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MTBLS25:  Urine Latin Square Spike-In for testing the NMR quantification tool, MetaboQuant.

 Authors: Matthias Klein , Peter Oefner , Wolfram Gronwald

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  Submitted: 03-Jan-2013 , Release date: 10-Jan-2013 , Update date: 17-Dec-2015

 Submitted by:  Matthias Klein  |   Study status: Public

Study Description

To test the quantification performance of MetaboQuant and other NMR quantification tools, a set of eight metabolites were added to replicates of a urine sample in varying concentrations. The metabolites were alanine, creatinine, histidine, betaine, acetate, phosphocreatine, β-hydroxybutyrate (BHBA) and lactate. These were added in concentrations of 6000, 3000, 1500, 750, 375, 188, 94, and 47 micromol/L in a Latin Square design. 2D 1H-13C-HSQC spectra were recorded on a 600 MHz Bruker Avance III spectrometer employing a triple-resonance cryoprobe and z-gradients.

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  Organism(s)

Homo sapiens

  Study Design Description

CHMO:nuclear magnetic resonance spectroscopy

targeted metabolites

MetaboQuant

NCIT:Latin Square Study

Protocol Description
Sample collection A spot urine specimen was collected from an apparently healthy volunteer and split into eight aliquots. A Latin Square spike-in experiment was prepared, where the sum of molar amounts of spike-in’s added to aliquots of a biological specimen is the same. Eight different metabolites, namely alanine, creatinine, histidine, betaine, acetate, phosphocreatine, β-hydroxybutyrate (BHBA) and lactate (Sigma-Aldrich, Steinheim, Germany), were added in varying amounts to the eight urine aliquots.
Extraction N/A
NMR sample 400 μl of this spiked urine were added to 200 μl of 0.1 M phosphate buffer (pH 7.4) and 50 μl of deuterium oxide containing TSP as reference substance.
NMR spectroscopy 2D 1H-13C HSQC spectra were acquired on an AVANCE III NMR spectrometer (Bruker BioSpin, Rheinstetten, Germany) using a 5 mm Triple-resonance cryoprobe with z-gradients probe at a proton frequency of 600 MHz and ambient temperature of 298 Kelvin.
NMR assay 2048 x 128 data points were collected using the pulse program r_hsqcetgppr (Bruker BioSpin) using 8 scans/increment, a relaxation delay of 3 secs, an acquisition time of 0.14 secs and 16 dummy scans. This resulted in a total acquisition time of 56 min/spectrum. The spectral widths were set to 12 and 165 ppm in the proton and carbon dimensions, respectively. Water suppression was achieved using presaturation during the relaxation delay.
Data transformation 2D spectra were semiautomatically processed employing a 90° shifted squared sine-bell window function in both dimensions. For increased resolution in the indirect carbon dimension, the number of data points was doubled prior to Fourier transform using complex forward linear prediction. All 2D spectra were manually phase corrected, and a polynomial baseline correction was applied excluding the region around the water artifact. For more details see [1].

Ref:
[1] Gronwald W, Klein MS, Kaspar H, Fagerer SR, Nürnberger N, Dettmer K, Bertsch T, Oefner PJ. Urinary metabolite quantification employing 2D NMR spectroscopy. Anal Chem. 2008 Dec 1;80(23):9288-97. doi: 10.1021/ac801627c. PMID:19551947
Metabolite identification For metabolite assignment, a urinary 2D 1H-13C HSQC spectrum was used. Amix 3.9.13 (Bruker BioSpin) was used for peak picking. An NMR peak search was performed using the Biological Magnetic Resonance Data Bank (available at http://www.bmrb.wisc.edu/), the Human Metabolome Database (available at http://www.hmdb.ca/) and the BBIOREFCODE database (Bruker BioSpin). To assign the signals, the spectrum was manually overlaid with reference spectra of pure compounds.
Software MetaboQuant (http://genomics.uni-regensburg.de/site/institute/software/metaboquant) is a tool developed by Matthias S. Klein that allows reliable quantification from nuclear magnetic resonance (NMR) spectrum peak integrals by using individual calibration factors for each peak. Additionally, several outlier detection methods have been implemented to exclude overlapped and corrupted peaks from quantification. Detailed information about MetaboQuant can be found in the manual that is included in the file MetaboQuant.zip (http://genomics.uni-regensburg.de/software/NMR/MetaboQuant_1.2.zip) and is also provided in this study folder (manual.pdf).
Installation MetaboQuant may be run as stand-alone application using Windows 2000 or newer. On other operating systems such as Linux or Mac OS, you may run MetaboQuant from within Matlab (The Mathworks, Natick, MA, USA). For this you need: Matlab version 7.1.0.246 (R14) Service Pack 3 or newer.

To install MetaboQuant:
Download the following files: MCRInstallerR2007b.exe and MetaboQuant.zip.
Run the file MCRInstallerR2007b.exe to install the required Matlab run-time routines.
Extract the file MetaboQuant.zip to a new folder.

