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Biogenic amines occurrence in fish meat sampled from restaurants in region of Czech Republic
Food Control 31 (2013) 49e52
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Food Control journal homepage: www.elsevier.com/locate/foodcont
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Biogenic amines occurrence in fish meat sampled from restaurants in region of Czech Republic ka a, *, Pavel Budinský b, Blanka Zimáková c, Marek Merhaut c, Radka Flasarová a, Frantisek Bun a, Leona Bun ková d Vendula Pachlová a, Vlastimil Kubán a
Department of Food Technology, Faculty of Technology, Tomas Bata University in Zlin, nam. T. G. Masaryka 5555, 76001 Zlin, Czech Republic Faculty Hospital in Motol, V Úvalu 84, 15006 Prague, Czech Republic Department of Hotel Management, The Institute of Hospitality Management, Svídnická 506, 18100 Prague, Czech Republic d Department of Environmental Protection Engineering, Faculty of Technology, Tomas Bata University in Zlin, nam. T. G. Masaryka 5555, 76001 Zlin, Czech Republic b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 27 May 2012 Received in revised form 16 September 2012 Accepted 25 September 2012
The aim of the work was to monitor the content of 8 biogenic amines (histamine, tyramine, phenylethylamine, tryptamine, putrescine, cadaverine, spermidine and spermine) in 112 samples of raw freshwater and see fish, and other animals living in the water which are offered in restaurants in the Czech Republic. In approx. 15% of the sampled bodies, increased levels of biogenic amines (practically more than 100 mg/kg) have been observed. In 6 samples of fish, the concentration of histamine was higher than the limit specified in Commission Regulation (EC) No 2073/2005 (more than 200 mg/kg). The results pointed out the necessity of keeping the storage conditions (cooling and/or freezing chain) during the distribution of products to consumers. Ó 2012 Elsevier Ltd. All rights reserved.
Keywords: Fish Biogenic amines Histamine Restaurants Health risk
1. Introduction Biogenic amines (BAs) are aliphatic (putrescine, cadaverine), aromatic (tyramine, 2-phenylethylamine) or heterocyclic (histamine, tryptamine) alkaline compounds formed in foodstuff mainly by microbial decarboxylation. BA precursors are free amino acids provided by proteolytic changes of proteins and/or peptides. Histamine is formed from histidine, tyramine from tyrosine, phenylethylamine from phenylalanine, tryptamine from tryptophane, cadaverine from lysine, and putrescine from arginine or ornitine. Apart from BAs mentioned above, the concentrations of polyamines, such as agmatine, spermine, and spermidine should be also observed. Agmatine is formed from arginine, spermine and spermidine from putrescine (Halász, Baráth, Simon-Sarkadi, & Holzapfel, 1994; Shalaby, 1996; Silla Santos, 1996). Contaminating bacteria from family Enterobacteriaceae (e.g. some strains from genera Salmonella, Shigella, Escherichia, Serratia, Yersinia, Morganella) and genera Pseudomonas are usually included in groups of microorganisms possessing decarboxylase enzymes.
* Corresponding author. Tel.: þ420 576 033 011; fax: þ420 577 210 172. ka). E-mail address: [email protected] (F. Bun 0956-7135/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodcont.2012.09.044
BAs are produced also by some lactic acid bacteria (e.g. strains from genera Lactobacillus, Enterococcus etc.). Lactic acid bacteria also occur in some food as contaminants failing the quality and safety of the product (e.g. in case of some meat products, beer, and wine). Presence of particular decarboxylases is not specific within the species and occurs only among certain number of strains in the ková et al., 2009; Halász et al., 1994; Shalaby, 1996; species (Bun Silla Santos, 1996). BAs and polyamines are endogenous compounds with the key functions in the metabolism of living organisms. Generally, low concentrations of BAs in food and drink (practically under 100 mg/kg) do not represent a significant risk for a healthy human. Human intestinal tract has detoxifying system consisting of monoaminooxidase (MAO), diaminooxidase (DAO), and histamine methyl-transpherase (HMT). However, higher amounts of BAs (generally above 100 mg/kg) may induce undesirable psychoactive and vasoactive effects (hypotension or hypertension, headache, nausea, breathing problems etc.). Histamine and tyramine may cause the latter mentioned undesirable effects directly. On the other hand, putrescine and cadaverine act as potentiators of effects of histamine or tyramine because putrescine and cadaverine could inhibit the detoxifying enzymatic system (Arnold & Brown, 1978; ková et al., 2009; Halász et al., 1994; Ten Brink, Damink, Joosten, Bun
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50
