Ec 150 3 nucleodur 100 3 c18ec column

LC-MS/MS analysis was performed on a Finnigan LCQ-Duo ion trap mass spectrometer with an ESI source (ThermoQuest, Hyland Scientific, Stanwood, WA, USA), coupled to a Thermo Scientific Accela HPLC system (MS pump plus, autosampler, and PDA detector plus) with an EC 150/3 Nucleodur 100-3 C18ec column (Macherey-Nagel, Dueren, Germany). A gradient of water and acetonitrile (ACN) with 0.1% formic acid each was applied at 30°C as follows: 0–15% ACN in 15 min; 15% ACN for 10 min; 15–35% ACN in 20 min; 35% ACN for 10 min; 35–50% ACN in 20 min; 50–100% ACN in 10 min; and 100% ACN for 15 min. The flow rate was 0.5 mL/min. The injection volume was 20 μL. All samples were measured in the positive and negative mode. The MS was operated with a capillary voltage of 10 V, source temperature of 240°C, and high purity nitrogen as a sheath and auxiliary gas at a flow rate of 80 and 40 (arbitrary units), respectively. The ions were detected in a mass range of 50-2000 m/z. Collision energy of 35% was used in MS/MS for fragmentation. Data acquisitions and analyses were executed by the Xcalibur™ 2.0.7 software (Thermo Scientific, Waltham, MA, USA).

HPLC-PDA-ESI-MS/MS analysis was carried out on a Finnigan LCQ-Duo ion trap mass spectrometer (Thermo Quest, San Jose, CA, USA) with an ESI source coupled to a Finnigan Surveyor HPLC system (Accela autosampler, MS pump plus, and PDA detector plus) (Thermo, San Jose, CA, USA) with an EC 150/3 Nucleodur 100-3 C18ec column (Macherey-Nagel, Düren, Germany). A gradient of water and acetonitrile (ACN) (with 0.1% formic acid for ESI⁺ mode and without formic acid for ESI⁻ mode) was applied from 2% to 100% ACN in 60 min at 30 °C at a flow rate of 0.5 mL/min. The injection volume was 20 µL. All samples were measured in the positive and negative ion mode. The MS was operated at +10 V capillary voltage for ESI⁺ and −10 V for ESI⁻, 240 °C source temperature, and high purity nitrogen as a sheath and auxiliary gas at a flow rate of 80 and 40 (arbitrary units), respectively. The ions were detected in a mass range of 50–2000 m/z. A collision energy of 35 eV was used in MS/MS fragmentation. Data acquisitions and analyses were executed by Xcalibur^TM 2.0.7 software (Thermo Scientific, Karlsruhe, Germany). Tentative metabolite assignments were achieved by comparison of mass and UV spectral data of the detected compounds in both negative and positive ionization modes with reported data alongside online public databases where references are added Table 2.

LC-MS/MS analysis was done on a Finnigan LCQ-Duo ion trap mass spectrometer with an ESI source (Thermo Quest, Austin, TX, USA) coupled to a Finnigan Surveyor HPLC system (MS pump plus, autosampler, and PDA detector plus) with an EC 150/3 NUCLEODUR 100-3 C18ec column (Macherey-Nagel, Düren, Germany).
A gradient of water and acetonitrile (ACN), without acid, was applied from 5% to 60% ACN in 45 min at 30 °C. Flow rate was 0.5 ml/min. Injection volume was about 20 µl. MS was operated in the negative mode with a capillary voltage of −10 V, source temperature of 220 °C, and high purity nitrogen as a sheath and auxiliary gas at a flow rate of 80 and 40 (arbitrary units), respectively. Ions were detected in a mass range of 50–2,000 m/z. Collision energy of 35% was used in MS/MS for fragmentation. Data acquisitions and analyses were done using Xcalibur™ 2.0.7 software (Thermo Fisher Scientific, Waltham, MA, USA).