• Title/Summary/Keyword: Production and Purification

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Novel Suspension-Phase Enzyme Reaction System Using Insoluble Extrusion Starch as Glycosyl Donor for Intermolecular Transglycosylation of L-Ascorbic Acid

  • Kim, Tae-Kwon;Jung, Se-Wook;Go, Young-Hoon;Lee, Yong-Hyun
    • Journal of Microbiology and Biotechnology
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    • v.16 no.11
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    • pp.1678-1683
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    • 2006
  • A novel suspension-phase enzyme reaction system for the intermolecular transglycosylation of L-ascorbic acid into 2-O-${\alpha}$-D-glucopyranosyl L-ascorbic acid supplementing extrusion starch as the glycosyl donor was developed using cyclodextrin glucanotransferase from Thermoanaerobacter sp. A high conversion yield compared to the conventional soluble-phase enzyme reaction system using cyclodextrins and soluble starch was achieved. The optimal reaction conditions were 2,000 units of cycIodextrin glucanotransferase, 20 g/l of L-ascorbic acid, and 50 g/l of extrusion starch at $50^{\circ}C$ for 24 h. The new suspension-phase enzyme reaction system also exhibited several distinct advantages other than a high conversion yield, including a lower accumulation of oligosaccharides and easily separable residual extrusion starch by centrifugation or filtration in the reaction mixture, which will facilitate the purification of 2-O-${\alpha}$-D-glucopyranosyl L-ascorbic acid. The new suspension-phase enzyme reaction system seems to be potentially applicable as the industrial process for the production of thermally and oxidatively stable 2-O-${\alpha}$-D-glucopyranosyl L-ascorbic acid.

Product of inulo-oligosaccharides from inulin by endo-inulinase activiting enzyme and Its deletion mutant protein from CFTase

  • Kim, Byeong-U;Ryu, Hye-Gyeong;Yu, Dong-Ju;Kim, Hyeon-Jeong
    • 한국생물공학회:학술대회논문집
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    • 2002.04a
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    • pp.528-530
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    • 2002
  • We have previously reported recombinant deletion mutant was constructed from cycloinulo-oligosaccharide fructanotransferase(CFTase) gene of Xanthmonas oryzae MGL21. Purification of the mutant protein from E. coli and reation condition for the production of inulo-oligosaccharide(ISO) were studied. The molecular mass of the CFTase deletion mutant protein was estimated to be 90kDa by SDS- Polyacrylamide gel electrophoresis. Optimum reaction pH and temperature were pH6.5 and $40^{\circ}C$, respectively. Under optimal reaction conditions, endo-inulinase activating enzyme was rapidly produced ISO from inulin. Components were DP(degree of polymerization) 3and 4 with trace amount of smaller oligosaccharides.

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Molecular Detection of Verotoxigenic Escherichia coli (VTEC) from Animal Feces for Screening VTEC-shedders

  • Kobayashi, Y.;Sato, M.;Taguchi, H.;Koike, S.;Nakatsuji, H.;Tanaka, K.
    • Asian-Australasian Journal of Animal Sciences
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    • v.17 no.3
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    • pp.423-427
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    • 2004
  • Seventy-six animals including cattle, sheep, horses, 6 species of zoo animals were employed for collection of fresh feces in order to detect verotoxigenic Esherichia coli (VTEC) by safe, quick and sensitive PCR-based molecular methods. Bacterial cell disruption with bead-beating followed by bacterial DNA purification with hydroxyapatide chromatography and gel filtration allowed DNA preparation from animal feces with high recovery and purity. A mountain goat was firstly shown by PCR and sequencing to shed verotoxin 2 gene (vt2) that was used to generate vt2 probe and second primer set for nested PCR to attempt more sensitive detection. Most sensitive nested PCR revealed that 45% of tested cattle and 47% of tested zoo animals were VTEC-positive, while least sensitive normal PCR detected VTEC from none of these animals except a mountain goat. Moderately sensitive detection by PCR in combination with hybridization suggested that the VTEC density varied between the VTEC-positive cattle.