& Huis in’t Veld, 1990). In Commission Regulation (EC) No. 2073 (2005), the concentration limit for histamine (100, resp. 200 mg/ kg) in fish and fishery products (especially in fish species of families Scombridae, Clupeidae, Engraulidae, Coryfenidae, Pomatomidae, Scombresosidae) is established. Thereby, the importance of BAs observation in these commodities is emphasized. Fish and fish products are consumed insufficiently in the region of Middle Europe. Fish meat could be considered to be functional food due to its nutritious characteristics (the high content of n-3 polyunsaturated fatty acids; high content of iron, zinc, and selenium; and simultaneously low content of sodium) (Dadáková, Krí zek, & Pelikánová, 2009; Zhang et al., 2011; Zhang, Xaio, Samaraweera, Lee, & Ahn, 2010). Nowadays, many common and/or special restaurants offer wide group of fish and fishery products. Salmon, tuna, mackerel, striped catfish, trout or carp are often consumed species of fish in Middle Europe. On the other hand, fish meat represents the system with very short shelf-life because postmortal changes are very fast. Therefore, the key requirements for food-safety maintenance of fish meat are quick chilling or even freezing and subsequently, keeping fish meat under desired temperature (in whole chilling or freezing chain). In fish meat, BAs may occur very often which correspond with fast postmortal changes (Arnold & Brown, 1978; Jaw et al., 2012; Krí zek, Vacha, & Pelikánová, 2011; Rawles, Flick, & Martin, 1996). Data of BAs occurrence in fish meat in restaurants in the region of Middle Europe have not been published in available literature. Only a few papers
dealing especially with modeling kinetics of BA production in model fish systems have been found (e.g. Krí zek et al., 2011). The aim of this work was to monitor the content of 8 BAs (histamine e HIM, tyramine e TYM, phenylethylamine e PHE, tryptamine e TRM, putrescine e PUT, cadaverine e CAD, spermidine e SPD, spermine e SPN) in the fish meat offered in restaurants in the Czech Republic. 2. Materials and methods 2.1. Samples One hundred and twelve raw samples were analyzed (details are shown in Table 1). Ninety-nine fish raw samples (10 freshwater species and 16 sea fish species) were obtained in the years 2010 and 2011 from restaurants and sushi bars in the Czech Republic. One cartilaginous fish sample (angler) and 12 other seafood (especially calamari, prawn and squid) were also sampled. Individual samples were stored frozen by the provider (at the temperature of 18 C and lower; totally 53 samples) or they were stored “on ice” (at the temperature of melting ice; totally 59 samples). Samples were taken from commonly consumed parts of animal bodies. During the transport to the laboratory, samples were kept in isothermal containers (approx. 2 h); and immediately, they were lyophilized (Christ Alpha 1e4). Lyophilized material was stored in 70 C until the analysis.
Table 1 Biogenic amines content in sampled bodies of fish, cartilaginous and other seafood animals (mg/kg). English name
Latin name
Na
Db
Alaska pollock Angler (sea devil) Atlantic bluefin tuna Atlantic cod Atlantic mackerel Atlantic pomfret Atlantic salmon Brown trout Bullet tuna Calamari Common carp Escolar (butterfish) European crayfish European hake European seabass European spiny lobster European squid Fleshy prawn Giant tiger prawn Gilthead seabream Grass carp Greenland halibut Hoki Nile tilapia Northern prawn Pike perch Pollock Rainbow trout Scallop Striped catfish Swordfish Tench Turbot Wels catfish Yellowfin tuna
Theragra chalcogramma Lophius piscatorius Thunnus thynnus Gadus morhua Scomber scombrus Brama brama Salmo salar Salmo trutta Auxis rochei Sepioteuthis sp. Cyprinus carpio Lepidocybium flavobrunneum Astacus astacus Merluccius merluccius Dicentrarchus labrax Palinurus vulgaris Loligo vulgaris Penaeus chinensis Penaeus monodon Sparus aurata Ctenopharyngodon idella Reinhardtius hippoglossoides Macruronus sp. Oreochromis niloticus Pandalus borealis Sander lucioperca Pollachius virens Oncorhynchus mykiss Pecten jacobaeus Pangasius hypophthalmus Xiphias gladius Tinca tinca Psetta maxima Silurus glanis Thunnus albacares
5 1 1 13 1 1 20 5 1 2 7 3 1 7 3 1 1 1 2 1 1 3 1 1 2 2 1 1 2 16 1 1 1 1 1
S O S S S S F F S O F S O S S O O O O S F S S F O F S F O F S F S F S
a
Biogenic amines and polyamines content (ND/þ/þþ/þþþ/þþþþ/þþþþþ)c HIM
PUT
CAD
SPD
SPN
5/0/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/11/1/0/0/1 0/0/0/0/0/1 0/0/0/0/0/1 12/3/3/2/0/0 5/0/0/0/0/0 1/0/0/0/0/0 1/1/0/0/0/0 6/0/1/0/0/0 3/0/0/0/0/0 0/1/0/0/0/0 7/0/0/0/0/0 2/0/1/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 2/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/2 1/0/0/0/0/0 1/0/0/0/0/0 2/0/0/0/0/0 1/1/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 2/0/0/0/0/0 14/0/1/0/0/1 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0
