Determination of 3 Limonin Analogues in Different Parts of Citrus Fruits of Different Varieties by Liquid Chromatography-Tandem Mass Spectrometry with Dispersed Solid Phase Extraction

The method of liquid chromatography-tandem mass spectrometry with dispersion solid phase extraction was proposed to simultaneously determine limonin, normillin, and phellosterone in 4 parts (oil cell layer, white layer, capsule, and pulp) of citrus fruits of different varieties. The 5.0 g of homogenized sample was taken, and 5 mL of water was added. The mixture was mixed well by vortex, and 20 mL of mixed solution composed of acetone and acetonitrile at volume ratio of 5∶5 was added. After mixing, ultrasonic extraction was made for 10 min, and centrifugation was made for 5 min. The 2 g of EMR-Lipid adsorbent was activated by 3 mL of water, which was added into the above supernatant. The mixture was vortexed for 30 s, ang 5 g of sodium chloride and magnesium sulfate at mass ratio of 1∶4 and 15 mg of graphitized carbon black were added. After quickly shaking well, the mixture was centrifuged for 1 min, and all the supernatant was collected, and blown to near dryness by nitrogen at 40 ℃. The residue was dissolved in 10.0 mL of mixed solution consisted of 0.1% (volume fraction, the same below) formic acid solution and acetonitrile at volume ratio of 9∶1, and the solution was passed through a 0.22 μm filter membrane. The filtrate was determined according to the optimized instrument working conditions. In chromatographic analysis, CAPCELL PAK ADME HR chromatographic column was used as the stationary phase, and mixed solutions composed of 0.1% formic acid solution and acetonitrile at different volume ratios were used as the mobile phase for gradient elution separation. In MS analysis, the modes of electric spray ion source positive ion (ESI⁺) and multiple reaction monitoring (MRM) were used for detection. It was shown that linear relationships between values of the mass concentration and peak area of 3 limonin analogues were kept in the range of 0.10-10.0 mg·L^-1, with detection limits (3S/N) of 0.01 mg·kg^-1. Test for recovery was made according to the standard addition method, giving recoveries in the range of 83.1%-90.5%, and RSDs (n=6) of the determined values ranged from 2.6% to 5.6%. The proposed method was used for the analysis of 30 actual samples. The distribution of limonin analogues in different parts of citrus fruits of different varieties showed significant differences. Among pummelo varieties, the total determined values of limonin analogues were higher and mainly distributed in the capsule part, while other varieties were mainly distributed in the white and oil layers.