Validation of a gas chromatography-electron capture detection of T-2 and HT-2 toxins in Chinese herbal medicines and related products after immunoaffinity column clean-up and pre-column derivatization

摘要

A sensitive, reproducible and accurate gas chromatography-electron capture detection (GC-ECD) method was developed for simultaneous determination of T-2 and HT-2 toxins in Chinese herbal medicines (CHMs) and related products. Then, gas chromatography-mass spectrometry (GC-MS) was applied to confirm the positive results and interfering peaks in some samples. All samples were injected into the Agilent GC-6890N gas system (Hewlett-Packard, Palo Alto, CA, USA), equipped with a 63Ni electron capture detector on a DB-1701 capillary column (30 m × 0.25 mm I.D., 0.25 urn film thickness, Agilent Technologies). The positive samples and interfering peaks in some samples were confirmed by GC-MS on a Varian 300 TQ Mass instrument (Agilent Varian Technologies, USA) coupled with a HP-5MS fused silica capillary column (30 m × 0.25 mm I.D., 0.25 μm film thickness, Agilent Technologies). The extraction solvent, clean-up column and pre-column derivatization conditions for T-2 and HT-2 toxins in all samples were optimized. Then, the established GC-ECD method used for T-2 and HT-2 toxins was validated for linearity, precision, selectivity and recovery. The optimized sampling methods were that extraction of samples with methanol/water (90:10, v/v), followed by immunoaffinity column (LAC) clean-up and pre-column derivatization with N-heptafluoro-butyryl imidazole (HFBI). And the optimized GC-ECD method was as follows: one microliter of the sample solution was injected into the chromatographic apparatus for analysis, the injector was operated in the splitless mode, the injector and detector temperatures were 270 °C and 300 °C, respectively, highpurity (over 99.99%) nitrogen at a flow-rate of 1.0 mL/min was used as the carrier gas, the column temperature was programmed as: 80 °C (hold for 2min), 10 °C/min to 200 °C, 5 °C /min to 220 °C, 2 °C/min to 230 °C (hold for 13 min), 3 °C/min to 270 °C, and 5 ° C/min up to 280 °C (hold for 5 min). The results showed that the limits of detection (LOD) of the validated GC-ECD method for T-2 and HT-2 toxins were 1.88 and 0.47 ng/g, respectively, and the recoveries for different CHMs spiked with 50, 100 and 1000 ng toxin/g ranged from 89.2% to 99.1% with relative standard deviations (RSD) < 6.0% for T-2 and from 85.9% to 99.0% with RSD < 8.8% for HT-2 toxin, respectively. The validated method was successfully applied for the determination of T-2 and HT-2 toxins in 89 Chinese herbal medicines and 10 related products including granula, pills and pulvises from different sources, where it was found that T-2 and HT-2 toxins were not detected (> LOD) in any of the tested samples. These results were reliable by confirmation using GC-MS. Some unknown peaks in Rhizoma Arisaematis and Raidix Scrophulariae were interfering peaks not the target toxins. This is a successful report for analyzing T-2 and HT2 toxins in CHMs and related products by GC-ECD. Future study would focus on the determination of more trichothecenes or mycotoxins in CHMs and related products to avoid health problems of these toxic materials in consumers.