• Journal of Microbiology and Biotechnology
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• v.17 no.1
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• pp.123-129
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• 2007

The trehalose $({\alpha}-D-glucopyranosyl-[1,1]-{\alpha}-D-glucopyranose)$ biosynthesis genes MhMTS and MhMTH, encoding a maltooligosyltrehalose synthase (MhMTS) and a maltooligosyltrehalose trehalohydrolase (MhMTH), respectively, have been cloned from the hyperthermophilic archaebacterium Metallosphaera hakonesis. The ORF of MhMTS is 2,142 bp long, and encodes 713 amino acid residues constituting a 83.8 kDa protein. MhMTH is 1,677 bp long, and encodes 558 amino acid residues constituting a 63.7 kDa protein. The deduced amino acid sequences of MhMTS and MhMTH contain four regions highly conserved for MTSs and three for MTHs that are known to constitute substrate-binding sites of starch-hydrolyzing enzymes. Recombinant proteins obtained by expressing the MhMTS and MhMTH genes in E. coli catalyzed a sequential reaction converting maltooligosaccharides to produce trehalose. Optimum pH of the MhMTS/MhMTH enzyme reaction was around 5.0 and optimum temperature was around 70 C. Trehalose-producing activity of the MhMTS/ MhMTH was notably stable, retaining 80% of the activity after preincubation of the enzyme mixture at $70^{\circ}C$ for 48 h, but was gradually abolished by incubating at above $85^{\circ}C$. Addition of thermostable $4-{\alpha}-glucanotransferase$ increased the yield of trehalose production from maltopentaose by 10%. The substrate specificity of the MhMTS/MhMTH-catalyzed reaction was extended to soluble starch, the most abundant maltodextrin in nature.

• Proceedings of the Korean Society of Postharvest Science and Technology of Agricultural Products Conference
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• 2003.04a
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• pp.135-136
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• 2003

알코올 분무, 셀로판지 부착 등의 표면처리가 포장김치의 표면갈변과, pH, 산도, 균수 및 관능적 품질에 미치는 영향을 조사하였다. 실험에 사용된 포장김치는 200 g 들이 플라스틱 용기에 김치를 담은 후 무처리, 70% 알코올로 표면을 1회 분무한 것, 김치표면에 폴리에틸렌필름(두께 0.04 mm)을 부착하여 저장실 (10$\pm$1$^{\circ}C$)에서 21일간 저장하면서 3일 간격으로 조사하였다. 그 결과, 무처리와 알코올을 분무한 경우는 저장 15일째에 pH가 4.0수준에 도달하였으나 김치표면에 polyethylene film을 부착한 경우는 21일째까지도 pH가 4.0 이상을 유지하였다. 산도의 경우도 pH와 동일한 경향을 나타내어 polyethylene film으로 표면처리함으로서 김치의 숙성이 지연되었다. 무처리와 알코올을 처리한 것은 12일째부터 김치표면이 갈변하기 시작하였으며 18일째는 그 갈변정도가 심하였다. 그러나 polyethylene film을 표면에 부착한 경우는 저장 21일째까지도 갈변정토가 미약하였다. 저장기간에 따른 총균수의 변화는 무처리와 알코올을 처리한 경우가 polyethylene film을 부착한 경우에 비하여 증가폭이 컸으나 젖산균수는 반대로 polyethylene film을 부착한 경우가 무처리 또는 알코올을 분무한 경우에 비하여 높아 김치의 품질이 양호하였다. 무처리와 알코올을 분무한 김치의 표면에는 저장 15일 이후에 산막표모 및 곰팡이의 균사체가 번식하는 김치도 발견되었다. 처리별 이미 및 이취의 발생정도를 조사한 결과 무처리와 알코올을 처리한 경우는 저장 15일경에 특히 용기 표면에 위치하는 김치에서 이미와 이취가 있었으나 polyethylene film을 부착한 경우는 이미와 이취가 전혀 나타나지 않았다.채반에 넣고 밀도를 측정한 결과 1.002g/㎤을 나타냈다. 즉 물의 이론밀도인 1g/㎤에 근접한 값을 보여 정확한 밀도 계측이 가능한 것으로 판단되었다. 또한 밀도 계측시스템의 측정 반복간 정밀도를 파악하기 위해 수박 6개를 임으로 선정하여 3반복 측정 시험한 결과, 측정표준편차가 0.001~0.004g/㎤로 해상도보다는 다소 높았으나 대체로 양호한 결과를 나타냈다. 수박 35개를 이용하여 개발 계측시스템과 사람이 직접 부력법으로 밀도를 측정 비교한 결과, 계측시스템에 의해 측정된 수박 밀도가 사람이 측정했을 때 보다 낮게 측정되었다. 수박의 외관인자(무게, 길이, 직경, 체적), 밀도와 당도의 상관관계 구명시험을 위해 원예연구소 시험포장에서 재배된 삼복꿀수박 총 74개를 공시재료로 하였고, 시험은 출수일별로 10~14개씩 수확하여 외관인자, 밀도, 당도를 각각 측정하고, 이들 인자들간의 상관관계를 구명하였다. 외관인자들간에는 높은 상관관계를 보였으나, 외관인자들과 밀도, 외관인자들과 당도, 밀도와 당도와는 매우 낮은 상관관계를 나타냈다. 점성에 영향을 미치는 가장 주요한 조건이라고 생각된다.환원당인 sucrose 함량은 계속 증가하였고 fructose, glucose, sorbitol의 함량(추황의 sorbitol을 제외)은 생장이 촉진됨에 따라 증가하다가 다시 점차적으로 감소하였다. 이러한 결과는 총당과 환원당의 측정결과와 일치한 것으로 나타났다. 결론적으로 배의 성장에 따라 산 함량은 감소하였고 당 함량은 증가하였다.luco-pyranoside, quercetin 7-O- -glucopyranoside, acacetin 7-O-$\beta$-D-glucuronide and apigenin-6-C-$\beta$-D-glucopyranosyl-8-C-$\b

