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1.
Artigo em Inglês | MEDLINE | ID: mdl-34499596

RESUMO

A Gram-stain-negative, coccus-shaped, obligately anaerobic, non-motile and non-spore-forming bacterium, designated strain JN500902T, was isolated from the mud in a fermentation cellar used continuously over 30 years for Chinese strong-flavour baijiu production. Colonies were white, circular, convex and smooth-edged. Growth was observed at 20-40 °C (optimum, 37 °C), at pH 5.0-10 (optimum, pH 7.5), with 0-2 % (w/v) NaCl and with 0-4 % (v/v) ethanol. The Biolog assay demonstrated positive reactions of strain JN500902T in the metabolism of l-fucose and pyruvate. The predominant cellular fatty acids (>10 %) consisted of C16 : 0 and C14 : 0. The major end metabolites of strain JN500902T were acetic acid and ethanol when incubated anaerobically in liquid reinforced clostridial medium. Acetate was the major organic acid end product. The complete genome size of strain JN500902T was 3 420 321 bp with 3327 identified genes. The G+C content was 43.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences affiliated strain JN500902T with the family Lachnospiraceae, having low sequence similarity (92.8 %) to the nearest type strain, Syntrophococcus sucromutans DSM 3224T and forming a clearly distinct branch. Core genome phylogenetic analysis of the isolate and 134 strains belonging to the family Lachnospiraceae also revealed that strain JN500902T was well-separated from other genera of this family as a monophyletic clade. The average nucleotide identity and amino acid identity values between strain JN500902T and 134 Lachnospiraceae strains were less than 74 and 65 %, respectively. Considering its polyphasic characteristics, strain JN500902T represents a novel genus and species within the family Lachnospiraceae, for which the name Novisyntrophococcus fermenticellae gen. nov., sp. nov. is proposed. The type strain is JN500902T (=CICC 24502T=JCM 33939T).


Assuntos
Clostridiales/classificação , Fermentação , Filogenia , Microbiologia do Solo , Anaerobiose , Técnicas de Tipagem Bacteriana , Composição de Bases , China , Clostridiales/isolamento & purificação , DNA Bacteriano/genética , Ácidos Graxos/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
2.
Front Microbiol ; 10: 1397, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31316481

RESUMO

Butyrate, one of the key aroma compounds in Luzhou-flavor baijiu, is synthesized through two alternative pathways: butyrate kinase (buk) and butyryl-CoA: acetate CoA-transferase (but). A lack of knowledge of butyrate-producing microorganisms hinders our ability to understand the flavor formation mechanism of baijiu. Here, temporal dynamics of microbial metabolic profiling in fermented grains (FG) was explored via PICRUSt based on 16S rRNA gene sequences. We found Bacilli and Bacteroidia were the major potential butyrate producers in buk pathway at the beginning of fermentation, while later Clostridia dominated the two pathways. Clone library analysis also revealed that Clostridia (~73% OTUs) was predominant in buk pathway throughout fermentation, followed by Bacilli and Bacteroidia, and but pathway was merely possessed by Clostridia. Afterward, Clostridia-specific 16S rRNA gene sequencing demonstrated Clostridium might be the major butyrate-producing genus in two pathways, which was subsequently evaluated using culture approach. Seventeen Clostridium species were isolated from FG based on 16S rRNA gene sequence-guided medium prediction method. Profiles of short-chain fatty acids and but and buk genes in these species demonstrated phylogenetic and functional diversities of butyrate-producing Clostridium in FG. These findings add to illustrate the diversity of potential butyrate producers during brewing and provide a workflow for targeting functional microbes in complex microbial community.

