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Extraction for LC-q-TOF-MS to detect secon … Extraction for LC-q-TOF-MS to detect secondary metabolites<br />
Each frozen sample was extracted with5 fold amount of solvent (methanol/water [8:2 v/v]) containing reference compounds (0.5 mg/L of lidocaine ([M+H]+, m/z 235.1804) and 10-camphorsulfonic acid ([M-H]-, m/z 231.0691) using a mixer mill MM301 (Retsch) at a frequency of 20 Hz for 5 min at 4°C. After centrifugation for 10 min at 15,000 × g, the supernatant was transferred into a 2 ml tube. Aliquot of the extracts was filtered using an Oasis® HLB μelusion plate (30 μm, Waters Co., Massachusetts, US). The extracts were transferred into a 2 ml tube.<br />
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LC-q-TOF-MS conditions to detect secondary metabolites<br />
After preparation of the extracts, the sample extracts (1 μl) were analyzed using an LC-MS system equipped with an electrospray ionization (ESI) interface (LC, Waters Acquity UPLC system; MS, Waters Xevo G2 Q-Tof). The analytical conditions were as follows. LC: column, Acquity bridged ethyl hybrid (BEH) C18 (pore size 1.7 μm, length 2.1× 100 mm, Waters); solvent system,acetonitrile(0.1% formic acid):water (0.1% formic acid); gradient program,
1 : 99 v/v at 0 min, 1 : 99 v/v at 0.1 min, 99.5 : 0.5 at 15.5 min,99.5 : 0.5 at 17.0 min, 1 : 99 v/v at 17.1 min and 1 : 99 at 20 min, flow rate, 0.3 ml/min,temperature, 40°C; MS detection: capillary voltage, +3.0 keV, cone voltage, 25.0 V, source temperature, 120°C, desolvation temperature, 450°C, cone gas flow, 50 l per h; desolvation gas flow, 800 l per h; collision energy, 6 V; mass range, m/z 100‒1500; scan duration, 0.1 sec; interscan delay, 0.014 sec; mode, centroid; polarity, positive; Lockspray (Leucineenkephalin): scan duration, 1.0 sec; interscan delay, 0.1 sec. The data were recorded using MassLynx version 4.1 software (Waters). ng MassLynx version 4.1 software (Waters).
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