• Mass Spectrometry Letters
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• 제5권3호
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• pp.63-69
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• 2014

In this study, we present a new isotope-coded carbamidomethylation (iCCM)-based quantitative proteomics, as a complementary strategy for conventional isotope labeling strategies, with providing the simplicity, ease of use, and robustness. In iCCM-based quantification, two proteome samples can be separately isotope-labeled by means of covalently reaction of all cysteinyl residues in proteins with iodoacetamide (IAA) and its isotope (IAA-$^{13}C_2$, $D_2$), denoted as CM and iCCM, respectively, leading to a mass shift of all cysteinyl residues to be + 4 Da. To evaluate iCCM-based isotope labeling in proteomic quantification, 6 protein standards (i.e., bovine serum albumin, serotransferrin, lysozyme, beta-lactoglobulin, beta-galactosidase, and alpha-lactalbumin) isotopically labeled with IAA and its isotope, mixed equally, and followed by proteolytic digestion. The resulting CM-/iCCM-labeled peptide mixtures were analyzed using a nLC-ESI-FT orbitrap-MS/MS. From our experimental results, we found that the efficiency of iCCM-based quantification is more superior to that of mTRAQ, as a conventional nonisobaric labeling method, in which both of a number of identified peptides from 6 protein standards and the less quantitative variations in the relative abundance ratios of heavy-/light-labeled corresponding peptide pairs. Finally, we applied the developed iCCM-based quantitative method to lung cancer serum proteome in order to evaluate the potential in biomarker discovery study.

• 한국자기공명학회논문지
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• 제15권2호
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• pp.115-127
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• 2011

A newly developed cost-effective approach to prepare $^{15}N/^{13}C$-labeled protein for NMR studies is presented. This method has been successfully applied to isotopically labeling of PTK6 SH2 domain and MTH 1880 protein. The production method generates cell density using a growing media containing $^{15}NH_4Cl$, $^{12}C_6$-D-glucose. Following a doubling time period for unlabeled metabolite exhaustion and then addition $^{13}C_6$-D-glucose into a M9 growing media, the cells are induced. Our results demonstrate that in order to get full incorporation of $^{13}C$, the isotopes are not totally required during the initial growth phase before induction. The addition of small amounts of $^{13}C_6$-D-glucose to the induction phase is sufficient to obtain more than 95% incorporation of isotopes into the protein. Our optimized protocol is two-thirds less costly than the classical method using $^{13}C$ isotope during the entire growth phase.

• 헤리티지:역사와 과학
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• 제36권
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• pp.241-266
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• 2003

Glass pieces excavated from Mireuksa Temple dated $7^{th}$ century A.D. were characterized by chemical composition, specific gravity and melting point. Lead isotope ratios of lead glasses were also compared with those of lead ore to attribute which lead ore was delivered for making lead glass. It was known that some lead glasses found in Japan were similar with those of Mireuksa Temple as comparing the data of chemical composition and lead isotope ratios. Characteristics of lead glass from Mireuksa Temple Thirty five glass pieces of Mireuksa Temple were analyzed for five oxides and found that all was lead glass system(PbO-$SiO_2$) with the range of 70~79% for PbO and 20~28% for $SiO_2$. The concentrations of oxides such as $Al_2O_3$, $Fe_2O_3$ and CuO were below 0.4%, 0.3% and 0.9%, respectively. Principal component analysis(PCA) as a statistical method was carried out to classify glasses with the similarities of chemical concentrations. The result of PCA has shown that three groups of glasses were created according to the excavation positions and two major oxides(PbO and $SiO_2$) greatly contributed to the dispersion of glasses on principal component 1(PC1) axis and trace element oxides($Al_2O_3$ and $Fe_2O_3$) for PC2 axis. Most of lead glasses were greenish by the efficacy of iron and copper oxides and some showed yellowish-green. The gravity of lead glasses was about 4.4~5.4 and estimated melting point was near $670^{\circ}C$. Lead isotope ratios of glasses were analyzed and found quite close to a lead ore from the Bupyeong mine in Gyeonggi-do. Comparison with lead glasses found in Japan Lead glasses of Mireuksa Temple were compared with those of Japan on the basis of chemical and physical data. Chemical compositions of Japanese lead glasses dated $7^{th}{\sim}8^{th}$ century A.D. were nearly similar with those of Mireuksa Temple but lead isotope ratios of those were separated into two groups. Three distribution maps of lead ores of Korea, Japan and China with lead isotope ratios were applied for lead glasses found in Japan. The result have shown that the locations of lead glasses from Fukuoka Prefecture coincided with the region of northen part of Korea and similar with those of Mireuksa Temple and lead glasses from Nara Prefecture dated $8^{th}$ century A.D. were located in the region of Japanese lead ore. This research has demonstrated that lead glasses of Mireuksa Temple conveyed to Miyajidake site, Fukuoka Prefecture around $7^{th}$ century A.D. and glass melting pots and glass beads excavated from Nara Prefecture confirmed the first use of Japanese lead ore for production of lead glasses from the end of $7^{th}$ century A.D.

