高山被孢霉淬灭方法及胞内代谢产物的气相色谱-质谱分析比较

在花生四烯酸生产菌高山被孢霉代谢组学研究中,需利用胞内代谢物的提取手段并基于气相色谱-质谱（GC-MS）分析方法对其进行检测。比较了3种胞内代谢物提取方法及不同色谱柱条件下GC-MS分析结果。研究表明：采用冷甲醇淬灭分别较液氮直接淬灭及真空过滤后,减少了胞内代谢物的泄露并更好地实现了胞外及胞内代谢物的分离。在对代谢物分析的比较中,极性色谱柱（DB-FFAP）检出的代谢物仅为11种,主要为有机酸、醛类;而代谢物经衍生化后采用非极性色谱柱（DB-5）共检出32种化合物,主要为糖、糖苷及醇类。     

Identifying decomposition products in extracts of cellular metabolites

Most methods of analyzing intracellular metabolites require extraction of metabolites from the cells. A concern in these methods is underestimation of metabolite levels due to incomplete extraction. In comparing extraction methods, then, it would seem that the best method for extracting a particular metabolite is the one that gives the largest yield. In extracting Escherichia coli with different methanol:water mixtures, we observed that >or=50% water gave an increased yield of nucleosides and bases compared with 相似文献   

Genetic algorithm based two-mode clustering of metabolomics data

Metabolomics and other omics tools are generally characterized by large data sets with many variables obtained under different environmental conditions. Clustering methods and more specifically two-mode clustering methods are excellent tools for analyzing this type of data. Two-mode clustering methods allow for analysis of the behavior of subsets of metabolites under different experimental conditions. In addition, the results are easily visualized. In this paper we introduce a two-mode clustering method based on a genetic algorithm that uses a criterion that searches for homogeneous clusters. Furthermore we introduce a cluster stability criterion to validate the clusters and we provide an extended knee plot to select the optimal number of clusters in both experimental and metabolite modes. The genetic algorithm-based two-mode clustering gave biological relevant results when it was applied to two real life metabolomics data sets. It was, for instance, able to identify a catabolic pathway for growth on several of the carbon sources. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. J. A. Hageman and R. A. van den Berg contributed equally to this paper.     

Molecular docking and GC-MS data for the inhibition of RAD51 expression by a compound from Clerodendrum inerme L.

Screening of potential inhibitors for RAD51 from petroleum ether extract of Clerodendrum inerme L. (C.inerme) is of interest. Presence of phytocompounds was identified using GC-MS analysis. Molecular docking and ADME properties were calculated for potential inhibitors for RAD51. A total of 25 phytocompounds were extracted from the petroleum ether extract of C.inerme. The compound 1,2,4-Trimethyl-3-nitrobicyclo [3.3.1]nonan-9-one shows binding features with the cancer target protein RAD51 similar to the FDA approved drug of 5-Flurouracil for further consideration in the context of pancreatic cancer drug discovery.     

玫瑰蜂花粉脂肪酸的GC-MS分析

用石油醚作溶剂,提取出玫瑰蜂花粉粗脂肪,然后进行甲酯化处理,用气相色谱-质谱联用技术进行分离鉴定,共鉴定出19种脂肪酸,并测定相对含量,不饱和脂肪酸相对含量为68.07%,单不饱和脂肪酸含量中异油酸相对含量最高,为52.88%。     

GC-MS metabolites profiling of anethole-rich oils by different extraction techniques: antioxidant,cytotoxicity and in-silico enzymes inhibitory insights

GC-MS profiling and metabolomics study of anise and star anise oils obtained by hydrodistillation, n-hexane, and microwave-assisted extraction methods were conducted herein. Trans-anethole was the major phenylpropanoid in both oils. Principal component and hierarchical cluster analyses revealed a clear separation of different extraction methods. Microwave-assisted star anise oil (MSA) revealed the highest anethole content (93.78%). MSA oil showed antioxidant activity using DPPH and ABTS assays, this was verified via an in-silico docking study of its major compounds on human tyrosinase and NAD(P)H oxidase. Trans-anethole displayed the best fitting scores (−8.9 and −10.1 Kcal/mole, respectively). MSA oil showed promising cytotoxic activity on different cell lines, mainly the cervical (HeLa) cell lines. Cell cycle inhibition at the G0–G1 phase was observed with an early apoptotic effect of the oil on HeLa cells. Trans-anethole achieved the best docking scores (−7.9, −9.3 and −9.9 Kcal/mole) for in-silico study on EGFR, CDK2 and CDK4 enzymes engaged in cancer, respectively.     

