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MB Sample ID: SA300454

Local Sample ID:C1_160
Subject ID:SU002904
Subject Type:Human
Subject Species:Homo sapiens
Taxonomy ID:9606

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Combined analysis:

Analysis ID AN004550 AN004551
Analysis type MS MS
Chromatography type Normal phase Normal phase
Chromatography system Agilent 6550 QTOF Agilent 6550 QTOF
Column MicroSolv Diamond Hydride (150 x 2.1mm, 4um) MicroSolv Diamond Hydride (150 x 2.1mm, 4um)
MS Type ESI ESI
MS instrument type QTOF QTOF
MS instrument name Agilent 6550 QTOF Agilent 6550 QTOF
Ion Mode POSITIVE NEGATIVE
Units intensity intensity

MS:

MS ID:MS004297
Analysis ID:AN004550
Instrument Name:Agilent 6550 QTOF
Instrument Type:QTOF
MS Type:ESI
MS Comments:MS acquisition comments: For aqueous normal phase LC/MS-based metabolomics, amniotic fluid metabolites were extracted by addition of 1 part amniotic fluid to 15 parts 70% acetonitrile in ddH2O (vol:vol). The mixture was briefly vortexed and then centrifuged for 5 min at 16,000 × g to pellet precipitated proteins. The protein pellet was solubilized in 0.2M NaOH and quantified by DC Protein Assay (Bio-Rad). The volume of metabolite extract was normalized by amniotic protein content. An aliquot of the resulting extract (3 mL) was subjected to LC/MS untargeted metabolite profiling in positive and negative ion modes as described previously described,64 using a platform comprised of an Agilent Model 1290 Infinity II liquid chromatography system coupled to an Agilent 6550 iFunnel time-of-flight MS analyzer. Chromatography of metabolites utilized aqueous normal phase (ANP) chromatography on a Diamond Hydride column (Microsolv). Mobile phases consisted of: (A) 50% isopropanol, containing 0.025% acetic acid, and (B) 90% acetonitrile containing 5 mM ammonium acetate. To eliminate the interference of metal ions on chromatographic peak integrity and electrospray ionization, EDTA was added to the mobile phase at a final concentration of 6 µM. The following gradient was applied: 0-1.0 min, 99% B; 1.0-15.0 min, to 20% B; 15.0 to 29.0, 0% B; 29.1 to 37min, 99% B. Mass spectrometer parameters used for both positive and negative ion mode data acquisition were as follows: drying gas temperature was 200°C with a flow rate of 14 L/min. Nebulizer pressure was at 35 psi. Sheath gas temperature was 350°C with a flow rate of 11 L/min. Capillary and nozzle voltage were at 3500V and 1000V, respectively. Mass spectra were acquired at a rate of 2 spectra/sec over the mass range of 100 to 1700 m/z. Data processing comments: Raw LC/MS data were analyzed using MassHunter Profinder 8.0 and MassProfiler Professional (MPP) 15.1 software (Agilent Technologies). To ascertain the identities of metabolites, LC/MS data were searched against an in-house annotated personal metabolite database created using MassHunter PCDL manager 8.0 (Agilent) based on monoisotopic neutral mass (<5 ppm mass accuracy) and chromatographic retention times of pure standards. A molecular formula generator (MFG) algorithm in MPP was used to generate and score empirical molecular formulae, based on a weighted consideration of monoisotopic mass accuracy, isotope abundance ratios, and spacing between isotope peaks. A tentative compound ID was assigned when the PCDL database and MFG scores concurred for a given candidate molecule. Software procedures used for feature assignments: Tentatively assigned molecules were confirmed based on a match of LC retention times and/or MS/MS fragmentation spectra for pure molecular standards.
Ion Mode:POSITIVE
  
MS ID:MS004298
Analysis ID:AN004551
Instrument Name:Agilent 6550 QTOF
Instrument Type:QTOF
MS Type:ESI
MS Comments:MS acquisition comments: For aqueous normal phase LC/MS-based metabolomics, amniotic fluid metabolites were extracted by addition of 1 part amniotic fluid to 15 parts 70% acetonitrile in ddH2O (vol:vol). The mixture was briefly vortexed and then centrifuged for 5 min at 16,000 × g to pellet precipitated proteins. The protein pellet was solubilized in 0.2M NaOH and quantified by DC Protein Assay (Bio-Rad). The volume of metabolite extract was normalized by amniotic protein content. An aliquot of the resulting extract (3 mL) was subjected to LC/MS untargeted metabolite profiling in positive and negative ion modes as described previously described,64 using a platform comprised of an Agilent Model 1290 Infinity II liquid chromatography system coupled to an Agilent 6550 iFunnel time-of-flight MS analyzer. Chromatography of metabolites utilized aqueous normal phase (ANP) chromatography on a Diamond Hydride column (Microsolv). Mobile phases consisted of: (A) 50% isopropanol, containing 0.025% acetic acid, and (B) 90% acetonitrile containing 5 mM ammonium acetate. To eliminate the interference of metal ions on chromatographic peak integrity and electrospray ionization, EDTA was added to the mobile phase at a final concentration of 6 µM. The following gradient was applied: 0-1.0 min, 99% B; 1.0-15.0 min, to 20% B; 15.0 to 29.0, 0% B; 29.1 to 37min, 99% B. Mass spectrometer parameters used for both positive and negative ion mode data acquisition were as follows: drying gas temperature was 200°C with a flow rate of 14 L/min. Nebulizer pressure was at 35 psi. Sheath gas temperature was 350°C with a flow rate of 11 L/min. Capillary and nozzle voltage were at 3500V and 1000V, respectively. Mass spectra were acquired at a rate of 2 spectra/sec over the mass range of 100 to 1700 m/z. Data processing comments: Raw LC/MS data were analyzed using MassHunter Profinder 8.0 and MassProfiler Professional (MPP) 15.1 software (Agilent Technologies). To ascertain the identities of metabolites, LC/MS data were searched against an in-house annotated personal metabolite database created using MassHunter PCDL manager 8.0 (Agilent) based on monoisotopic neutral mass (<5 ppm mass accuracy) and chromatographic retention times of pure standards. A molecular formula generator (MFG) algorithm in MPP was used to generate and score empirical molecular formulae, based on a weighted consideration of monoisotopic mass accuracy, isotope abundance ratios, and spacing between isotope peaks. A tentative compound ID was assigned when the PCDL database and MFG scores concurred for a given candidate molecule. Software procedures used for feature assignments: Tentatively assigned molecules were confirmed based on a match of LC retention times and/or MS/MS fragmentation spectra for pure molecular standards.
Ion Mode:NEGATIVE
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