@article {540, title = {A method for comparative metabolomics in urine using high resolution mass spectrometry.}, journal = {J Chromatogr A}, volume = {1443}, year = {2016}, month = {2016 Apr 22}, pages = {83-92}, abstract = {

Developing a workflow for metabolite profiling from biological fluids using mass spectrometry is imperative to extract accurate information. In this study, urine samples from smokers (n=10) and nonsmokers (n=10) were analyzed using an ultrahigh performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) system. For the analysis, two different chromatographic methods [Reversed phase chromatography (RPC) and Hydrophilic interaction liquid chromatography (HILIC)], in two ionization modes (positive and negative) were used. Spiked reserpine (positive ion mode) or taurocholate (negative ion mode) were used for data extraction and normalization. Quality controls (QCs), prepared by pooling urine samples from both smokers and non-smokers (each n=10), were used to assess the reproducibility of the method. The final data output from SIEVE 2.2 after applying a cut-off for QC coefficient of variation (CV) \<20\% and p-value \<0.05 showed 165, 83, 177 and 100 unique components in RP positive/negative, HILIC positive/negative modes, respectively. Statistical analysis showed clustering of the two groups and the QCs, while the variable importance in projection (VIP) scores for the top fifteen metabolites in each of the four modes indicated the metabolites most responsible for the differences. Application of the developed workflow for comparative metabolomic analysis of urine in different diseased models will be of great use in the field of clinical metabolomics.

}, keywords = {Chromatography, Liquid, Chromatography, Reverse-Phase, Hydrophobic and Hydrophilic Interactions, Mass Spectrometry, Metabolomics, Reproducibility of Results, Urinalysis}, issn = {1873-3778}, doi = {10.1016/j.chroma.2016.02.080}, author = {Ramakrishnan, Padma and Nair, Sreenath and Rangiah, Kannan} } @article {501, title = {A quantitative metabolomics peek into planarian regeneration. [Mass spectrometry - Metabolomics]}, journal = {Analyst}, volume = {140}, year = {2015}, month = {2015 May 21}, pages = {3445-64}, abstract = {

The fresh water planarian species Schmidtea mediterranea is an emerging stem cell model because of its capability to regenerate a whole animal from a small piece of tissue. It is one of the best model systems to address the basic mechanisms essential for regeneration. Here, we are interested in studying the roles of various amines, thiols and nucleotides in planarian regeneration, stem cell function and growth. We developed mass spectrometry based quantitative methods and validated the differential enrichment of 35 amines, 7 thiol metabolites and 4 nucleotides from both intact and regenerating planarians. Among the amines, alanine in sexual and asparagine in asexual are the highest (\>1000 ng/mg) in the intact planarians. The levels of thiols such as cysteine and GSH are 651 and 1107 ng mg(-1) in planarians. Among the nucleotides, the level of cGMP is the lowest (0.03 ng mg(-1)) and the level of AMP is the highest (187 ng mg(-1)) in both of the planarian strains. We also noticed increasing levels of amines in both anterior and posterior regenerating planarians. The blastema from day 3 regenerating planarians also showed higher amounts of many amines. Interestingly, the thiol (cysteine and GSH) levels are well maintained during planarian regeneration. This suggests an inherent and effective mechanism to control induced oxidative stress because of the robust regeneration and stem cell proliferation. Like in intact planarians, the level of cGMP is also very low in regenerating planarians. Surprisingly, the levels of amines and thiols in head regenerating blastemas are \~{}3 times higher compared to those for tail regenerating blastemas. Thus our results strongly indicate the potential roles of amines, thiols and nucleotides in planarian regeneration.

}, keywords = {Animals, Calibration, Chromatography, High Pressure Liquid, Limit of Detection, Metabolomics, Planarians, Reference Standards, Regeneration, Reproduction, Asexual, Species Specificity, Tandem Mass Spectrometry}, issn = {1364-5528}, doi = {10.1039/c4an02037e}, author = {Natarajan, Nivedita and Ramakrishnan, Padma and Lakshmanan, Vairavan and Palakodeti, Dasaradhi and Rangiah, Kannan} }