三角梅属四个品种花挥发性组分的GC-MS分析

三角梅不仅观赏性强,而且具有多方面的保健作用。为了揭示三角梅的次生代谢机制,本研究采用水蒸气蒸馏法提取三角梅属四个品种(Bougainvillea glabra ‘Magnifica’、Bougainvillea glabra ‘Pink Pixie’、Bougainvillea glabra ‘Formosa’和Bougainvillea×buttiana ‘Mahara’)花的挥发性成分;通过GC-MS法对所提取的成分进行定性和定量分析。共分离出85种成分,经NIST数据库检索,并与标准谱图比较共鉴定出72个化学组分,占总挥发性组分含量的90%以上。结果表明,4个三角梅品种的花中含有植醇、丁子香酚、亚油酸、亚麻酸以及维生素E等对人类有益的化合物。     

Integrated pathway-based and network-based analysis of GC-MS rice metabolomics data under diazinon stress to infer affected biological pathways

Diazinon insecticide is widely applied in rice (Oryza sativa L.) fields in Iran. However, concerns are now being raised about its potential adverse impacts on rice. In this study, a time-course metabolic change in rice plants was investigated after diazinon treatment using gas chromatography-mass spectrometry (GC-MS) and subsequently three different methods, MetaboAnalyst, MetaboNetwork, and analysis of reporter reactions, as a potential multivariate method were used to find the underlying changes in metabolism with stronger evidence in order to link differentially expressed metabolites to biological pathways. Results clearly showed the similarity of acetylcholinesterase (AChE) of rice plants to that of animals in terms of its inhibitability by diazinon and emphasized that subsequent accumulation of AChE mainly affects the metabolism of osmolites and tricarboxylic acid intermediates subsequent accumulation of ACh mainly affects the metabolism of osmolites and TCA intermediates.     

鲜姜化学成分的GC—MS分析

本文采用鲜姜榨汁-过滤的方法提取鲜姜内成分。然后对其化学成分进行GC／MS分析。从鲜姜汁中分离确定了33中成分,其中萜类物质相对含量较高。     

藏药短管兔耳草脂溶性化学成分的GC-MS分析

采用毛细管色谱-质谱联用技术对青海省短管兔耳草石油醚浸提物的化学成分进行了分析,共分离出42个峰,鉴定出32种不同化合物。用气相色谱面积归一化法测定了各化学成分的相对含量,主要成分有亚油酸（39．037％）、正三十一烷（10．827％）、正二十九烷（6．683％）、棕榈酸（5．234％）、正十八烷（2．485％）、γ-谷甾醇（2．264％）等。     

Identifying and quantifying sources of variation in microarray data using high-density cDNA membrane arrays.

Microarray experiments involve many steps, including spotting cDNA, extracting RNA, labeling targets, hybridizing, scanning, and analyzing images. Each step introduces variability, confounding our ability to obtain accurate estimates of the biological differences between samples. We ran repeated experiments using high-density cDNA microarray membranes (Research Genetics Human GeneFilters Microarrays Version I) and 33P-labeled targets. Total RNA was extracted from a Burkitt lymphoma cell line (GA-10). We estimated the components of variation coming from: (1) image analysis, (2) exposure time to PhosphorImager screens, (3) differences in membranes, (4) reuse of membranes, and (5) differences in targets prepared from two independent RNA extractions. Variation was assessed qualitatively using a clustering algorithm and quantitatively using a version of ANOVA adapted to multivariate microarray data. The largest contribution to variation came from reusing membranes, which contributed 38% of the total variation. Differences in membranes and in exposure time each contributed about 10%. Differences in target preparations contributed less than 5%. The effect of image quantification was negligible. Much of the effect from reusing membranes was attributable to increasing levels of background radiation and can be reduced by using membranes at most four times. The effects of exposure time, which were partly attributable to variation in the scanning process, can be minimized by using the same exposure time for all experiments.     

Placental tissue metabolome analysis by GC-MS: Oven-drying is a viable sample preparation method

Abstract

Analysis of the human placenta metabolome has great potential to advance the understanding of complicated pregnancies and deleterious fetal outcomes in remote populations, but samples preparation can present unique challenges. Herein, we introduce oven-drying as a simple and widely available method of sample preparation that will facilitate investigations of the placental metabolome from remote and under-studied populations. Placentae from complicated and uncomplicated pregnancies were prepared in three ways (oven-dried at 60?°C, fresh, lyophilized) for metabolome analysis via gas chromatography-mass spectrometry (GC-MS). Multiple computer models (e.g. PLS-DA, ANN) were employed to classify and determine if there was a difference in placentae metabolome and a group of metabolites with high variable importance in projection scores across the three preparations and by complicated vs. control groups. The analyses used herein were shown to be thorough and sensitive. Indeed, significant differences were detected in metabolomes of complicated vs. uncomplicated pregnancies; however, there were no statistical differences in the metabolome of placentae prepared by oven-drying vs. lyophilization vs. fresh placentae. Oven-drying is a viable sample preparation method for placentae intended for use in metabolite analysis via GC-MS. These results open many possibilities for researching metabolome patterns associated with fetal outcomes in remote and resource-poor communities worldwide.     

