Determination of Six Selenium Species in Ganoderma Lucidum by HPLC-ICP-MS After Microwave-Assisted Extraction with Enzyme

0.5 mL of Ganoderma lucidum sample was mixed with 10 mL of water and 20 mg of protease XIV, and 6 selenium species in the sample, including selenic acidSe(Ⅵ), seleniteSe(Ⅳ), selenomethionine (SeMet), selenocystine (SeCys₂), selenoethionine (SeEt) and methylselenose semi Cystine (SeMeCys), were extracted at 37℃ and the microwave power of 300 W for 30 min. After the extract was centrifuged for 10 min, the supernatant was passed through a 0.22 μm filtering membrane and the filtrate was analyzed by HPLC-ICP-MS. The Agilent ZORBAX SB-Aq C₁₈ chromatographic column was selected as the stationary phase, and the 20 mmol·L^-1 citric acid solution (pH 4.5) containing 5 mmol·L^-1 sodium heptane sulfonate was used as the mobile phase for isocratic elution. Under optimized instrument conditions, retention times of the six selenium species were found to be 3.21, 4.06, 6.41, 4.38, 7.38, 6.05 min, respectively. 3 mL of commercially available hydrogen peroxide solution and 5 mL of nitric acid were mixed with 0.2 g of Ganoderma lucidum sample,and the mixture solution was digested under 1 200 W microwave power at 160℃ for 30 min. The volume of digestion solution was made up to 25.0 mL with 5% (volume fraction) hydrochloric acid solution, and then the total selenium in the above solution was determined by ICP-MS with the detected isotope of ⁸²Se. The results showed that the mass concentrations of the 6 selenium species were linearly related to their corresponding peak areas in the range of 0.5-100.0 μg·L^-1, with detection limits (3S/N) in the range of 0.03-0.15 μg·kg^-1. The recovery test was made by addition of standard solutions to the Ganoderma samples at three concentration levels, giving recoveries ranged from 91.7% to 98.7%, with RSDs (n=6) of the measured values in the range of 0.031% to 3.2%. The prescribed method was used to analyze the selenium species of commercially available Ganoderma lucidum, Ganoderma sinense, stalkless Ganoderma sinense and selenium-enriched Ganoderma lucidum, and the highest selenium content and the most complete species were found in the selenium-enriched Ganoderma lucidum of the 4 samples.