Hp 5975c

Freeze-dried soil (Christ-Alpha 1–4 LD plus) was subjected to ultrasonic extraction with a 30 mL mixture of n-hexane and acetone (2:1, v/v). The contents of Phe, Pyr and Bap in soil were quantitatively analyzed by the internal standard method [22 (link)]. Determination of PAHs content by Agilent HP 7890 gas chromatograph combined with Agilent HP 5975C inert mass selective detector (7890/5975C).

As described previously^{4 (link)}, for each sample, a known quantity of trophallactic fluid was collected into a graduated glass capillary tube and blown into an individual glass vial containing 5 µl of 100% ethanol. This biological sample was added to a 1:1 mixture of isooctane and methanol, processed and stored at –80 °C until analysis. Before analysis, 50% acetonitrile (HPLC grade) was added. Prior to purification, farnesol (Sigma-Aldrich, St Louis, MO) was added to each sample to serve as an internal standard. Briefly, samples were derivatized in a solution of methyl-d alcohol (Sigma-Aldrich, St Louis, MO) and trifluoroacetic acid (Sigma-Aldrich) then analyzed using an HP 7890 A Series GC (Agilent Technologies, Santa Clara, CA) coupled to an HP 5975 C inert mass selective detector monitoring at m/z 76 and 225 to ensure specificity for the d3-methoxyhydrin derivative of JH III. Total abundance was quantified against a standard curve of derivatized JH III, and adjusted for the starting volume of trophallactic fluid. The detection limit of the assay is approximately one pg.

A divinylbenzene/carboxen/polydimethylsiloxane-coated fiber (DVB/CAR/PDMS 50/30 μm, Supelco, Bellefonte, PA, USA) was used for SPME. The GC-MS analysis was performed on an HP 7890A series II GC (Agilent Technologies, Wilmington, DE, USA) coupled to a mass spectrometer (HP 5975C, Agilent Technologies). Helium (99.999%) was used as the carrier gas with a column flow rate of 1.9 mL min⁻¹, and the capillary column used was HP-5 (50 m × 0.32 mm inner diameter, 0.52 μm film thickness, Agilent Technologies). An MPS2 autosampler (Gerstel, Linthicum, MD, USA) was used for automatic sample feeding. The injection of the sample and the data reading were according to [21 (link)]. The presumptive identification of volatile compounds was achieved by comparing the GC retention times and mass spectra with the data system library (NIST, 2005 software, Mass Spectral Search Program V.2.0d; NIST 2005, Washington, DC, USA).