• Title/Summary/Keyword: LC-MS/MS analysis

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Analysis of Nutrient Content by Digestion Phase of Legumes using an In Vitro Digestion Model (In Vitro Digestion Model을 활용한 두류 소화 단계별 영양성분 변화 분석)

  • Da Bin Lee;Kyeong A Jang;In Seon Hwang;Min Sook Kang;Mi-Kyung Seo;Haeng Ran Kim;Seon Mi Yoo
    • The Korean Journal of Food And Nutrition
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    • v.36 no.5
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    • pp.368-378
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    • 2023
  • Changes in contents of free sugars, amino acids, and fatty acids of legumes were analyzed for each phase of in vitro digestion. In addition, contents of resistant starch in raw and digested pulses were compared. Soybeans, kidney beans, cowpeas, and chickpeas were analyzed. An in vitro digestion model was used to analyze contents of nutrients using LC-MS and GC-MS. Stachyose in kidneybean, cowpea, and chickpea increased as the digestion phase progressed. In four types of legumes, raffinose slightly decreased or showed no significant difference between the Oral phase and the BBMV phase. Content of glucose, a monosaccharide, increased during the BBMV phase. During the digestion phase, levels of free amino acids and free fatty acids also increased. Content of resistant starch was reduced compared to that in the raw material. It was 0.01g/100 g food in soybean, 1.06 g/100 g food in red kidney bean, 0.77g/ 100g food in cowpea, and 0.76 g/100 g food in chickpea. It was confirmed that nutrients in the in vitro digestion model were liberated at each digestion phase with changes in the content of resistant starch. These results are expected to be used as fundamental data for obtaining bioavailability of nutrients.

Identification of an ISR-Related Metabolite Produced by Pseudomonas chlororaphis O6 against the Wildfire Pathogen Pseudomonas syringae pv. tabaci in Tobacco

  • Park, Myung-Ryeol;Kim, Young-Cheol;Park, Ju-Yeon;Han, Song-Hee;Kim, Kil-Yong;Lee, Sun-Woo;Kim, In-Seon
    • Journal of Microbiology and Biotechnology
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    • v.18 no.10
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    • pp.1659-1662
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    • 2008
  • Pseudomonas chlororaphis O6 exhibits induced systemic resistance (ISR) against P. syringae pv. tabaci in tobacco. To identify one of the ISR metabolites, O6 cultures were extracted with organic solvents, and the organic extracts were subjected to column chromatography followed by spectroscopy analyses. The ISR bioassay-guided fractionation was carried out for isolation of the metabolite. High-resolution mass spectrometric analysis of the metabolite found $C_{9}H_{9}O_{3}N$ with an exact mass of 179.0582. LC/MS analysis in positive mode showed an $(M+H)^{+}$ peak at m/z 180. Nuclear magnetic resonance ($^{1}H,\;^{13}C$) analyses identified all protons and carbons of the metabolite. Based on the spectroscopy data, the metabolite was identified as 4-(aminocarbonyl) phenylacetate (4-ACPA). 4-ACPA applied at 68.0 mM exhibited ISR activity at a level similar to 1.0 mM salicylic acid. This is the first report to identify an ISR metabolite produced by P. chlororaphis O6 against the wildfire pathogen P. syringae pv. tabaci in tobacco.

Genomics and LC-MS Reveal Diverse Active Secondary Metabolites in Bacillus amyloliquefaciens WS-8

  • Liu, Hongwei;Wang, Yana;Yang, Qingxia;Zhao, Wenya;Cui, Liting;Wang, Buqing;Zhang, Liping;Cheng, Huicai;Song, Shuishan;Zhang, Liping
    • Journal of Microbiology and Biotechnology
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    • v.30 no.3
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    • pp.417-426
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    • 2020
  • Bacillus amyloliquefaciens is an important plant disease-preventing and growth-promoting microorganism. B. amyloliquefaciens WS-8 can stimulate plant growth and has strong antifungal properties. In this study, we sequenced the complete genome of B. amyloliquefaciens WS-8 by Pacific Biosciences RSII (PacBio) Single Molecule Real-Time (SMRT) sequencing. The genome consists of one chromosome (3,929,787 bp) and no additional plasmids. The main bacteriostatic substances were determined by genome, transcriptome, and mass spectrometry data. We thereby laid a theoretical foundation for the utilization of the strain. By genomic analysis, we identified 19 putative biosynthetic gene clusters for secondary metabolites, most of which are potentially involved in the biosynthesis of numerous bioactive metabolites, including difficidin, fengycin, and surfactin. Furthermore, a potential class II lanthipeptide biosynthetic gene cluster and genes that are involved in auxin biosynthesis were found. Through the analysis of transcriptome data, we found that the key bacteriostatic genes, as predicted in the genome, exhibited different levels of mRNA expression. Through metabolite isolation, purification, and exposure experiments, we found that a variety of metabolites of WS-8 exert an inhibitory effect on the necrotrophic fungus Botrytis cinerea, which causes gray mold; by mass spectrometry, we found that the main substances are mainly iturins and fengycins. Therefore, this strain has the potential to be utilized as an antifungal agent in agriculture.

