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MB Sample ID: SA136953
Local Sample ID: | 171009bBG_8_005_1 |
Subject ID: | SU001690 |
Subject Type: | Bacteria |
Subject Species: | Rothia mucilaginosa |
Taxonomy ID: | 43675 |
Genotype Strain: | RmFLR01 |
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Sample Preparation:
Sampleprep ID: | SP001696 |
Sampleprep Summary: | For the 29 unlabeled standards mixture, 10 µL of the final solution was dried down for GC-MS measurement. Derivatization and data acquisition of mixture aliquots by GC-MS were reproduced on three days for inter-day precision. Dried mixtures were derivatized by adding 10 µL of 40 mg/mL methoxyamine hydrochloride (Sigma-Aldrich, St. Louis, MO, USA) in pyridine (Sigma-Aldrich, St. Louis, MO, USA) and shaking at 30°C for 1.5 h. Subsequently, 90 µL MTBSTFA (Sigma-Aldrich, St. Louis, MO, USA) was added with 13 fatty acid methyl esters (FAMEs) as retention index markers and shaken at 80°C for 30 min. Samples were immediately transferred to crimp top vials and injected onto each GC-MS instrument. Same samples of R. mucilaginosa cultures were extracted using published methods [19]. Samples were added 1 mL pre-chilled, degassed acetonitrile: isopropanol: water (v/v/v 3:3:2, Fisher Scientific) followed by vortexing 30 s and shaking at 4°C for 5 min. Samples were centrifuged for 2 min at 12,210 × g to precipitate debris from extracts. Supernatants were collected and split into two equal portions. One aliquot was dried to completeness in a Labconco cold trap centrifuge evaporator and then resuspended in 0.5 mL degassed acetonitrile: water (v/v 1:1, Fisher Scientific) to remove triacylglycerides. Resuspension solutions were vortexed for 30s and centrifuged for 2 min. Supernatants were transferred into clean Eppendorf tubes and dried down completely. Dried extracts were derivatized as given above. |