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Harvested sample is frozen by liquid N2 an … Harvested sample is frozen by liquid N2 and resulting powder (100mg) are solved in 300uL methanol solution. 20uL sample is injected into HPLC after 0.2um membrane filter treatment. HPLC conditions: Agilent 1100 series (Agilent), Column: TSKgel-100V (4.6 x 250 mm, 5 micrometer; TOSOH), Solvent: A; 0.1% formic acid aq. B; ACN (addition 0.1% formic acid fc.), Gradient: (B);3% (0.0 to 1.0 min), 3 to 30% (1.0 to 26.0 min), 50 to 90% (26.0 to 41.0 min), 90% to 100% (41.1 to 46.0 min), 100% (46.1 to 51.0 min), 3% (51.1 to 58.0 min), Column temp.: 40 degree C, Flow rate=0.5mL/min, PDA: 200-650 nm (2 nm step). FT-ICR-MS conditions: Filter 1: FTMS + c norm !corona !pi res=100000 o(150.0-2000.0); 2: ITMS + c norm !corona !pi Dep MS/MS Most intense ion from (1); 3: ITMS + c norm !corona !pi Dep MS/MS 2nd most intense ion from (1); 4: ITMS + c norm !corona !pi Dep MS/MS 3rd most intense ion from (1). i Dep MS/MS 3rd most intense ion from (1).
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