High Performance Liquid Chromatographic Method for Determination of Metazosulfuron Residue in Representative Crops

  • Lee, Hyeri;Kim, Eunhye;Lee, Young Deuk;Kim, Jeong-Han
    • Korean Journal of Environmental Agriculture
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    • v.32 no.2
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    • pp.128-135
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    • 2013
  • BACKGROUND: This study was performed to develop a single residue analytical method for new herbicide metazosulfuron in crops. METHODS AND RESULTS: Brown rice, apple, mandarin, Kimchi cabbage and soybean were selected as representative crops, and clean-up system, partition solvent and extraction solvent were optimized. Instrumental limit of quantitation (ILOQ), linearity of calibration curve and method limit of quantitation (MLOQ) were determined based on the chromatography and whole procedures. For recovery tests, brown rice, apple, mandarin, Kimchi cabbage and soybean samples were macerated and fortified with metazosulfuron standard solution at three levels (MLOQ, 10 MLOQ and 100 MLOQ). And then those were extracted with acetonitrile, concentrated, and partitioned with ethyl acetate. Then the extracts were concentrated again and cleaned-up through $NH_2$ (aminopropyl) SPE cartridge with acetone : dichloromethane (1% acetic acid) (20 : 80, v/v) before concentration and analysis with HPLC. CONCLUSION(S): ILOQ of metazosulfuron was 2 ng (S/N${\geq}$10) and good linearity was achieved between 0.05 and 12.5 mg/Kg of metazosulfuron standard solutions, with coefficients of determination of 0.9999. MLOQ was 0.02 mg/Kg. Good recoveries from 74.1 to 116.9% with coefficients of variation (C.V.) of less than 10% were obtained, regardless of sample type, which satisfies the criteria of Korea Food and Drug Administration (KFDA). Those results were reconfirmed with LC-MS (SIM). The method established in this study is simple, economic and efficient to be applied to most of crops as an official and general method for residue analysis of metazosulfuron.

Pyrophen Produced by Endophytic Fungi Aspergillus sp Isolated from Piper crocatum Ruiz & Pav Exhibits Cytotoxic Activity and Induces S Phase Arrest in T47D Breast Cancer Cells

  • Astuti, Puji;Erden, Willy;Wahyono, Wahyono;Wahyuono, Subagus;Hertiani, Triana
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.2
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    • pp.615-618
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    • 2016
  • Ethyl acetate extracts obtained from culture of endophytic fungi Aspergillus sp isolated from Piper crocatum Ruiz & Pav, have been shown to possess cytotoxic activity against T47D breast cancer cells. Investigations were here conducted to determine bioactive compounds responsible for the activity. Bioassay guided fractionation was employed to obtain active compounds. Structure elucidation was performed based on analysis of LC-MS, $^1H$-NMR, $^{13}C$-NMR, COSY, DEPT, HMQC, HMBC data. Cytotoxity assays were conducted in 96 well plates against T47D and Vero cell lines. Bioassay guided isolation and chemical investigation led to the isolation of pyrophen, a 4-methoxy-6-(1'-acetamido-2'-phenylethyl)-2H-pyran-2-one. Further analysis of its activity against T47D and Vero cells showed an ability to inhibit the growth of T47D cells with IC50 values of $9.2{\mu}g/mL$ but less cytotoxicity to Vero cells with an $IC_{50}$ of $109{\mu}g/mL$. This compound at a concentration of 400 ng/mL induced S-phase arrest in T47D cells.