0/0/5/0/0/0 0/0/0/1/0/0 0/0/1/0/0/0 0/0/11/2/0/0 0/1/0/0/0/0 1/0/0/0/0/0 1/11/3/4/0/1 0/0/4/1/0/0 1/0/0/0/0/0 0/1/1/0/0/0 0/2/4/1/0/0 2/1/0/0/0/0 0/0/1/0/0/0 0/0/5/2/0/0 0/1/2/0/0/0 0/0/0/1/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/0/2/0/0/0 1/0/0/0/0/0 0/0/1/0/0/0 1/2/0/0/0/0 0/0/1/0/0/0 0/0/1/0/0/0 0/0/1/1/0/0 0/2/0/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0 1/0/0/0/0/1 4/2/10/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 1/0/0/0/0/0 0/0/0/0/1/0 1/0/0/0/0/0
5/0/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 1/7/5/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 10/5/5/0/0/0 1/3/1/0/0/0 1/0/0/0/0/0 1/1/0/0/0/0 3/2/2/0/0/0 3/0/0/0/0/0 0/1/0/0/0/0 4/3/0/0/0/0 1/1/1/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/1/1/0/0/0 1/0/0/0/0/0 0/1/0/0/0/0 3/0/0/0/0/0 1/0/0/0/0/0 0/0/1/0/0/0 0/2/0/0/0/0 0/1/1/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/1/0/0 9/7/0/0/0/0 0/1/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0
0/5/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/12/1/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 1/15/3/1/0/0 0/4/1/0/0/0 1/0/0/0/0/0 0/1/1/0/0/0 0/0/7/0/0/0 0/1/2/0/0/0 0/1/0/0/0/0 0/7/0/0/0/0 1/1/1/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/2/0/0/0/0 1/0/0/0/0/0 0/0/1/0/0/0 1/2/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/2/0/0/0/0 1/1/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 2/0/0/0/0/0 1/13/2/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0
0/5/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/12/1/0/0/0 0/0/1/0/0/0 0/0/1/0/0/0 1/16/3/0/0/0 0/5/0/0/0/0 1/0/0/0/0/0 0/1/1/0/0/0 0/7/0/0/0/0 0/0/2/1/0/0 0/1/0/0/0/0 0/7/0/0/0/0 1/1/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/0/1/0/0/0 0/1/1/0/0/0 1/0/0/0/0/0 0/1/0/0/0/0 1/1/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/2/0/0/0/0 1/1/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 1/0/1/0/0/0 1/13/1/1/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0
N e the number of samples in the certain species of freshwater or see fish or other water animals. D e the group of samples: F e freshwater fish, S e see fish, O e other water animals. The contents of biogenic amines were expressed using the magnitude: “ND” e not detected; “þ” <10 mg/kg; “þþ” 10e50 mg/kg; “þþþ” 50e100 mg/kg; “þþþþ” 100e200 mg/kg; “þþþþþ” >200 mg/kg. b c
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2.2. Determination of biogenic amines content Biogenic amines were extracted from the lyophilized matter threefold with the use of 0.6 mol l1 perchloric acid. Content of 8 biogenic amines (HIM, TYM, PHE, TRM, PUT, CAD, SPD, SPN) was determined by the method of high performance liquid chromatography (LabAlliance, State College, USA; Agilent Technologies, Agilent, Paolo Alto, USA) after the preceding derivatization by dansylchloride. Derivatization, chromatographic separation (Cogent column HPS C18, 150 4.6 mm, 5 mm, Cogent, Eatontown, USA) and detection (spectrophotometrically l ¼ 254 nm) were performed lá, Pechová, Komprda, according to Dadáková et al. (2009) and Sme (2004). An example of chromatogram of a stanKlejdus, and Kubán dard mixture of biogenic amines and chromatogram of extract from a body of Atlantic salmon is shown in Fig. 1. Every sample was extracted three times, every extract was derivatized twice and every derivatized mixture was spread on the column (n ¼ 18) three times. Results were expressed for the fresh matter before lyophilization. Differences between biogenic amines concentrations in tested bodies were statistically evaluated by KruskaleWallis and Wilcoxon tests. KruskaleWallis test was applied also for the analysis of variance (the evaluation of effect of storage methods and samples groups on BA occurrence). Statistical programme UnistatÒ 5.5 (Unistat Ltd., London, UK) and the significance level of 0.05 were used for the tests. 3. Results and discussion TRM was detected only in two fish samples (carp and pike perch) in the amounts between 0.9 and 2.5 mg/kg (data not shown). PHE was recorded in 13 fish samples (5 Atlantic cods, 4 Atlantic salmons, 2 Alaska pollocks and a sample of European hake and striped catfish). In above mentioned cases, the concentrations of PHE between 0.5 and 4.9 mg/kg were recorded (data not shown). Silla Santos (1996) and Ten Brink et al. (1990) suggest (from point of view of health risk) the limit for PHE income at the level of 30 mg/ kg. The latter mentioned limit was not exceeded in tested bodies. Relatively low content of PHE (<4 mg/kg) in fish is presented also by Hwang et al. (2012) and Zhai et al. (2012). In 107 samples, TYM was not detected or the concentrations ranged below 10 mg/kg (data not shown). Only in 5 bodies (4 Atlantic salmons and 1 carp), TYM concentrations between 10 and 50 mg/kg were recorded, which could be evaluated as safe for healthy consumers (Halász et al., 1994; Silla Santos, 1996; Ten Brink et al., 1990). Low TYM content (practically < 50 mg/kg) in most of the fish samples have