Hp 5ms quartz capillary column

NaBH₄ and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) was used to carboxyl-reduce LAP-1 according to the methods described in the previous literature.^{13 (link)} The complete methylation was verified by the FT-IR spectrum with O–H absorption band disappearance.
Then, formic acid (88%) and trifluoroacetic acid were used to hydrolyze the methylated samples for 6 h and 8 h at 100 °C, separately. After being reduced by NaBH₄ and acetylated, the products were determined on a gas chromatography-mass spectrometer (6890N-5975B, Agilent, USA) that was equipped with an HP-5MS quartz capillary column (30 m × 0.25 mm × 0.25 μm). Temperature rising program: 1 °C min⁻¹, 160–180 °C, 3 °C min⁻¹ to 220 °C, held for 10 min. The mass charge ratio range was set at 25–500.

QL-9a was inoculated in MSM medium containing OHHL at a concentration of 0.5 mmol·L⁻¹, incubated at 30 °C and 200 rpm, and sampled at 12 h intervals, with three samples collected simultaneously from each group. The sample processing method is mentioned in Section 3.5. and was subjected to gas chromatography–mass spectrometry (GC–MS) (Agilent 6890 N/5975, Santa Clara, CA, USA) for degradation product detection. The following were the detection requirements: an Agilent HP-5MS quartz capillary column (30 m × 250 μm × 0.25 μm) with high-purity helium as the carrier gas, flowing at a rate of 1.0 mL/min. The temperature increased from 200 °C to 280 °C at a rate of 25 °C per minute. Electron energy of 70 eV, a split injection volume of 1 μL, and a split ratio of 5:1 was used. The quadrupole was heated to 150 °C, while the ion source was heated to 230 °C [21 (link)].

Headspace SPME was employed to collect the volatile compounds from flower tissues, which were absorbed by a 75 μm CAR/PDMS fiber (Sigma-Aldrich) for 2 h at 25°C. Total trapped volatile compounds were subsequently thermally desorbed and transferred to an Agilent 5975-6890N gas chromatography-mass spectrometry (GC-MS) apparatus (Agilent Technologies) equipped with HP5-MS quartz capillary column (250 μm diameter, 60 m length, and 0.25 μm film thickness). The instrument used for the gas chromatography–mass spectrometry analysis was an Agilent 7890B-7000B triple quadrupole gas-mass spectrometer. The carrier gas was helium with 1 ml/min of flow rate. The temperature of the electron ionization (EI) ion source is 230°C, and the electron energy was 70 eV. The temperature of the quadrupole is 150°C for 280°C of interface temperature, and the mass scanning range was 50–400 amu. The temperature program was isothermal at 60°C for 3 min, then increased at a rate of 5°C min^–1 to 300°C, and was then mainteined at 300°C for 5 min.
Through the NIST (National Institute of Standards and Technology) 2011 standard library, the volatile compounds detected during the experiment were identified and qualitatively analyzed. The obtained compounds were compared with the literature to obtain the determined volatile terpenoids.

An Agilent 7820A GC system equipped with automated sample injector model 7693A and MS detector 5977B (Agilent Technologies, Waldbronn, Germany) was used for the GC experiments. The GC apparatus was equipped with a split/splitless inlet, working in a split ratio of 10:1 mode to a 60 m × 0.25 mm HP-5MS quartz capillary column with a 0.25 µm film thickness (Agilent Technologies, Waldbronn, Germany). Data acquisition and analysis were performed using a MassHunter 5977B MSD Bundle with 7820 GC software NIST MS Spectral Library version 2.3.
For sample shaking, Multi-Speed Vortex MSV-3500 (Biosan, Riga, Latvia) was used. During the study, a Mikro 220R centrifuge with fast cool function (Hettich Zentrifugen, Tuttlingen, Germany), and a FiveEasy F-20 pH-meter (Mettler Toledo, Greifensee, Switzerland) were also used. Samples were stored in an ultra-low-temperature freezer (Panasonic Healthcare Co., Ltd., Sakata, Japan). Water was purified using a Milli-QRG system (Millipore, Vienna, Austria).

The methylation analysis of PSP2-1 was determined based on the previously reported method [57 (link)]. A total of 5 mg of the polysaccharide sample PSP2-1 was dissolved in dimethyl sulfoxide, and 20 mg of sodium hydroxide was added for 1 h. Then, 10 μL iodomethane (CH₃I) and 500 μL dichloromethane were added for 20 min, respectively, after being centrifuged, discarded after the aqueous phase, and the dichloromethane was evaporated. Furthermore, the sample was hydrolyzed with 2.5 mol/L trifluoroacetic acid at 121 °C for 90 min, Then, 50 μL of 2 mol/L ammonia and 50 μL of 1 mol/L NaBD₄ was added for 2.5 h at room temperature, and acetic acid was added to terminate the reaction. Lastly, the sample was acetylated by adding 250 μL of acetic anhydride for 2.5 h at 100 °C, and the methylated alditol acetate derivatives were analyzed with an HP-5 ms quartz capillary column (30 m × 0.25 mm× 0.25 μm, Agilent Technology, USA), which was connected to the GC-MS system (GC7890A-MS5975C, Agilent Technology, USA).