Identifying and quantifying in vivo methylation sites by heavy methyl SILAC

Protein methylation is a stable post-translational modification (PTM) with important biological functions. It occurs predominantly on arginine and lysine residues with varying numbers of methyl groups, such as mono-, di- or trimethyl lysine. Existing methods for identifying methylation sites are laborious, require large amounts of sample and cannot be applied to complex mixtures. We have previously described stable isotope labeling by amino acids in cell culture (SILAC) for quantitative comparison of proteomes. In heavy methyl SILAC, cells metabolically convert [(13)CD(3)]methionine to the sole biological methyl donor, [(13)CD(3)]S-adenosyl methionine. Heavy methyl groups are fully incorporated into in vivo methylation sites, directly labeling the PTM. This provides markedly increased confidence in identification and relative quantitation of protein methylation by mass spectrometry. Using antibodies targeted to methylated residues and analysis by liquid chromatography-tandem mass spectrometry, we identified 59 methylation sites, including previously unknown sites, considerably extending the number of in vivo methylation sites described in the literature.     

皱皮木瓜果实中有机酸成分的GC-MS分析

木瓜为常用中药,其原植物为木瓜属（Chaenomeles Lindl．）植物皱皮木瓜[Chaenomeles speciosa（Sweet）Nakai],又称贴梗海棠、贴梗木瓜等,中国南方各地均有栽培。其花可供观赏;果实入药,能驱风强壮、舒筋、镇痛消肿,民间常用于浸泡药酒;种仁富含油酸和亚油酸。对皱皮木瓜有机酸的脂肪性成分已有研究报道,但尚无其水溶性有机酸成分的研究报道。皱皮木瓜的药用功效主要为镇痛解痉,但至今未明确其主效成分,有机酸虽具有一定的镇痛功效,但有机酸的种类不同,功效也不相同。鉴于长期以来皱皮木瓜还是作为汤药服用,所以就其水溶性有机酸组成及含量开展研究,特别是与其有机酸中的脂肪性成分进行比较研究尤为必要,对于揭示皱皮木瓜主要功效成分具有重要的意义。     

莱菔子脂肪酸成分的GC-MS分析

采用索氏提取法提取莱菔(Raphanussativus)种子中的脂肪酸成分,进行甲酯化处理后用气相色谱-质谱联用技术分离和鉴定脂肪酸成分的组成和含量。结果表明,从莱菔子中共分离鉴定出12种脂肪酸成分,占总量的99.32%,主要包括芥酸(32.47%)、油酸(29.07%)、亚油酸(9.45%)、亚麻酸(8.41%)、棕榈酸(4.31%)和硬脂酸(2.08%)等脂肪酸,其中饱和脂肪酸占9.25%,不饱和脂肪酸高达90.07%。     

3种红枣的挥发性化学成分的乙醇提取及测定

新鲜成熟的去核油枣、木枣及团枣用95％乙醇浸泡,低温蒸除乙醇、膏状物用乙醚萃取,用气相色谱－质谱联用仪测定乙醚溶液。结果表明,3种新鲜成熟的红枣的挥发性成分主要为棕榈烯酸乙酯、棕榈酸乙酯、肉豆蔻乙酯、亚油酸乙酯、油酸乙脂、月桂酸乙酯、硬脂酸乙酯、苹果酸二乙酯、葵酸乙酸等16种酯类,以及乙酸和棕榈烯酸等2种酸和3－羟基－2－丁酮。不同品种的红枣中所含酯类的多少及含量的高低均有所差异。在同一种红枣的果肉中检测到16种酯类尚属首次报道。     

蒙椴树叶挥发油成分的气相色谱-质谱分析

采用水蒸气蒸馏法对蒙椴树叶挥发油进行提取,挥发油回收率约为0.6%(w);用气相色谱—质谱联用技术对挥发性成分进行分析,鉴定了45个化合物,占挥发油成分的80.35%以上。其中主要有己醇,己烯醇,贝壳杉烯和苯甲酸等     

Influence of ethanol concentration of extraction solvent on metabolite profiling for Salviae Miltiorrhizae Radix et Rhizoma extract by 1H NMR spectroscopy and multivariate data analysis

Salviae Miltiorrhizae Radix et Rhizoma (Danshen in China) and its related preparations are widely used in clinical practice due to its high medicinal value. In recent years, ¹H NMR technology has made great progress and demonstrated its unique advantages in the field of botanical metabolomics. In this study, ¹H NMR-based metabolomics was used to investigate the dissolution of various metabolites in Danshen as a function of ethanol concentration. ¹H NMR spectroscopy of Danshen extract identified 28 metabolites including 6 sugars, 11 amino acids, 3 organic acids, 4 salvianolic acids, and 4 tanshinones. Multivariate statistical analysis was used to classify and compare various Danshen extracts. PCA and HCA were used to obtain a global overview of the similarity in the samples and two-class OPLS-DA models were established for identifying characteristic metabolites. Then, ¹H-qNMR method was used to estimate the concentration of 22 metabolites, which is helpful to further describe the changes in metabolite ratios of various Danshen extracts. The result of this study laid the foundation for further biological activity research, and also provided an important reference for subsequent process research and quality control of Danshen related preparations.     

白木香花和果实挥发油成分的GC-MS分析

采用溶剂萃取法提取白木香(Aquilaria sinensis(Lourl.)Gilg)花和果实的挥发油,经GC-MS分析,从花挥发油中鉴定出26个化合物,占总油量的92.07%;从果实挥发油中鉴定出26个化合物,占总油量的93.66%.其中11个化合物为共有成分,且二者均含壬酸等致香成分.