Establishment and Selection of Indicator Materials for Cervi Parvum Cornu Pharmacopuncture

  • Yang, Kyu-Jin;Lee, Ki-Beom;Kim, No-Hyeon;Kim, Tae-Gyu;Gi, Yu-Mi;Joo, Hwan-Soo;Suh, Chang-Yong;Lee, In-Hee;Chung, Hwa-Jin;Ha, In-Hyuk;Lee, Jae-Woong
    • Journal of Acupuncture Research
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    • v.34 no.4
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    • pp.180-184
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    • 2017
  • Background: Recently, Cervi Parvum Cornu pharmacopuncture has been widely used. But no studies on the indicator materials for Cervi Parvum Cornu pharmacopuncture have been conducted. The aim of this study was to select indicator materials that would aid in the uniform preparation of standardized Cervi Parvum Cornu pharmacopuncture. Methods: Three lots of Cervi Parvum Cornu pharmacopuncture were analysed. Each lot was prepared using the same methods and materials. Chondroitin sulfate, alanine, and leucine were selected as the indicator materials for Cervi Parvum Cornu. For standardization, chondroitin sulfate analysis was performed using the colorimetric method, while alanine and leucine were analyzed using liquid chromatography-mass spectrometry (LC-MS). Results: Analysis of the three lots of Cervi Parvum Cornu pharmacopuncture found chondroitin sulfate levels of $108.9{\pm}17.3ug/ml$, $118.8{\pm}5.0ug/ml$ and $112.3{\pm}11.9ug/ml$. Alanine levels were $44.9{\pm}2.8ug/ml$, $44.6{\pm}0.3ug/ml$, and $43.9{\pm}0.2ug/ml$. Leucine levels were $29.6{\pm}0.7ug/ml$, $29.0{\pm}0.1ug/ml$, and $29.4{\pm}0.1ug/ml$. Conclusion: These results suggest that chondroitin sulfate, alanine, and leucine may be useful for the standardization of Cervi Parvum Cornu pharmacopuncture.

Effect of medis composition on the Cordycepin and content Nutritional Components of Cordyceps militaris (배지 조성이 번데기 동충하초의 영양성분 및 Cordycepin 함량에 미치는 영향)

  • Cho, Soo-Muk;Park, Hong-Ju;Seo, Geon-Sik;Hong, Jong-Deok
    • The Korean Journal of Mycology
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    • v.37 no.2
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    • pp.161-166
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    • 2009
  • The purpose of the present study is to develop a simple, fast and sensitive LC/MS method for simultaneous separation and the determination of an active component in the oriental medicinal mushroom Cordyceps militaris. Based on this work, the contents of cordycepin in Cordyceps militaris fruiting cultivated on various media were determined and compared. And also, the nutritional components such as minerals and vitamins were determined in order to provide useful information to consumer as a food material. The analysis methods of nutritional components were chosen on the basis of AOAC. The optimum separation for cordycepin was achieved using a solvent gradient consisting of the mixture of 0.1% formic acid in methanol (solvent B) in a background of 0.1% formic acid in water (solvent A) as a mobile phase and a 3.0${\times}$150 Waters XTera column. Selective ion monitoring (SIR) mode ([M+H]+ at m/z 252) was used for quantitative analysis of cordycepin. The cultivated Cordyceps militaris on various media contained 1~14 /g of cordycepin, 0.65~1.08% of thiamine, 0.86~7.17% of riboflavin, and 3.01~5.26% of niacin. The content of mineral components varied on categories, especially contained 500~3500% of potassium as a major mineral. Cordycepin, niacin and potassium were found much higher in the fruiting cultivated with soy power media (gold 10) than other media.

Microbiota, co-metabolites, and network pharmacology reveal the alteration of the ginsenoside fraction on inflammatory bowel disease

  • Dandan Wang;Mingkun Guo;Xiangyan Li;Daqing Zhao;Mingxing Wang
    • Journal of Ginseng Research
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    • v.47 no.1
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    • pp.54-64
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    • 2023
  • Background: Panax ginseng Meyer (P. ginseng) is a traditional natural/herbal medicine. The amelioration on inflammatory bowel disease (IBD) activity rely mainly on its main active ingredients that are referred to as ginsenosides. However, the current literature on gut microbiota, gut microbiota-host co-metabolites, and systems pharmacology has no studies investigating the effects of ginsenoside on IBD. Methods: The present study was aimed to investigate the role of ginsenosides and the possible underlying mechanisms in the treatment of IBD in an acetic acid-induced rat model by integrating metagenomics, metabolomics, and complex biological networks analysis. In the study ten ginsenosides in the ginsenoside fraction (GS) were identified using Q-Orbitrap LC-MS. Results: The results demonstrated the improvement effect of GS on IBD and the regulation effect of ginsenosides on gut microbiota and its co-metabolites. It was revealed that 7 endogenous metabolites, including acetic acid, butyric acid, citric acid, tryptophan, histidine, alanine, and glutathione, could be utilized as significant biomarkers of GS in the treatment of IBD. Furthermore, the biological network studies revealed EGFR, STAT3, and AKT1, which belong mainly to the glycolysis and pentose phosphate pathways, as the potential targets for GS for intervening in IBD. Conclusion: These findings indicated that the combination of genomics, metabolomics, and biological network analysis could assist in elucidating the possible mechanism underlying the role of ginsenosides in alleviating inflammatory bowel disease and thereby reveal the pathological process of ginsenosides in IBD treatment through the regulation of the disordered host-flora co-metabolism pathway.

