1H NMR metabolomics analysis of glioblastoma subtypes: correlation between metabolomics and gene expression characteristics

Glioblastoma multiforme (GBM) is the most common form of malignant glioma, characterized by unpredictable clinical behaviors that suggest distinct molecular subtypes. With the tumor metabolic phenotype being one of the hallmarks of cancer, we have set upon to investigate whether GBMs show differences in their metabolic profiles. (1)H NMR analysis was performed on metabolite extracts from a selection of nine glioblastoma cell lines. Analysis was performed directly on spectral data and on relative concentrations of metabolites obtained from spectra using a multivariate regression method developed in this work. Both qualitative and quantitative sample clustering have shown that cell lines can be divided into four groups for which the most significantly different metabolites have been determined. Analysis shows that some of the major cancer metabolic markers (such as choline, lactate, and glutamine) have significantly dissimilar concentrations in different GBM groups. The obtained lists of metabolic markers for subgroups were correlated with gene expression data for the same cell lines. Metabolic analysis generally agrees with gene expression measurements, and in several cases, we have shown in detail how the metabolic results can be correlated with the analysis of gene expression. Combined gene expression and metabolomics analysis have shown differential expression of transporters of metabolic markers in these cells as well as some of the major metabolic pathways leading to accumulation of metabolites. Obtained lists of marker metabolites can be leveraged for subtype determination in glioblastomas.     

HORA suite: a database and software for human metabolomics

HORA suite (Human blOod Range vAlidator) consists of a Java application used to validate the metabolomic analysis of human blood against a database that stores the normal plasma and serum range concentrations of metabolites. The goal of HORA is to find the metabolites that are outside the normal range and to show those not present in the list provided by the user, for different thresholds of concentration. Moreover it supplies a graphical interface to manage the data. The software can also be used to compare different metabolomic techniques. HORA is open-source software and it can be accessed at . A separate file contains instructions for the installation and a brief tutorial.     

Analytical metabolomics: nutritional opportunities for personalized health

Nutrition is the cornerstone of health; survival depends on acquiring essential nutrients, and dietary components can both prevent and promote disease. Metabolomics, the study of all small molecule metabolic products in a system, has been shown to provide a detailed snapshot of the body's processes at any particular point in time, opening up the possibility of monitoring health and disease, prevention and treatment. Metabolomics has the potential to fundamentally change clinical chemistry and, by extension, the fields of nutrition, toxicology and medicine. Technological advances, combined with new knowledge of the human genome and gut microbiome, have made and will continue to make possible earlier, more accurate, less invasive diagnoses, all while enhancing our understanding of the root causes of disease and leading to a generation of dietary recommendations that enable optimal health. This article reviews the recent contributions of metabolomics to the fields of nutrition, toxicology and medicine. It is expected that these fields will eventually blend together through development of new technologies in metabolomics and genomics into a new area of clinical chemistry: personalized medicine.     

Plant metabolomics: large-scale phytochemistry in the functional genomics era

Metabolomics or the large-scale phytochemical analysis of plants is reviewed in relation to functional genomics and systems biology. A historical account of the introduction and evolution of metabolite profiling into today's modern comprehensive metabolomics approach is provided. Many of the technologies used in metabolomics, including optical spectroscopy, nuclear magnetic resonance, and mass spectrometry are surveyed. The critical role of bioinformatics and various methods of data visualization are summarized and the future role of metabolomics in plant science assessed.     

Alignment of high resolution mass spectra: development of a heuristic approach for metabolomics

One of the challenges of using mass spectrometry for metabolomic analyses of samples consisting of thousands of compounds is that of peak identification and alignment. This paper addresses the issue of aligning mass spectral data from different samples in order to determine average component m/z peak values. The alignment scheme developed takes the instrument m/z measurement error into consideration in order to heuristically align two or more samples using a technique comparable to automated visual inspection and alignment. The results obtained using mass spectral profiles of replicate human urine samples suggest that this heuristic alignment approach is more efficient than other approaches using hierarchical clustering algorithms. The output consists of an average m/z and intensity value for the spectral components together with the number of matches from the different samples. One of the major advantages of using this alignment strategy is that it eliminates the boundary problem that occurs when using predetermined fixed bins to identify and combine peaks for averaging and the efficient runtime allows large datasets to be processed quickly.     

