• 약학회지
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• 제45권1호
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• pp.46-54
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• 2001

Protein carboxyl Ο-methylation is a kind of enzymatic reaction producing carboxyl methylester catalyzed by protein carboxyl Ο-methyltransferases at the carboxyl group of amino acid residues in polypeptide. Since the finding of carboxyl methylesterl many studies have been focused on the under-standing of biological functions in eukaryotes but still not clear except for roles in Ras attachment to membrane and protein repair. In this study, we investigated the protein carboxyl methylation in porcine liver and testis in respect of identification and characterization of carboxyl methylesters and natural proteinous substrates using pH stability of the esters and electrophoresis under acidic and basic conditions. We detected several kinds of methyl esters, 3 kinds each in cytosolic fractions from liver and testis. Under the treatment of strong acid and base, the ratio between base-stable substrates and unstable ones in liver (4 : 6) was different from the ratio obtained in testis (6 : 4). The methyl accepting capacities were affected by enzymatic proteolysis between the range of 55 to 65% in liver and of 35 to 45% in testis. Separation of the methylated proteins by acidic electrophoresis in the presence of urea and SDS revealed distinctively natural substrates of 26, 33 and 80 kD in the cytosol from liver and of 14, 25, 32 and 86 kD from testis. Most of the labelling, however were lost following electrophoresis under moderate alkaline condition, except for molecules of newly detected 7 and 17 kD in livers and 15, 29, 40 and 80 kD in testis. From these results, it was proposed that protein carboxyl Ο-methylation in each organs may be catalyzed by different classes of protein carboxyl Ο-methyltransferases. In addition, it is suggested that the protein carboxyl methylation in liver and testis may have different patterns in respect of natural substrates.

• 한국수산과학회지
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• 제35권5호
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• pp.524-528
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• 2002

국내에서 양식되는 가시파래의 당 결합 양식을 조사하기 위하여 가시파래의 수용성 산성 다당을 황산기 제거 및 우론산의 환원처리를 한 후 methylation하여 GC/MSD로 분석을 하였다. 가시파래의 수용성 산성 다당의 우론산은 대부분이 glucuronic acid 였으며, 주로 황산기를 함유한 rhamnose, xylose, glucuronic acid로 이루어진 다당이었다. 이 수용성 산성 다당을 화학적 수식한 후 GC/MSD로 분석시 황산기는 rhamnose의 C-3과 xylose의 C-2에 결합되어 있었으며, 주 결합구조는 C-3에 황산기가 결합한 1,4-, 1,2,4-결합된 rhamnose, C-2에 황산기가 결합한 1,4-결합된 xylose, 1,4-결합된 xylose, 그리고 1,4-결합된 glucuronic ac:김였다. 그리고 일부 미량의 1,4-결합된 rhamnose, 1,4,6-결합된 galactose도 분리되었다.

• Journal of Ginseng Research
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• 제44권4호
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• pp.570-579
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• 2020

Background: Many researchers reported that the various immune activities of red ginseng are due to acid polysaccharides. But, the exact structural characteristics of the acidic polysaccharide in red ginseng have not been fully elucidated. Therefore, we isolated the acidic polysaccharide from red ginseng and characterized the structural property of the active moiety of this polysaccharide, which contributes to the immunostimulatory activity of red ginseng. Methods: A polysaccharide (RGP-AP-I) was purified from red ginseng via size-exclusion chromatography using Sephadex G-100. Immunostimulatary activity of RGP-AP-I was investigated via anti-complementory and macrophage stimulatory activity. The structure of RGP-AP-I was characterized by HPLC, sugar composition, β-glucosyl Yariv reagent and methylation analysis. Results: Peritoneal macrophages stimulated using RGP-AP-I significantly augmented the production of various cytokines such as interleukin (IL)-6, IL-12, and tumor necrosis factor (TNF)-α. The primary structure of RGP-AP-I was elucidated by assessing its sugar composition and methylation analysis. RGP-AP-I is a 96 kDa acidic polysaccharide, and comprises nine different monosaccharides, which mainly include sugars such as rhamnose (Rha, 9.5%), galacturonic acid (GalA, 18.4%), galactose (Gal, 30.4%), and arabinose (Ara, 35.0%). RGP-AP-I exhibited an considerable reaction with the β-glucosyl Yariv reagent, revealing the presence of arabino-β-3,6-galactan. Methylation analysis indicated that RGP-AP-I comprises 21 different glycosyl linkages, such as 3-, 4-, 6- and 3,6-linked Galp; 5-linked Araf; 2,4-linked Rhap; and 4-linked GalAp, which are characteristics of rhamnogalacturonan I (RG-I). Conclusion: we assumed that the immunostimulatory activity of RGP-AP-I may be due to the RG-I structure, which comprises a main chain with a repeating linkage unit, [→2)-Rhap-(1→4)-GalAp-(1→] and three groups of side chains such as (1→5)-linked arabinan, (1→4)-linked galactan, and arabino-β-3,6-galactan, which branch at the C(O)4 positions of Rha residues in the main chain of RGP-AP-I.

