Approximately 2.5 g of wet T. hoshinota tissue from each sample was centrifuged at 2000 × g for 30 s to remove excess water. Sponge tissues were placed in mortars and ground with 4 ml of 100% acetone to homogenize the tissues. The homogeneous samples were transferred into a 5 ml centrifuge tube and centrifuged at 2000 × g at room temperature for 30 s. The supernatant was transferred into a new 5‐ml centrifuge tube. The samples were covered with aluminium foil paper for being shaded and stored at 4°C until pigment analysis.
Pigment content analysis was performed by UPLC‐MS. The UPLC‐MS system used the Agilent 1290 Infinity II ultra‐performance liquid chromatography (UPLC) system (Agilent Technologies, Palo Alto, CA, USA) coupled online to the Dual AJS electrospray ionization (ESI) source of an Agilent 6545 quadrupole time‐of‐flight (Q‐TOF) mass spectrometer (Agilent Technologies). The samples were separated using a Kinetex XB‐C18 column (2.6 μm, 4.6 × 100 mm, Phenomenex, Torrance, CA, USA) at 40°C. The chromatogram was acquired; mass spectral peaks were detected and their waveform processed using Agilent Qualitative Analysis 10.0 software (Agilent, USA).
Pigment content analysis was performed by UPLC‐MS. The UPLC‐MS system used the Agilent 1290 Infinity II ultra‐performance liquid chromatography (UPLC) system (Agilent Technologies, Palo Alto, CA, USA) coupled online to the Dual AJS electrospray ionization (ESI) source of an Agilent 6545 quadrupole time‐of‐flight (Q‐TOF) mass spectrometer (Agilent Technologies). The samples were separated using a Kinetex XB‐C18 column (2.6 μm, 4.6 × 100 mm, Phenomenex, Torrance, CA, USA) at 40°C. The chromatogram was acquired; mass spectral peaks were detected and their waveform processed using Agilent Qualitative Analysis 10.0 software (Agilent, USA).