• Molecules and Cells
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• 제37권2호
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• pp.172-177
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• 2014

A glycosyltransferase, YjiC, from Bacillus licheniformis has been used for the modification of the commercially available isoflavonoids genistein, daidzein, biochanin A and formononetin. The in vitro glycosylation reaction, using UDP-${\alpha}$-D-glucose as a donor for the glucose moiety and aforementioned four acceptor molecules, showed the prominent glycosylation at 4' and 7 hydroxyl groups, but not at the $5^{th}$ hydroxyl group of the A-ring, resulting in the production of genistein 4'-O-${\beta}$-D-glucoside, genistein 7-O-${\beta}$-D-glucoside (genistin), genistein 4',7-O-${\beta}$-D-diglucoside, biochanin A-7-O-${\beta}$-D-glucoside (sissotrin), daidzein 4'-O-${\beta}$-D-glucoside, daidzein 7-O-${\beta}$-D-glucoside (daidzin), daidzein 4', 7-O-${\beta}$-D-diglucoside, and formononetin 7-O-${\beta}$-D-glucoside (ononin). The structures of all the products were elucidated using high performance liquid chromatography-photo diode array and high resolution quadrupole time-of-flight electrospray ionization mass spectrometry (HR QTOF-ESI/MS) analysis, and were compared with commercially available standard compounds. Significantly higher bioconversion rates of all four isoflavonoids was observed in both in vitro as well as in vivo bioconversion reactions. The in vivo fermentation of the isoflavonoids by applying engineered E. coli $BL21(DE3)/{\Delta}pgi{\Delta}zwf{\Delta}ushA$ overexpressing phosphoglucomutase (pgm) and glucose 1-phosphate uridyltransferase (galU), along with YjiC, found more than 60% average conversion of $200{\mu}M$ of supplemented isoflavonoids, without any additional UDP-${\alpha}$-D-glucose added in fermentation medium, which could be very beneficial to large scale industrial production of isoflavonoid glucosides.

• ALGAE
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• 제35권2호
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• pp.177-187
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• 2020

Cyanobacteria have been widely reported to produce a variety of UV-absorbing mycosporine-like amino acids (MAAs). Herein, we reported production of the unusual MAA, mycosporine-glycine-alanine (MGA) in the cyanobacterium Sphaerospermopsis torques-reginae ITEP-024 using a newly developed UHPLC-DAD-MS/HRMS (ultra-high performance liquid chromatography-diode array detection-high resolution tandem mass spectrometry) method. MGA had previously been first identified in a red-algae, but S. torques-reginae strain ITEP-024 is the first cyanobacteria to be reported as an MGA producer. Herein, the chemical structure of MGA is fully elucidated from one-dimensional / two-dimensional nuclear magnetic resonance and HRMS data analyses. MAAs are unusually produced constitutively in S. torques-reginae ITEP-024, and this production was further enhanced following UV-irradiance. It has been proposed that MAA biosynthesis proceeds in cyanobacteria from the pentose phosphate pathway intermediate sedoheptulose 7-phosphate. Annotation of a gene cluster encoded in the genome sequence of S. torques-reginae ITEP-024 supports these gene products could catalyse the biosynthesis of MAAs. However, addition of glyphosate to cultures of S. torques-reginae ITEP-024 abolished constitutive and ultra-violet radiation induced production of MGA, shinorine and porphyra-334. This finding supports involvement of the shikimic acid pathway in the biosynthesis of MAAs by this species.

• Mycobiology
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• 제49권4호
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• pp.434-437
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• 2021

In our ongoing search for new secondary metabolites from fungi, a basidiomycete fungus Irpex consors was selected for mycochemical investigation, and three new zwitterionic alkaloids (1-3) and five known compounds (4-8) were isolated from the culture broth (16 l) of I. consors. The culture filtrate was fractionated by a series of column chromatography including Diaion HP-20, silica gel, and Sephadex LH-20, Sep-Pak C₁₈ cartridge, medium pressure liquid chromatography (MPLC), and high pressure liquid chromatography (HPLC) to yield eight compounds (1-8). The structures of the isolated compounds were elucidated by the interpretation of nuclear magnetic resonance (NMR) spectra and high-resolution mass spectrometry (HR-MS). Their antioxidant and antibacterial activities were examined. The zwitterionic structures of three new sesquiterpene alkaloids (1-3) were determined together with five known compounds identified as stereumamide E (4), stereumamide G (5), stereumamide H (6), stereumamide D (7), and sterostrein H (8). This is the first report of the zwitterionic alkaloids in the culture broth of I. consors. Three new zwitterionic alkaloids were named as consoramides A-C (1-3).

