Agilent 1100 series lc msd trap xct

MK₈(H₂) (10 mg/L, 0.1 mL in acetone) was mixed with 0.5 mL of MGO (10 mmol/L, in 0.01 mol/L PBS, pH 7.4) and then incubated at 37 °C for 6 h. MK₈(H₂) with the PBS buffer solution at the same volume was served as a control. After incubation, 0.5 mL of o-phenylenediamine (50 mmol/L in acetone) was added to terminate the reaction. The HPLC analysis was performed to evaluate the reaction process of MK₈(H₂) and MGO. The HPLC analysis of MGO before and after reaction was performed according to the published article using a C18 column [33 (link)]. The HPLC analysis of MK₈(H₂) before and after reaction was performed according to the published article using a C30 column [20 (link)].
To analyze the reaction product of MK₈(H₂) and MGO, Agilent 1100 Series LC-MSD-Trap-XCT and GS-120-5-C18-A column (250 × 4.6, 5 μm) were used here. The elution program was based on the published article [33 (link)]. Injection Volume was set at 40 μL. The column temperature was set at 25 °C and the samples were detected at 315 nm. MS conditions: APCI ion source, positive ion mode, the ion source temperature: 280 °C, capillary voltage: 1500 V, drying air-flow rate: 5 L/min, atomizing chamber temperature: 280 °C, atomizing chamber voltage: 60 psi, the scanning range of mass-to-charge ratio: 30–2000.

In-gel digestion by trypsin and the following LC-MS/MS analysis using the Agilent 1100 series LC/MSD Trap XCT (Agilent Technologies, Palo Alto, CA) were performed as described previously [46] (link). Proteins were identified by the Spectrum Mill MS Proteomics Workbench against the Swiss-Prot protein database search engine, and the NCBI protein database search engine. We also manually incorporated protein information from seven H. pylori-specific genome databases (five Japanese strains 35A, F16, F30, F32, F52, a Korean strain Hp51, and an Amerindian strain Shi470) that were downloaded from NCBI Genome site of Helicobacter pylori.