Nitrofuran analysis

Particularly, in a residue analysis, identification of detected analytes is required to confirm a result. For nitrofurans analysis, there are a few confirmatory methods reported, including the use of the photodiode-array system and MS detection. The use of a photodiode array detector coupled to HPLC (HPLC-DAD) offers advantages that the target peak can be identified by its retention time and absorption spectrum. In this case, the continuous spectral data generated during the analysis are collected to check for interfering substances by comparing the spectra of samples with those of the standards. However, the specificity and the limit of detection are not sufficient to determine or identify... 

Koziarz JWP, J Veall, N Sandhu, P Kumar, B Hoecher, IB Lambert (1998) Oxygen-insensitive nitroreductases analysis of the roles of nfsA and nfsB in development of resistance to 5-nitrofuran derivatives in Escherichia coli. J Bacteriol 180 5529-5539. 

In liquid chromatographic analysis of nitrofuran antibacterials, the most popular detector is the ultraviolet visible (UV-vis) spectrophotometer. Nitrofurans exhibit strong absorption at wavelengths around 365 nm and are, therefore, ideal for direct determination (Table 29.5). Detection wavelengths of 275 nm (56, 57) and 400 nm (175) have also been suggested. Electrochemical detection is also frequently applied in liquid chromatographic methods for the determination of various nitrofuran antibacterials in edible animal products (172, 173, 179). 

Confirmatory analysis of suspected liquid chromatographic peaks is usually accomplished by a photodiode array detector that continuously collects spectral data during the chromatographic separation and further compares the spectrum (200-550 nm) of the eluted suspected compound with that of a standard (37, 38, 66, 161, 163, 166-168, 178, 180, 181). Online absorbance ratio techniques combined witlr off-line thin-layer chromatography have been also reported (171). Although tliese confirmation techniques are relatively simple, their sensitivity is not generally adequate to identify trace levels of residual nitrofurans in edible animal products. 

Coupling of liquid chromatography with mass spectrometry allows unequivocal online spectrometric identification of all nitrofurans at the very low residue concentrations required by regulatory agencies for confirmatory analysis in animal-derived foods. Typical examples of mass spectrometry applications in confirming nitrofuran residues in edible animal products employ thermospray (174, 176), ionspray (166), or atmospheric pressure chemical ionization (157) interfaces. 

A Posyniak, S Semeniuk, J Niedzielska, J Zmudzki. Solid-phase extraction and liquid-chromatography analysis of nitrofuran compounds in meat. Chem Anal (Warsaw) 39 289-294, 1994. 

A.K. Debnath et al., Mechanistic interpretation of the genotoxicity of nitrofurans (antibacterial agents) using quantitative structure-activity relationships and comparative molecular field analysis. J. Med. Chem. 36, 1007-1016 (1993)... 

Debnath, A.K., Hansch, C., Kim, K.H. and Martin, Y.C. (1993). Mechanistic Interpretation of the Genotoxicity of Nitrofurans (Antibacterial Agents) Using Quantitative Structure-Activity Relationships and Comparative Molecular Field Analysis. J.Med.Chem.,36,1007-1016. 

Since chemical analysis of nitrofurans is indispensable their characteristic ultraviolet absorption has been recommended for assayi A6i. 

Vass M, Hruska K, Franek M, Nitrofuran anti-biotics A review on the application, prohibition and residual analysis. Vet. Med. CzechPraha. 2008 53 469-500. 

A common procedure for the analysis of nitrofuran metabolites involves hydrolysis of the protein-bound metabolites under acidic conditions followed by deriva-tization with 2-nitrobenzaldehyde (Eig. 7.5). After neutralization of the digest, solvent extraction is carried out with ethyl acetate. Residues are detected by LC-UV or LC-MS/MS In some cases an additional liquid-liquid extraction or solid-phase extraction step is applied to remove excessive matrix compounds. A broad overview of applied methods was published by Vass et al. in 2008. ... 

Nitrofurans are banned substances within the EU and in some other countries because of their mutagenic and geno-toxic characteristics. Nitrofuran metabolites are still found, primarily in aquaculture products originating from Southeast Asia, with SEM (the metabolite of nitrofurazone) having the highest incidence. Methods for detecting residues of nitrofurans aim for protein-bound metabolites that may persist in tissues for considerable periods after treatment. Methods are reported for the detection and identification of nitrofuran metabolites in many different food products. The main difficulty in nitrofuran metabolite analysis is the low selectivity of SEM as a marker metabolite of nitrofurazone. Several other possible sources of SEM have been identified and investigated, the most important of which is the use of... 

Numerous analytical methods have been developed for the analysis of nitrofurans in various samples. These include microbiological, thin-layer chromatography (TLC), and high-performance liquid chromatography (HPLC) techniques. HPLC is the most widely used technique for determining nitrofurans of food sample origin, such as shrimps, fish, ° milk, egg and chicken... 

The separations of nitrofurans have been carried out, in most instances, with a reversed phase (RP) column. As summarized in Table 1, Cig columns were selected as the analytical column for LC in most reports however, the use of a Cg, CN, and cyanopropyl-modified silica analytical column has also been described. The parent compounds, as well as their metabolites, are satisfactorily separated by Cig columns with commonly used mobile phases, such as mixtures of acetonitrile with phosphate buffer, acetate buffer, or water. The mobile phase elution for HPLC analysis can be isocratic or as a gradient system. Acetonitrile-0.1 M aqueous solution of sodium perchlorate (28 72), with 0.5% glacial acetic acid, was reported by Galeano Diaz " as an optimum mobile phase for the separation of the three nitrofuran derivatives nitrofurantoin, furazolidone, and furaltadone in milk. Lin et al." used an acetonitrile gradient with an initial hold time of 1 min at 0% acetonitrile. 

UV detection and diode-array detection (DAD) have been widely used, coupled to HPLC, for the analysis of the parent compounds of nitrofurans. The detection wavelengths of nitrofiirans have been set at 368 nm," 365 nm, " 360 nm," 270 nm, or at 254 nm. " An HPLC method, with electrochemical detection, has also been described for the monitoring of furazolidone and furaltadone hydrochloride in chickoi tissues and eggs. " ... 

Table 1 High-performance liquid chromatography (HPLC) conditions of the analysis of nitrofurans.

Edder, P. Vargas, S. Ortelli, D. Claude, C. Analysis of 15. nitrofuran metabolites in food by high-performance liquid... 

Degroodt, J.M. Wyhowski de Bukanski, B. De Groof, J. Beemaert, H. Srebmik, S. Chloramphenicol and nitrofuran residue analysis hy HPLC and photodiode array detection in meat and fish. J. Liq. Chromatogr. 1992, 75, 2355-2371. 

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