• 한국근적외분광분석학회:학술대회논문집
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• 한국근적외분광분석학회 2001년도 NIR-2001
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• pp.1516-1516
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• 2001

During the recent years, wine analysis has played an increasing role due the health benefits of phenolic ingredients in red wine [1]. On the other hand there is the need to be able to distinguish between different wine varieties. Consumers want to know if a wine is an adulterated one or if it is based on the pure grape. Producers need to certificate their wines in order to ensure compliance with legal regulations. Up to now, the attempts to investigate the origin of wines were based on high-performance liquid chromatography (HPLC), gas chromatography (GC) and pyrolysis mass spectrometry (PMS) [l,2,3]. These methods need sample pretreatment, long analysis times and therefore lack of high sample throughput. In contradiction to these techniques using near infrared spectroscopy (NIRS), no sample pretreatment is necessary and the analysis time for one sample is only about 10 seconds. Hence, a near infrared spectroscopic method is presented that allows a fast classification of wine varieties in bottled red wines. For this, the spectra of 50 bottles of Cabernet Sauvignon, Lagrein and Sangiovese (Chianti) were recorded without any sample pretreatment over a wavelength range from 1000 to 2500 nm with a resolution of 12 cm$\^$-1/. 10 scans were used for an average spectrum. In order to yield best reproducibility, wines were thermostated at 23$^{\circ}C$ and a optical layer thickness of 3 mm was used. All recorded spectra were partitioned into a calibration and validation set (70% and 30%). Finally, a 3d scatter plot of the different investigated varieties allowed to distinguish between Cabernet Sauvignon, Lagrein and Sangiovese (Chianti). Considering the short analysis times this NRS-method will be an interesting tool for the quality control of wine verification and also for experienced sommeliers.

• FOCUS: LIFE SCIENCE
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• 제1호
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• pp.3.1-3.7
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• 2024

The article discusses the critical role of chromatography in the analysis and purification of proteins in biopharmaceuticals, emphasizing the importance of comprehensive characterization for ensuring their safety and efficacy. It highlights the use of Avantor® ACE® HPLC columns for the separation and purification of proteins, focusing on the analysis of intact proteins using reversed-phase liquid chromatography (RPLC) with fully porous particles. This article also details the application of different mobile phase additives, such as TFA and formic acid, and emphasizes the advantages of using type B ultra-pure silica-based columns for efficiency and peak shape in biomolecule analysis. Additionally, it addresses the challenges of analyzing intact proteins due to slow molecular diffusion and introduces the concept of solid-core (or superficially porous) particles, emphasizing their benefits over traditional porous particles for the analysis of therapeutic proteins. Furthermore, it discusses the development of Avantor® ACE® UltraCore BIO columns, specifically designed for the high-efficiency separation of large biomolecules, such as proteins, and demonstrates their effectiveness in achieving high-resolution separations, even for higher molecular weight proteins like monoclonal antibodies (mAbs). In addition, it underscores the complexity of analyzing and characterizing intact protein biopharmaceuticals, requiring a range of analytical techniques and the use of wide-pore stationary phases, operated at elevated temperatures and with relatively shallow gradients. It highlights the comprehensive range of options offered by Avantor® ACE® wide pore columns, including both fully porous and solid-core particles, bonded with a variety of complementary stationary phase chemistries to optimize selectivity during method development. The use of ultrapure and highly inert base silica is emphasized for enabling the use of lower concentrations of mobile phase modifiers without compromising analyte peak shape, particularly beneficial for LC-MS applications. Then the article concludes by emphasizing the significance of reversed-phase liquid chromatography and its compatibility with mass spectrometry as a valuable tool for the separation and analysis of intact proteins and their closely related variants in biopharmaceuticals.

• 한국식품과학회지
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• 제43권5호
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• pp.544-552
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• 2011

식품 중 존재하는 PAHs를 분석하기 위해 GC-MS/MS를 이용하여 분쇄원두커피를 대상으로 15종의 PAHs를 동시분석 하였다. 전처리법은 soxhlet법, 비누화분해 후 액체-액체 추출법, 초음파후 고체-액체추출법으로 비교실험하여 재현성과 불순물제거효율이 가장 좋은 초음파 후 고체-액체 추출법을 선택하여 실험하였다. 실험법의 회수율은 PAHs종에 따라 52.6-93.3%, 검출한계는 0.002-0.1 ${\mu}g/kg$, 정량한계는 0.006-0.2 ${\mu}g/kg$로 PAHs분석에 가장 좋은 감도를 나타내는 HPLC-FLD나 GC-MS-SIM과 비슷하거나 낮은 수준의 검출한계를 나타내었다. 적합한 전처리 방법을 선택한 후 시중 유통되는 원두커피를 대상으로 15종의 PAHs 함량을 조사하였을 때 46종의 총 PAHs농도는 불검출-5.988 ${\mu}g/kg$이었다. 분쇄원두커피를 커피메이커로 추출하여 PAHs의 이행계수를 계산 한 후 WHO와 U.S.EPA에서 설정한 상대독성계수(TEF)를 근거로 음용되는 원두커피의 인체노출량(mg/kg b.w./day)을 구한 결과 $5.24{\times}10^{-8}$ mg/kg b.w./day였다.

