• Journal of Microbiology and Biotechnology
• /
• 제31권4호
• /
• pp.540-549
• /
• 2021

The Wnt/β-catenin signaling pathway is involved in breast cancer and Myxococcus fulvus KYC4048 is a myxobacterial strain that can produce a variety of bioactive secondary metabolites. Although a previous study revealed that KYC4048 metabolites exhibit anti-proliferative effects on breast cancer, the biochemical mechanism involved in their effects remains unclear. In the present study, KYC4048 metabolites were separated into polar and non-polar (ethyl acetate and n-hexane) fractions via liquid-liquid extraction. The effects of these polar and non-polar KYC4048 metabolites on the viability of breast cancer cells were then determined by MTT assay. Expression levels of Wnt/β-catenin pathway proteins were determined by Western blot analysis. Cell cycle and apoptosis were measured via fluorescence-activated cell sorting (FACS). The results revealed that non-polar KYC4048 metabolites induced cell death of breast cancer cells and decreased expression levels of WNT2B, β-catenin, and Wnt target genes (c-Myc and cyclin D1). Moreover, the n-hexane fraction of non-polar KYC4048 metabolites was found most effective in inducing apoptosis, necrosis, and cell cycle arrest, leading us to conclude that it can induce apoptosis of breast cancer cells through the Wnt/β-catenin pathway. These findings provide evidence that the n-hexane fraction of non-polar KYC4048 metabolites can be developed as a potential therapeutic agent for breast cancer via inhibition of the Wnt/β-catenin pathway.

• Mass Spectrometry Letters
• /
• 제11권2호
• /
• pp.36-40
• /
• 2020

Untargeted metabolomics is a useful tool for drug development focusing on novel chemotherapeutic and chemopreventative agents against cancer cells. In recent years, quadrupole time of flight liquid chromatography-mass spectrometry (Q-TOF LC/MS)-based untargeted metabolomic approaches have gained importance to evaluate the effect of these agents at the molecular level. The researchers working on cell culture studies still do not apply standardized methodologies on sample preparation for untargeted metabolomics approaches. In this study, the rough and wet lysis techniques performed on MCF-7 breast cancer cells were compared with each other via the Q-TOF LC/MS-based metabolomic approach. The C18 and hydrophilic interaction liquid chromatography (HILIC) columns were used for the separation of the metabolites in MCF-7 cell lysates. 505 peaks were detected through the HILIC column and 551 peaks were found through the C18 column for the wet lysis technique. This situation supported by the base peak chromatograms showed that the wet lysis technique allowed us to extract higher number of non-polar metabolites. Almost equal number of metabolites was found for the C18 and HILIC columns (697 peaks for the HILIC column and 695 peaks for the C18 column) when the rough lysis technique was used. However, the intensities of polar metabolites were higher for the rough lysis technique on base peak chromatograms for both the HILIC and C18 columns. Although cell lysis technique, which is the first step in the sample preparation for cell culture studies, does not cause dramatic differences in the number of the detected metabolite peaks, it affects the polar and non-polar metabolite ratio significantly. Therefore, it must be considered carefully especially for in vitro drug development studies.

• 한국축산식품학회지
• /
• 제39권1호
• /
• pp.1-12
• /
• 2019

The objective of this study was to establish an optimized 1D $^1H$ quantitative nuclear magnetic resonance (qNMR) analytical method for analyzing polar metabolites in meat. Three extraction solutions [0.6 M perchloric acid, 10 mM phosphate buffer, water/methanol (1:1)], three reconstitution buffers [20 mM 3-morpholinopropane-1-sulfonic acid, 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid, phosphate buffer], and two pulse programs (zg30, noesypr1d) were evaluated. Extraction with 0.6 M perchloric acid and 20 mM phosphate resulted in a stable baseline and no additional overlap for quantifying polar metabolites in chicken breast. In qNMR analysis, zg30 pulse program (without water-suppression) showed smaller relative standard deviation (RSD) and faster running time than noesypr1d (water-suppression). High-performance liquid chromatography was compared with qNMR analyses to validate accuracy. The zg30 pulse program showed good accuracy and lower RSD. The optimized qNMR method was able to apply for beef and pork samples. Thus, an optimized 1D $^1H$ qNMR method for meat metabolomics was established.