3.
Biomed Pharmacother ; 116: 109007, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31170663

RESUMO

It's critical for tube formation and angiogenesis to repair ischemic myocardium or stroke. This study aimed to investigate role of microRNA-126 (miR-126) in tube formation in human umbilical vein endothelial cells (HUVECs) and associated mechanisms. Primary neural stem cells (NSCs) and HUVECs were cultured and transfected with microRNA-126 mimics and miR-126 inhibitor. Cell counting kit-8 (CCK-8) and cell cycle assay were conducted for evaluating NSCs viability. Transwell assay was conducted to observe invasive ability of HUVECs. Quantitative real-time PCR (qRT-PCR) assay was used to examine epidermal growth factor like domain 7 (EGFL7) and miR-126 mRNA both in vitro and animal models. Tube forming capability was evaluated in HUVECs. Dual luciferase assay was performed to evaluate interaction between miR-126 and EGFL7 gene. Western blot assay was used to determine phosphoinositide-3-kinase/protein kinase-B (PI3K/AKT) signaling molecules and EGFL7. The results indicated that miR-126 significantly decreased cell viability, inhibited invasive ability and modulated cell cycle of NSCs compared to miR-NC group (p < 0.05). miR-126 significantly inhibited tube formation of HUVECs compared to miR-NC group (p < 0.05). miR-126 significantly down-regulated EGFL7 mRNA and protein expression compared to miR-NC (p < 0.05). Atorvastatin significantly increased CD34 and enhanced EGFL7 expression in traumatic brain injury (TBI) rats brain tissues compared to Model group (p < 0.05). miR-126 significantly down-regulated and atorvastatin up-regulated PI3K/AKT signaling pathway (p < 0.05). Atorvastatin significantly increased EGFL7 and down-regulated miR-126 expression in TBI rats brain tissues compared to Model group (p < 0.05). miR-126 interacted with and negatively correlated with EGFL7 gene both in vitro and in TBI models. In conclusion, microRNA-126 inhibited tube formation of HUVECs by interacting with EGFL7 and down-regulating PI3K/AKT signaling pathway.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Regulação para Baixo/genética , Família de Proteínas EGF/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , MicroRNAs/metabolismo , Neovascularização Fisiológica , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Animais , Antígenos CD34/metabolismo , Atorvastatina/administração & dosagem , Atorvastatina/farmacologia , Sequência de Bases , Lesões Encefálicas Traumáticas/genética , Lesões Encefálicas Traumáticas/patologia , Proteínas de Ligação ao Cálcio/genética , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Regulação para Baixo/efeitos dos fármacos , Família de Proteínas EGF/genética , Fatores de Crescimento Endotelial/metabolismo , Humanos , MicroRNAs/genética , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/genética , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
4.
Int J Food Microbiol ; 297: 41-50, 2019 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-30878841

RESUMO

Butyrate and its derivates pertain to the key aroma contributors of strong-flavour baijiu, a kind of Chinese liquors, that is produced from grains by solid-state multispecies anaerobic fermentation in a mud cellar. Microbes inhabiting in the fermentation pit mud largely determines baijiu's flavour and quality. In order to shed light on the microbial functional groups driving butyrate production in pit mud, clone library analysis was firstly performed and the results demonstrated that Clostridia (relative abundance: 50%) and Bacilli (37%) were major groups possessing butyrate kinase (buk) pathway and Clostridia (98%) dominated butyryl-CoA:acetate CoA-transferase (but) pathway. According to Clostridial specific-16S rRNA gene sequencing analysis, we found the resilience character of Clostridial community in pit mud. Amongst Clostridial groups, 32.0% of the sequences were grouped into Clostridiales incertae sedis, followed by Heliobacteriaceae (18.3%) and Clostridiaceae 1 (8.4%). Moreover, Hydrogenispora, Sedimentibacter and Clostridium were the top three abundant genera. Relative abundance of Hydrogenispora was higher in the late days of fermentation, while Sedimentibacter exhibited higher proportion in the early days. Different from the previous studies using universal bacterial primer sets, Hydrogenispora was first reported as one dominant genus in pit mud. As for the reported potential butyrate producer Clostridium, nineteen species were obtained and ten of them were first isolated from the pit mud. Amongst them, buk was identified in eleven species by PCR analysis, while but was identified in the other seven, indicating the species-specific butyrate synthesis pathways of Clostridium. This study provides a perspective on targeting and isolating specific functional microbes in baijiu microbiota with the gene sequence-based medium prediction method.