• Proceedings of the Korean Society of Postharvest Science and Technology of Agricultural Products Conference
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• 2003.04a
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• pp.22-39
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• 2003

Biological activity of phenolic compounds in seeds and leaves of safflower (Carthamu tinctorius L.) were evaluated using several in vitro and in vivo assays. Six phenolic constituents were isolated from the seeds and identified as N-feruloylserotonia, N- (p-coumaroyl)serotonin, matairesinol, 8′-hydroxyarctigenin, acacetin 7-O-$\beta$-D-glucoside (tilianine) and acacetin. Six phenolic compounds exhibited considerable antioxidative activity, and especially two serotonins showed potent DPPH radical scavenging activity and antiperoxidative activity against rat liver microsomal lipid peroxidation induced by the hydroxyl radical generated via a Fenton-type reaction. Additionally, six phenolic compounds possessed comparable cytotoxicity against three cancer cells, Hela cell, MCF-7 and HepG2 cell, and particularly acacetin and its glycosides had the most potent cytotoxicity. Moreover, we found that feeding safflower seeds attenuated bone loss, and lowered levels of plasma and liver lipids in ovariectomized rats. Serotonins, lignans and flavones stimulated proliferation of the osteoblast-like cells in a dose-dependent manner (10$^{-15}$ ~10$^{-6}$ M), as potently as E$_2$ (17$\beta$-estradiol). Particularly, serotonins were mainly responsible for bone-protecting and lipid lowering effects in ovariectomized rats. Meanwhile, eight flavonoids, including a novel quercetin-7-O-(6"-O-acetyl)-$\beta$-D-glucopyranoside and seven kown flavonoids, luteolin quercetin, luteolin 7-O-$\beta$-D-glucopyranoside, luteolin-7-O-(6"-O-acetyl)-$\beta$-D-gluco-pyranoside, quercetin 7-O- -glucopyranoside, acacetin 7-O-$\beta$-D-glucuronide and apigenin-6-C-$\beta$-D-glucopyranosyl-8-C-$\beta$-D-glucopyranoside were first isolated and identified from safflower leaf. Among these flavonoids, luteolin-acetyl-glucoside and $\beta$quercetin- acetyl-glucoside showed potent antioxidative activities against 2-deoxyribose degradation and lipid peroxidation in rat liver microsomes. Luteolin, quercetin and their corresponding glycosides also exhibited strong antioxidative activity, while acacetin glucuronide and apigenin-6, 8-di-C-glucoside were relatively less active. Finally, changes in phenolic compositions were also determined by HPLC in the safflower seed and leaf during growth stages and roasting process to produce standardized supplement powerds. These results suggest that phenolic compounds in the roasted safflower seed and leaf may be useful as potential sources of therapeutic agents against several pathological disorders such as carcinogenesis, atherosclerosis and osteoporosis.

