• Title, Summary, Keyword: $^3H$-leucine

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Preparation of Glutamic Acid-Leucine Copolymer Containing Indomethacin for Controlled Delivery

  • Yeom, Young-Il;Kim, Hyun-Pyo;Kim, Hack-Joo;Byun, Si-Myung;Kim, Nam-Deuk
    • Bulletin of the Korean Chemical Society
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    • v.7 no.3
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    • pp.213-217
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    • 1986
  • A series of copolypeptides of glutamic acid and leucine have been synthesized by N-carboxy-${\alpha}$-amino acid anhydride procedure and cast to form injectable microparticulate monolithic devices in which indomethacin was physically dispersed. With these devices, various release properties and possible clinical application were studied. The release rate of the drug had a close relationship with the monomer composition of the copolymer matrix as well as the environmental pH condition. The monolithic device of glutamic acid/leucine = 50/50 was found to be the most promising one as a ploymeric delivery system of indomethacin. The intrinsic viscosity of this copolymer was 4.35 dl/g and the release rate was 18.5${\mu}g/g/day$.

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Effect of Amino Acids and Dissolved Oxygen on Expression of Invertase in Recombinant Saccharomyces cerevisiae (재조합 Saccharomyces cerevisiae의 Invertase 발현에 미치는 아미노산과 용존산소의 영향)

  • 신해헌;조정섭;변유량;박혜영
    • Microbiology and Biotechnology Letters
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    • v.20 no.3
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    • pp.348-354
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    • 1992
  • In order to improve the productivity of invertase by recombinant Saccharomyces cerevisiae containing SUC2 gene, the effect of amino acids and dissolved oxygen concentration on the gene expression was investigated. Optimal concentrations of leucine and histidine for cell growth and cloned gene expression were 0.03 gig and 0.04 gig, respectively, expressed as the ratio of amino acid/glucose. The lack or excess of leucine and histidine has inhibitory effect on cell growth and invertase expression. In batch culture, the less aeration was, the higher invertase activity was. In continuous culture at a dilution rate of 0.09 h 1 with controlled dissolved oxygen tension, invertase activity increased dramatically at DOT levels below 5% air saturation, and a maximum activity of 215.54 KUlg cell was obtained under unaerated condition.

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Comparative Studies on the Dietary Fiber, Amino Acids and Lipid Components of Yullmoo and Yeomjoo (율무와 염주의 식이섬유, 아미노산 및 지질 성분의 비교)

  • Woo, Ja-Won;Lee, Mi-Suck;Lee, Hee-Ja;Kim, Hyong-Soo
    • Korean Journal of Food Science and Technology
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    • v.21 no.2
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    • pp.269-275
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    • 1989
  • The study was conducted to compare the components such as proximate composition, total dietary fiber(TDF) content, acid detergent fiber(ADF) content, lignin, water binding capacity(WBC), amino acid composition and lipid components from brown Job's tears, dehulled Job's tears, brown Yeomjoo and bran of Job's tears. The results from this study are summerized as follows: Total dietary fiber(TDF) content of Job's tears, brown Job's tears, brown Yeomjoo and bran of Job's tears were 2.70%, 3.86%, 4.33% and 13.3% each. Water-binding capacity(WBC) of TDF and ADF were $2.63{\pm}0.02g\;H_2O/g$ TDF and $5.89{\pm}0.15g\;H_2O/g$ ADF each. In amono acids composition of samples, glutamic acid content was the highest and the next was leucine. Chemical score of leucine in dehulled Job's tears was very high(189), in contrast lysine was very low$(22{\sim}23)$ So lysine was a first limitting amino acid in Job's tears and Yeomjoo. Neutral lipid contents were 90.89%-96.55%, glycolipid contents were 2.35%-7.48% and phospholipid contents were very low. The major fatty acids of lipid fractions were palmitic acid. oleic acid and linoleic acid.

