Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA174177

Local Sample ID:	P H-501
Subject ID:	SU001930
Subject Type:	Mammal
Subject Species:	Mesocricetus auratus
Taxonomy ID:	10036

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Sample Preparation:

Sampleprep ID:	SP001936
Sampleprep Summary:	Hamster plasma samples were diluted 1:4 with methanol (v/v), vortexed for 30 seconds, and incubated at -20 C for 2 hours. Samples were centrifuged for 10 minutes at 13,500 x g at 4°C and supernatant was transferred to a new centrifuge tube, concentrated, and stored at -80 C until reconstitution. Hamster plasma was thawed on ice. A 50 µL aliquot was transferred onto the solid-phase-extraction (SPE)-system CAPTIVA-EMR Lipid 96-wellplate (Agilent Technologies) before addition of 250 µL of acetonitrile containing 1% formic acid (v/v) and 10 µM internal standard (consisting of uniformly 13C and 15N labeled amino acids from Cambridge Isotope Laboratories, Inc). The samples were mixed for 1 min at 360 rpm on an orbital shaker at room temperature prior to a 10 min incubation period at 4 C. Afterwards, 200 µL 80% acetonitrile in water (v/v) were added to the samples. The samples were mixed on an orbital shaker (360 rpm) for an additional 10 min at room temperature. The samples were then eluted into a 96-deepwell collection plate by centrifugation (10 min, 57 x g, 4 C followed by 2 min, 1000 x g, 4 C). Polar eluates were stored at -80 C until the day of LC/MS analysis. The SPE-plates were then washed twice with 500 µL 80% acetonitrile in water (v/v). Lipids still bound to the SPE-material were then released into a second elution plate, in two elution steps applying 2x 500 µL 1:1 methyl tert-butyl ether:methanol (v/v) onto the SPE cartridge and centrifuging for 2 min at 1000 g and 4 C. The combined eluates were dried under a stream of nitrogen (Biotage SPE Dry Evaporation System) at room temperature and reconstituted with 100 µL 1:1 2-propanol:methanol (v/v) prior to LC/MS analysis.

Sampleprep ID:

SP001936

Sampleprep Summary:

Hamster plasma samples were diluted 1:4 with methanol (v/v), vortexed for 30 seconds, and incubated at -20 C for 2 hours. Samples were centrifuged for 10 minutes at 13,500 x g at 4°C and supernatant was transferred to a new centrifuge tube, concentrated, and stored at -80 C until reconstitution. Hamster plasma was thawed on ice. A 50 µL aliquot was transferred onto the solid-phase-extraction (SPE)-system CAPTIVA-EMR Lipid 96-wellplate (Agilent Technologies) before addition of 250 µL of acetonitrile containing 1% formic acid (v/v) and 10 µM internal standard (consisting of uniformly 13C and 15N labeled amino acids from Cambridge Isotope Laboratories, Inc). The samples were mixed for 1 min at 360 rpm on an orbital shaker at room temperature prior to a 10 min incubation period at 4 C. Afterwards, 200 µL 80% acetonitrile in water (v/v) were added to the samples. The samples were mixed on an orbital shaker (360 rpm) for an additional 10 min at room temperature. The samples were then eluted into a 96-deepwell collection plate by centrifugation (10 min, 57 x g, 4 C followed by 2 min, 1000 x g, 4 C). Polar eluates were stored at -80 C until the day of LC/MS analysis. The SPE-plates were then washed twice with 500 µL 80% acetonitrile in water (v/v). Lipids still bound to the SPE-material were then released into a second elution plate, in two elution steps applying 2x 500 µL 1:1 methyl tert-butyl ether:methanol (v/v) onto the SPE cartridge and centrifuging for 2 min at 1000 g and 4 C. The combined eluates were dried under a stream of nitrogen (Biotage SPE Dry Evaporation System) at room temperature and reconstituted with 100 µL 1:1 2-propanol:methanol (v/v) prior to LC/MS analysis.