Jian-Jun Zhong,Ningbo Liao,Mingfeng He,Yunfeng Pu,Donghong Liu
Urocanic acid (UA), existing in trans - or cis -isoform, is of fairly recent interest to food researchers because of its potential public health hazards of scombrotoxicity and immunotoxicity, as well as associating with fish spoilage. This work is among the first efforts to study the analytical chemistry of UA in fish. With 0.6 M perchloric acid UA was extracted, and co-extracted fish matrix components were efficiently removed through a reactive extraction of UA. The optimum conditions for the reactive extraction, which allowed an 80% recovery of UA, were sample pH adjustment to 9, twice extractions with 32% (w/w) di (2-ethylhexyl) phosphate in hexanol, and a back-extraction with 0.1 M hydrochloric acid at 1:1 phase ratio. A chaotropic hexafluorophosphate salt was added to acidic water-acetonitrile mobile phases to improve the reversed-phase chromatography of UA, which otherwise was poorly retained. Optimum separation conditions were obtained for fish samples and enabled a fast (10 min), convenient-to-use chromatography that clearly outperforms cumbersome legacy ion-pair chromatography. Intended for routine use in our laboratory, the proposed method passed an in-house validation test for linearity, matrix effect (on reactive extraction), accuracy, precision, and detectability.
Jian-Jun Zhong,Liping Xia,Liuru Xue,Boyu Liu,Lingling Zhao,Hua Ye,Zhanming Li
Histamine food poisoning (HFP) or scombrotoxic fish poisoning is a major public health concern associated with seafood and seafood products containing copious amounts of free histidine , which may be decarboxylated into scombrotoxic histamine, the main culprit of HFP. Moreover, the deamination product of the histidine, trans -urocanic acid, is assumed to be scombrotoxic by degranulating mast cells to release endogenous histamine after isomerizing to the cis -form, as well as known to have immunosuppressive effects. In recognition of the potential health risks associated with the histidine degradatives, this work aimed to provide a method for the simultaneous determination of histidine, histamine, and urocanic acid isomers in food, based on precolumn dansylation and subsequent reversed-phase HPLC analysis. The dansylation conditions were optimized for yield with the design of experiments approach consisted of a center point-enhanced 2 4 factorial design followed by sequential steepest ascent searches. Satisfactory chromatographic separation of fish sauce was achieved within 9 min by retention modeling of both the analytes and potentially interfering peaks. The method was fully validated with limits of detection between 0.004 and 0.017 mg 100 g −1 fish sauce, and successfully applied to several fish sauce samples, in which the histamine and urocanic acid levels were found low and unlikely of concern, with correspondingly low levels of the precursor histidine.
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