• Journal of Microbiology
• /
• 제39권2호
• /
• pp.133-138
• /
• 2001

We investigated the antagonism of indigenous bacteria isolated from stressed mussels and their extracellular metabolites on the adult zebra mussel, Dreissena polymorpha. Selective bacterial isolates including Aeromonas media, A. salmonicida, A. veronii, and Shewanella putrefaciens, showed strong lethality against adult mussels and 100% mortality was observed within 5 days of incubation. Bacterial metabolites, fractionated and concentrated from stationary-phase culture supernatants of these bacterial isolates, displayed varying degrees of antagonistic effects on zebra mussels. Among the three size fractions examined, <5, 5-10, and >10 kDa, the mast lethal fraction seems to be >10 kDa for three of the four isolates tested. Further chemical analyses of these size fractions revealed that the predominant constituents were polysaccharides and proteins. No 2-keto-3-deoxyoctanoic acid (2-KDO), deoxyribonucleic acids (DNA) or uranic acid were detectable. Extraction of supernatants of two antagonistic isolates with polar solvent suggested that polar molecules are present in the active fraction. Our data suggest that extracellular metabolites produced by antagonistic bacteria are also involved in disease development in zebra mussels and elucidation of the mechanisms involved may offer a novel strategy for control of biofouling invertebrates.

• Journal of Applied Biological Chemistry
• /
• 제56권2호
• /
• pp.109-112
• /
• 2013

Azole fungicides are one of the most wide-spread antifungal compounds in agriculture and pharmaceutical applications. Their major mode of action is the inhibition of ergosterol biosynthesis, giving depletion of ergosterol, precursors and abnormal steroids. However, metabolic consequences of such inhibition, other than steroidal metabolitesare not well established. Comprehensive metabolic profiles of Saccharomyces cerevisiae has been presented in this study. Wild type yeast was treated either with glucose as control or azole fungicide (ketoconazole). Both polar metabolites and lipids were analyzed with gas chromatography-mass spectrometry. Approximately over 180 metabolites were characterized, among which 18 of them were accumulated or depleted by fungicide treatment. Steroid profile gives the most prominent differences, including the accumulation of lanosterol and the depletion of zymosterol and ergosterol. However, the polar metabolite profile was also highly different in pesticide treatment. The concentration of proline and its precursors, glutamate and ornithine were markedly reduced by ketoconazole. Lysine and glycine level was also decreased while the concentrations of serine and homoserine were increased. The overall metabolic profile indicates that azole fungicide treatment induces the depletion of many polar metabolites, which are important in stress response.

• Journal of Microbiology and Biotechnology
• /
• 제20권4호
• /
• pp.809-816
• /
• 2010

Microbial biotransformations can be used to predict mammalian drug metabolism. The present investigation deals with microbial biotransformation of valdecoxib using microbial cultures. Thirty-nine bacterial, fungal, and yeast cultures were used to elucidate the biotransformation pathway of valdecoxib. A number of microorganisms metabolized valdecoxib to various levels to yield nine metabolites, which were identified by HPLC-DAD and LC-MS-MS analyses. HPLC analysis of biotransformed products indicated that a majority of the metabolites are more polar than the substrate valdecoxib. Basing on LC-MS-MS analysis, the major metabolite was identified as a hydroxymethyl metabolite of valdecoxib, whereas the remaining metabolites were produced by carboxylation, demethylation, ring hydroxylation, N-acetylation, or a combination of these reactions. The hydroxymethyl and carboxylic acid metabolites were known to be produced in metabolism by mammals. From the results, it can be concluded that microbial cultures, particularly fungi, can be used to predict mammalian drug metabolism.