Metabolic engineering of flavonoids in tomato (Solanum lycopersicum): the potential for metabolomics

Flavonoids comprise a large and diverse group of polyphenolic plant secondary metabolites. In plants, flavonoids play important roles in many biological processes such as pigmentation of flowers, fruits and vegetables, plant-pathogen interactions, fertility and protection against UV light. Being natural plant compounds, flavonoids are an integral part of the human diet and there is increasing evidence that dietary polyphenols are likely candidates for the observed beneficial effects of a diet rich in fruits and vegetables on the prevention of several chronic diseases. Within the plant kingdom, and even within a single plant species, there is a large variation in the levels and composition of flavonoids. This variation is often due to specific mutations in flavonoid-related genes leading to quantitative and qualitative differences in metabolic profiles. The use of such specific flavonoid mutants with easily scorable, visible phenotypes has led to the isolation and characterisation of many structural and regulatory genes involved in the flavonoid biosynthetic pathway from different plant species. These genes have been used to engineer the flavonoid biosynthetic pathway in both model and crop plant species, not only from a fundamental perspective, but also in order to alter important agronomic traits, such as flower and fruit colour, resistance, nutritional value. This review describes the advances made in engineering the flavonoid pathway in tomato (Solanum lycopersicum). Three different approaches will be described; (I) Increasing endogenous tomato flavonoids using structural or regulatory genes; (II) Blocking specific steps in the flavonoid pathway by RNA interference strategies; and (III) Production of novel tomato flavonoids by introducing novel branches of the flavonoid pathway. Metabolite profiling is an essential tool to analyse the effects of pathway engineering approaches, not only to analyse the effect on the flavonoid composition itself, but also on other related or unrelated metabolic pathways. Metabolomics will therefore play an increasingly important role in revealing a more complete picture of metabolic perturbation and will provide additional novel insights into the effect of the introduced genes and the role of flavonoids in plant physiology and development.     

A gender-specific discriminator in Sprague-Dawley rat urine: the deployment of a metabolic profiling strategy for biomarker discovery and identification

The use of nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS) as complementary analytical techniques for open metabolic profiling is illustrated in the context of defining urinary biochemical discriminators between male and female Sprague-Dawley rats. Subsequent to the discovery of a female-specific urinary discriminator by LC-MS, further LC, MS, and NMR methods have been applied in a coordinated effort to identify this urinary component. Thereafter, the biological relevance and context of the identified component, in this case a steroid metabolite, has been achieved. This approach will be deployed in future studies of disease, drug efficacy, and toxicity to discover and identify biologically relevant markers.     

A hitchhiker’s guide to parasite transmission: The phoretic behaviour of feather lice

Transmission to new hosts is a fundamental challenge for parasites. Some species meet this challenge by hitchhiking on other, more mobile parasite species, a behaviour known as phoresis. For example, feather-feeding lice that parasitise birds disperse to new hosts by hitchhiking on parasitic louse flies, which fly between individual birds. Oddly, however, some species of feather lice do not engage in phoresis. For example, although Rock Pigeon (Columba livia) “wing” lice (Columbicola columbae) frequently move to new hosts phoretically on louse flies (Pseudolynchia canariensis), Rock Pigeon “body” lice (Campanulotes compar) do not. This difference in phoretic behaviour is puzzling because the two species of lice have very similar life cycles and are equally dependent on transmission to new hosts. We conducted a series of experiments designed to compare the orientation, locomotion and attachment capabilities of these two species of lice, in relation to louse flies. We show that wing lice use fly activity as a cue in orientation and locomotion, whereas body lice do not. We also show that wing lice are more capable of remaining attached to active flies that are walking, grooming or flying. The superior phoretic ability of wing lice may be related to morphological adaptations for life on wing feathers, compared to body feathers.     

Proposed minimum reporting standards for chemical analysis

There is a general consensus that supports the need for standardized reporting of metadata or information describing large-scale metabolomics and other functional genomics data sets. Reporting of standard metadata provides a biological and empirical context for the data, facilitates experimental replication, and enables the re-interrogation and comparison of data by others. Accordingly, the Metabolomics Standards Initiative is building a general consensus concerning the minimum reporting standards for metabolomics experiments of which the Chemical Analysis Working Group (CAWG) is a member of this community effort. This article proposes the minimum reporting standards related to the chemical analysis aspects of metabolomics experiments including: sample preparation, experimental analysis, quality control, metabolite identification, and data pre-processing. These minimum standards currently focus mostly upon mass spectrometry and nuclear magnetic resonance spectroscopy due to the popularity of these techniques in metabolomics. However, additional input concerning other techniques is welcomed and can be provided via the CAWG on-line discussion forum at or . Further, community input related to this document can also be provided via this electronic forum. The contents of this paper do not necessarily reflect any position of the Government or the opinion of the Food and Drug Administration Sponsor: Metabolomics Society http://www.metabolomicssociety.org/ Reference: http://msi-workgroups.sourceforge.net/bio-metadata/reporting/pbc/ http://msi-workgroups.sourceforge.net/chemical-analysis/ Version: Revision: 5.1 Date: 09 January, 2007     

OrFin: A web tool for detection of putative orthologs

Identification of ortholog is one of the important tasks to understand a novel genome. It helps to assign functional annotations, from one organism to another organism. To identify the putative ortholog, Reciprocal Best BLAST hit (RBBH) method is known to be an efficient approach. OrFin makes use of the same approach to identify pair of orthologous proteins for a given set of sequences of two species. It is a user-friendly web tool which works with user defined parameters to search RBBHs. Results are produced in both html and text format.

