• Title/Summary/Keyword: Crude extracts

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Genes Encoding Ribonucleoside Hydrolase 1 and 2 from Corynebacterium ammoniagenes

  • Lee, Jin-Ho;Kim, Hyun-Soo;Lee, Won-Sik;Park, Young-Hoon;Bang, Won-Gi
    • Proceedings of the Microbiological Society of Korea Conference
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    • 2006.05a
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    • pp.105-107
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    • 2006
  • Two kinds of nucleoside hydrolases (NHs) encoded by rih1 and rih2 were cloned from Corynebacterium ammoniagenes using deoD- and gsk-defective Escherichia coli. Sequence analysis revealed that NH 1 was a protein of 337 aa with a deduced molecular mass of 35,892 Da, whereas NH 2 consisted of 308 aa with a calculated molecular mass of 32,310 Da. Experiments with crude extracts of IPTG-induced E. coli CGSC 6885(pTNU23) and 6885(pTNI12) indicated that the Rihl enzyme could catalyse the hydrolysis of uridine and cytidine and showed pyrimidine-specific ribonucleoside hydrolase activity. Rih2 was able to hydrolyse both purine and pyrimidine ribonucleosides with the following order of activity-inosine>adenosine>uridine>guanosine>xanthosine>cytidine-and was classified in the non-specific NHs family. rih1 and rih2 deletion mutants displayed a decrease in cell growth on minimal medium supplemented with pyrimidine and purine/pyrimidine nucleosides, respectively, compared with the wild-type strain. Growth of each mutant was substantially complemented by introducing rih1 and rih2, respectively. Furthermore, disruption of both rih1 and rih2 led to the inability of the mutant to utilize purine and pyrimidine nucleosides as sole carbon source on minimal medium. These results indicated that rih1 and rih2 play major roles in the salvage pathways of nucleosides in this micro-organism.

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Expression and DNA Sequence of the Gene Coding for the lux-specific Fatty Acyl-CoA Reductase from photobacterium phosphoreum

  • Lee, Chan-Yong;Edward A. Meighen
    • Journal of Microbiology
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    • v.38 no.2
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    • pp.80-87
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    • 2000
  • The nucleotide sequence of the luxC gene coding for lux-specific fatty acyl-CoA reductase and the upstream DNA (325bp)of the structural gene from bioluminescent bacterium, Photobacterium phosphoreum, has been deternubed. An open reading frame extending for more than 20 codons in 325 bp DNA upstream of luxC was not present in both directions. The lux gene can be translated into a polypeptide of 54 kDa and the amino acid sequences of lux specific reductases of P. phosphoreum shares 80, 65, 58, and 62% identity with those of the Photobacterium leiognathi, Vibrio fischeri, Vibrio harveyi, and Xehnorhabdus luminescenens reductases, respectively. Analyses of codon usage, showing that a high frequency (2.3%) of the isoleucine codon, AUA, in the luxC gene compared to that found in Escherichia coli genes (0.2%) and its absence in the luxA and B genes, suggested that the AUA codon may play a modulator role in the expression of lux gene in E. coli. The structural genes (luxC, D, A, B, E) of the P. phosphoreum coding for luciferase (${\alpha}$,${\beta}$) and fatty acid reductase (r, s, t) polypeptides can be expressed exclusively in E. coli under the T7 phage RNA polymerase/promoter system and identificationof the [35S]methionine labelled polypeptide products. The degree of expression of lux genes in analyses of codon usage. High expression of the luxC gene could only be accomplished in a mutant E. coli 43R. Even in crude extracts, the acylated acyl-CoA reductase intermediate as well as acyl-CoA reductrase activities could be readily detected.

