• Title/Summary/Keyword: HPLC-ESI-MS/MS

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Production, Purification, and Characterization of Taxol and 10-DABIII from a new Endophytic Fungus Gliocladium sp. Isolated from the Indian Yew Tree, Taxus baccata

  • Sreekanth, D.;Syed, A.;Sarkar, S.;Sarkar, D.;Santhakumari, B.;Ahmad, A.;Khan, M.I.
    • Journal of Microbiology and Biotechnology
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    • v.19 no.11
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    • pp.1342-1347
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    • 2009
  • We have isolated endophytic fungi from the Indian yew tree, Taxus baccata, and then screened for taxol production. Out of the 40 fungal cultures screened, one fungus Gliocladium sp. was found to produce taxol and 10-DABIII (10-deacetyl baccatin III). These compounds were purified by TLC and HPLC and characterized using UV-spectroscopy, ESI-MS, MS/MS, and proton NMR. One liter of Gliocladium sp. culture yielded $10\;{\mu}g$ of taxol and $65\;{\mu}g$ of 10-DABIII. The purified taxol from the fungus showed cytotoxicity towards cancer lines HL-60 (leukemia), A431 (epidermal carcinoma), and MCF-7 (breast cancer).

Polyphenolic Profile of Fermented Houttuynia cordata Thunb. and Overall Contribution to Antioxidant and Lipolytic Activities

  • Lee, Soo Jung;Hu, Wensi;Lee, Eun Jung;Choi, Jin Young;Koo, Ok Kyung
    • Food Engineering Progress
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    • v.22 no.4
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    • pp.295-303
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    • 2018
  • Polyphenol profiles, physicochemical properties, antioxidant activities, and inhibitory effect of adipocyte differentiation of Houttuynia cordata fermented with Lactobacillus brevis B84 were evaluated. Six polyphenols were characterized for this plant by using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), and the results were compared with total phenolic content by a spectrophotometric method. The total amount of the identified polyphenols was lower than that determined by the spectrophotometric method. However, the fermentation process influenced polyphenol composition such as content of vanillic acid and caffeic acid. The phytochemical profiles were evaluated by high-performance liquid chromatography with UV and electrospray ionization mass spectrometry detection ($HPLC-DAD-ESI-MS^n$). Total sugar and reducing sugar contents decreased after fermentation. Antioxidant activities such as DPPH, ABTS, and superoxide anion radical scavenging and reducing power were evaluated to compare the beneficial effect after fermentation. Fermented H. cordata increased the lipolytic effect in 3T3-L1 adipocytes. Overall, the results indicate that the fermentation of H. cordata with L. brevis B84 produces changes of phenolic compounds, antioxidant activity, and lipolytic effect.

Analysis of Peanut and Peanut Butter Retailed in Korea for Aflatoxin $B_1$ (국내 시판 땅콩 및 땅콩버터 중 Aflatoxin $B_1$ 오염 분석)

  • Park, Je-Won
    • Korean Journal of Food Science and Technology
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    • v.38 no.2
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    • pp.309-312
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    • 2006
  • Aflatoxin $B_1$ in 70 retail samples, including 40 food-grade peanut (28 domestic, 12 imported) and 30 peanut butter (12 domestic, 18 imported) samples, was analyzed by high-performance liquid chromatography (HPLC) with fluorescence detection (FD), and positive samples were confirmed using HPLC with mass spectrometry (MS). Recoveries of aflatoxin $B_1$ spiked at 2 ppb exceeded 80% in both commodities. Detection limits for aflatoxin $B_1$ by HPLC-FD and MS analysis were 0.8 and 0.1 ppb, respectively. Four domestic and six imported peanut samples contained detectable levels of aflatoxin $B_1$ with means of 19 and 32 ppb, respectively. Aflatoxin $B_1$ was found in two domestic and three imported peanut butter samples with mean aflatoxin $B_1$ of 10 and 12 ppb, respectively. Peanut commodity showed more frequent aflatoxin $B_1$ contamination compared to its processed peanut butter product, and levels of aflatoxin $B_1$, especially in imported peanuts, were significantly (p<0.05) higher than those of other commodities. These results suggest peanut and peanut butter are not major contributors to dietary intake of aflatoxin $B_1$ in South Korea.