Assuntos
Bebidas Alcoólicas , Clostridium/enzimologia , Clostridium/genética , Fermentação , Microbiologia do Solo , Bebidas Alcoólicas/microbiologia , Bacillus/enzimologia , Bacillus/genética , Butiratos/metabolismo , Microbiota/genética , RNA Ribossômico 16S/genética
5.
Int J Syst Evol Microbiol ; 69(3): 859-865, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30735112

RESUMO

A novel Gram-stain-positive, rod-shaped, obligately anaerobic, non-motile, spore-forming and binary fission encapsulated bacterium, designated strain JN500901T, was isolated from a mud cellar which has been continuously used for the fermentation of Chinese strong-flavour baijiu for over 100 years. Growth of JN500901Toccurred at pH 4.5-8.0 (optimum, pH 5.0), 20-40 °C (37 °C), 0-2 % (w/v) NaCl and 0-10 % (v/v) ethanol. The Biolog assay revealed that strain JN500901T metabolized d-fructose, l-fucose, isomaltulose and l-rhamnose among the 95 studied carbon sources. p-Cresol was the predominant volatile metabolite in the fermentation broth of strain JN500901T incubated in liquid reinforced clostridial medium under anaerobic conditions. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain JN500901T belongs to Clostridiumsensu stricto, and shared the highest sequence similarity to Clostridiumcarboxidivorans DSM 15243T (94.2 %), followed by Clostridiumscatologenes DSM 757T (94.1 %). The dominant cellular fatty acids (>10 %) were C16 : 0 FAME (36.6 %), C19 : 0 cyc 9,10 DMA (19.8 %) and C16 : 1 cis 9 DMA (11.8 %). The complete genome of strain JN500901T contained a circular chromosome of 2.812 Mb with 2611 genes and 31.0 mol% G+C content. Comparative genome analysis of the strain JN500901T, Clostridiumcarboxidivorans DSM 15243T and Clostridiumscatologenes DSM 757T revealed 74.5 and 74.8 % average nucleotide identity, respectively. Based on the phenotypic, biochemical and phylogenetic analyses presented here, strain JN500901T is considered to be a novel species of the genus Clostridiumsensustricto, for which the name Clostridium fermenticellae sp. nov. is proposed. The type strain is JN500901T (=CICC 24501T=JCM 32827T).


Assuntos
Bebidas Alcoólicas , Clostridium/classificação , Filogenia , Microbiologia do Solo , Técnicas de Tipagem Bacteriana , Composição de Bases , China , Clostridium/isolamento & purificação , DNA Bacteriano/genética , Etanol , Ácidos Graxos/química , Fermentação , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
6.
Rapid Commun Mass Spectrom ; 29(7): 645-53, 2015 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-26212282

RESUMO

RATIONALE: Glycine is the smallest amino acid used in protein synthesis, but it is also a very important precursor for the biosynthesis of other nitrogen-containing metabolites, such as purine nucleosides and nucleotides for synthesis of RNA, DNA etc. Abnormalities in glycine metabolism therefore cause diseases such as cancer. A quick and unambiguous method to trace the metabolites arising from glycine is required for targeting defect points within metabolic networks. METHODS: This paper describes a method for using (15)N-glycine to culture A549 cancer cells for use with high-resolution mass spectrometry (HRMS) and tandem mass spectrometry (HRMS(2)) that can detect the (M+1)/M pair peaks appearing in the cell metabolites. The 1 Da difference in the pair peaks can be used to point out and identify the nitrogen metabolites of glycine. RESULTS: Thirteen nitrogen-containing metabolites derived from glycine were confirmed. Among them were metabolites containing purine, such as adenine, adenosine, AMP, ADP, ATP, S-adenosylmethionine and γ-glutathione; these were the most sensitive to the (15)N-glycine-enrichment technique. Therefore, they are promising biomarkers for monitoring the glycine metabolism network. CONCLUSIONS: The method developed here could be applied to investigations of metabolism of other amino acids, and for drug discovery studies targeting the enzymes related to amino acid metabolism.


