A simple and rapid thin-layer chromatography (TLC)-densitometry method for histamine analysis in fish and fishery products was established. Histamine was extracted from seafood with 80% ethanol. The sample extracts and standard solutions of histamine were applied to cellulose TLC plate and developed by ascending chromatography with ammonia-ethanol 1:3 (v/v) as mobile phase. Histamine was visualized with pauly's reagent, the intensity of the colored spots was digitized and calculated by image processing method software, Lane & Spot Analyzer (ATTO, Tokyo, Japan). Successful separation of histamine from other (imidazole compounds) pauly's reagent-positive components in 80% ethanol fish extracts was achieved. The linearity of the histamine estimation using this method was good within the range from 30 ng to 1000 ng of histamine (r2 = 0.9997). Histamine can be satisfactorily detected and completely separated from histidine, 4-methyl-imidazole and other pauly's reagent-positive compounds. This method does not need expensive instrument, any tedious pretreatment to eliminate potential interference other imidazole compounds such as histidine or carnosine. It is also less reagents compared with HPLC method. Moreover, it is a simple, rapid, sensitive, reproducible. Therefore this method is suitable for monitor of histamine in multiple fish and fishery products simultaneously that contain as little as 20 ppm histamine (2.0 mg/100 g).
- Pauly's reagent development