Moniliformin in Norwegian grain S. Uhlig{*, M. Torp{, J. Jarp{, A. Parich{, A. C. Gutleb{§ and R. Krska{ { National Veterinary Institute, Department of Feed and Food Hygiene, Ullevaalsveien 68, PO Box 8156 Dep., N-0033 Oslo, Norway; { Center for Analytical Chemistry, Institute for Agrobio- technology (IFA Tulln), Konrad Lorenz Str. 20, A-3430 Tulln, Austria (Received 21 November 2003; revised 15 March 2004; accepted 26 March 2004) Norwegian grain samples (73 oats, 75 barley, 83 wheat) from the 2000–02 growing seasons were examined for contamination with moniliformin, and the association between the fungal metabolite and the number of kernels infected with common Fusaria was investigated. Before quantification of moniliformin using ion pairing reversed-phase high-performance liquid chromato- graphy with diode array ultraviolet light detection, all samples were extracted using acetonitrile/water (84/16) and disposable strong anion exchange columns used for clean up. The limit of detection was 40 g kg 1 . Moniliformin was found in 25, 32 and 76% of the barley, oats and wheat samples, respectively. The maximum concentrations of moniliformin in barley, oats and wheat were 380, 210 and 950 g kg 1 , respec- tively. At the same time, the prevalence and infection level of the moniliformin-producing F. avenaceum/ arthrosporioides was as high as 100 and >53% on average, respectively. Moniliformin concentrations were significantly correlated to the variables grain spe- cies, growing season and infection with F. avenaceum/ arthrosporioides and F. culmorum. The survey indi- cates that the prevalence of moniliformin in Norwegian grain is high, especially in wheat. On the other hand, field conditions in Norway do not seem to favour contamination of grain with high levels of moniliformin. Keywords: food, Fusarium, high-performance liquid chromatography (HPLC), mycotoxin, validation Introduction In 1972, a toxic compound was purified from extracts of a strain of Fusarium moniliforme Sheldon isolated from southern leaf blight-damaged corn seed (Cole et al. 1973). The compound was given the name moniliformin and its structure was subsequently determined to be the sodium or potassium salt of 1-hydroxycyclobut-1-ene-3,4-dione (figure 1) (Springer et al. 1974), which was already known as semisquaric acid (Hoffmann et al. 1971). Among biomolecules this compound is characterized by an unusually high acidity, with reported pK a values between 0.0 and 1.7 (Bellus et al. 1978, Scharf et al. 1978, Steyn et al. 1978). Moniliformin was found to inhibit several enzyme systems and gluconeogenesis in vitro (Burka et al. 1982, Chen et al. 1990, Deruiter et al. 1993, Pirrung et al. 1996), and was shown to be toxic to several cell lines at higher concentrations (Vesonder et al. 1993, Wu et al. 1995, Reams et al. 1996). The compound causes muscular weakness and induces acidosis (Nagaraj et al. 1997, Wu and Vesonder 1997), the main target organ being the cardiac muscle (Zhao et al. 1993, Ledoux et al. 1995, Nagaraj et al. 1996, Harvey et al. 2001). The oral LD 50 of the pure compound is similar to those of potent trichothecenes such as diacetoxyscirpenol and T-2 toxin for poultry (Kriek et al. 1977, Ueno 1983). Several analytical methods for the determination of moniliformin, employing many types of chromatog- raphy, have been published and were summarized by Munimbazi and Bullerman (2001). More recently developed methods use ion-exchange chromatog- raphy (Kandler et al. 2002) or ion pairing chroma- tography (Parich et al. 2003). The method of Parich et al., with minor modifications, was applied for the analysis of grain samples in the present survey because of its significant advantages for easier sample preparation, a lower limit of detection and a recovery as good as, or better, than that obtained using older methods. Food Additives and Contaminants, Vol. 21, No. 6 (June 2004), pp. 598–606 * To whom correspondence should be addressed. e-mail: silvio.uhlig@vetinst.no § Present address: Institute for Environmental Studies, Vrije Universiteit Amsterdam, De Boelelaan 1115, NL-1081HV Amsterdam, the Netherlands. Food Additives and Contaminants ISSN 0265–203X print/ISSN 1464–5122 online # 2004 Taylor & Francis Ltd http://www.tandf.co.uk/journals DOI: 10.1080/02652030410001704258