Availability

This web tool is freely available at http://bifl.uohyd.ac.in/orfin     

E-probe Diagnostic Nucleic acid Analysis (EDNA): A theoretical approach for handling of next generation sequencing data for diagnostics

Plant biosecurity requires rapid identification of pathogenic organisms. While there are many pathogen-specific diagnostic assays, the ability to test for large numbers of pathogens simultaneously is lacking. Next generation sequencing (NGS) allows one to detect all organisms within a given sample, but has computational limitations during assembly and similarity searching of sequence data which extend the time needed to make a diagnostic decision. To minimize the amount of bioinformatic processing time needed, unique pathogen-specific sequences (termed e-probes) were designed to be used in searches of unassembled, non-quality checked, sequence data. E-probes have been designed and tested for several selected phytopathogens, including an RNA virus, a DNA virus, bacteria, fungi, and an oomycete, illustrating the ability to detect several diverse plant pathogens. E-probes of 80 or more nucleotides in length provided satisfactory levels of precision (75%). The number of e-probes designed for each pathogen varied with the genome size of the pathogen. To give confidence to diagnostic calls, a statistical method of determining the presence of a given pathogen was developed, in which target e-probe signals (detection signal) are compared to signals generated by a decoy set of e-probes (background signal). The E-probe Diagnostic Nucleic acid Analysis (EDNA) process provides the framework for a new sequence-based detection system that eliminates the need for assembly of NGS data.     

A metabolomic analysis of medicinal diversity in Huang-qin (<Emphasis Type="Italic">Scutellaria baicalensis</Emphasis> Georgi) genotypes: discovery of novel compounds

In vitro manipulation of plant regeneration in the Chinese medicinal species Scutellaria baicalensis Georgi (Huang-qin) resulted in 26 chemically distinct germplasm lines. Antioxidant potential, growth rate and concentration of baicalin, baicalein, melatonin, and wogonin were the selective markers used to identify elite lines. Metabolomic analysis of a subset of the most distinct lines revealed that Huang-qin extracts contained over 2,000 compounds including 781 determined to be of putative medicinal importance as determined by a database search, as well as previously unidentified amino-derivatives of baicalin and wogonin. Huang-qin also contained a metabolite with the same net formula as hyperforin, previously thought to be unique to Hypericum perforatum L. Together these results provide new insights into the biochemical complexity of an important medicinal species and demonstrate the power of in vitro manipulation in combination with untargeted metabolomic screening for the production of new germplasm.     

Preliminary assays to elucidate the structure of oxytetracycline’s degradation products in sediments: Determination of natural tetracyclines by high-performance liquid chromatography–fast atom bombardment mass spectrometry

A very specific high-performance liquid chromatography–mass spectrometric method for the determination of natural tetracyclines was developed in order to characterise the degradation products of oxytetracycline in sediments. First, extraction used a clean up step with a Bond Elut Certify^® LRC cartridge. A 3 μm Spherisorb^® ODS1 column was then used with a methanol, acetonitrile and oxalic acid mobile phase gradient. Chromatographic resolution in these conditions was 3.31 between oxytetracycline and tetracycline. Two liquid chromatography–mass spectrometry methodologies based on a particle beam and a frit fast atom bombardment interface were developed. In the first approach, ionisation was performed in the negative chemical mode using methane as reacting gas. In the other case, glycerol–thioglycerol mixture was used as matrix to ensure good sensitivity. MS–MS experiment was performed to determinate oxytetracycline fragmentation pattern in the perspective of degradation product study.     

A scheme for multiple sequence alignment optimization—an improvement based on family representative mechanics features

As a basic tool of modern biology, sequence alignment can provide us useful information in fold, function, and active site of protein. For many cases, the increased quality of sequence alignment means a better performance. The motivation of present work is to increase ability of the existing scoring scheme/algorithm by considering residue-residue correlations better. Based on a coarse-grained approach, the hydrophobic force between each pair of residues is written out from protein sequence. It results in the construction of an intramolecular hydrophobic force network that describes the whole residue-residue interactions of each protein molecule, and characterizes protein's biological properties in the hydrophobic aspect. A former work has suggested that such network can characterize the top weighted feature regarding hydrophobicity. Moreover, for each homologous protein of a family, the corresponding network shares some common and representative family characters that eventually govern the conservation of biological properties during protein evolution. In present work, we score such family representative characters of a protein by the deviation of its intramolecular hydrophobic force network from that of background. Such score can assist the existing scoring schemes/algorithms, and boost up the ability of multiple sequences alignment, e.g. achieving a prominent increase (∼50%) in searching the structurally alike residue segments at a low identity level. As the theoretical basis is different, the present scheme can assist most existing algorithms, and improve their efficiency remarkably.