulolytic fungus isolated from Agave plantation in northeastern Thailand was identified as Acrophialophora nainiana. The fungus was capable of growing at pH between 3 - 7 and 25 - 45 $^{\circ}C$, with the optimum conditions at pH 5.0 and 40 $^{\circ}C$. The wild isolate produced cellulases, comprising of exoglucanase (0.019 U/mg protein), endoglucanase (0.366 U/mg protein), and ${\beta}$-glucosidase (0.001 U/mg protein). Mutations with UV and NTG produced the UV 10-2 mutant with cellulases activities including exoglucanase (0.093 U/mg protein), endoglucanase (0.585 U/mg protein), and ${\beta}$-glucosidase (0.013 U/mg protein). Purification of the enzymes with ultrafiltration, ammonium sulfate precipitation, and ion-exchange chromatography yielded the maximal cellulase specific activities of 2.736 U/mg protein (exoglucanase), 0.235 U/mg protein (endoglucanase), and 0.008 U/mg protein (${\beta}$-glucosidase). The mutant's cellulases were the most active at pH 5.0 and 60 $^{\circ}C$. Ion-exchange chromatography revealed that A. nainiana UV 10-2 cellulases were comprised of two peaks with one peak showing the single endoglucanase activity while the other peak showed a mixture of the three enzyme activities. Production of A. nainiana UV 10-2 cellulases using banana leaf stalk as the sole carbon source gave comparable yields to that of the pure ${\alpha}$-cellulose. The enzymes were used in the simultaneous saccharification and fermentation (SSF) of plant residue (Coix aquatica) along with Kluveromyces marxianus to produce ethanol. Moreover, when the enzymes were used in the bioscouring process of fabric, the desiravle traits of textile processing including immediate water absorbency, increased in whiteness and reduction of yellowness of the treated fabric were observed.

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Distribution of poly-${\gamma}$-glutamate (${\gamma}$-PGA) producers in Korean fermented foods, Cheongkukjang, Doenjang, and Kochujang

  • Kang, Seong-Eun;Rhee, Joo-Hyung;Park, Chung;Sung, Moon-Hee;Lee, In-Hyung
    • Food Science and Biotechnology
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    • v.14 no.5
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    • pp.704-708
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    • 2005
  • Poly-y-glutamate (${\gamma}$-PGA) has great potential as a biodegradable polymer in a broad range of industrial fields such as food, cosmetics, medicine and water treatment. In order to isolate ${\gamma}$-PGA producers that are suitable for specific industrial applications, 653 Bacillus-like strains were isolated from 439 varieties of three Korean fermented foods, Cheongkukjang, Doenjang, and Kochujang, which were collected from different regions across Korea. A very high level of ${\gamma}$-PGA production was demonstrated in 4.7%, 1.8%, and 3.0% of the Bacillus-like strains isolated from Cheongkukjang, Doenjang, and Kochujang samples, respectively, which produced a viscous substance to such extent that it overflowed to the lid of the plate on the glutamate-dependent ${\gamma}$-PGA production plates. On glutamate-independent ${\gamma}$-PGA production plates, 5.1%, 5.9%, and 6.1% of Bacillus-like strains isolated from Cheongkukjang, Doenjang, and Kochujang samples, respectively, showed high production. The maximum ${\gamma}$-PGA production yields were 32.5 g/L and 5 g/L, depending on the purification methods in the glutamate-dependent media, with the higher yield resulting from a simple precipitation of ${\gamma}$-PGA by either methanol or ethanol and dialysis. The viscous substance produced by each strain showed different morphological characteristics, suggesting that isolated ${\gamma}$-PGA producers could produce various types of ${\gamma}$-PGA.

DNA Extraction from Protozoan Oocysts/Cysts in Feces for Diagnostic PCR

  • Hawash, Yousry
    • Parasites, Hosts and Diseases
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    • v.52 no.3
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    • pp.263-271
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    • 2014
  • PCR detection of intestinal protozoa is often restrained by a poor DNA recovery or by inhibitors present in feces. The need for an extraction protocol that can overcome these obstacles is therefore clear. $QIAamp^{(R)}$ DNA Stool Mini Kit (Qiagen) was evaluated for its ability to recover DNA from oocysts/cysts directly from feces. Twenty-five Giardia-positive, 15 Cryptosporidium-positive, 15 Entamoeba histolytica-positive, and 45 protozoa-free samples were processed as control by microscopy and immunoassay tests. DNA extracts were amplified using 3 sets of published primers. Following the manufacturer's protocol, the kit showed sensitivity and specificity of 100% towards Giardia and Entamoeba. However, for Cryptosporidium, the sensitivity and specificity were 60% (9/15) and 100%, respectively. A series of optimization experiments involving various steps of the kit's protocol were conducted using Cryptosporidium-positive samples. The best DNA recoveries were gained by raising the lysis temperature to the boiling point for 10 min and the incubation time of the InhibitEX tablet to 5 min. Also, using a pre-cooled ethanol for nucleic acid precipitation and small elution volume ($50-100{\mu}l$) were valuable. The sensitivity of the amended protocol to Cryptosporidium was raised to 100%. Cryptosporidium DNA was successfully amplified by either the first or the second primer set. When applied on parasite-free feces spiked with variable oocysts/cysts counts, ${\approx}2$ oocysts/cysts were theoretically enough for detection by PCR. To conclude, the Qiagen kit with the amended protocol was proved to be suitable for protozoan DNA extraction directly from feces and support PCR diagnosis.

Screening of Anticoagulant PoIysaccharides from Edible Plants (식물로 부터 혈액 항응고 활성 다당류의 검색)

  • Kweon, Mee-Hyang;Park, Mee-Kyung;Ra, Kyung-Soo;Sung, Ha-Chin;Yang, Han-Chul
    • Applied Biological Chemistry
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    • v.39 no.2
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    • pp.159-164
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    • 1996
  • Screening of anticoagulant activity was conducted for the hot water extracts of 73 kinds of medicinal herbs, 41 kinds of Korean edible plants, and 5 kinds of sea weeds using plasma recalcification test(Tr). In the first screening several extracts of the plants, Alisma calndiculatum, Corydalis ternata Panax notoginseng, Allium sativum, Ganderma luidum, Codium fragile, showed high activities. When the plants were reextracted with various solvent conditions, acidic water extracts of Codium fragile showed the highest activity in APTT. A crude polysaccharide fraction(CF-1) was prepared by methanol reflux, ethanol precipitation, dialysis and Iyophilization of the acid extracts. CF-1 comprised 80.8% total sugar consisting of arabinose, galactose and glucose as the main monomers, 8.7% protein, and 13.3% sulfate. The anticoagulant activity of CF-1 was not changed by pronase digestion, but decreased by periodate oxidation, and this indicated that the anticoagulant activity was attributed to the polysaccharide portion.

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