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Compositions of Fatty Acid, Free Amino Acid and Total Amino Acid of Lespedeza x chiisanensis T. LEE (지리산 싸리의 지질 구성 지방산과 유리 아미노산 및 구성 총 아미노산 조성에 관하여)

  • 김종균
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.22 no.5
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    • pp.586-591
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    • 1993
  • The compositions of amino acid in the protein and fatty acid in the lipid of Lespedeza x chiisanensis T. LEE were analyzed by HPLC and GC, respectively. The contents of crude oil and protein from the extracts were 11.13% and 5.18%, respectively. The amount of free anino acids in the protein was 443.14mg/100g, and 94.84mg/100g of essential amino acid were contained in the free amino acid. The amount of total amino acids in the protein was 3159.85mg/100g, and 1068.18mg/100g of essential amino acid were contained in the total amino acid. The compositions of fatty acid in the lipid were $C_{18\;:\;2}=45.05%,\;C_{18\;:\;3}=18.71%,\;C_{19}=14.70%,C_{18\;:\;1}=6.81%,\;C_{16}=4.35%,\;C_{16\;:\;1}=1.59%$ in order, respectively. 72.44% of unsaturated fatty acids were contained in the lipid.

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Antioxidant and Immunoenhancement Activities of Ginger (Zingiber officinale Roscoe) Extracts and Compounds in In Vitro and In Vivo Mouse and Human System

  • Rungkat, F-Zakaria;Nurahman;E Prangdimurt;Tejasari
    • Preventive Nutrition and Food Science
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    • v.8 no.1
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    • pp.96-104
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    • 2003
  • Ginger (Zingiber officinale Roscoe) is traditionally used as appetite enhancer, improver of the digestive system, antithusive, anti-cold, antipyretic, analgesic, and antiinflammation. In vitro evaluation using human lymphocyte cultures showed almost similar indication with those in in vivo mouse study, NK cell lysing activity was improved significantly. Proliferation activity of B and T cells, and CD3$^{+}$ and CD3$^{+}$CD4$^{+}$T cell subset were better observed using oleoresin or gingerol and shogaol fractions. Although there were higher activities in gingerol, the improvement was almost equal to that by oleoresin. Shogaol did not show better improvement except at higher concentration. It could be concluded that treatment with single bioactive compound, such as gingerol, did not show significant effects compared to oleoresin, the crude extract. In human study, involving healthy male adult, the improvement of NK cell lysing activity was again demonstrated and even more apparent. The mechanism involved in the protection seemed to be through the antioxidant activity of gingerol. However, other mechanism underlying the improvement of NK cell lysing activity must be involved since this improvement seemed to be specifically toward NK cell activity. Since NK cells ave specific for the elimination of virus-infected cell and mutated cells, this positive effect on the immune system are very interesting. This work has also scientifically proved that the traditional beliefs that ginger had preventive effects on common cold appeared to be reasonable.

Antioxidative Activity of Some Solvent Extract from Caesalpinia sappan L. (소목 추출물의 항산화 효과)

  • Lim, Dae-Kwan;Choi, Ung;Shin, Dong-Hwa
    • Korean Journal of Food Science and Technology
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    • v.28 no.1
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    • pp.77-82
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    • 1996
  • Antioxidative activity of the extract from Caesalpinia sappan L. by various solvent was compared with several commercial antioxidants, using the Rancimat method. AI (antioxidative index; induction period of oil containing extract/induction period of control oil) of all extracts were higher than commercial antioxidants, such as BHA, ${\delta}-tocopherol$ and ascorbic acid. The ethanol extract was fractionated by liquid liquid extraction. Ethyl acetate fraction showed higher AI than the whole crude extract. When comparing POV and TBA value of palm oil and lard containing different level of each fraction, the oxidative stability of ethyl acetate fraction at 200 ppm level on palm oil and lard were similar to that of BHT at 200 ppm level, and better than BHA, ${\delta}-tocopherol$ and control.

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Enhancing immune responses to inactivated foot-and-mouth virus vaccine by a polysaccharide adjuvant of aqueous extracts from Artemisia rupestris L.