Analysis of Fluoroquinolone Antibiotics in Foods (식품 중 플루오로퀴놀론계 항생제의 분석)

  • Kim, Hee-Yun;Shin, Min-Su;Choi, Hee-Ju;Park, Se-Jong;Song, Jae-Sang;Cheong, So-Young;Choi, Sun-Hee;Lee, Hwa-Jeong;Kim, Young-Seon;Choi, Jae-Chun
    • Korean Journal of Food Science and Technology
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    • v.41 no.6
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    • pp.636-643
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    • 2009
  • Residual fluoroquinolone levels in animal foods retailed in Korea were monitored according to the method outlined in Korea Food Code using HPLC-FLD and HPLC-ESI-MS/MS for confirmation. The optimum ion transitions were $360{\rightarrow}316$, 342 m/z for enrofloxacin, $332{\rightarrow}314$, 288 m/z for ciprofloxacin, $320{\rightarrow}301$, 230 m/z for norfloxacin, $334{\rightarrow}315$, 290 m/z for pefloxacin, $362{\rightarrow}318$, 261, 334 m/z for ofloxacin, and $262{\rightarrow}201$, 126 m/z for flumequin. Enrofloxacin, ciprofloxacin and norfloxacin residues were found in 12 out of 388 samples. These antibiotics were only found in chicken samples, while no residues were found in beef, pork, milk and egg samples. Using this monitoring method, detection rates of 3.1, 1.3, and 0.3% were obtained for enrofloxacin, ciprofloxacin and norfloxacin, respectively. The levels of enrofloxacin, ciprofloxacin and norfloxacin detected in food samples ranged from 0.01 to 0.73 mg/kg in 12 samples, 0.01-0.03 mg/kg in 5 samples, and 0.12 mg/kg in only a sample, respectively.

Simultaneous Analysis of Bioactive Metabolites from Rehmannia glutinosa by HPLC-DAD-MS/MS

  • Won, Tae-Hyung;Ryu, Sung-Kwang;Kang, Sam-Sik;Shin, Jong-Heon
    • Natural Product Sciences
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    • v.16 no.2
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    • pp.116-122
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    • 2010
  • A high-performance liquid chromatography (HPLC) with diode array detector (DAD) and electrospray ionization mass spectrometry (ESI-MS) was established for the simultaneous determination of five representative metabolites of the iridoid and phenolic classes from Rehmannia glutinosa. The optimal chromatographic conditions were obtained on an ODS column (5 mm, $4.6{\times}250\;mm$) with the column temperature at $25^{\circ}C$. The mobile phase was composed of water and acetonitrile using a gradient elution with the flow rate 0.3 mL/min. Detection wavelength was set at 205 nm. All calibration curves showed good linear regression ($r^2$ > 0.997) within test ranges. Limits of detection (LOD) and quantitation (LOQ) values were lower than 0.123 and $0.373\;{\mu}g/mL$, respectively. The developed method provided satisfactory precision and accuracy with overall intra-day and inter-day variations of 0.09 - 0.76% and 0.16 - 1.41%, respectively, and the overall recoveries of 99.03 - 102.67% for all of the compounds analyzed. In addition, effectiveness of diverse extraction methods was compared to each other for the development of standard analytic method. The verified method was successfully applied to the quantitative determination of five representative metabolites in twenty-one commercial Rehmannia glutinosa samples from different markets in Korea and China. The analytical results showed that the contents of the five analytes vary significantly with sources.

Determination of Veterinary Antibiotic Residues: IV. Comparable Analytical Methods with EPA Methods 1694_A Review (시료 중 잔류 항생제 분석 방법: IV. EPA method 1694와 비교 가능한 기기 분석 방법)

  • Kim, Chansik;Ryu, Hong-Duck;Chung, Eu Gene;Kim, Yongseok;Rhew, Doug Hee
    • Journal of Korean Society on Water Environment
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    • v.32 no.6
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    • pp.670-699
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    • 2016
  • In this study, 16 antibiotics were selected from among the top 30 veterinary antibiotics sold in South Korea in 2014, as well as from among the pharmaceuticals targeted by EPA method 1694, in order to review analytical methods for the detection of trace levels of antibiotics in environmental samples: surface water, soils, animal origin foods, and manures. LC-MS/MS was heavily used. In the chromatography for the detection of the selected antibiotics, the $C_{18}$ column was mostly used at the temperature of $30{\sim}40^{\circ}C$. Water and methanol/acetonitrile were commonly chosen as a nonpolar and a polar mobile phase, respectively. Gradient elution was applied to separate multiclass antibiotics. Volatile additives, such as formic acid, acetic acid, and ammonium acetate were mixed with the mobile phase to improve the ionization efficiency of analytes and the sensitivity in MS detection. Electrospray ionization (ESI) was widely used in the LC-MS/MS and positive ionization was preferred to determine the selected antibiotics. A protonated $[M+H]^+$ molecule was selected as a precursor ion, and its two transitions were analyzed, one for quantitative measurement and the other for confirmation. This study reviewed linearity of the calibration curve, recovery, repeatability, method detection limits (MDLs), and method quantification limits (MQLs) for each target compound used to validate the developed analytical methods.