Characterization of γ-Aminobutyric acid(GABA) produced by a lactic acid bacterium from button mushroom bed

  • Lee, Yun-Seok;Song, Tae-Young;Kong, Won-Sik;Yoon, Min-Ho
    • Journal of Mushroom
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    • v.11 no.4
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    • pp.181-186
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    • 2013
  • ${\gamma}$-Aminobutyric acid(GABA) is a four carbon non-protein amino acid that has several well-known physiological functions, such as a postsynaptic inhibitory neurotransmitter in the brain and induction of hypotensive and tranquilizer effects. A lactic acid bacterium was isolated from button mushroom bed, which is showing high GABA productivity by TLC or HPLC analysis. The strain was identified as Lactobacillus hilgardii by analysis of 16S rDNA gene sequence. When the maximum production of GABA by L. hilgardii was investigated with various concentration of monosodium glutamate, the yield of GABA reached to be 53.65 mM at 1% mono sodium glutamate (MSG) in flask cultivation. A Glutamate decarboxylase (GAD) enzyme, which was known to convert MSG to GABA, was purified from a cell-free extract of L. hilgardii and the molecular weights of purified GAD was estimated to 60,000 by SDS-PAGE. The optimum pH and temperature of GAD were at pH4.6 and at $37^{\circ}C$, respectively. The GAD activity was increased by the addition of sulfate ions such as ammonium sulfate, sodium sulfate and magnesium sulfate, indicating that the increase of hydrophobic interaction causes the increase of GAD activity.

Optimization of Ammonia Decomposition and Hydrogen Purification Process Focusing on Ammonia Decomposition Rate (암모니아 반응기의 분해 효율 최적화를 통한 암모니아 분해 및 수소 정제 공정 모델 연구)

  • DAEMYEONG CHO;JONGHWA PARK;DONSANG YU
    • Transactions of the Korean hydrogen and new energy society
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    • v.34 no.6
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    • pp.594-600
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    • 2023
  • In this study, a process model and optimization design direction for a hydrogen production plant through ammonia decomposition are presented. If the reactor decomposition rate is designed to approach 100%, the amount of catalyst increases and the devices that make up the entire system also have a large design capacity. However, if the characteristics of the hydrogen regeneration process are reflected in the design of the reactor, it becomes possible to satisfy the total flow rate of fuel gas with the discharged tail gas flow rate. Analyzing the plant process simulation results, it was confirmed that when an appropriate decomposition rate is maintained in the reactor, the phenomenon of excess or shortage of fuel gas disappears. In addition, it became possible to reduce the amount of catalyst required and design the optimized capacity of the relevant processes.

Developmental Changes of Serum IgA, IgG and IgM Concentrations in Broiler Chicks - II. Isolation of IgA and Developmental Changes of Serum IgA Levels (육계의 혈청중 면역글로부린(IgA, IgG, IgM)농도의 발육시기별 변화상 - II. IgA 분리 및 발육시기별 농도수준)

  • 김정우;이민호;김춘수;김상희;박근식
    • Korean Journal of Poultry Science
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    • v.21 no.3
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    • pp.169-174
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    • 1994
  • An experiment was conducted to establish a large scale production method of anti-serum against chicken IgA and to profile the developmental changes of serum IgA levels during the feeding period(from hatching to 7 weeks of age) in broiler chicks. Blood samples were taken from Hubbard chicken at the age of hatching, 3 days of age, and weekly thereafter till to 7 weeks of age. The pure IgA was isolated from ammonium sulfate treated chicken bile juice by gel filtration chromatography ( Sepharose GL-6B) - The quantitative assay of serum IgA were carried by RID method. Developmental changes of serum IgA concentrations were 0.42 mg /mL at hatching, thereafter dicreased gradually, lowest at 1 week of age(0.17 mg /mL), and gradually increased to 7 weeks of age(2.73 mg /mL). There was no sexual difference in serum IgA level, but female chicks showed higher IgA levels than male chicks during the experimental period.

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Recovery of Useful Components from Rice-washing Water Using Membranes (분리막을 이용한 쌀뜨물내 유효성분의 회수)

  • 정건용;박성희
    • Membrane Journal
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    • v.12 no.3
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    • pp.165-170
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    • 2002
  • Membrane process was investigated to recover useful components, such as protein from rice-washing water generated in the production of the washed-rice. The filtration experiments were carried out using not only a dead-end Amicon cell to determine suitable membranes but also a hollow fiber ultrafiltration, spiral wound nanofiltration and reverse osmosis modules for home water purification. Ultrafiltration module(molecular weight cut-off : 10,000 dalton) was not suitable for recovery of useful components or protein in the rice-washing water, but nanofiltration and reverse osmosis modules showed a good performance. in the case of 250% concentration of the rice-washing water contained about 9% protein the proteins in concentrates of nanofiltration and reverse osmosis were 18% and 22%, which were about 2 and 2.4 times higher protein concentrations than those of feed, respectively.

