Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA325930

Local Sample ID:	M1605
Subject ID:	SU003106
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606

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Sample Preparation:

Sampleprep ID:	SP003112
Sampleprep Summary:	For LC-MS analysis, 40 µL of serum was mixed with 800 µL of a cold mixture (-20ºC) of methanol:MTBE:Chloroform (1.33:1:1, v/v/v) with Sphinganine (D17:0) and palmitic acid-d31 as internal standards. Samples were vortexed for 30 s and shaken for 20 min at maximum speed at room temperature. Next, samples were centrifuged (13,200 rpm, room temperature, 5 min). After centrifugation, supernatant was directly injected into the system. For GC-MS analysis, protein precipitation was achieved by mixing 1 volume of serum with 3 volumes of cold (-20ºC) acetonitrile with 25 ppm of palmitic acid-d31 as internal standard, followed by methoximation with O-methoxyamine hydrochloride (15 mg/mL) in pyridine, and sylation with BSTFA: TMCS (99:1). Finally, 20 ppm of tricosane in heptane was added as second internal standard. For CE-MS analysis, 100 µL of serum was mixed with 100 µL of 0.2 M formic acid containing 5% acetonitrile and 0.4 mM methionine sulfone, 2 mM paracetamol and 0.5 mM 4-Morpholineethanesulfonic acid, 2-(N-Morpholino) ethanesulfonic acid (MES) as internal standards. The sample was transferred to an ultracentrifugation device (Millipore Ireland Ltd., Carrigtohill, Ireland) with a 30 kDa protein cutoff for deproteinization through centrifugation (2000 × g, 4 °C, 90 min).

Sampleprep ID:

SP003112

Sampleprep Summary:

For LC-MS analysis, 40 µL of serum was mixed with 800 µL of a cold mixture (-20ºC) of methanol:MTBE:Chloroform (1.33:1:1, v/v/v) with Sphinganine (D17:0) and palmitic acid-d31 as internal standards. Samples were vortexed for 30 s and shaken for 20 min at maximum speed at room temperature. Next, samples were centrifuged (13,200 rpm, room temperature, 5 min). After centrifugation, supernatant was directly injected into the system. For GC-MS analysis, protein precipitation was achieved by mixing 1 volume of serum with 3 volumes of cold (-20ºC) acetonitrile with 25 ppm of palmitic acid-d31 as internal standard, followed by methoximation with O-methoxyamine hydrochloride (15 mg/mL) in pyridine, and sylation with BSTFA: TMCS (99:1). Finally, 20 ppm of tricosane in heptane was added as second internal standard. For CE-MS analysis, 100 µL of serum was mixed with 100 µL of 0.2 M formic acid containing 5% acetonitrile and 0.4 mM methionine sulfone, 2 mM paracetamol and 0.5 mM 4-Morpholineethanesulfonic acid, 2-(N-Morpholino) ethanesulfonic acid (MES) as internal standards. The sample was transferred to an ultracentrifugation device (Millipore Ireland Ltd., Carrigtohill, Ireland) with a 30 kDa protein cutoff for deproteinization through centrifugation (2000 × g, 4 °C, 90 min).