been published by other authors (e.g. Hwang et al., 2012; Krí zek et al., 2011; Zhai et al., 2012). HIM, PUT, CAD, SPD and SPN content determined in the samples of bodies of freshwater and sea fish and other seafood are shown in Table 1. The only controlled biogenic amine (by the national food inspection authority) is HIM. HIM was not detected in 78 samples. In 19 tested bodies, HIM concentrations below 10 mg/kg were recorded. In other 9 cases, HIM concentrations of were below 100 mg/kg (P < 0.05) which is still considered as the safe limit for a healthy human (Halász et al., 1994; Silla Santos, 1996; Ten Brink et al. 1990). On the other hand, in 6 bodies (two Greenland halibuts, and a sample of Atlantic mackerel, Atlantic cod, striped catfish and Atlantic pomfret) the limit of 100 mg/kg was significantly exceeded (P < 0.05). Although the all latter mentioned fish samples were stored at freezing temperatures (the fish bodies were obtained from six different restaurants), the concentrations reached even the level above 1000 mg/kg (P < 0.05), which should be considered as dangerous (Halász et al., 1994; Silla Santos, 1996; Ten Brink et al. 1990). These results emphasize the necessity of BAs monitoring and evaluation of health risks.
Fig. 1. Separation of mixtures of biogenic amines as dansylamides. Part A: a standard mixture (tryptamine, phenylethylamine, putrescine, cadaverine, histamine, ISTD e internal standard e 1,7eheptandiamine, tyramine, spermidine and spermine) in concentration of 50 mg/l in acetonitrile. Part B: a derivatized sample extracted from the body of Atlantic salmon.
PUT was the most abundant biogenic amine observed in our samples (Table 1). Only in 14 cases, PUT was not detected. PUT concentrations up to 10 mg/kg and between 10 and 100 mg/kg were recorded in 26 and 69 samples, respectively. Three bodies (Atlantic salmon, Wels catfish and scallop) possessed PUT concentration >100 mg/kg (P < 0.05), which should be pointed out as serious. In 92 bodies, CAD was not detected or the concentrations were up to 10 mg/kg. More significant CAD concentrations were observed in 20 cases, where only once the amount exceeded the level 50 mg/kg (P < 0.05). PUT and CAD could act as potentiators of toxicological effects of HIM and TYM (Arnold & Brown, 1978; Halász
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ka et al. / Food Control 31 (2013) 49e52 F. Bun
et al., 1994; Prester, 2011; Silla Santos, 1996; Ten Brink et al. 1990). The occurrence of PUT and CAD is usually related to the hygiene conditions in the process of meat treatment and could point out to the presence of contaminating microflora, especially gramnegative strains of family Enterobacteriaceae and genus Pseudomonas (Halász et al., 1994; Hwang et al., 2012; Rawles et al., 1996; Zhai et al., 2012). In nearly 80% of samples, SPD and SPN concentrations were very low (up to 10 mg/kg) or these polyamines were not detected. SPD concentration ranging between 50 and 100 mg/kg was determined in the only case of Atlantic salmon. In two fish samples (striped catfish and escolar) SPN concentration above 50 mg/kg were observed. Apart from the concentrations of individual BAs, the sum of determined BAs amounts was also evaluated. In almost 85% of tested samples, the sum of BAs was below 100 mg/kg. On the other hand, in 7% of observed bodies, the BAs concentrations between 100 and 200 mg/kg were recorded. Surprisingly, in 8% of samples, the BAs concentrations above 200 mg/kg and in 5% of bodies even the amounts exceeded 1000 mg/kg. The latter mentioned concentrations represent amounts which could cause serious health problems even for healthy consumers. Unfortunately, potentially dangerous concentrations of BAs were also observed in fish bodies which are widely served in restaurants in the Middle Europe (e.g. Atlantic mackerel, Atlantic salmon and striped catfish). Analysis of variance showed no significant difference between the samples stored frozen and “on ice” (P 0.05). Similarly, the BAs concentration in freshwater and sea fish did not differ significantly (P 0.05). The BAs concentrations of the other seafood samples were not significantly higher in comparison with the bodies of freshwater and sea fish (P 0.05). 4. Conclusion Monitoring of the occurrence of 8 biogenic amines in the bodies of 112 fish and other seafood obtained from restaurants in the Czech Republic has been performed. In more than three quarters of samples, the sum of concentrations of biogenic amines was under 100 mg/kg, which could be considered to be a safe limit for healthy consumers. On the other hand, in 8% of tested samples, dangerous amounts of BAs were found out and in 6 cases even legislatively set limits were exceeded. Although fish and fish products should be a part of human diet, it is important to deal with their short shelf-life which could be determined also by biogenic amines concentrations.