• 분석과학
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• 제16권4호
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• pp.309-319
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• 2003

본 연구에서는 전주시 대기 시료를 대상으로 PCBs 209개 전 이성체를 분석하였다. 추출한 시료는 황산 실리카겔 및 활성탄 컬럼으로 정제하였으며 DB-5 컬럼 ($60m{\times}0.25mm{\times}0.25{\mu}m$)을 이용하여 고분해능 가스크로마토그래피 및 고분해능 질량분석기 (HRGC/HRMS)로 분석을 수행하였다. 대기 시료 중 PCBs 농도는 $0.003{\sim}0.163pg-TEQ/m^3$의 범위를 보였다. DB-5 컬럼을 사용하여 분석한 결과 총 162개의 피크가 검출되었고, 그 중 37개의 피크는 하나 이상의 이성체가 중복된 피크로 정성되었다. 또한, 독성을 가지고 있는 coplanar-PCB 중에서도 IUPAC No. 118/106과 105/127번의 분리가 어려워 coplanar-PCB 농도를 과대평가 할 가능성이 있는 것으로 나타났다. 발생원 시료 (Kanechlor와 소각로 배가스)와 대기 시료와의 coplanar-PCB 이성체 분포를 비교한 결과, PCB-81, 77, 126와 169이성체는 소각로 시료에서 높은 비율을 나타냈다.

• 한국축산식품학회지
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• 제30권1호
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• pp.73-86
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• 2010

The current study was designed to optimize solid phase microextraction (SPME)-GC-MS conditions for extraction and analysis of volatile components for Hanwoo beef and to establish a tentative database of flavor components. Samples were taken from Hanwoo longissimus muscle (30 mon old steer, $1^+B$ carcass grade) at 24 h postmortem. Results indicated that the optimum adsorption time for $75{\mu}m$ CAR/PDMS fiber was 60 min at $60^{\circ}C$. Thermal cleaning at $250^{\circ}C$ for 60 min was the best practice for decontamination of the fiber. A short analysis program with a sharp oven temperature ramp resulted in a better resolution and higher number of measurable volatile components. With these conditions, 96 volatile compounds were identified with little variation including 22 aldehydes, 8 ketones, 31 hydrocarbons, 12 alcohols, 8 nitrogen- and sulfurcontaining compounds, 5 pyrazines and 10 furans. A noticeable observation was the high number of hydrocarbons, aldehydes, ketones, alcohols and 2-alkylfurans which were generated from lipid decomposition especially the oxidation and degradation of unsaturated and saturate fatty acids. This implies that these compounds can be candidates for flavor specification of highly marbled beef such as Hanwoo flavor.

• 미생물학회지
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• 제46권1호
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• pp.1-8
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• 2010

미생물 군집과 기능연구를 위한 최근의 새로운 분석기술의 발전은 다양한 유전 관련 정보를 제공해왔다. mRNA를 포함하는 핵산을 기초로 한 연구를 뛰어넘어서 메타프로테오믹스는 미생물 군집의 유전형 및 표현형의 특징적인 정보를 보다 정교하게 제공할 수 있다. 이미 서로 다른 미생물 생태계인 해수, 인간의 배설물, 활성 슬러지, 산성 광산 폐수 생물막, 토양 등에 메타프로테오믹스 기술이 유용하게 사용되었다. 이들 연구는 여러 측면에서 상당히 다르지만 미생물 군집의 구조, 기능, 생리, 상호관계, 생태, 진화적 측면을 결정적으로 상호 연결한다는 것을 밝혀냈다. 본 총설은 메타프로테오믹스에 대한 현재까지의 가장 최신의 정보를 요약하여 제공함으로써 메타프로테오믹스에 대한 정확한 이해와 활용을 통해 다방면의 메타프로테오믹스가 가능하도록 하고자 하였다.

• Journal of Microbiology and Biotechnology
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• 제27권8호
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• pp.1472-1482
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• 2017

Bacterial cytochrome P450 (CYP) steroid hydroxylases are effectively useful in the pharmaceutical industry for introducing hydroxyl groups to a wide range of steroids. We found a putative CYP steroid hydroxylase (BaCYP106A2) from the bacterium Bacillus sp. PAMC 23377 isolated from Kara Sea of the Arctic Ocean, showing 94% sequence similarity with BmCYP106A2 (Bacillus megaterium ATCC 13368). In this study, soluble BaCYP106A2 was overexpressed to evaluate its substrate-binding activity. The substrate affinity ($K_d$ value) to 4-androstenedione was $387{\pm}37{\mu}M$. Moreover, the crystal structure of BaCYP106A2 was determined at $2.7{\AA}$ resolution. Structural analysis suggested that the ${\alpha}8-{\alpha}9$ loop region of BaCYP106A2 is intrinsically mobile and might be important for initial ligand binding. The hydroxyl activity of BaCYP106A2 was identified using in vitro enzyme assays. Its activity was confirmed with two kinds of steroid substrates, 4-androstenedione and nandrolone, using chromatography and mass spectrometry methods. The main products were mono-hydroxylated compounds with high conversion yields. This is the second study on the structure of CYP106A steroid hydroxylases, and should contribute new insight into the interactions of bacterial CYP106A with steroid substrates, providing baseline data for studying the CYP106A steroid hydroxylase from the structural and enzymatic perspectives.