• 자원환경지질
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• 제33권6호
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• pp.491-504
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• 2000

Environmental isotope $^{18}O$, $^{2}H$, $^{3}H$,$^{13}C$, $^{34}S$and $^{87}Sr/^{86}Sr$) studies on ${CO_2}$-rich waters in the Kangwon Province were carried out to elucidate the origin, residence time, water-rock interaction and mixing process of their. ${\delta}^{18}O$ and ${\delta}D$ data indicate that ${CO_2}$-rich waters were derived from the local meteoric water. It also shows that each type of ${CO_2}$-rich water has distinct isotopic composition and Na-${HCO_3}$ type water (-10.8 to -12.1${\textperthousand}$, ${\delta}^{18}O$ ) is lighter than other type waters. These depleted isotopic values supposedly indicate that, considering the altitude effect of isotope in Korea, the recharge area of Na-${HCO_3}$ type water can be estimated to be relatively higher in elevation than those of Ca-${HCO_3}$ and Ca-Na-${HCO_3}$ type waters. Tritium contents close to zero are observed in the Na-${HCO_3}$ type water, confirming a long residence time and the possibility of a ${CO_2}$ inflow into the aquifer at great depth. These isotope data also show that the Ca-${HCO_3}$ type water has undergone mixing process with surface water during ascending at depth, whereas Na-${HCO_3}$ type water was less mixed with surface waters. The carbon isotope data (-8.8 to ＋0.8 ${\textperthousand}$ ${\delta}^{13}C$) indicate that dissolved carbon in the ${CO_2}$-rich waters was possibly derived from deep seated ${CO_2}$ gas. The high ${\delta}^{34}S$ values (up to 38.1${\textperthousand}$) of dissolved sulfates suggest that sulfate reduction by microbial activity had occurred at depth. Strontium isotopic data ($^{87}Sr/^{86}Sr$) of ${CO_2}$-rich waters indicate that the chemistry of the ${CO_2}$-rich waters is determined by water-rock (granite) interaction.

• 생명과학회지
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• 제8권4호
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• pp.410-415
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• 1998

HPLC 및 electrospary mass spectrum으로부터 L-L dipeptide의 존제하에서 pepsin은 hexapeptide인 L-S-pNF-Nle-A-OMe를 가수분해하여 가수분해물외의 새로운 생성물을 합성하는 것이 확인되었다. 이 생성물은 254nm에서 p-nitro-Phe 잔기를 포함하는 peptide였다. 실험결과로부터 E(L-S-pNF)와 L-L 사이의 acyl transpeptidation에 의해 L-S-pNF-L-L가 생성됨을 뒷받침한다. 이러한 transpeptidation 결과는 product 저해실험에 의한 결과에 기초한 것과는 반대로 L-S-pNF가 해리되기전에 Nle-A-L-OMe가 먼저 한다는 것을 보여준다. 그리고, electrospray mass spectrum 으로부터 위에서 검출된 새로운 펩티드에 해당하는 peak (MW 636.1)을 얻었는데, 이는 새 펩티드의 생성을 확실히 증명하는 증거이다. 한편, Nle-A-L-OMe 생성에 대한 solvent isotope effect는 1.736$\pm$0.121이며 L-S-pNF는 2.28$\pm$0.184 그리고 L-S-pNF-L-L의 생성에는 inverse isotope effect로서 0.576$\pm$0.045였는데, 이는 상기 생성물 해리 순서를 확인시켜 준다. D$_{2}$에서 transpeptidation은 더 빠르기 때문에 isotopically-sensitive단계는 Nle-A-L-OMe해리후에 존재하는 것을 알 수 있다. 본 실험결과는, Rebholz and Northrop$^{1)}$ 및 Cho등의 $^{2)} iso-mechanism이론의 타당성을 제시한다.