• Journal of Ginseng Research
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• v.42 no.1
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• pp.42-49
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• 2018

Background: Ginsenoside F1 has been described to possess skin-whitening effects on humans. We aimed to synthesize a new ginsenoside derivative from F1 and investigate its cytotoxicity and melanogenesis inhibitory activity in B16BL6 cells using recombinant glycosyltransferase enzyme. Glycosylation has the advantage of synthesizing rare chemical compounds from common compounds with great ease. Methods: UDP-glycosyltransferase (BSGT1) gene from Bacillus subtilis was selected for cloning. The recombinant glycosyltransferase enzyme was purified, characterized, and utilized to enzymatically transform F1 into its derivative. The new product was characterized by NMR techniques and evaluated by MTT, melanin count, and tyrosinase inhibition assay. Results: The new derivative was identified as (20S)-$3{\beta},6{\alpha},12{\beta}$,20-tetrahydroxydammar-24-ene-20-O-${\beta}$-D-glucopyranosyl-3-O-${\beta}$-D-glucopyranoside(ginsenoside Ia), which possesses an additional glucose linked into the C-3 position of substrate F1. Ia had been previously reported; however, no in vitro biological activity was further examined. This study focused on the mass production of arduous ginsenoside Ia from accessible F1 and its inhibitory effect of melanogenesis in B16BL6 cells. Ia showed greater inhibition of melanin and tyrosinase at $100{\mu}mol/L$ than F1 and arbutin. These results suggested that Ia decreased cellular melanin synthesis in B16BL6 cells through downregulation of tyrosinase activity. Conclusion: To our knowledge, this is the first study to report on the mass production of rare ginsenoside Ia from F1 using recombinant UDP-glycosyltransferase isolated from B. subtillis and its superior melanogenesis inhibitory activity in B16BL6 cells as compared to its precursor. In brief, ginsenoside Ia can be applied for further study in cosmetics.

• Korean Journal of Microbiology
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• v.44 no.1
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• pp.74-79
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• 2008

The purpose of this study is to clone cgt gene from Paenibacillus sp. JB-13 and to overexpress the protein in E. coli. For this purpose, the cgt gene was amplified from Paenibacillus sp. JB-13 genomic DNA by PCR using degenerate oligonucleotide primers. The sequence analysis results showed that the cgt gene from Paenibacillus sp. JB-13 has 98% homology with the cgt gene of Bacillus sp. To overexpress the protein, the cgt gene was cloned into pEXP7 expression vector and transformed into E. coli. The production of CGTase by recombinant E. coli was optimized under following conditions: 0.5% glucose, 3.0% polypeptone, 0.3% $K_2HPO_4$, 0.5% NaCl, and 7.0 of initial pH, 2.0% of inoculum, $37^{\circ}C$ of culture temperature for 14 hr. And the optimal agitation was found at 0.1 vvm. The synthesis of 2-O-${\alpha}$-D-Glucopyranosyl L-Ascorbic acid (AA-2G) using the CGTase expressed in E. coli was identified as AA-2G by HPLC and HPLC confirmed that treating AA-2G made by cloned CGTase with ${\alpha}$-glucosidase substantially produced AA and glucose.