Purification and Characterization of Oxidase Induced from Thiobacillus neapolitanus R-10 (Thiobacillus neapolitanus R-10으로 부터 산화 효소의 분리 정제와 특성)

  • Weon, Yong-Don;Kim, Dong-Suck;Ryu, Beung-Ho
    • Journal of Environmental Science International
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    • v.5 no.1
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    • pp.43-50
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    • 1996
  • Thiobacillus neopolitanus R-10, which produces a active thiosulfate oxidase was isolated from nightsoil. The optimal culture conditions of Thiobacillus neopolitanus R-10 for the production of enzyme was determined as followed: 0.8% $Na_2S_2O_3$, 0.2% $KH_2PO_4$, 0.2% $K_2HPO_4$, 0.04% $Na_2CO_3$, 0.02% $MgSO_4$.$7H_2O$, 2ml trace elements solution, ann PH 6.5 at 3$0^{\circ}C$ and 72hr cultivation. The oxidase was successively purified 83 folds yield by ${(NH_2)}_2SO_4$ fractionation, DEAE-Cellulose, Sephadex A-50 column chromatogrophy and gel Sephadex G-150 gel filteration with yield of 5.9%. The molecular weight of purified enzyme was estimated to be 43.000 dalton by SDS-polyacrylamide gel electrophoresis and gel filteration column chromatography The enzyme activity was highest at 40t and PH 7.0 The enzyme activity was relatively high by $\beta$-mercaptoethanol but strongly inhibited by cysteine.

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Development of an Automated System for the Routine Preparation of Carbon-11 Labeled Radiopharmaceuticals

  • 오승준;최연성;최용;김상은;이경한;김병태;김영서;하현준
    • Bulletin of the Korean Chemical Society
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    • v.19 no.9
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    • pp.952-956
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    • 1998
  • An automated system was developed for the routine preparation of carbon-11 ($^11C$) labeled radiopharmaceuticals, which consisted of three major parts including [$^11C$]methylation of the precursor with [$^11C$] iodomethane ($[^11C]CH_3I)$, purification of the desired product and formulation of the final $^11C$ labeled radiopharmaceutical. The whole system included seven three-way slider valves, eleven solenoid valves, four pneumatic cylinders, a HPLC (High Performance Liquid Chromatography) system and a rotary evaporator. Using this system, we investigated the radiochemical synthesis of L-[$methyl-^11C$]methionine, which is the most widely used amino acid in tumor PET (Positron Emission Tomography) studies. The overall operation took 3035 min including the production of $[^11C]CH_3I$ (10.5 min) and decay-corrected radiochemical yield was 25%. The automated system we described herein can be widely utilized for the preparation of many $^11C$ labeled radiopharinaceuticals and has been shown to be efficient, reliable and easy to operate.

Escherichia coli에서 발현된 재조합 인간 상피세포 증식인자의 정제 및 특성

  • 박세철;유광현
    • Microbiology and Biotechnology Letters
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    • v.24 no.4
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    • pp.478-484
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    • 1996
  • Recombinant human epidermal growth factor (rhEGF) was produced by E. coli BL21 harboring a plasmid pYHB101. The maximum production was 68.7 mg/l when the E. coli strain was cultured at 25$\circ$C for 48 hours in the modified MBL medium containing 10 g/l glucose with 1 mM IPTG induction at 2 hours after inoculation. The rhEGF was purified upto 267 folds by Amberlite XAD- 7 chromatography, ultrafiltration, and DEAE Sepharose fast flow ion exchange chromatography with an overall yield of 66.6%. The purified rhEGF was further separated into two fractions by HPLC. The N-terminal amino acid sequence of the second fraction was Asn-Ser-Asp-Ser-Glu-Cys-Pro-Leu-Ser-His. The effect of rhEGF on the DNA synthesis was examined using in vitro biological assay based on the incorporation of 5'-bromo-2'- deoxy-uridine (BrdU). The purified rhEGF shows no difference with natural human epidermal growth factor (nhEGF) in N-terminal amino acids residues and biological activity. From the results, we concluded that rhEGF produced from E. coli harboring the plasmid pYHB101 was apparently the same as nhEGF.

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