This study established a validated analytical method for the first time on the determination of nitrofuran metabolites, including semicarbazide (SEM), 1-aminohydantoin (AHD), 3-amino-2-oxazolidinone (AOZ) and 3-amino-5-morpholinomethyl-2-oxazolinone (AMOZ) in gelatin Chinese medicine. A C18 column with the mobile phase consisting of acetonitrile and 5 mmol/L ammonium acetate in water was used to separate these nitrofuran metabolites. The limit of detection of SEM, AHD, AOZ and AMOZ were found to be 0.2 µg/kg, 0.3 µg/kg, 0.2 µg/kg and 0.2 µg/kg, whereas their limit of quantification were 0.6 µg/kg, 0.8 µg/kg, 0.6 µg/kg and 0.5 µg/kg. These nitrofuran metabolites exhibited a good linear standard curve (regression coefficients above 0.99) with a concentration range of 2 µg/L to 100 µg/L. Regarding extraction procedure, gelatin Chinese medicine was pre-treated with pepsin and then extracted using 5% formic acid (v/v) in acetonitrile. The resultant extract was purified through dispersive solid phase extraction using 1000 mg anhydrous sodium sulfate, 300 mg octadecyl carbon silica gel sorbent absorbent and 500 mg ethylenediamine-N-propyl carbon silica gel absorbent, and then further purified on Oasis PRiME HLB cartridges. The matrix effect was effectively eliminated after the clean-up procedure as confirmed by comparing the ratio of standard curves prepared by standards dissolved in both matrix solvent and 5 mmol/L ammonium acetate in water: acetonitrile (95:5, v/v). The recoveries of these nitrofuran metabolites under the 1 µg/kg, 2 µg/kg and 10 µg/kg spiking levels were between 77.4% and 95.6%. These metabolites after the extraction were stable at 4 °C for 24 h. The validated method was used to analyze the residue level of these nitrofuran metabolites in 25 gelatin Chinese medicines. Results showed that only one Colla Corii Asini sample contained SEM (2.52 µg/kg) and AOZ (6.27 µg/kg), whereas one Testudinis Carapacis et Plastri sample had SEM (1.27 µg/kg) and AMOZ (9.53 µg/kg).

中文翻译：

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分散固相萃取和直通固相萃取与超高效液相色谱-串联质谱联用测定明胶中药中的四种硝基呋喃代谢物。

这项研究首次建立了一种有效的分析方法，用于测定硝基呋喃代谢物，包括氨基脲（SEM），1-氨基乙内酰脲（AHD），3-氨基-2-恶唑烷酮（AOZ）和3-氨基-5-吗啉代甲基-明胶中药中的2-恶唑啉酮（AMOZ）。使用流动相由乙腈和5 mmol / L乙酸铵在水中组成的C18柱分离这些硝基呋喃代谢物。SEM，AHD，AOZ和AMOZ的检出限分别为0.2 µg / kg，0.3 µg / kg，0.2 µg / kg和0.2 µg / kg，而其定量限分别为0.6 µg / kg，0.8 µg / kg。 kg，0.6 µg / kg和0.5 µg / kg。这些硝基呋喃代谢物的浓度范围为2 µg / L至100 µg / L，显示出良好的线性标准曲线（回归系数大于0.99）。关于提取程序，明胶中药先用胃蛋白酶预处理，然后用5％甲酸（v / v）的乙腈萃取。使用1000 mg无水硫酸钠，300 mg十八烷基碳硅胶吸附剂和500 mg乙二胺-N-丙基碳硅胶吸附剂，通过分散固相萃取法纯化所得提取物，然后在Oasis PRiME HLB柱上进一步纯化。通过比较溶解在基质溶剂和5 mmol / L乙酸铵中的标准品在水：乙腈（95：5，v / v）中制备的标准曲线的比例，证实了净化程序后基质效应被有效消除。在1 µg / kg，2 µg / kg和10 µg / kg峰值水平下，这些硝基呋喃代谢物的回收率在77.4％至95.6％之间。提取后这些代谢产物在4°C下稳定24 h。经验证的方法用于分析25种明胶中药中这些硝基呋喃代谢物的残留量。结果表明，只有一个Colla Corii Asini样品含有SEM（2.52 µg / kg）和AOZ（6.27 µg / kg），而一个Testudinis Carapacis et Plastri样品的SEM（1.27 µg / kg）和AMOZ（9.53 µg / kg）。