Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA352094

Local Sample ID:	AWT-IR_2
Subject ID:	SU003341
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

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Subject:

Subject ID:	SU003341
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

Factors:

Local Sample ID	MB Sample ID	Factor Level ID	Level Value	Factor Name
AWT-IR_2	SA352094	FL041093	Heart	Sample source
AWT-IR_2	SA352094	FL041093	Aged	Age
AWT-IR_2	SA352094	FL041093	WT	Genotype
AWT-IR_2	SA352094	FL041093	I/R	Treatment

Collection:

Collection ID:	CO003334
Collection Summary:	The sample collection method was: All mice groups underwent anesthesia with 2%-3% isoflurane and 100% O2. The mouse hearts were excised and rinsed in ice-cold PBS. Both atriums were removed; only the left ventricular myocardium, including the infarct area, was collected and was quickly frozen by using liquid nitrogen. Samples were preserved below -80 °C and were sent out for metabolomic and lipidomic analysis.
Sample Type:	Heart

Treatment:

Treatment ID:	TR003350
Treatment Summary:	YWT: young wild type; Sham: no treatment; MIF20: treatment with MIF20; AWT: aged wild type; MIFff: with MIF; cMIFKO: knock out MIF; I/R: "ischemia-reperfusion"; For I/R, the myocardium underwent 45 minutes long ischemic condition, then followed by 24 hours long reperfusion. MIF20 was administered 5 minutes before the onset of reperfusion by I.V. injection via the jugular vein. MIF20 was administered until the MIF20 concentration in blood reached 8nM. Procedure is : We injected 100 uL of MIF20 solution (160 nM) into a mouse by I.V. Consider the total blood volume of a mouse is about 2 mL. Therefore, the MIF20 solution was diluted 20 times, reaching 8 nM in blood.

Treatment ID:

TR003350

Treatment Summary:

YWT: young wild type; Sham: no treatment; MIF20: treatment with MIF20; AWT: aged wild type; MIFff: with MIF; cMIFKO: knock out MIF; I/R: "ischemia-reperfusion"; For I/R, the myocardium underwent 45 minutes long ischemic condition, then followed by 24 hours long reperfusion. MIF20 was administered 5 minutes before the onset of reperfusion by I.V. injection via the jugular vein. MIF20 was administered until the MIF20 concentration in blood reached 8nM. Procedure is : We injected 100 uL of MIF20 solution (160 nM) into a mouse by I.V. Consider the total blood volume of a mouse is about 2 mL. Therefore, the MIF20 solution was diluted 20 times, reaching 8 nM in blood.

Sample Preparation:

Sampleprep ID:	SP003348
Sampleprep Summary:	The lipidomic sample preparation method was: The samples were thawed on ice. Approximately 10 mg of each sample was weighed and homogenized by ball-mill in 1mL of extraction solution (MTBE:Methanol = 3:1, V/V) with internal standards. The mixture was vortexed for 15 min. Next, the mixture was added with 200 µL of water and vortexed for 1 min and incubating at 4 °C for 10 min. After centrifugation at 12000 rpm for 10 min (4 °C), 200 µL of the upper phase was collected for complete solvent drying under 20 °C. The residue was reconstituted using 200 µL of reconstitution solution (ACN:IPA = 1:1, V/V), followed by vortex for 3 min and centrifugation at 12000 rpm for 3 min. A 120 µL of the final supernatant was used for LC-MS analysis. The data acquisition instruments consisted of Ultra Performance Liquid Chromatography (UPLC) (Nex-era LC-40) and tandem mass spectrometry (MS/MS) (Triple Quad 6500+).

Sampleprep ID:

SP003348

Sampleprep Summary:

The lipidomic sample preparation method was: The samples were thawed on ice. Approximately 10 mg of each sample was weighed and homogenized by ball-mill in 1mL of extraction solution (MTBE:Methanol = 3:1, V/V) with internal standards. The mixture was vortexed for 15 min. Next, the mixture was added with 200 µL of water and vortexed for 1 min and incubating at 4 °C for 10 min. After centrifugation at 12000 rpm for 10 min (4 °C), 200 µL of the upper phase was collected for complete solvent drying under 20 °C. The residue was reconstituted using 200 µL of reconstitution solution (ACN:IPA = 1:1, V/V), followed by vortex for 3 min and centrifugation at 12000 rpm for 3 min. A 120 µL of the final supernatant was used for LC-MS analysis. The data acquisition instruments consisted of Ultra Performance Liquid Chromatography (UPLC) (Nex-era LC-40) and tandem mass spectrometry (MS/MS) (Triple Quad 6500+).

Combined analysis:

Analysis ID	AN005283
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Nexera LC-40
Column	Thermo Accucore C30 (150 x 2.1mm,2.6um)
MS Type	ESI
MS instrument type	Triple quadrupole
MS instrument name	ABI Sciex 6500+
Ion Mode	UNSPECIFIED
Units	lipid content in the sample (nmol/g)

Chromatography:

Chromatography ID:	CH003996
Chromatography Summary:	Solvent A: acetonitrile /water (60/40, V/V) (0.1% formic acid added, 10 mmol/L ammonium formate) Solvent B: acetonitrile / Isopropyl alcohol (10/90, V/V) (0.1% formic acid added, 10 mmol/L ammonium formate)
Instrument Name:	Nexera LC-40
Column Name:	Thermo Accucore C30 (150 x 2.1mm,2.6um)
Column Temperature:	45
Flow Gradient:	A:B; 80:20(V/V) at 0 min, 70:30(V/V) at 2 min, 40:60(V/V) at 4 min , 15:85(V/V) at 9 min, 10:90(V/V) at 14 min, 5:95(V/V) at 15.5 min, 5:95(V/V) at 17.3 min, 80:20(V/V) at 17.5 min, 80:20(V/V) at 20 min
Flow Rate:	0.35ml/min
Solvent A:	60% acetonitrile:40% water; 0.1% formic acid; 10 mmol/L ammonium formate
Solvent B:	10% acetonitrile:90% isopropyl alcohol; 0.1% formic acid added; 10 mmol/L ammonium formate
Chromatography Type:	Reversed phase

MS:

MS ID:	MS005013
Analysis ID:	AN005283
Instrument Name:	ABI Sciex 6500+
Instrument Type:	Triple quadrupole
MS Type:	ESI
MS Comments:	LIT and triple quadrupole (QQQ) scans were acquired on a triple quadrupole-linear ion trap mass spectrometer (QTRAP), QTRAP® 6500+ LC-MS/MS System, equipped with an ESI Turbo Ion-Spray interface, operating in positive and negative ion mode and controlled by Analyst 1.6.3 software (Sciex). The ESI source operation parameters were as follows: ion source, turbo spray; source temperature 500 ℃; ion spray voltage (IS) 5500 V(Positive),-4500 V(Neagtive); Ion source gas 1 (GS1), gas 2 (GS2), curtain gas (CUR) were set at 45, 55, and 35 psi, respectively. Instrument tuning and mass calibration were performed with 10 and 100 μmol/L polypropylene glycol solutions in QQQ and LIT modes, respectively. QQQ scans were acquired as MRM experiments with collision gas (nitrogen) set to 5 psi. DP and CE for individual MRM transitions was done with further DP and CE optimization. A specific set of MRM transitions were monitored for each period according to the lipids eluted within this period.
Ion Mode:	UNSPECIFIED