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C10 September

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This folder contains mass spectra of extracellular extracts of Chaetoceros tenuissimus exposed to the protist grazer Oxyrrhis marina. C. tenuissimus was grown to different growth stages, specifically log (“L1-3”) and stationary (“S1-3”) growth phases before exposed to O. marina (n = 3). One treatment also included C. tenuissimus exposed to filtrate of O. marina only (“SF”). Filtrates of the now grazed diatoms (and media-only controls (“log BLK” and "STA blk")) were collected at T = 0, 2 and 4 days, frozen and then extracted using ABN column (30mg/mL; Biotage) with the following SPE method:
       Conditioning: 1 mL methanol
       Equilibration: 1 mL water, 0.1% formic acid
       Load: 10 mL filtrate
       Wash: 1 mL 95:5 water/methanol
       Elution: 1 mL 95:5 methanol/water
       Eluent (i.e. extracts) collected, dried under vacuum

Extracts profiled using Agilent HPLC 1260 coupled to time of flight mass spectrometer with ESI source (Agilent Technologies, ToF LC/MS 6230). All samples run in positive ion mode.

Samples re-dissolved in 20 uL of methanol then diluted 1:10 in methanol. For each sample, 15 uL were injected. HPLC solvents were solvent A: acetonitrile 0.1% formic acid and Solvent B: water 0.1% formic acid.  The following was used as the HPLC solvent scheme: 0-5.0 min hold at 5% A, 95% B; 5.1-10.0 min ramp to 40% A, 60% B; 10.1-12.00 min hold at 40% A, 60% B;12.1-17.0 min ramp to 95% A, 5% B; 17.1-24.0 min hold at 95% A, 5 % B; 24-24.1 drop to 5% A, 95% B; 24.1-28.0 min hold at 5% A; 95% B.

Column: Phenomenex C18, 150x4.6mm; 2.6 um particle size; column temperature held at 40 *C during run.

Mass spectrometer settings:
Gas Temp: 350 *C
Drying gas: 8 L/min
Nebulizer: 40 psig
Sheath gas: 350 *C
Sheath gas flow: 10 L/min
Capillary voltage: 3500V
Nozzle voltage: 1000V
Acquisition: 20- 3000 m/z
Reference mass calibrant used: 121.0509 m/z and 922.0098 m/z

Files are available at this ftp site: ftp://ftp.whoi.edu/pub/science/MCG/gbmf/Mincer/C10_September2013

Originally published: April 24, 2014

Last updated: April 24, 2014
 


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