• Bulletin of the Korean Chemical Society
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• 제8권3호
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• pp.193-196
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• 1987

The gas phase pyrolyses of eight esters have been studied by MNDO-MO method. In the ethylformate pyrolysis, ${\alpha}$-methylation had a steric releasing effect whereas ${\beta}$-methylation had a steric crowding in the transition state; the latter, however, is over-compensated by a greater electronic repulsion resulting in a net steric releasing effect. Considerations of formal charges and geometrical changes involved in the activation led us to propose a pyrolysis mechanism in which a preequilibrium of acidic proton transfer is followed by the rate-limiting bond polarization of $C_{\alpha}$-O bond in a cyclic transition state.

• Archives of Pharmacal Research
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• 제27권2호
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• pp.206-216
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• 2004

The functional role of protein carboxylmethylation (PCM) has not yet been clearly elucidated in the tissue level. The biochemical feature of PCM in porcine spleen was therefore studied by investigating the methyl accepting capacity (MAC) of natural endogenous substrate proteins for protein carboxyl O-methyltransferase (PCMT) in various conditions. Strong acidic and alkaline-conditioned (at pH 11.0) analyses of the MAC indicated that approximately 65％ of total protein methylation seemed to be mediated by spleen PCMT. The hydrolytic kinetics of the PCM products, such as carboxylmethylesters (CMEs), under mild alkaline conditions revealed that there may be three different kinds of CMEs [displaying half-times (T$_{1}$2/) of 1.1 min (82.7％ of total CMEs), 13.9 min (4.6％), and 478.0 min (12.7％)], assuming that the majority of CME is base-labile and may be catalyzed by class I PCMT. In agreement with these results, several natural endogenous substrate proteins (14, 31 and 86 kDa) were identified strikingly by acidic-conditioned electrophoresis, and their MAC was lost upon alkaline conditions. On the other hand, other proteins (23 and 62 kDa) weakly appeared under alkaline conditions, indicating that PCM mediated by class II or III PCMT may be a minor reaction. The MAC of an isolated endogenous substrate protein (23-kDa) was also detected upon acidic-conditioned electrophoresis. Therefore, our date suggest that most spleen PCM may be catalyzed by class I PCMT, which participates in repairing aged proteins.

• Environmental Analysis Health and Toxicology
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• 제15권3호
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• pp.107-113
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• 2000

Chlorophenoxy acid류는 제초제로 폭넓게 사용되고 있는 농약 중의 하나이다. 특히 2, 4-D와 2, 4, 5-T는 내분비계 장애 물질 중의 하나로 알려져 있다. 본 연구에서는 이 두 가지 물질을 산성하에서 액체-액체 추출법으로 추출한 후 유도체화하여 GC/MS로 분석하였다. 추출용매로 methylene chloride와 diethyl ether를 사용하여 추출율을 비교한 결과 ether를 사용한 경우 2, 4-D는 약 98% 2, 4, 5-T는 약 82%의 좋은 회수율을 보였다. 또한 산성기를 유도체화하기 위하여 $CH_3$I/Acetone -K$_2$CO$_3$와 황산/메탄올로 각각 methylation실험을 행하였고, 산성조건에서 triflouroethanol (TFAA/TEE)과 반응시켜 각각의 검출검도를 비교하였다. 그 결과 TPAA/TEE를 사용하여 반응시킨 결과감도가 좋은 것으로 나타났다.

• 한국균학회지
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• 제27권1호통권88호
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• pp.76-81
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• 1999

영지 균사체의 액체 배양에 의하여 생산된 세포의 다당시료 분획 중 우수한 항암활성을 나타내었던 산성분획인 BWS-DA-GI의 구조를 균사체 유래의 산성 다당 분획(MWS-DA-GI)과 비교하면서 조사하였다. BWS-DA-GI의 구성성분을 검토한 결과, 다량의 glucose, mannose 및 galactose와 미량의 xylose 및 fucose를 함유하였으며, glucose, galactose 및 mannose의 몰비는 2.5:2.1:2.5이었다. 반면, 균사체 유래의 산성 다당 분획인 MWS-DA-GI는 다량의 galactose, fucose, mannose 및 glucose와 미량의 xylose를 함유하였다. 또 BWS-DA-GI의 구조를 균사체 유래의 다당 분획인 MWS-DA-GI와 비교하면서 과요오드 산화, Smith분해, affinity chromatography 및 메칠화 분석으로 조사한 결과, 두 시료 모두 ${\beta}-1,3-glucan$이었고, BWS-DA-GI는 1,6분지를 보유하고 있는 반면, MWS-DA-GI는 1,4 및 1,6분지를 함께 보유하는 것으로 추정되었으며, 분자량은 각각 1,200,000 및 1,000,000 dalton이었다.

