Goenaga Infante Heidi, Ovejero Bendito María del Carmen, Cámara Carmen, Evans Linda, Hearn Ruth, Moesgaard Sven
LGC Limited, Queens Road, Teddington, Middlesex, TW11 OLY, UK.
Anal Bioanal Chem. 2008 Apr;390(8):2099-106. doi: 10.1007/s00216-008-1979-y. Epub 2008 Mar 5.
A method for the accurate determination of ultratrace selenium species of relevance to cancer research, such as gamma-glutamyl-Se-methylselenocysteine (gamma-glutamyl-SeMC), using species-specific double isotope dilution analysis (IDA) with HPLC-ICP-MS is reported for the first time. The (77)Se-enriched gamma-glutamyl-SeMC spike was produced in-house by collecting the fraction at the retention time of the gamma-glutamyl-SeMC peak from a chromatographed aqueous extract of (77)Se-enriched yeast, pooling the collected fractions and freeze-drying the homogenate. The Se content of this spike was characterised using reverse isotope dilution mass spectrometry (IDMS) and the isotopic composition of this spike was checked prior to quantification of the natural abundance dipeptide species in garlic using speciated IDA. The extraction of the gamma-glutamyl-SeMC species in water was performed in a sonication bath for 2 h after adding an appropriate quantity of (77)Se-enriched gamma-glutamyl-SeMC to 50 mg of garlic to give optimal (78)Se/(77)Se and (82)Se/(77)Se ratios of 1.5 and 0.6, respectively. The effect of ultrasonic nebulisation, in comparison with the loading of the ICP with carbon (through the addition of methane gas on-line), on the detection of Se associated with gamma-glutamyl-SeMC using collision/reaction cell ICP-MS with hydrogen as collision gas was investigated. Sensitivity enhancements of approximately fourfold and twofold were achieved using USN and methane mixed plasma, respectively, in comparison with conventional nebulisation and conventional Ar ICP-MS. However, an approximately twofold improvement in the detection limit was achieved using both approaches (42 ng kg(-1) for (78)Se using peak height measurements). The use of species-specific IDMS enabled quantification of the dipeptide species at ng g(-1) levels (603 ng g(-1) Se) in the complex food matrix with a relative standard deviation (RSD, n = 4) of 4.5%, which was approximately half that obtained using standard addition as a confirmatory technique. Furthermore, good agreement was found between the gamma-glutamyl-SeMC species concentrations obtained using both calibration methods.
首次报道了一种使用高效液相色谱 - 电感耦合等离子体质谱联用的物种特异性双同位素稀释分析（IDA）准确测定与癌症研究相关的超痕量硒物种的方法，例如γ-谷氨酰 - 硒甲基硒代半胱氨酸（γ-谷氨酰 - SeMC）。通过从富含(^{77}Se)酵母的色谱水提取物中在γ-谷氨酰 - SeMC峰的保留时间收集馏分、合并收集的馏分并冻干匀浆，在内部制备富含(^{77}Se)的γ-谷氨酰 - SeMC加标物。使用反向同位素稀释质谱法（IDMS）对该加标物的硒含量进行表征，并在使用特定物种IDA对大蒜中天然丰度二肽物种进行定量之前检查该加标物的同位素组成。在向50 mg大蒜中加入适量富含(^{77}Se)的γ-谷氨酰 - SeMC后，在超声浴中进行2 h水相中γ-谷氨酰 - SeMC物种的萃取，以使(^{78}Se/^{77}Se)和(^{82}Se/^{77}Se)的最佳比例分别为1.5和0.6。研究了与通过在线添加甲烷气体向ICP中加载碳相比，超声雾化对使用以氢气作为碰撞气体的碰撞/反应池ICP-MS检测与γ-谷氨酰 - SeMC相关的硒的影响。与传统雾化和传统氩ICP-MS相比，使用超声雾化（USN）和甲烷混合等离子体分别实现了约四倍和两倍的灵敏度增强。然而，两种方法均实现了约两倍的检测限改善（使用峰高测量时(^{78}Se)的检测限为42 ng kg(^{-1})）。使用物种特异性IDMS能够在复杂食品基质中以ng g(^{-1})水平（603 ng g(^{-1})硒）对二肽物种进行定量，相对标准偏差（RSD，n = 4）为4.5%，这大约是使用标准加入作为确证技术所获得值的一半。此外，两种校准方法获得的γ-谷氨酰 - SeMC物种浓度之间具有良好的一致性。