• Molecules and Cells
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• v.19 no.1
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• pp.137-142
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• 2005

We showed recently that ginsenosides inhibit the activity of various types of ion channel. Here we have investigated the role of the carbohydrate component of ginsenoside $Rg_3$ in the inhibition of $Na^+$ channels. The channels were expressed in Xenopus oocytes by injecting cRNAs encoding rat brain Nav1.2 ${\alpha}$ and ${\beta}1$ subunits, and analyzed by the two-electrode voltage clamp technique. Treatment with $Rg_3$ reversibly inhibited the inward $Na^+$ peak current ($I_{Na}$) with an $IC_{50}$ of $32.2{\pm}4.5{\mu}M$, and the inhibition was voltage-dependent. To examine the role of the sugar moiety, we prepared a straight chain form of the second glucose and a conjugate of this glucose with 3-(4-hydroxyphenyl) propionic acid hydrazide (HPPH). Neither derivative inhibited $I_{Na}$. Treatment with the carbohydrate portion of ginsenoside $Rg_3$, sophorose [${\beta}-D-glucopyranosyl$ ($1{\rightarrow}2$)-${\beta}-glucopyranoside$], or the aglycone (protopanaxadiol), on their own or in combination had no effect on $I_{Na}$. These observations indicate that the carbohydrate portion of ginsenoside $Rg_3$ plays an important role in its effect on the $Na^+$ channel.

• Microbiology and Biotechnology Letters
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• v.26 no.2
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• pp.179-186
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• 1998

Dextransucrase hyper-producing Leuconostoc mesenteroides B-512FMCM and dextransucrase constitutive mutants B-742CB and B-1355C catalyzed the transfer of glucose from sucrose to other carbohydrates which were present or were added to the reaction digests. When the acceptor was a maltose, gentiobiose, lactose or raffinose, there was produced a series of oligosaccharide acceptor products or single product based on the kinds of enzymes and reaction conditions. To obtain the quantitative information about the yield and the distribution of acceptor products and dextran two experimental parameters were studied: a) the ratio of acceptor to sucrose and b) the amount of enzyme at constant carbohydrate concentration (100 mM). As the amount of enzyme increased, the synthesis of acceptor products (of maltose or gentiobiose) increased, and the formation of dextran decreased. As the ratio of acceptor to sucrose increased, the amount of dextran and the number of acceptor-products decreased and the amount of acceptor-products increased. When maltose or gentiobiose was an acceptor, the glucose from sucrose was transferred to the C-6 hydroxyl group of the nonreducing-end glucose residue of accepters to give a homologous series of isomaltosyl dextrins. In case of lactose or raffinose, there was produced only one acceptor product from B-512FMCM dextransucrase reaction. In the lactose acceptor reaction, the glucose from sucrose was transferred to the C-2 hydroxyl of the reducing end glucose residue of lactose. To get a series of oligosaccharides from lactose or raffinose acceptor reaction we used B-742CB dextransucrase or B-1355C alternansucrase with 500 mM sucrose in reaction digest.

• Proceedings of the Korea Society of Poultry Science Conference
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• 2003.11a
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• pp.91-92
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• 2003