• Journal of Ginseng Research
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• 제48권5호
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• pp.454-463
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• 2024

Background: Panax quinquefolius L, widely recognized for its valuable contributions to medicine, has aroused considerable attention globally. Different from the extensive research has been dedicated to the root of P. quinquefolius, its berry has received relatively scant focus. Given its promising medicinal properties, this study was focused on the structural characterizations and anti-inflammatory potential of acidic polysaccharides from the P. quinquefolius berry. Materials and methods: P. quinquefolius berry was extracted with hot water, precipitated by alcohol, separated by DEAE-52-cellulose column to give a series of fractions. One of these fractions was further purified via Sephadex G-200 column to give three fractions. Then, the main fraction named as AGBP-A3 was characterized by methylation analysis, NMR spectroscopy, etc. Its anti-inflammatory activity was assessed by RAW 264.7 cell model, zebrafish model and molecular docking. Results: The main chain comprised of α-L-Rhap, α-D-GalAp and β-D-Galp, while the branch consisted mainly of α-L-Araf, β-D-Glcp, α-D-GalAp, β-D-Galp. The RAW264.7 cell assay results showed that the inhibition rates against IL-6 and IL-1β secretion at the concentration of 625 ng/mL were 24.83 %, 11.84 %, while the inhibition rate against IL-10 secretion was 70.17 % at the concentration of 312 ng/mL. In the zebrafish assay, the migrating neutrophils were significantly reduced in number, and their migration to inflammatory tissues was inhibited. Molecular docking predictions correlated well with the results of the anti-inflammatory assay. Conclusion: The present study demonstrated the structure of acidic polysaccharides of P. quinquefolius berry and their effect on inflammation, providing a reference for screening anti-inflammatory drugs.

• 한국식품과학회지
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• 제29권6호
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• pp.1105-1112
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• 1997

약산성 다당 GL-4IIb2'에 대해 연속적 부분 산가수 분해 과정을 거쳐, 2종의 oligo당 획분 PA-2' 및 PA-1-III를 분리하고 그들의 구조분석을 행하였다. PA-2'는 주요 구성당으로 Rha 와 특이당 Kdo를 동일한 비율로 함유하고 있었으며, permethylated oligosaccharide-alditol로 전환시켜, GC-MS로 분석한 결과, m/z 189$(bA_1)$및 m/z 308 $(aJ_2)$에서 6-deoxyhexose와 Kdo에 기인하는 fragment ion이 관찰되었다. 이 peak는 m/z 162에서 특징적인 ion을 나타낸 반면 m/z 177 ion이 관찰되지 않음으로써 Kdo의 C4위치가 아닌 C5위치가 치환되어 있음을 알 수 있었다. 한 methyl화 분석 결과, PA-2'은 환원말단 Rhap와 5-substituted Kdo가 높은 비율로 검출되었으며 환원 형태의 PA-2'를 대상으로 $^1M-NMR$ 분석을 하였을 때 ${\delta}$ 5.09 ppm에서 ${\alpha}-L-Rha$의 anomeric proton에 기인한 단일 signal이 관찰되었다. 이상의 결과로 부터 PA-2'는 주로 ${\alpha}-L-Rhap-(1{\rightarrow}5)-Kdo$로 구성되었다는 사실을 알 수 있었다 한편. PA-1-III는 소량의 Ara 믹 Dha와, 다량의 Rha 및 Kdo가 주요 구성당으로 나타났으며, PA-1-III 획분의 permethylated oligosaccharide-alditol 유도체는 GC상에서 3개의 peak (1P, 2P 및 3P)로 분리되었다. 이들의 MS 분석 결과, 높은 비율로 검출된 1P는 $Rhap-(1{\rightarrow}5)-Kdo$의 구조로 나타났으며 반면 낮은 비율의 2P및 3P는 $Araf-(1{\rightarrow}5)-Dha$의 동일구조를 갖는 것으로 확인되었는데 이는 Dha의 carbonyl 환원과정 중 생성된 epimer로 판단되었다.

• 분석과학
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• 제22권6호
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• pp.480-487
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• 2009

토양 및 식물 중 잔류하는 제초제 디캄바를 기체크로마토그래피-질량검출법으로 측정하는 방법을 개발하였다. 토양 또는 식물 시료를 pH 2로 조절한 후 diethyl ether로 추출한 다음 0.1 N HCl로 정제한 후 증발 건조시켰다. 잔류물에 10% 황산 메탄올 용액 1 mL를 가한 후 $80^{\circ}C$에서 2 시간 반응시켰다. 반응 후 탄산수소소듐 포화용액 4 mL를 서서히 가하여 중화시킨 후 diethyl ether 5 mL로 재 추출 한 다음 추출액을 농축시켜 GC/MS에 주입하여 분석하였다. 그 결과 $1.0{\sim}100{\mu}g/kg$의 정량구간 내에서 $R^2$=0.999 이상의 좋은 직선성을 보였다. 분석결과 총 32개 토양 시료 중 15개 토양에서 디캄바가 $2.9-123.9{\mu}g/kg$의 농도범위로 검출되었다. 한편 식물에서는 총 10개의 시료 중 5 개 시료에서 디캄바가 $43-33,252{\mu}g/kg$의 농도범위로 검출되었다. 따라서 소나무가 고사한 이유는 디캄바를 직접 나무에 또는 주변에 뿌렸기 때문으로 판단된다.