Assuntos
Glicina/química , Glicina/metabolismo , Isótopos de Nitrogênio/análise , Isótopos de Nitrogênio/metabolismo , Espectrometria de Massas em Tandem/métodos , Linhagem Celular , Glicina/análise , Humanos , Íons/análise , Íons/química , Metabolômica , Isótopos de Nitrogênio/química
7.
Int J Clin Exp Pathol ; 8(10): 12234-42, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26722408

RESUMO

OBJECTIVE: To investigate the role and mechanism of epidermal growth factor like domain 7 (EGFL7) in glioma angiogenesis by cell co-culture and RNA interference. METHODS: NSCs-HUVECs co-culture system was established using Transwell culturing techniques. The interactions between glioma and endothelial cells were simulated in-vitro. Cellular expression of EGFL7 in NSCs and HUVEC was targeted and suppressed by lentiviral vector carrying siRNA. The effect of EGFL7 on angiogenesis in glioma in-vitro micro-environment was detected by endothelial cell proliferation, adhesion and tube formation assay. RESULTS: Following EGFL7 gene silencing, expression of EGFL7 in HUVECs was reduced and cell adhesion capability was inhibited significantly. Endothelial cells failed to form a lumen-like structure after EGFL7 gene silencing, shown by the tube formation assay. CONCLUSION: By regulating endothelial cell adhesion, EGFL7 plays a key role in the regulation of glioma angiogenesis.


Assuntos
Fatores de Crescimento Endotelial/metabolismo , Glioma/patologia , Neovascularização Patológica/patologia , Proteínas de Ligação ao Cálcio , Adesão Celular , Proliferação de Células , Técnicas de Cocultura , Família de Proteínas EGF , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Fatores de Crescimento Endotelial/genética , Glioma/metabolismo , Células-Tronco Embrionárias Humanas , Células Endoteliais da Veia Umbilical Humana , Humanos , Neovascularização Patológica/metabolismo , Interferência de RNA
8.
Chem Asian J ; 8(8): 1854-62, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23712930

RESUMO

A large amount of interesting information on the alternating copolymerization of CO2 with terminal epoxides has already been reported, such as the regiochemistry of epoxide ring-opening and the stereochemistry of the carbonate unit sequence in the polymer chain. Moreover, the microstructures of CO2 copolymers from propylene oxide and cyclohexene oxide have also been well-studied. However, the microstructure of the CO2 copolymer from styrene oxide (SO), an epoxide that contains an electron-withdrawing group, has not yet been investigated. Herein, we focus on the spectroscopic assignment of the CO2 copolymer from styrene oxide at the diad level by using three kinds of model dimer compounds, that is, T-T, H-T, and H-H. By comparing the signals in the carbonyl region, we concluded that the signals at δ=154.3, 153.8, and 153.3 ppm in the (13)C NMR spectrum of poly(styrene carbonate) were due to tail-to-tail, head-to-tail, and head-to-head carbonate linkages, respectively. Moreover, various isotactic and syndiotactic model compounds based on T-T, H-T, and H-H (dimers (R,R)-T-T, (S,S)-T-T, and (R,S)-T-T; (R,R)-H-T, (S,S)-H-T, and (R,S)-H-T; (R,R)-H-H, (S,S)-H-H, and (R,S)-H-H) were synthesized for the further spectroscopic assignment of stereospecific poly(styrene carbonate)s. We found that the carbonate carbon signals were sensitive towards the stereocenters on adjacent styrene oxide ring-opening units. These discoveries were found to be well-matched to the microstructures of the stereoregular poly(styrene carbonate)s that were prepared by using a multichiral Co(III)-based catalyst system.

9.
Acta Crystallogr Sect E Struct Rep Online ; 65(Pt 2): o270, 2009 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-21581884

RESUMO

The title compound, C(16)H(17)NO(3), adopts a folded conformation in the crystal structure. The crystal packing is stabilized by inter-molecular O-H⋯O and N-H⋯O hydrogen-bonding inter-actions. The absolute configuration was assigned assuming that the absolute configuration of the starting material l-tyrosine was retained during the synthesis.