비태인의 급여(0, 600 ppm)가 사료의 단백질 수준(14, 16 ％)에 따라 산란계의 생산성과 난품질, 혈액성상, 간과 가슴육의 일반성분에 미치는 영향을 구명하기 위하여 83주령 하이라인 산란계 192수를 이용하여 12주간 사양실험을 실시하였다. 기초사료는 에너지 수준이 2,800 kcal/kg, methionine과 lysine, cystine의 수준은 단백질 수준에 비례하도록 하였다. 조사항목으로 산란율과 난중, 사료섭취량, 사료요구율, 계란품질은 4주 간격으로 측정했다. 실험 종료시 복강지방과 혈액중 total protein과 albumin, BUN, triglyceride, total cholesterol, HDL-cholesterol, LDL-cholesterol 함량, 간의 methionine, choline 함량, 간과 가슴육의 수분, 조단백질, 조지방 함량을 측정하였다. 산란율은 단백질 수준에 따라 증가하였으며(P＜0.05) 비태인 급여에 의한 차이는 없었다. 난중은 단백질과 비태인 급여로 증가하였으며(P＜0.05) 산란양은 단백질 수준이 증가할수록 감소하였다(P＜0.05). 비태인 급여로 사료 요구율은 현저하게 개선되었지만(P＜0.05), 계란의 품질은 차이가 없었다. 혈청 total protein은 비태인 급여로 현저히 증가하였으며 특히 단백질 14 ％ 급여구에서 크게 증가하였다(P＜0.05). 복강지방 함량은 단백질 수준과 비태인 급여에 따라 감소하는 경향을 보였다. 가슴육과 간의 조단백질 함량은 사료의 단백질 수준에 따라 증가하였다(P＜0.05). 간의 조지방 함량은 비태인의 급여로 감소하는 경향을 보였다. 간의 methionine 함량은 단백질과 비태인 수준에 따라 증가하는 경향을 보였으며 choline 함량은 비태인의 급여에 의해서만 증가하는 경향을 보였다. 따라서 비태인의 급여는 단백질 수준이 높은 조건에서 산란율을 개선시키고 난중을 증가시키며 사료 요구율을 개선한다.록 산가는 증가하는 것을 볼 수 있다. 홍국주의 전자 공여능에 의한 항산화력은 25.6%, 아질산염 소거능은 27.6%, Total phenolic compound 함량은 12.34mg%, ACE저해작용은 38%의 항산화력을 나타냈다.과 $O_2$와 $CO_2$의 농도에서 평균오차 0.2%로 정밀한 것으로 나타났으며 호흡속도 측정용 챔버의 혼합기체 공급측과 배기측의 가스 농도를 3회 반복 측정한 결과 재현성에서는 0.1%이하의 편차로 나타났다. 개방계 호흡속도 자동 측정 시스템을 이용하여 환경기체조성하에서 토마토의 호흡속도를 측정하는 실측 실험을 수행한 결과 2$0^{\circ}C$에서 12.7~42.1mg$CO_2$/kg.hr였으며 12$^{\circ}C$에서 2.5~8.2mg$CO_2$/kg.hr로 일반적으로 보고되고 있는 토마토 호흡속도와 일치하는 결과를 나타내었다.다.환원당인 sucrose 함량은 계속 증가하였고 fructose, glucose, sorbitol의 함량(추황의 sorbitol을 제외)은 생장이 촉진됨에 따라 증가하다가 다시 점차적으로 감소하였다. 이러한 결과는 총당과 환원당의 측정결과와 일치한 것으로 나타났다. 결론적으로 배의 성장에 따라 산 함량은 감소하였고 당 함량은 증가하였다.luco-pyranoside, quercetin 7-O- -glucopyranoside, acacetin 7-O-$\beta$-D-glucuronide and apigenin-6-C-$\beta$-D-glucopyranosyl-8-C-$\beta$-D-glucopyranoside were first isolated and identified from safflower leaf. Among these flavonoids, luteolin

• Journal of Applied Biological Chemistry
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• v.61 no.4
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• pp.371-377
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• 2018

The fruits of Panax ginseng were extracted with 80% aqueous MeOH and the concentrates were partitioned into EtOAc, n-BuOH, and $H_2O$ fractions. The repeated $SiO_2$ and octadecyl $SiO_2$ column chromatographies for the EtOAc fraction led to isolation of five ginsenosides. The chemical structures of these compounds were determined as ginsenoside F1 (1), ginsenoside F2 (2), ginsenoside F3 (3), ginsenoside Ia (4), notoginsenoside Fe (5) based on spectroscopic analyses including nuclear magnetic resonance, MS, and infrared. Compounds 2-5 were isolated for the first time from the fruits of P. ginseng in this study. All isolated compounds were evaluated for cytotoxic activities against human cancer cell lines such as HCT-116, SK-OV-3, human cervix adenocarcinoma (HeLa), HepG2, and SK-MEL-5. Among them compounds 2, 4, and 5 showed significant cytotoxicity on cancer cells. Compound 2 exhibited cytotoxicity on SK-MEL-5, HepG2, and HeLa cells with $IC_{50}$ values of 82.8, 86.8, and $78.3{\mu}M$, respectively. Compound 4 showed cytotoxicity on HCT-116, SK-MEL-5, SK-OV-3, HepG2, and HeLa cells with $IC_{50}$ values of 24.5, 25.4, 26.3, 22.0, and $24.9{\mu}M$, respectively. Compound 5 did on SK-MEL-5 cell with $IC_{50}$ value of $81.7{\mu}M$. The cytotoxicity of ginsenoside 2, 4, and 5 isolated from the fruits of Panax ginseng showed strong inhibition effect against on cancer cells, all of which have a glucopyranosyl moiety on C-3.