• Bulletin of the Korean Chemical Society
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• 제34권12호
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• pp.3811-3816
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• 2013

Nucleophilic substitution reactions of ethyl methyl (2), ethyl propyl (4) and diisopropyl (7) chlorothiophosphates with substituted anilines and deuterated anilines are investigated kinetically in acetonitrile at $55.0^{\circ}C$. A concerted mechanism is proposed based on the selectivity parameters. The deuterium kinetic isotope effects (DKIEs; $k_H/k_D$) are secondary inverse ($k_H/k_D=0.66-0.99$) with 2, primary normal and secondary inverse ($k_H/k_D=0.78-1.19$) with 4, and primary normal ($k_H/k_D=1.06-1.21$) with 7. The primary normal and secondary inverse DKIEs are rationalized by frontside attack involving hydrogen bonded, four-center-type transition state, and backside attack involving in-line-type transition state, respectively. The anilinolyses of ten chlorothiophosphates are examined based on the reactivity, steric effect of the two ligands, thio effect, reaction mechanism, DKIE and activation parameter.

• Animal cells and systems
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• 제14권4호
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• pp.297-303
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• 2010

To document the basic ecological aspects of Chaoborus species, which has never been reported in Korea, we attempted to identify the species, to monitor seasonal and vertical dynamics, and to elucidate trophic relations of the species in Sang-Chun reservoir. Using morphological characteristics, we identified the collected samples as C. flavicans. Also, we compared the distribution of C. flavicans, Daphnia rosea and chlorophyll a to observed seasonal dynamics. The increase of C. flavicans was observed 1-2 weeks after the increase of D. rosea. Survey of diel vertica migration patterns in the summer season showed that C. flavicans were in hypolimnion at daytime, but moved to the epilimnion at night. Finally, to determine trophic relationships in Sang-Chun reservoir, additional studies on the food web were undertaken by stable isotope analysis. Chaoborus flavicans I-II instars appear to be filter feeders based on carbon isotope values. Trophic levels of C. flavicans III-IV instars were shown to be higher than other zooplankto based on nitrogen isotope values.

• Bulletin of the Korean Chemical Society
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• 제29권9호
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• pp.1769-1773
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• 2008

fThe kinetics and mechanism of the reactions of Y-O-aryl phenylphosphonochloridothioates with X-pyridines are investigated in acetonitrile at 35.0 ${^{\circ}C}$. The negative value of the cross-interaction constant, $\rho$XY = −0.46, indicates that the reaction proceeds by concerted $S_N2$ mechanism. The observed $k_H/k_D$ values involving d-5 pyridine ($C_5D_5N$) nucleophiles are greater than unity (1.05-1.11). The net primary deuterium kinetic isotope effects, $(k_H/k_D)_{net}$ = 1.28-1.35, excluding the increased $pK_a$ effect of d-5 pyridine are obtained. The transition state with a hydrogen bond between the leaving group Cl and the hydrogen (deuterium) atom in the C-H(D) is suggested for the studied reaction system.

• Bulletin of the Korean Chemical Society
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• 제32권7호
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• pp.2306-2310
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• 2011

The nucleophilic substitution reactions of diethyl thiophosphinic chloride with substituted anilines ($XC_6H_4NH_2$) and deuterated anilines ($XC_6H_4ND_2$) are investigated kinetically in acetonitrile at 55.0 $^{\circ}C$. The values of deuterium kinetic isotope effects (DKIEs; $k_H/k_D$) invariably increase from secondary inverse ($k_H/k_D$ < 1) to primary normal (kH/kD > 1) as the nucleophiles change from the strongly basic to weakly basic anilines. The secondary inverse with the strongly basic anilines and primary normal DKIEs with the weakly basic anilines are rationalized by the gradual transition state (TS) variation from a predominant backside attack, via invariably increasing the fraction of a frontside attack, to a predominant frontside attack, in which the reaction mechanism is a concerted $S_N2$ pathway. A frontside attack involving a hydrogen bonded, four-center-type TS is substantiated by the primary normal DKIEs.

• Bulletin of the Korean Chemical Society
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• 제33권10호
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• pp.3274-3278
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• 2012

The kinetic studies on the reactions of phenyl N-phenyl phosphoramidochloridate (8) with substituted anilines ($XC_6H_4NH_2$) and deuterated anilines ($XC_6H_4ND_2$) have been carried out in acetonitrile at $60.0^{\circ}C$. The obtained deuterium kinetic isotope effects (DKIEs; $k_H/k_D$) are huge secondary inverse ($k_H/k_D$ = 0.52-0.69). A concerted mechanism is proposed with a backside attack transition state (TS) on the basis of the secondary inverse DKIEs and the variation trends of the $k_H/k_D$ values with X. The degree of bond formation in the TS is really extensive taking into account the very small values of the DKIEs. The steric effects of the two ligands on the rates are extensively discussed for the aminolyses of the chlorophosphate-type substrates on the basis of the Taft equation.