Acknowledgment The financial support from the Grant Agency of the Czech 503/11/1417) and Internal Grant of TBU in Republic (Grant No. GACR Zlin (No. IGA/FT/2012/027) is greatly acknowledged. References Arnold, S. H., & Brown, W. D. (1978). Histamine toxicity from fish products. Advances in Food Research, 24, 113e154. ková, L., Bun ka, F., Hlobilová, M., Van átková, Z., Nováková, D., & Dráb, V. (2009). Bun Tyramine production of technological important strains of Lactobacillus, Lactococcus and Streptococcus. European Food Research and Technology, 229, 533e538. Commission regulation (EC) No 2073/2005 of 15 November 2005 on microbiological criteria for foodstuffs. Dadáková, E., Krí zek, P., & Pelikánová, T. (2009). Determination of biogenic amines in foods using ultra-performance liquid chromatography (UPLC). Food Chemistry, 116, 365e370. Halász, A., Baráth, Á., Simon-Sarkadi, L., & Holzapfel, W. (1994). Biogenic amines and their production by microorganisms in food. Trends in Food Science and Technology, 51, 42e49. Hwang, C. C., Lin, C. M., Huang, C. Y., Huang, Y. L., Kang, F. C., Hwang, D. F., et al. (2012). Chemical characterisation, biogenic amines contents, and identification of fish species in cod and escolar steaks, and salted escolar roe products. Food Control, 25, 415e420. Jaw, Y. M., Chen, Y. Y., Lee, Y. C., Lee, P. H., Jiang, C. M., & Tsai, Y. H. (2012). Histamine content and isolation of histamine-forming bacteria in fish meal and fish soluble concentrate. Fisheries Science, 78, 155e162. Krí zek, M., Vacha, F., & Pelikánová, T. (2011). Biogenic amines in carp roe (Cyprinus caprio) preserved by four different methods. Food Chemistry, 126, 1493e1497. Prester, L. (2011). Biogenic amines in fish, fish products and shellfish: a review. Food Additives and Contaminants Part A e Chemistry Analysis Control Exposure and Risk Assessment, 28, 1547e1560. Rawles, D. D., Flick, G. J., & Martin, R. E. (1996). Biogenic amines in fish and shellfish. Advances in Food Nutrition and Research, 39, 329e365. Shalaby, A. R. (1996). Significance of biogenic amines to food safety and human health. Food Research International, 29, 675e690. Silla Santos, M. H. (1996). Biogenic amines: their importance in foods. International Journal of Food Microbiology, 29, 213e231. lá, D., Pechová, P., Komprda, T., Klejdus, B., & Kubán , V. (2004). Chromatographic Sme determination of biogenic amines in dry salami during the fermentation and storage (in Czech). Chemical Papers, 98, 432e437. Ten Brink, B., Damink, C., Joosten, H. M. L. J., & Huis in’t Veld, J. H. J. (1990). Occurrence and formation of biologically active amines in foods. International Journal of Food Microbiology, 11, 73e84. Zhai, H., Yang, X., Li, L., Xia, G., Cen, J., Huang, H., et al. (2012). Biogenic amines in commercial fish products sold in southern China. Food Control, 25, 303e308. Zhang, J., Liu, Z., Hu, Y., Fang, Z., Chen, J., Wu, D., et al. (2011). Effect of sucrose on the generation of free amino acids and biogenic amines in Chinese traditional dry-cured fish during processing and storage. International Journal of Food Science and Technology, 48, 69e75. Zhang, W., Xiao, S., Samaraweera, H., Lee, E. J., & Ahn, D. U. (2010). Improving functional value of meat products. Meat Science, 86, 15e31.
Contents lists available at SciVerse ScienceDirect
Food Control journal homepage: www.elsevier.com/locate/foodcont
Short communication
Biogenic amines occurrence in fish meat sampled from restaurants in region of Czech Republic ka a, *, Pavel Budinský b, Blanka Zimáková c, Marek Merhaut c, Radka Flasarová a, Frantisek Bun a, Leona Bun ková d Vendula Pachlová a, Vlastimil Kubán a
Department of Food Technology, Faculty of Technology, Tomas Bata University in Zlin, nam. T. G. Masaryka 5555, 76001 Zlin, Czech Republic Faculty Hospital in Motol, V Úvalu 84, 15006 Prague, Czech Republic Department of Hotel Management, The Institute of Hospitality Management, Svídnická 506, 18100 Prague, Czech Republic d Department of Environmental Protection Engineering, Faculty of Technology, Tomas Bata University in Zlin, nam. T. G. Masaryka 5555, 76001 Zlin, Czech Republic b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 27 May 2012 Received in revised form 16 September 2012 Accepted 25 September 2012
The aim of the work was to monitor the content of 8 biogenic amines (histamine, tyramine, phenylethylamine, tryptamine, putrescine, cadaverine, spermidine and spermine) in 112 samples of raw freshwater and see fish, and other animals living in the water which are offered in restaurants in the Czech Republic. In approx. 15% of the sampled bodies, increased levels of biogenic amines (practically more than 100 mg/kg) have been observed. In 6 samples of fish, the concentration of histamine was higher than the limit specified in Commission Regulation (EC) No 2073/2005 (more than 200 mg/kg). The results pointed out the necessity of keeping the storage conditions (cooling and/or freezing chain) during the distribution of products to consumers. Ó 2012 Elsevier Ltd. All rights reserved.