  • Wang, Danyang;Yang, Yu;Li, Jinyu;Wang, Bin;Zhang, Ailian
    • Journal of Veterinary Science
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    • v.22 no.3
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    • pp.30.1-30.15
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    • 2021
  • Background: New-generation adjuvants for foot-and-mouth disease virus (FMDV) vaccines can improve the efficacy of existing vaccines. Chinese medicinal herb polysaccharide possesses better promoting effects. Objectives: In this study, the aqueous extract from Artemisia rupestris L. (AEAR), an immunoregulatory crude polysaccharide, was utilized as the adjuvant of inactivated FMDV vaccine to explore their immune regulation roles. Methods: The mice in each group were subcutaneously injected with different vaccine formulations containing inactivated FMDV antigen adjuvanted with three doses (low, medium, and high) of AEAR or AEAR with ISA-206 adjuvant for 2 times respectively in 1 and 14 days. The variations of antibody level, lymphocyte count, and cytokine secretion in 14 to 42 days after first vaccination were monitored. Then cytotoxic T lymphocyte (CTL) response and antibody duration were measured after the second vaccination. Results: AEAR significantly induced FMDV-specific antibody titers and lymphocyte activation. AEAR at a medium dose stimulated Th1/Th2-type response through interleukin-4 and interferon-γ secreted by CD4+ T cells. Effective T lymphocyte counts were significantly elevated by AEAR. Importantly, the efficient CTL response was remarkably provoked by AEAR. Furthermore, AEAR at a low dose and ISA-206 adjuvant also synergistically promoted immune responses more significantly in immunized mice than those injected with only ISA-206 adjuvant and the stable antibody duration without body weight loss was 6 months. Conclusions: These findings suggested that AEAR had potential utility as a polysaccharide adjuvant for FMDV vaccines.

Heterologous Expression and Characterization of a Laccase from Laccaria bicolor in Pichia pastoris and Arabidopsis thaliana

  • Wang, Bo;Yan, Ying;Xu, Jing;Fu, Xiaoyan;Han, Hongjuan;Gao, Jianjie;Li, Zhenjun;Wang, Lijuan;Tian, Yongsheng;Peng, Rihe;Yao, Quanhong
    • Journal of Microbiology and Biotechnology
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    • v.28 no.12
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    • pp.2057-2063
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    • 2018
  • Laccases can oxidize a variety of phenolic and non-phenolic substrates including synthetic dyes. In this research, a laccase gene Lcc9 from Laccaria bicolor was chemically synthesized and optimized to heterogeneous expression in Pichia pastoris and Arabidopsis thaliana. The properties of recombinant laccase expressed by P. pastoris were investigated. The laccase activity was optimal at 3.6 pH and $40^{\circ}C$. It exhibited $K_m$ and $V_{max}$ values of $0.565mmol\;l^{-1}$ and $1.51{\mu}mol\;l^{-1}\;min^{-1}$ for ABTS respectively. As compared with untransformed control plants, the laccase activity in crude extracts of transgenic lines exhibited a 5.4 to 12.4-fold increase. Both laccases expressed in transgenic P. pastoris or A. thaliana could decolorize crystal violet. These results indicated that L. bicolor laccase gene may be transgenically exploited in fungi or plants for dye decolorization.

Antioxidant Compounds and Antioxidant Activities of Ethanolic Extracts from Brown Rice Cultivars (현미 품종별 에탄올 추출물의 항산화 성분 및 항산화 활성)

  • Kim, Hyun-Joo;Lee, Ji Hae;Lee, Byong Won;Lee, Yu Young;Lee, Byoung Kyu;Woo, Koan Sik
    • The Korean Journal of Food And Nutrition
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    • v.31 no.6
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    • pp.949-956
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    • 2018
  • This study investigated chemical components, antioxidant compounds, and activity of brown rice cultivars, to select good cultivar to be used for processing of mixed-rice in the food industry. Proximate compositions, phytic acid, phenolic compounds, and antioxidant activity of brown rice were significantly different among cultivars. Moisture, crude ash, fat, protein, and carbohydrate contents of brown rice were 9.51~12.82, 1.05~1.93, 1.84~6.24, 5.90~9.60 and 71.75~80.34 g/100 g, respectively. Phytic acid content of brown rice cultivars was 7.39~0.87 mg/g. Total polyphenol content of Joeunheukmi and Geonganghongmi cultivars, were 615.25 and $311.14{\mu}g\;GAE/g$, total flavonoid content was 267.75 and $100.67{\mu}g\;CE/g$, respectively. DPPH radical scavenging activity of Geonganghongmi, Joeunheukmi and Hyeugkwang cultivars was 89.17, 87.94 and 43.17%, ABTS radical scavenging activity was 113.57, 113.34, and $93.53{\mu}mol\;TE/g$, and ferric reducing antioxidant potential was 951.67, 1,075.75, and $508.33{\mu}M/g$, respectively. As a result, phenolic compounds and antioxidant activities of pigmented brown rice were high, and it could be used as a functional material.

