Peng Hanyong, Hu Bin, Liu Qingqing, Yang Zonglin, Lu Xiufen, Huang Rongfu, Li Xing-Fang, Zuidhof Martin J, Le X Chris
J Chromatogr A. 2014 Nov 28;1370:40-9. doi: 10.1016/j.chroma.2014.10.012.
Human exposure to high concentrations of arsenic from water and food is an important health concern. Although 3-nitro-4-hydroxyphenylarsonic acid (Roxarsone) was used for more than 60 years as a feed additive to feed chickens, little is known about the metabolism of this arsenic species in chicken. Determination of potential arsenic metabolites present at trace concentrations is an analytical challenge, requiring efficient separation and sensitive detection. The primary objective of this research is to develop a method that enables the identification and quantification of various arsenic species in chicken liver. This report describes a method of high performance liquid chromatography (HPLC) separation with both inductively coupled plasma mass spectrometry (ICPMS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS) detection. Anion exchange HPLC enabled the separation of Roxarsone and other arsenic species within 12 min. Detection with both ICPMS and ESI-MS/MS allowed for identification and quantification of eight arsenic species in chicken livers, including arsenobetaine, inorganic arsenite, dimethylarsinic acid, monomethylarsonic acid, inorganic arsenate, 3-amino-4-hydroxyphenylarsonic acid, N-acetyl-4-hydroxyphenylarsonic acid (N-AHPAA), and Roxarsone. The concentrations of these arsenic species, with the exception of arsenobetaine, are significantly higher in the Roxarsone-fed chickens than in the control chickens. The simultaneous detection by both ICPMS and ESIMS from the same HPLC separation allowed for comparison of peaks in both ICPMS and ESIMS chromatograms. This is advantageous over two separate analyses, particularly when HPLC retention times might fluctuate due to sample matrix effect. HPLC separation with the complementary atomic and molecular mass spectrometry detection prevented misidentification of co-eluting compounds, as demonstrated by the determination of two possible metabolites of Roxarsone, N-AHPAA and 4-amino-phenylarsonic acid (4-APAA). N-AHPAA was confirmed by HPLC separation with simultaneous arsenic-specific detection by ICPMS and multiple reaction monitoring by ESIMS. Although an arsenic-containing compound had identical retention time as 4-APAA in the HPLC–ICPMS chromatogram, it was ruled out as 4-APAA from the simultaneous detection by ESIMS. The identification and quantitation of trace arsenic species present in complex samples demonstrate the potential of HPLC separation with simultaneous ICPMS and ESIMS detection for other speciation applications.
人类通过水和食物接触高浓度砷是一个重要的健康问题。尽管3-硝基-4-羟基苯胂酸（洛克沙胂）作为鸡饲料添加剂使用了60多年，但人们对这种砷化合物在鸡体内的代谢知之甚少。测定痕量浓度下潜在的砷代谢物是一项分析挑战，需要高效分离和灵敏检测。本研究的主要目的是开发一种能够鉴定和定量鸡肝脏中各种砷化合物的方法。本报告描述了一种采用电感耦合等离子体质谱（ICPMS）和电喷雾电离串联质谱（ESI-MS/MS）检测的高效液相色谱（HPLC）分离方法。阴离子交换HPLC能够在12分钟内分离洛克沙胂和其他砷化合物。通过ICPMS和ESI-MS/MS检测可鉴定和定量鸡肝脏中的八种砷化合物，包括砷甜菜碱、无机亚砷酸盐、二甲基胂酸、一甲基胂酸、无机砷酸盐、3-氨基-4-羟基苯胂酸、N-乙酰-4-羟基苯胂酸（N-AHPAA）和洛克沙胂。除砷甜菜碱外，这些砷化合物在喂食洛克沙胂的鸡中的浓度显著高于对照鸡。通过同一HPLC分离同时进行ICPMS和ESIMS检测，可比较ICPMS和ESIMS色谱图中的峰。这比两次单独分析更具优势，特别是当HPLC保留时间可能因样品基质效应而波动时。HPLC分离结合互补的原子和分子质谱检测可防止共洗脱化合物的错误鉴定，如对洛克沙胂的两种可能代谢物N-AHPAA和4-氨基苯胂酸（4-APAA）的测定所示。通过HPLC分离结合ICPMS的砷特异性检测和ESIMS的多反应监测，证实了N-AHPAA。尽管一种含砷化合物在HPLC–ICPMS色谱图中的保留时间与4-APAA相同，但通过ESIMS同时检测排除了它是4-APAA的可能性。复杂样品中痕量砷化合物的鉴定和定量证明了HPLC分离结合ICPMS和ESIMS同时检测在其他形态分析应用中的潜力。