A Plant Metabolomic Approach to Identify the Difference of the Seeds and Flowers Extracts of Carthamus tinctorius L.

  • Ozan Kaplan;Nagehan Saltan;Arzu Kose;Yavuz Bulent Kose;Mustafa Celebier
    • Mass Spectrometry Letters
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    • v.14 no.2
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    • pp.42-47
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    • 2023
  • Carthamus tinctorius L. (known as safflower) is a valuable oil plant whose importance is increasing rapidly in the world due to its high adaptation to arid regions. The seeds of this unique plant are especially used in edible oil, soap, paint, varnish and lacquer production. Its flowers are used in vegetable dye production and medicinal purposes beside its features as a coloring and flavoring in food. After the oil is removed, the remaining pulp and plant parts are used as animal feed, and dry straw residues are used as fuel. Beside all these features, its usage as a herbal medicinal plants for various diseases has gained importance on recent years. In this study, it was designed a plant metabolomic approach which transfers all the recent data processing strategies of untargeted metabolomics in clinical applications to the present study. Q-TOF LC/MS-based analysis of the extracts (70% ethanol, hexane, and chloroform) for both seed and flowers was performed using a C18 column (Agilent Zorbax 1.8 µM, 100 × 2.1 mm). Differences were observed in seed and fruit extracts and these differences were visualized using principal component analysis (PCA) plots. The total number and intersections of the peaks in the extracts were visualized using peak count comparison graph. Based on the experimental results, the number of the detected peaks for seeds was higher than the ones for the flowers for all solvent systems to extract the samples.

In vitro anti-Trypanosoma cruzi activity of methanolic extract of Bidens pilosa and identification of active compounds by gas chromatography-mass spectrometry analysis

  • Gabriel Enrique Cazares-Jaramillo;Zinnia Judith Molina-Garza;Itza Eloisa Luna-Cruz;Luisa Yolanda Solis-Soto;Jose Luis Rosales-Encina;Lucio Galaviz-Silva
    • Parasites, Hosts and Diseases
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    • v.61 no.4
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    • pp.405-417
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    • 2023
  • Chagas disease, caused by Trypanosoma cruzi parasite, is a significant but neglected tropical public health issue in Latin America due to the diversity of its genotypes and pathogenic profiles. This complexity is compounded by the adverse effects of current treatments, underscoring the need for new therapeutic options that employ medicinal plant extracts without negative side effects. Our research aimed to evaluate the trypanocidal activity of Bidens pilosa fractions against epimastigote and trypomastigote stages of T. cruzi, specifically targeting the Brener and Nuevo León strains-the latter isolated from Triatoma gerstaeckeri in General Terán, Nuevo León, México. We processed the plant's aerial parts (stems, leaves, and flowers) to obtain a methanolic extract (Bp-mOH) and fractions with varying solvent polarities. These preparations inhibited more than 90% of growth at concentrations as low as 800 ㎍/ml for both parasite stages. The median lethal concentration (LC50) values for the Bp-mOH extract and its fractions were below 500 ㎍/ml. Tests for cytotoxicity using Artemia salina and Vero cells and hemolytic activity assays for the extract and its fractions yielded negative results. The methanol fraction (BPFC3MOH1) exhibited superior inhibitory activity. Its functional groups, identified as phenols, enols, alkaloids, carbohydrates, and proteins, include compounds such as 2-hydroxy-3-methylbenzaldehyde (50.9%), pentadecyl prop-2-enoate (22.1%), and linalool (15.4%). Eight compounds were identified, with a match confirmed by the National Institute of Standards and Technology (NIST-MS) software through mass spectrometry analysis.