3 4 5 thba

2,4,6-THBA, 3,4-DHBA, 3,4,5-THBA, 4-HBA and trypsin-EDTA were obtained from Sigma Aldrich (St. Louis, MO, USA); H1 Histones and Immobilon membranes from EMD Millipore (Billerica, MA, USA); ³²P γ-ATP from MP Biochemicals (Solon, OH, USA); RT-PCR reagents from New England Biolabs (NEB, Ipswich, MA, USA); qPCR reagents from Applied Biosystems (Foster City, CA, USA); Annexin V/7-AAD kit from Beckman Coulter (Miami, FL, USA); Super Signal™ West Pico Chemiluminescent Substrate, protease inhibitor tablets and all other chemicals were obtained from Thermo Fisher Scientific, Inc. (Waltham, MA, USA).

Dried G. japonicum (300 g) was extracted with 70% ethanol (3000 mL) at 80 °C for 6 h. The extract was filtered and lyophilized to dry powder. The yield of G. japonicum extract was 9.1%. The analysis of 3, 4, 5-trihydroxybenzaldehyde (3, 4, 5-THBA)(Sigma-Aldrich, MO, USA), an active or a standard compound [16 (link)] in G. japonicum extract, was performed on a high-performance liquid chromatography system (Jasco, Hachioji, Tokyo, Japan) equipped with a PU-980 pump, AS-950-10 autosampler, and MD-2010 PDA detector. An analytical chromatogram was obtained at 30 °C on a Waters Sunfire™ C18 5 μm column (4.6 mm × 250 mm) through gradient elution using a mobile phase composed of water, 0.2% (v/v) formic acid, and acetonitrile/MeOH (60:40, v/v). We performed gradient separation using 0.2% formic acid. The gradient was decreased by 90% from 0 to 10 min, 77% from 10 to 15 min, 40% from 15 to 35 min, and was increased by 90% from 35 to 40 min. The run time and flow rate were set at 40 min and 1.0 mL/min, respectively, while the samples were detected at 330 nm. The concentration of 3, 4, 5-THBA in extract powder was found to be 3.5 ± 0.12 mg/g using the peak area in the standard chromatogram (Figure 1).