Agilent 7890b gas chromatograph with 5977a inert mass selective detector

Leaves collected from different individual trees of C. burmannii were ground into fine powder in liquid nitrogen. Powder samples (0.05–0.10 g) were weighed accurately, soaked in 2.0 mL petroleum ether using an ultrasonic cleaner for 30 min, and centrifuged at 16,000 rpm for 5 min in preparation for GC/MS analysis. Three technical replicates were performed for each of the three biological samples. An Agilent 7890B Gas Chromatograph with 5977A Inert Mass Selective Detector (Agilent, USA) was used to analyze the extraction. Helium was used as the carrier gas with a flow rate of 1 mL/min. A Cyclosil-B GC column (30 m × 0.25 mm × 0.25 µm) with initial temperature of 50 °C was used to separate samples. The GC column temperature program was as follows: oven temperature was increased from 50 °C to 180 °C at 2 °C/min, and then from 180 °C to 300 °C at 4 °C/min, then held at 300 °C for 4 min. Injector: split mode with split ratio of 20:1, injection volume of 1.0 µL and inlet temperature of 250 °C. D-Borneol was confirmed by comparing its retention time with that of the known standard, which was determined under the same conditions. The concentration of D-borneol in the plant sample was calculated from its respective standard curves.

200 mg leaf tissues were ground frizzed in liquid nitrogen, 1.5 mL hexane was added into centrifuge tubes, ultrasonic for 30 min with 60 Hz and then heated under a 56 °C water bath for 1 h. After centrifugation, the supernatant was taken and passed through a 0.22-µm organic membrane as the test solution for GC-MS analysis using Agilent 7890B Gas Chromatograph with 5977A inert Mass Selective Detector (Agilent, California, USA). The gas chromatograph was equipped with an HP-5MS capillary column (30 m × 250 mm × 0.25 mm). The instrument was set to an initial temperature of 50 °C and maintained for 0 min. Then, the oven temperature was increased to 130 °C at a rate of 20 °C/min and then increased to 150 °C at a rate of 2 °C/min. The temperature was maintained at 150 °C for 5 min and later increased to 230 °C at a rate of 20 °C/min. The injection volume was 1 µL, and the injection port temperature was 230 °C. In addition, patchouli alcohol standards were purchased from NanTong FeiYu, China. All reagents used were analytical grade.