Keywords: Fish Biogenic amines Histamine Restaurants Health risk
1. Introduction Biogenic amines (BAs) are aliphatic (putrescine, cadaverine), aromatic (tyramine, 2-phenylethylamine) or heterocyclic (histamine, tryptamine) alkaline compounds formed in foodstuff mainly by microbial decarboxylation. BA precursors are free amino acids provided by proteolytic changes of proteins and/or peptides. Histamine is formed from histidine, tyramine from tyrosine, phenylethylamine from phenylalanine, tryptamine from tryptophane, cadaverine from lysine, and putrescine from arginine or ornitine. Apart from BAs mentioned above, the concentrations of polyamines, such as agmatine, spermine, and spermidine should be also observed. Agmatine is formed from arginine, spermine and spermidine from putrescine (Halász, Baráth, Simon-Sarkadi, & Holzapfel, 1994; Shalaby, 1996; Silla Santos, 1996). Contaminating bacteria from family Enterobacteriaceae (e.g. some strains from genera Salmonella, Shigella, Escherichia, Serratia, Yersinia, Morganella) and genera Pseudomonas are usually included in groups of microorganisms possessing decarboxylase enzymes.
* Corresponding author. Tel.: þ420 576 033 011; fax: þ420 577 210 172. ka). E-mail address: [email protected] (F. Bun 0956-7135/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodcont.2012.09.044
BAs are produced also by some lactic acid bacteria (e.g. strains from genera Lactobacillus, Enterococcus etc.). Lactic acid bacteria also occur in some food as contaminants failing the quality and safety of the product (e.g. in case of some meat products, beer, and wine). Presence of particular decarboxylases is not specific within the species and occurs only among certain number of strains in the ková et al., 2009; Halász et al., 1994; Shalaby, 1996; species (Bun Silla Santos, 1996). BAs and polyamines are endogenous compounds with the key functions in the metabolism of living organisms. Generally, low concentrations of BAs in food and drink (practically under 100 mg/kg) do not represent a significant risk for a healthy human. Human intestinal tract has detoxifying system consisting of monoaminooxidase (MAO), diaminooxidase (DAO), and histamine methyl-transpherase (HMT). However, higher amounts of BAs (generally above 100 mg/kg) may induce undesirable psychoactive and vasoactive effects (hypotension or hypertension, headache, nausea, breathing problems etc.). Histamine and tyramine may cause the latter mentioned undesirable effects directly. On the other hand, putrescine and cadaverine act as potentiators of effects of histamine or tyramine because putrescine and cadaverine could inhibit the detoxifying enzymatic system (Arnold & Brown, 1978; ková et al., 2009; Halász et al., 1994; Ten Brink, Damink, Joosten, Bun
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& Huis in’t Veld, 1990). In Commission Regulation (EC) No. 2073 (2005), the concentration limit for histamine (100, resp. 200 mg/ kg) in fish and fishery products (especially in fish species of families Scombridae, Clupeidae, Engraulidae, Coryfenidae, Pomatomidae, Scombresosidae) is established. Thereby, the importance of BAs observation in these commodities is emphasized. Fish and fish products are consumed insufficiently in the region of Middle Europe. Fish meat could be considered to be functional food due to its nutritious characteristics (the high content of n-3 polyunsaturated fatty acids; high content of iron, zinc, and selenium; and simultaneously low content of sodium) (Dadáková, Krí zek, & Pelikánová, 2009; Zhang et al., 2011; Zhang, Xaio, Samaraweera, Lee, & Ahn, 2010). Nowadays, many common and/or special restaurants offer wide group of fish and fishery products. Salmon, tuna, mackerel, striped catfish, trout or carp are often consumed species of fish in Middle Europe. On the other hand, fish meat represents the system with very short shelf-life because postmortal changes are very fast. Therefore, the key requirements for food-safety maintenance of fish meat are quick chilling or even freezing and subsequently, keeping fish meat under desired temperature (in whole chilling or freezing chain). In fish meat, BAs may occur very often which correspond with fast postmortal changes (Arnold & Brown, 1978; Jaw et al., 2012; Krí zek, Vacha, & Pelikánová, 2011; Rawles, Flick, & Martin, 1996). Data of BAs occurrence in fish meat in restaurants in the region of Middle Europe have not been published in available literature. Only a few papers
dealing especially with modeling kinetics of BA production in model fish systems have been found (e.g. Krí zek et al., 2011). The aim of this work was to monitor the content of 8 BAs (histamine e HIM, tyramine e TYM, phenylethylamine e PHE, tryptamine e TRM, putrescine e PUT, cadaverine e CAD, spermidine e SPD, spermine e SPN) in the fish meat offered in restaurants in the Czech Republic. 2. Materials and methods 2.1. Samples One hundred and twelve raw samples were analyzed (details are shown in Table 1). Ninety-nine fish raw samples (10 freshwater species and 16 sea fish species) were obtained in the years 2010 and 2011 from restaurants and sushi bars in the Czech Republic. One cartilaginous fish sample (angler) and 12 other seafood (especially calamari, prawn and squid) were also sampled. Individual samples were stored frozen by the provider (at the temperature of 18 C and lower; totally 53 samples) or they were stored “on ice” (at the temperature of melting ice; totally 59 samples). Samples were taken from commonly consumed parts of animal bodies. During the transport to the laboratory, samples were kept in isothermal containers (approx. 2 h); and immediately, they were lyophilized (Christ Alpha 1e4). Lyophilized material was stored in 70 C until the analysis.