To run MetaboQuant:
Double-click on the file MetaboQuant.exe. Starting MetaboQuant may take some minutes, as the Matlab run-time environment takes some time to be loaded. Extract MetaboQuant.zip to a new folder.
Start Matlab and change the working directory to the MetaboQuant installation folder, e.g. using the command cd.
To run MetaboQuant, type MetaboQuant in the Matlab command line.
Source Name Organism Organism part Protocol REF Sample Name
Urine Homo sapiens urine Sample collection Latin_Square_1_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_2_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_3_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_4_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_5_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_6_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_7_2012_04_05
Urine Homo sapiens urine Sample collection Latin_Square_8_2012_04_05
Validations marked with (*) have been allowed by the MetaboLights Curators.
Click here for the detailed description of Validations.
Condition Status Description Requirement Group Message
PASSES Study Title MANDATORY STUDY OK
PASSES Study Description MANDATORY STUDY OK
PASSES Study text successfully parsed OPTIONAL STUDY OK
PASSES Study Contact(s) have listed email MANDATORY CONTACT OK
PASSES Sample(s) MANDATORY SAMPLES OK
PASSES Sample Name consistency check MANDATORY ASSAYS OK
PASSES Publication(s) associated with this Study MANDATORY PUBLICATION OK
PASSES Minimal Experimental protocol MANDATORY PROTOCOLS OK
PASSES Comprehensive Experimental protocol OPTIONAL PROTOCOLS OK
PASSES Sample Collection protocol MANDATORY PROTOCOLS OK
PASSES Protocols text successfully parsed OPTIONAL PROTOCOLS OK
PASSES Organism name MANDATORY ORGANISM OK
PASSES Organism part MANDATORY ORGANISM OK
* PASSES Study Factors MANDATORY FACTORS No Study Factor information is provided
PASSES Assay platform information OPTIONAL ASSAYS OK
PASSES Assay has raw files referenced MANDATORY FILES OK
PASSES Assay referenced raw files detection in filesystem MANDATORY FILES OK
PASSES Raw files in the Assay(s) have the correct format MANDATORY FILES OK
PASSES Assay(s) MANDATORY ASSAYS OK
PASSES All Assays have Metabolite Assignment File (MAF) referenced OPTIONAL FILES OK
PASSES Metabolite Assignment File (MAF) is present in Study folder MANDATORY FILES OK
PASSES Metabolite Assignment File (MAF) has correct format MANDATORY FILES OK
PASSES Metabolite Identification File (MAF) content MANDATORY FILES OK
PASSES ISA-Tab investigation file check MANDATORY ISATAB OK

Assay 

Assay file name: a_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy.txt
Measurement: metabolite profiling
Technology: NMR spectroscopy
Platform: Bruker

Data

Sample Name Protocol REF Extraction Method Extract Name Protocol REF NMR tube type Solvent Sample pH Temperature Unit Labeled Extract Name Label Protocol REF Instrument NMR Probe Number of transients Pulse sequence name Magnetic field strength Unit Acquisition Parameter Data File Protocol REF NMR Assay Name Free Induction Decay Data File Protocol REF Normalization Name Derived Spectral Data File Protocol REF Data Transformation Name Metabolite Assignment File
Latin_Square_1_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_1_2012_04_05 Latin_Square_1_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_2_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_2_2012_04_05 Latin_Square_2_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_3_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_3_2012_04_05 Latin_Square_3_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_4_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_4_2012_04_05 Latin_Square_4_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_5_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_5_2012_04_05 Latin_Square_5_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_6_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_6_2012_04_05 Latin_Square_6_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_7_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_7_2012_04_05 Latin_Square_7_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
Latin_Square_8_2012_04_05 Extraction NMR sample 5 mm short 0.1 M phosphate buffered D2O 7.4 298 UO:kelvin hydrogen molecular entity NMR spectroscopy Bruker AVANCE III 600 MHz spectrometer 5 mm triple-resonance cryoprobe with z-gradients 128 2D 1H-13C HSQC (hsqcetgppr) 14.1 tesla NMR assay Latin_Square_8_2012_04_05 Latin_Square_8_2012_04_05.zip Data transformation Metabolite identification m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv

Pathways - Assay 



MetExplore Pathways Mapping

Name DB Identifier Mapped Metabolite(s)
  Download whole study (SLOW)  |  Download study (FTP)  |    Download metadata    

Aspera Download Details:

List of study files   Subset

File
audit
metexplore_mapping.json
Latin_Square_1_2012_04_05.zip
Latin_Square_2_2012_04_05.zip
Latin_Square_3_2012_04_05.zip
Latin_Square_4_2012_04_05.zip
Latin_Square_5_2012_04_05.zip
Latin_Square_6_2012_04_05.zip
Latin_Square_7_2012_04_05.zip
Latin_Square_8_2012_04_05.zip
MetaboQuant_1.2.zip
i_Investigation.txt
a_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy.txt
manual.pdf
m_urine latin square spike-in for testing metaboquant_metabolite profiling_NMR spectroscopy_v2_maf.tsv
s_Urine Latin Square Spike-In for Testing MetaboQuant.txt

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