• 생약학회지
• /
• 제50권3호
• /
• pp.232-238
• /
• 2019

Phytochemical study of Ganoderma lucidum (Polyporaceae) let to the isolation of five lanostane triterpenoid (1-5), along with two nitrogen derived phenolic compounds, N-phenylethylformamide (6) and N-acetylphenethylamine (7). The structures of the compounds were determined by 1D and 2D NMR, and MS experiments, as well as by comparison of their data with published values. Compounds 6 and 7 were isolated for the first time from the genus Ganoderma and this species. All the compounds were evaluated for cancer chemopreventive potential based on their ability to inhibit nitric oxide (NO) production induced by lipopolysaccharide (LPS) in mouse macrophage RAW 264.7 cells in vitro. Among the isolates, compounds 2 and 3 showed moderate inhibitory activity against NO production.

• Journal of Ginseng Research
• /
• 제13권1호
• /
• pp.37-41
• /
• 1989

Benzo(a)pyrene(BP)의 monooxygenase(AHH)에 의해서 생성된 반응성 대사물질들의 in vitro DNA와의 결합 및 BP 대사에 관여하는 효소들의 활성도에 미치는 인삼 물추출물의 영향을 조사하였으며, DNA-BP metabolite adduct들은 Sephadex LH-20 column으로 chromatography하여 5개의 major peak 들을 얻었다. 이 peak 들을 극성이 큰 순서대로 A부터 E까지 임의로 정하고 5개의 peak들을 7,8-diol-9,10-oxide(A), 7,8-oxide(B). 4,5-oxide(C), 9-HO-BP(D & E) adduct들로 잠정적으로 확인하였다. Peak A, C, D 그리고 E는 각각 대조군의 30, 15, 20 그리고 30%로 감소되었으며 peak B는 의미있는 변화를 보이지 않았다. DNA-BP 결합 억제와 관련하여 in vitro와 in vivo 투여시의 경향이 유사하여 EH의 활성도만 BP투여 대조군보다 38%정도 의미있게 유도되었다.

• 한국토양비료학회지
• /
• 제45권6호
• /
• pp.936-942
• /
• 2012

This review was to elucidate the fate of Bentazon(3-isopropyl-1H-2,1,3-benzothiadiazin-4(3H)-one-2,2-dioxide) and its metabolites in soil. Bentazon is rapidly degraded to form polar metabolites which are mostly adsorbed to soil components, such as humin or fulvic acid, as non extractable forms and mineralized into $CO_2$ by light or micro-organisms in both aerobic or nonaerobic condition. The degradation of Bentazon is dependent on the rate of organic matters in soil and the use of land for the tillage. The degradation rate is decreased as the amount of organic matters in soil increases and if the land is under use for tillage. Sorption and mobility of Bentazon depends on soil pH and the content of organic matters in soil. Usually, the sorption of the metabolites of Bentazon is decreased with increase in the mobility and pH. Almost all of Bentazon is degraded within rhizosphere or forms conjugate bonds with soil organic matters before it reaches to the ground water.

• Journal of Microbiology and Biotechnology
• /
• 제29권7호
• /
• pp.1144-1154
• /
• 2019

There have been several studies regarding lichen-associated bacteria obtained from diverse environments. Our screening process identified 49 bacterial species in two lichens from the Himalayas: 17 species of Actinobacteria, 19 species of Firmicutes, and 13 species of Proteobacteria. We discovered five types of strong antimicrobial agent-producing bacteria. Although some strains exhibited weak antimicrobial activity, NP088, NP131, NP132, NP134, and NP160 exhibited strong antimicrobial activity against all multidrug-resistant strains. Polyketide synthase (PKS) fingerprinting revealed results for 69 of 148 strains; these had similar genes, such as fatty acid-related PKS, adenylation domain genes, PfaA, and PksD. Although the association between antimicrobial activity and the PKS fingerprinting results is poorly resolved, NP160 had six types of PKS fingerprinting genes, as well as strong antimicrobial activity. Therefore, we sequenced the draft genome of strain NP160, and predicted its secondary metabolism using antiSMASH version 4.2. NP160 had 46 clusters and was predicted to produce similar secondary metabolites with similarities of 5-100%. Although NP160 had 100% similarity with the alkylresorcinol biosynthetic gene cluster, our results showed low similarity with existing members of this biosynthetic gene cluster, and most have not yet been revealed. In conclusion, we expect that lichen-associated bacteria from the Himalayas can produce new secondary metabolites, and we found several secondary metabolite-related biosynthetic gene clusters to support this hypothesis.