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Effect of Individual Fatty Acids on Synthesis and Secretion of Apolipoprotein and Lipoprotein in hep-G2 Cells

  • Ryowon Choue
    • Journal of Nutrition and Health
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    • v.27 no.9
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    • pp.910-923
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    • 1994
  • The effects of individual fatty acids, differing in their degree of unsaturation(18:0, 18:1, 18:2 and 18:3) on the biosynthesis and secretion and lipids were investigated in Hep-G2 cells. Synthesis of apolipoprotein was measured by the incorporation of 3H-leucine into apolipoprotein(d<1.21g/ml) and synthesis of lipids was measured by the incorporation of 3H-glycerol and 14C-acetate into various lipid classes. Inclusion of 1.0mM of each fatty acids into the culture medium significantly increased the synthesis of total apolipoprotein and Apo B(p<0.05). However, addition of fatty acid did not affect the synthesis of cellular and medium protein. Among different fatty acids tested, oleic acid had the greatest effect on Apo B synthesis. While stearic, linoleic and linolenic acid, all had similar effects. The secretion of triglyceride into the medium markedly increased in all fatty acid groups being 5-6 times over the albumin control. The triglyceride secretion was the highest int he oleic acid group. The secretion of phospholipid and cholesterol also increased with triglyceride output. A positive relationship existed between the output of lipoprotein-triglyceride and Apo B. Since the synthesis of Apo B was significantly increased when various fatty acids were included into the culture medium, part of the apparently stimulated synthesis of the apolipoprotein may be in response to the increased formation and secretion of lipoprotein lipids.

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Analysis of quaternary structure of leucine-responsive regulatory protein (Lrp) by crosslink experiments (교차결합 실험을 통한 루신 대응 조절 단백질의 4차 구조 분석)

  • Lee, Euiho;Pokoo, Robert;Nguyen, Loi Thuan;Lee, Chan Yong
    • Korean Journal of Microbiology
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    • v.53 no.4
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    • pp.297-303
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    • 2017
  • Leucine-responsive regulatory protein (LRP) is a regulatory protein of molecular weight 18.8 kDa and is widely known to regulate many metabolic and functional activities of operons in Escherichia coli. The gene for Lrp from Escherichia coli in pQE system of 6 ${\times}$ His-tagging was expressed and $^3H$-labeled protein, as well as the wild type Lrp, was purified. The crosslink experiments were performed to analyze the quaternary structure of Lrp at high of $5{\mu}M$ and at low concentrations below $0.3{\mu}M$ with cross linkers, such as glutaraldehyde, 1, 2, 3, 4-diepoxy-butane (DEB), and ethylene glycol bis (succinimidyl succinate) (EGS). In the experiments, we found that the Lrp protein can be formed higher conformation states of tetramer, hexamer, octamer, as well as dimeric state when incubated with the above cross linkers.

Inhibition of Enzymatic Degradation of Leucine Enkephalin and $[D-Ala^2]$-Leucine Enkephalinamide in Various Rabbit Mucosal Extracts by Inhibitors (효소 억제제에 의한 토끼의 점막 추출액중 로이신엔케팔린 및 [D-알라$^2$-로이신엔케팔린아미드의 분해 억제)

  • Chun, In-Koo;Park, In-Sook;Hyun, Jeen
    • Journal of Pharmaceutical Investigation
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    • v.26 no.3
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    • pp.175-185
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    • 1996
  • To inhibit the enzymatic degradation of leucine enkephalin (Leu-Enk) and its synthetic analog. $[D-ala^2]$-leucine enkephalinamide (YAGFL), in the nasal, rectal and vaginal mucosal and serosal extracts of rabbits, effects of enzyme inhibitors such as amastatin (AM), puromycin (PM), thiorphan (TP), thimerosal (TM), EDTA, N-carboxymethyl-Phe-Leu (CPL), phenylethyl alcohol (PEA), phenylmercuric acetate (PMA), benzalkonium chloride (BC) and modified cyclodextrins, alone or in combination, were observed by assaying the pentapeptides staying intact during incubation. Mucosa extracts were prepared by exposing freshly-excised mucosal specimens mounted on Valia-Chien cells to isotonic phosphate buffer while stirring. The degradation of Leu-Enk and YAGFL followed the apparent first-order kinetics. The half-lives (mean) in the nasal, rectal and vaginal mucosal extracts were found to be 1.07, 0.33 and 1.14 hr for Leu-Enk, and 16.9, 6.2 and 6.8 hr for YAGFL, respectively. AM or PM, which is an aminopeptidase inhibitor, did not show a sufficient inhibition of Leu-Enk $(50\;{\mu}g/ml)$ degradation in all kinds of extracts. $Dimethyl-{\beta}-cyclodextrin\;(DM-{\beta}-CyD)$ decreased the degradation rate constants of Leu-Enk about 2 or 3 times, comparing with no additive. However, the use of mixed inhibitors of AM $(50\;{\mu}M)$/TM (0.25 mM)/EDTA (5 mM) resulted in a full stabilization of Leu-Enk by decreasing the degradation rate constants 67.3, 161.3 and 113.8 times far the nasal, rectal and vaginal mucosal extracts, respectively, comparing with no inhibitor. With mixed inhibitors, Leu-Enk remained intact more than 90% after 6 hr-incubation. In the stabilization of YAGFL, hM, TP or CPL alone showed little efffct, and some additives demonstrated a considerable inhibition of YAGFL degradation in the rank order of TM > BC > EDTA. However, the addition of mixed inhibitors such as TM (0.5 mM) and EDTA (5 mM) into the extracts protected YAGFL from the degradation by more than 85% even after 24 hr-incubation, suggesting almost complete inhibition of YAGFL degradation in the extract. On the other hand, $DM-{\beta}-CyD\;or\;hydroxypropyl-{\beta}-cyclodextrin$ (10%) were also found to retard enzymatic degradation rates of YAGFL markedly, and resulted in staying intact more than 80% of YAGFL in the nasal and vaginal mucosal extracts, and more than 60% in the rectal mucosal extract after 16 hr-incubation.