Table 1 Biogenic amines content in sampled bodies of fish, cartilaginous and other seafood animals (mg/kg). English name
Latin name
Na
Db
Alaska pollock Angler (sea devil) Atlantic bluefin tuna Atlantic cod Atlantic mackerel Atlantic pomfret Atlantic salmon Brown trout Bullet tuna Calamari Common carp Escolar (butterfish) European crayfish European hake European seabass European spiny lobster European squid Fleshy prawn Giant tiger prawn Gilthead seabream Grass carp Greenland halibut Hoki Nile tilapia Northern prawn Pike perch Pollock Rainbow trout Scallop Striped catfish Swordfish Tench Turbot Wels catfish Yellowfin tuna
Theragra chalcogramma Lophius piscatorius Thunnus thynnus Gadus morhua Scomber scombrus Brama brama Salmo salar Salmo trutta Auxis rochei Sepioteuthis sp. Cyprinus carpio Lepidocybium flavobrunneum Astacus astacus Merluccius merluccius Dicentrarchus labrax Palinurus vulgaris Loligo vulgaris Penaeus chinensis Penaeus monodon Sparus aurata Ctenopharyngodon idella Reinhardtius hippoglossoides Macruronus sp. Oreochromis niloticus Pandalus borealis Sander lucioperca Pollachius virens Oncorhynchus mykiss Pecten jacobaeus Pangasius hypophthalmus Xiphias gladius Tinca tinca Psetta maxima Silurus glanis Thunnus albacares
5 1 1 13 1 1 20 5 1 2 7 3 1 7 3 1 1 1 2 1 1 3 1 1 2 2 1 1 2 16 1 1 1 1 1
S O S S S S F F S O F S O S S O O O O S F S S F O F S F O F S F S F S
a
Biogenic amines and polyamines content (ND/þ/þþ/þþþ/þþþþ/þþþþþ)c HIM
PUT
CAD
SPD
SPN
5/0/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/11/1/0/0/1 0/0/0/0/0/1 0/0/0/0/0/1 12/3/3/2/0/0 5/0/0/0/0/0 1/0/0/0/0/0 1/1/0/0/0/0 6/0/1/0/0/0 3/0/0/0/0/0 0/1/0/0/0/0 7/0/0/0/0/0 2/0/1/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 2/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/2 1/0/0/0/0/0 1/0/0/0/0/0 2/0/0/0/0/0 1/1/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 2/0/0/0/0/0 14/0/1/0/0/1 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0
0/0/5/0/0/0 0/0/0/1/0/0 0/0/1/0/0/0 0/0/11/2/0/0 0/1/0/0/0/0 1/0/0/0/0/0 1/11/3/4/0/1 0/0/4/1/0/0 1/0/0/0/0/0 0/1/1/0/0/0 0/2/4/1/0/0 2/1/0/0/0/0 0/0/1/0/0/0 0/0/5/2/0/0 0/1/2/0/0/0 0/0/0/1/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/0/2/0/0/0 1/0/0/0/0/0 0/0/1/0/0/0 1/2/0/0/0/0 0/0/1/0/0/0 0/0/1/0/0/0 0/0/1/1/0/0 0/2/0/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0 1/0/0/0/0/1 4/2/10/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 1/0/0/0/0/0 0/0/0/0/1/0 1/0/0/0/0/0
5/0/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 1/7/5/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 10/5/5/0/0/0 1/3/1/0/0/0 1/0/0/0/0/0 1/1/0/0/0/0 3/2/2/0/0/0 3/0/0/0/0/0 0/1/0/0/0/0 4/3/0/0/0/0 1/1/1/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/1/1/0/0/0 1/0/0/0/0/0 0/1/0/0/0/0 3/0/0/0/0/0 1/0/0/0/0/0 0/0/1/0/0/0 0/2/0/0/0/0 0/1/1/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/1/0/0 9/7/0/0/0/0 0/1/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0 1/0/0/0/0/0
0/5/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/12/1/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 1/15/3/1/0/0 0/4/1/0/0/0 1/0/0/0/0/0 0/1/1/0/0/0 0/0/7/0/0/0 0/1/2/0/0/0 0/1/0/0/0/0 0/7/0/0/0/0 1/1/1/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/2/0/0/0/0 1/0/0/0/0/0 0/0/1/0/0/0 1/2/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/2/0/0/0/0 1/1/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 2/0/0/0/0/0 1/13/2/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/1/0/0/0/0