Studies on Improved Amylases Developed by Protoplast Fusion of Aspergillus species

  • Adeleye, Tolulope Modupe;Kareem, Sharafadeen Olateju;Olufunmilayo, Bankole Mobolaji;Atanda, Olusegun;Osho, Michael Bamitale;Dairo, Olawale
    • Microbiology and Biotechnology Letters
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    • v.49 no.1
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    • pp.45-56
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    • 2021
  • Improved amylases were developed from protoplast fusants of two amylase-producing Aspergillus species. Twenty regenerated fusants were screened for amylase production using Remazol Brilliant Blue agar. Crude enzyme extracts produced by solid state fermentation of rice bran were assayed for activity. Three variable factors (temperature, pH and enzyme type) were optimized to increase the amylase activity of the parents and selected fusants using rice bran medium and solid state fermentation. Analysis of this optimization was completed using the Central Composite Design (CCD) of the Response Surface Methodology (RSM). Amylase activity assays conducted at room temperature and 80℃ demonstrated that Aspergillus designates, T5 (920.21 U/ml, 966.67 U/ml), T13 (430 U/ml, 1011.11 U/ml) and T14 (500.63 U/ml, 1012.00 U/ml) all exhibited improved function making them the preferred fusants. Amylases produced from these fusants were observed to be active over the entire pH range evaluated in this study. Fusants T5 and T14 demonstrated optimal activity under acidic and alkaline conditions, respectively. Fusants T13 and T14 produced the most amylase at 72 h while parents TA, TC and fusant T5 produced the most amylase after 96 h of incubation. Response surface methodology examinations revealed that the enzyme from fusant T5 was the optimal enzyme demonstrating the highest activity (1055.17 U/ml) at pH 4 and a temperature of 40℃. This enzyme lost activity with further increases in temperature. Starch hydrolysis using fusant T5 gave the highest yield of glucose (1.6158 g/100 ml). The significant activities of the selected fusants at 28 ± 2℃ and 80℃ and the higher sugar yields from cassava starch hydrolysis over their parental strains indicate that it is possible to improve amylase activity using the protoplast fusion technique.

Synthesis of ginsenoside Rb1-imprinted magnetic polymer nanoparticles for the extraction and cellular delivery of therapeutic ginsenosides

  • Liu, Kai-Hsi;Lin, Hung-Yin;Thomas, James L.;Shih, Yuan-Pin;Yang, Zhuan-Yi;Chen, Jen-Tsung;Lee, Mei-Hwa
    • Journal of Ginseng Research
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    • v.46 no.5
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    • pp.621-627
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    • 2022
  • Background: Panax ginseng (ginseng) is a traditional medicine that is reported to have cardioprotective effects; ginsenosides are the major bioactive compounds in the ginseng root. Methods: Magnetic molecularly imprinted polymer (MMIP) nanoparticles might be useful for both the extraction of the targeted (imprinted) molecules, and for the delivery of those molecules to cells. In this work, plant growth regulators were used to enhance the adventitious rooting of ginseng root callus; imprinted polymeric particles were synthesized for the extraction of ginsenoside Rb1 from root extracts, and then employed for subsequent particle-mediated delivery to cardiomyocytes to mitigate hypoxia/reoxygenation injury. Results: These synthesized composite nanoparticles were first characterized by their specific surface area, adsorption capacity, and magnetization, and then used for the extraction of ginsenoside Rb1 from a crude extract of ginseng roots. The ginsenoside-loaded MMIPs were then shown to have protective effects on mitochondrial membrane potential and cellular viability for H9c2 cells treated with CoCl2 to mimic hypoxia injury. The protective effect of the ginsenosides was assessed by staining with JC-1 dye to monitor the mitochondrial membrane potential. Conclusion: MMIPs can play a dual role in both the extraction and cellular delivery of therapeutic ginsenosides.