10.
Rapid Commun Mass Spectrom ; 22(19): 2977-83, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18763263

RESUMO

Several O-2',3'-isopropylideneuridine-O-5'-phosphoramidate monoesters were synthesized and analyzed by negative ion electrospray ionization tandem mass spectrometry (ESI-MS(n)). Two kinds of novel rearrangement reactions were observed due to the difference in the amino acid in the nucleoside phosphoramidate monoesters, and possible mechanisms were proposed. One involves a five-membered cyclic transition state. The other is formation of a stable five-membered ring intermediate by Michael addition. Results were confirmed by tandem mass spectrometry and isotopically labeled hydrogen atoms. Furthermore, the internal hydrogen exchange between active hydrogen and methyl acrylate in the heated capillary of the mass spectrometer was found. The characteristic fragmentation behavior in ESI-MS may be used to monitor this kind of compounds in the biological metabolism.


Assuntos
Amidas/química , Deutério/química , Modelos Químicos , Modelos Moleculares , Nucleosídeos/química , Ácidos Fosfóricos/química , Espectrometria de Massas por Ionização por Electrospray/métodos , Ânions , Simulação por Computador , Ésteres , Conformação Molecular , Coloração e Rotulagem
11.
Acta Crystallogr Sect E Struct Rep Online ; 64(Pt 7): o1326, 2008 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-21202950

RESUMO

The asymmetric unit of the title compound, C(12)H(11)NO(5), contains two independent mol-ecules with similar geometric parameters but different orientations of the phenyl rings. The mol-ecular packing is stabilized by weak nonclassical C-H⋯O hydrogen-bonding inter-actions.

12.
Environ Monit Assess ; 130(1-3): 281-7, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17072553

RESUMO

HPLC and HPLC-MS are the fastest and most accurate techniques for analysis of organic phosphorus pesticide (OPP) at the present time. Using these techniques, 14 strains of methamidopho (MAP) degrading-bacteria from the area contaminated with MAP have been identified. The results from HPLC and HPLC-MS analyses showed that the highest degradation rate was 73% after 7 days. In order to determine what metabolites will be formed after degradation, a key issue that has been neglected for a long time, we used ((31)P) NMR to track the degradation process. The results showed that different strains produced different metabolites. Ten strains were divided into three groups (groups A, B and C) by their metabolic profiling. Strains in group A degraded MAP into phosphor acid by breaking down all P-N, P-O and P-S bonds in 7 days. Strains in groups B and C had only broken down partially P-N and P-S bonds at the same time. Therefore, the bacterial strains in group A had a greater application potential than the other two groups. In addition, most metal phosphates are unsolvable in water. The analysis of X-ray showed, that the phosphate radicals generated by bacterial degradation induce crystallogenesis of heavy metal salts in water phase and also cause the chemical sedimentation of their crystals. Furthermore, these crystals are hydrogen phosphates. The results suggested that the MAP-degrading bacteria could be used for cleaning up not only the organic phosphorous pesticide contamination but also the phosphorous and heavy metal contamination in water environment simultaneously.


Assuntos
Bactérias , Biodegradação Ambiental , Espectroscopia de Ressonância Magnética/métodos , Compostos Organofosforados/análise , China , Ecossistema , Compostos Organofosforados/efeitos adversos , Compostos Organofosforados/classificação , Resíduos de Praguicidas , Rios
13.
J Mass Spectrom ; 40(10): 1382-6, 2005 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-16220467

RESUMO

Phosphoryl chloride was able to promote the formation of peptide and the ringopening of tetrahydrofuran (THF) followed by a sequence of successive reactions by simply refluxing. ESI multistage tandem mass spectrometry was applied to trace the reaction and elucidate the product structures, dipeptide 4-chlorobutyl ester.


Assuntos
Aminoácidos/química , Furanos/química , Compostos de Fósforo/química , Solventes/química , Espectrometria de Massas por Ionização por Electrospray/métodos , Dipeptídeos/química
14.
Eur J Mass Spectrom (Chichester) ; 11(3): 319-24, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-16107746

RESUMO

Phosphorus oxychloride was found to induce ring opening of tetrahydrofuran (THF) to synthesise THF oligomer incorporated phosphate. ESI multi-stage tandem mass spectrometry was used to study the products and confirm the structures.

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