Modi®cation of a colorimetric method for histamine analysis in ®sh meal Sevim KoÈse a, *, George Hall b a Black Sea University, Faculty of Marine Sciences and Technology, 61530, C Ë amburnu, Trabzon, Turkey b Loughborough University, Chemical Engineering Department, Loughborough, Leicestershire, LE11 3TU, UK Received 22 October 1999; accepted 4 April 2000 Abstract This study describes the modi®cations necessary to apply the colorimetric method for histamine detection in ®sh meal. Although the original colorimetric method works well with ®sh ¯esh, diculties were observed during ®ltration of the homogenised sample and column separation when it was applied to ®sh meal. In this study, the reason for the problems encountered was studied by processing ®sh meal using dierent parts of ®sh. It was found that the problem was caused by the presence of bone in the sample, which suggested that calcium in bone reacted with potassium hydroxide (KOH) used to adjust pH during the experiment. This was con®rmed by a chemical test reacting calcium carbonate with KOH solutions at dierent concentrations. The reaction was rever- sible in acidic conditions with trichloracetic acid (TCA) or hydrochloric acid (HCl). Several tests were applied before the ®ltration and neutralization stage of the method to solve the problems occurred. Centrifugation of the homogenised sample  20 min at 23 000 g and ®ltration of the aliquot after neutralisation gave the best result among the tests applied. The method was further modi®ed by evaluating the suitable sample size, TCA volume and TCA concentration used as well as simplifying the moisture content application for ®sh meal. Other minor alterations were also suggested for both original and modi®ed method at the preparation stage of diazotizing solution and absorption measurement. # 2000 Elsevier Science Ltd. All rights reserved. Keywords: Histamine; Colorimetric method; Fish meal; Modi®cation 1. Introduction Histamine poisoning is a widely reported food-borne chemical intoxication resulting from ingestion of food which contains unusually high levels of histamine (Taylor, 1983). The foods commonly involved in histamine poi- soning of humans are routinely analysed for histamine and many countries have regulations on histamine content in certain foods such as scombroid ®sh. Although other his- tamine-like amines such as cadaverine, putrescine and spermidine play an important role in histamine poisoning (Kim & Bjeldanes, 1979), the presence of histamine in foods is still used an indicator in histamine food poisoning. Histamine food poisoning has been observed in poul- try where it is called gizzard erosion or black vomiting due to the presence of black vomit in the gizzards of aected chickens (KoÈse, 1993). This was ®rst reported by Shifrine, Ousterhout, Grau and Vaghan (1959) who carried out feeding trials with 12 day old-chicks fed spoiled tuna meal with known levels of histamine in their diet. The results showed that there was a reduction in the rate of growth and depression in total growth of chicks that were proportional to the amount of hista- mine in the feed. Later, several other researchers sup- ported these ®ndings with extended feeding trials. They agreed that certain types of ®sh meal contain high levels of histamine which cause the illness (Harry, Tucker & Lauresen, 1975; Itakura, Kazama & Goto, 1982; Jans- sen, 1971; Miyazaki & Umemura, 1987; Umemura et al., 1981). It has been reported that mortality may go as high as 15% and morbidity is higher than mortality, perhaps up to 30% (KoÈse, 1993). Therefore, analysis of histamine in ®sh meal is also necessary to avoid such illness in poultry or other animals which are fed ®sh meal. Colorimetric, ¯uorometric, thin layer chromatography (TLC) and high performance liquid chromatography 0963-9969/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved. PII: S0963-9969(00)00103-4 Food Research International 33 (2000) 839±845 www.elsevier.com/locate/foodres * Corresponding author at: Karadeniz Teknik Universitesi, Deniz Bilimleri FakuÈltesi, 61530, CË amburnu, Trabzon, Turkey. Tel.: +90- 462-752-2158; fax: +90-462-752-2805 ext. 120.