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A Trial for Utilizing Flounder Skin Gelatin as an Emulsifier through Enzymatic Modification (가자미피 젤라틴의 효소적 수식에 의한 유화제의 시제)

  • KIM Se-Kwon;JEON You-Jin
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.24 no.5
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    • pp.345-355
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    • 1991
  • In order to effectively utilize the by-products of sea-food, the utilization of enzyme-modified flounder(Limanda aspera) skin gelatin as an emulsifier was investigated. In the experiment, the gelatin was extracted from the flounder skin with the heat-treatment at $60^{\circ}C$ and in pH 5.0 for 3 hrs with four volumes of distilled water and emulsifiers were enzymatically modified L-leucine alkyl esters$(L-leucine-OC_n$ : n= 2, 4, 6, 8 and 10) to the gelatin$(EMFSG-C_2,\;EMFSG-C_4,\;EMFSG-C_6,\;EMFSG-C_8,\;EMFSG-C_{10})$ for improving the functional properties such as emulsifying activity, emulsifying viscosity, whippability, electric conductivity, critical micelle concentration and interface tension, etc. Also, the functional properties of the L-leucine alkyl ester modified gelatins were compared with those of Tween-60 as reference. Molecular weights of the enzymatically modified flounder skin gelatin(EMFSG) were 20.5kDa. in $EMFSG-C_2.\;19.5 kDa.\;in\;EMFSG-C_4\;and\;16.5kDa.\;in\;EMFSG-C_6,\;EMFSG-C_8$ and $EMFSG-C_{10}$. respectively. Emulsifying activity and emulsifying viscosity in the modified gelatins were risen with increase of carbon number of the introduced L-leucine alkyl esters. Among the modified gelatins, $EMFSG-C_6$ exhibited the highest emulsifying stability and foaming stability, whereas $EMFSG-C_8$ showed the highest whippability. The electric conductivities of the all $EMFSG-C_n$ were linearly risen to critical micelle concentration(CMC) , therefore $EMFSG-C_{10}$ exhibited the lowest CMC value and interface tension, and dense particles in the microscopic observation. In conclusion, the best quality in functional properties was assured on $EMFSG-C_{10}$.

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Inhibition of Angiotensin II-Induced Vascular Smooth Muscle Cell Hypertrophy by Different Catechins