Qualitative Analysis of the Major Constituents in Traditional Oriental Prescription Bang-poong-tong-sung-san by Liquid Chromatography/Ultraviolet Detector/Ion-Trap Time-of-Flight Mass Spectrometry

  • Eom, Han Young;Kim, Hyung-Seung;Han, Sang Beom
    • Mass Spectrometry Letters
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    • v.5 no.1
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    • pp.24-29
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    • 2014
  • An advanced and reliable high performance liquid chromatography (HPLC)/ultraviolet detector (UV)/ion-trap time-of-flight (IT-TOF) mass spectrometry was developed for the simultaneous quantification of 19 marker compounds in Bang-poong-tong-sung-san (BPTS), a traditional oriental prescription. Various parameters affecting HPLC separation and IT-TOF detection were investigated, and optimized conditions were identified. The separation was achieved on a Capcell PAK C18 column ($1.5mm{\times}250mm$, $5{\mu}m$ particle size) using a gradient elution of acetonitrile and water containing 0.1% formic acid at a flow rate of 0.1 mL/min. The column temperature was maintained at $40^{\circ}C$ and the injection volume was $2{\mu}L$. IT-TOF system was equipped with an electrospray ion source (ESI) operating in positive or negative ion mode. The optimized electrospray ionization parameters were as follows: ion spray voltage, +4.5 kV (positive ion mode), or -3.5 kV (negative ion mode); drying gas ($N_2$), 1.5 L/min; heat block temperature, $200^{\circ}C$. Automatic $MS^n$ (n = 1~3) analyses were carried out to obtain structural information of analytes. Elemental compositions and their mass errors were calculated based on their accurate masses obtained from a formula predictor software. The marker compounds in BPTS were identified by comparisons between $MS^n$ spectra from standards and those from extracts. Moreover, the libraries of $MS^2$ and $MS^3$ spectra and accurate masses of parent and fragment ions for marker compounds were constructed. The developed method was successfully applied to the BPTS extracts and identified 17 out of 19 marker compounds in the BPTS extracts.

Dibenzocyclooctene lignan compounds isolated from the fruits of Schisandra chinensis Baill

  • Piao, LongZhu;Lee, Yu-Joung;PhamPhu, ThanhTruc;Shin, Jae-Kyoon
    • Natural Product Sciences
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    • v.11 no.4
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    • pp.248-252
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    • 2005
  • Schisandra chinensis Baill. (Shisandraceae) is used commonly in traditional Korean herbal medicine. One new dibenzocyclooctene lignan compound, benzoylgomisin Q, was found, for the first time, in the fruits of Schisandra chinensis Baill., together with five known lignan compounds; schisandrin, gomisin A, gomisin G, deoxyschisandrin, and gomisin N, using an ESI-positive MS-preparative HPLC auto-purification system. The structures of these compounds were determined via UV, MS, and NMR spectral studies.

Biotransformation of Flavone by CYP105P2 from Streptomyces peucetius

  • Niraula, Narayan Prasad;Bhattarai, Saurabh;Lee, Na-Rae;Sohng, Jae Kyung;Oh, Tae-Jin
    • Journal of Microbiology and Biotechnology
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    • v.22 no.8
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    • pp.1059-1065
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    • 2012
  • Biocatalytic transfer of oxygen in isolated cytochrome P450 or whole microbial cells is an elegant and efficient way to achieve selective hydroxylation. Cytochrome P450 CYP105P2 was isolated from Streptomyces peucetius that showed a high degree of amino acid identity with hydroxylases. Previously performed homology modeling, and subsequent docking of the model with flavone, displayed a reasonable docked structure. Therefore, in this study, in a pursuit to hydroxylate the flavone ring, CYP105P2 was co-expressed in a two-vector system with putidaredoxin reductase (camA) and putidaredoxin (camB) from Pseudomonas putida for efficient electron transport. HPLC analysis of the isolated product, together with LC-MS analysis, showed a monohydroxylated flavone, which was further established by subsequent ESI/MS-MS. A successful 10.35% yield was achieved with the whole-cell bioconversion reaction in Escherichia coli. We verified that CYP105P2 is a potential bacterial hydroxylase.