  • Zheng, Ying;Song, Hye-Jin;Yun, Seok-Hee;Chae, Yeon-Jeong;Jia, Hao;Kim, Chan-Hyung;Ha, Tae-Sun;Sachinidis, Agapios;Ahn, Hee-Yul;Davidge, Sandra T.
    • The Korean Journal of Physiology and Pharmacology
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    • v.9 no.2
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    • pp.117-123
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    • 2005
  • A cumulative evidence indicates that consumption of tea catechin, flavan-3-ol derived from green tea leaves, lowers the risk of cardiovascular diseases. However, a precise mechanism for this cardiovascular action has not yet been fully understood. In the present study, we investigated the effects of different green tea catechins, such as epigallocatechin-3 gallate (EGCG), epigallocatechin (EGC), epicatechin-3 gallate (ECG), and epicatechin (EC), on angiotensin II (Ang II)-induced hypertrophy in primary cultured rat aortic vascular smooth muscle cell (VSMC). [$^3H$]-leucine incorporation was used to assess VSMC hypertrophy, protein kinase assay, and western blot analysis were used to assess mitogen-activated protein kinase (MAPK) activity, and RT-PCR was used to assess c-jun or c-fos transcription. Ang II increased [$^3H$]-leucine incorporation into VSMC. However, EGCG and ECG, but not EGC or EC, inhibited [$^3H$]-leucine incorporation increased by Ang II. Ang II increased phosphorylation of c-Jun, extracellular-signal regulated kinase (ERK) 1/2 and p38 MAPK in VSMC, however, EGCG and ECG , but not EGC or EC, attenuated c-Jun phosphorylation increased by Ang II. ERK 1/2 and p38 MAPK phosphorylation induced by Ang II were not affected by any catechins. Ang II increased c-jun and c-fos mRNA expression in VSMC, however, EGCG inhibited c-jun but not c-fos mRNA expression induced by Ang II. ECG, EGC and EC did not affect c-jun or c-fos mRNA expression induced by Ang II. Our findings indicate that the galloyl group in the position 3 of the catechin structure of EGCG or ECG is essential for inhibiting VSMC hypertrophy induced by Ang II via the specific inhibition of JNK signaling pathway, which may explain the beneficial effects of green tea catechin on the pathogenesis of cardiovascular diseases observed in several epidemiological studies.

Characterization of Calcium-Activated Bifunctional Peptidase of the Psychrotrophic Bacillus cereus

  • Kim Jong-Il;Lee Sun-Min;Jung Hyun-Joo
    • Journal of Microbiology
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    • v.43 no.3
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    • pp.237-243
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    • 2005
  • The protease purified from Bacillus cereus JH108 has the function of leucine specific endopeptidase. When measured by hydrolysis of synthetic substrate (N-succinyl-Ala-Ala-Pro-Leu-p-nitroanilide), the enzyme activity exhibited optimal activity at pH 9.0, $60^{\circ}C$. The endopeptidase activity was stimulated by $Ca^{++},\;Co^{++},\;Mn^{++},\;Mg^{++},\;and\;Ni^{++}$, and was inhibited by metal chelating agents such as EDTA, 1,10-phenanthroline, and EGTA. Addition of serine protease inhibitor, PMSF, resulted in the elimination of the activity. The endopeptidase activity was fully recovered from the inhibition of EDTA by the addition of 1 mM $Ca^{++}$, and was partially restored by $Co^{++}\;and\;Mn^{++}$, indicating that the enzyme was stabilized and activated by divalent cations and has a serine residue at the active site. Addition of $Ca^{++}$ increased the pH and heat stability of endopeptidase activity. These results show that endopeptidase requires calcium ions for activity and/or stability. A Lineweaver-Burk plot analysis indicated that the $K_m$ value of endopeptidase is 0.315 mM and $V_{max}$ is 0.222 ) is $0.222\;{\mu}mol$ of N-succinyl-Ala-Ala-Pro-Leu-p-nitroanilide per min. Bestatin was shown to act as a competitive inhibitor to the endopeptidase activity.

Organic Constituents in Kimchis (Ixeris sonchifolia H.) -On free amino acids- (고들빼기 김치의 유기성분(有機成分)(I) - 유이(遊離)아미노산(酸)에 관(關)하여 -)

  • Kang, Dong Hee;Woo, Young Sook;Lee, Young Kyoung;Chung, Seung Yong
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.12 no.3
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    • pp.225-229
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    • 1983
  • The change of free amino acids was identified during fermentation of kimchis (Ixeris sonchifoliaH.) added fermented anchovy. The results were summarized as follows; In fresh roots and leaves of Ixeris sonchifolia H., 8 kinds of free amino were determined respectively. Among them, argine, cysteine and glutamic acid were abundant in fresh roots, while arginine, valine, isoleucine and phenylalanine in fresh leaves, especially arginine was dominant in fresh both roots and leaves. The amount of total amino acids in fresh leaves was about 2.5 times of that of roots. After fermentation, 15 kinds of free amino acids were determined in kimchis, and the characteristic favor of in was attributed to such amino acids as threonine, glutamic acid, alanine, leucine and cysteine. The content of total free amino acids in kimchi leaves was increased to about 5 times of that in fresh (9,435,6mg % on dry base), but in kimchi roots, 11 times of that in fresh was contained (7,079,1mg % on dry base) In kimchi'es extract, 16 kinds of free amino acids were determined, and threonine, glutamic acid, alanine, cysteine and leucine were abundant.

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