0/5/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/12/1/0/0/0 0/0/1/0/0/0 0/0/1/0/0/0 1/16/3/0/0/0 0/5/0/0/0/0 1/0/0/0/0/0 0/1/1/0/0/0 0/7/0/0/0/0 0/0/2/1/0/0 0/1/0/0/0/0 0/7/0/0/0/0 1/1/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/0/1/0/0/0 0/1/1/0/0/0 1/0/0/0/0/0 0/1/0/0/0/0 1/1/1/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 0/2/0/0/0/0 1/1/0/0/0/0 0/1/0/0/0/0 0/0/1/0/0/0 1/0/1/0/0/0 1/13/1/1/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0 0/1/0/0/0/0
N e the number of samples in the certain species of freshwater or see fish or other water animals. D e the group of samples: F e freshwater fish, S e see fish, O e other water animals. The contents of biogenic amines were expressed using the magnitude: “ND” e not detected; “þ” <10 mg/kg; “þþ” 10e50 mg/kg; “þþþ” 50e100 mg/kg; “þþþþ” 100e200 mg/kg; “þþþþþ” >200 mg/kg. b c
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2.2. Determination of biogenic amines content Biogenic amines were extracted from the lyophilized matter threefold with the use of 0.6 mol l1 perchloric acid. Content of 8 biogenic amines (HIM, TYM, PHE, TRM, PUT, CAD, SPD, SPN) was determined by the method of high performance liquid chromatography (LabAlliance, State College, USA; Agilent Technologies, Agilent, Paolo Alto, USA) after the preceding derivatization by dansylchloride. Derivatization, chromatographic separation (Cogent column HPS C18, 150 4.6 mm, 5 mm, Cogent, Eatontown, USA) and detection (spectrophotometrically l ¼ 254 nm) were performed lá, Pechová, Komprda, according to Dadáková et al. (2009) and Sme (2004). An example of chromatogram of a stanKlejdus, and Kubán dard mixture of biogenic amines and chromatogram of extract from a body of Atlantic salmon is shown in Fig. 1. Every sample was extracted three times, every extract was derivatized twice and every derivatized mixture was spread on the column (n ¼ 18) three times. Results were expressed for the fresh matter before lyophilization. Differences between biogenic amines concentrations in tested bodies were statistically evaluated by KruskaleWallis and Wilcoxon tests. KruskaleWallis test was applied also for the analysis of variance (the evaluation of effect of storage methods and samples groups on BA occurrence). Statistical programme UnistatÒ 5.5 (Unistat Ltd., London, UK) and the significance level of 0.05 were used for the tests. 3. Results and discussion TRM was detected only in two fish samples (carp and pike perch) in the amounts between 0.9 and 2.5 mg/kg (data not shown). PHE was recorded in 13 fish samples (5 Atlantic cods, 4 Atlantic salmons, 2 Alaska pollocks and a sample of European hake and striped catfish). In above mentioned cases, the concentrations of PHE between 0.5 and 4.9 mg/kg were recorded (data not shown). Silla Santos (1996) and Ten Brink et al. (1990) suggest (from point of view of health risk) the limit for PHE income at the level of 30 mg/ kg. The latter mentioned limit was not exceeded in tested bodies. Relatively low content of PHE (<4 mg/kg) in fish is presented also by Hwang et al. (2012) and Zhai et al. (2012). In 107 samples, TYM was not detected or the concentrations ranged below 10 mg/kg (data not shown). Only in 5 bodies (4 Atlantic salmons and 1 carp), TYM concentrations between 10 and 50 mg/kg were recorded, which could be evaluated as safe for healthy consumers (Halász et al., 1994; Silla Santos, 1996; Ten Brink et al., 1990). Low TYM content (practically < 50 mg/kg) in most of the fish samples have been published by other authors (e.g. Hwang et al., 2012; Krí zek et al., 2011; Zhai et al., 2012). HIM, PUT, CAD, SPD and SPN content determined in the samples of bodies of freshwater and sea fish and other seafood are shown in Table 1. The only controlled biogenic amine (by the national food inspection authority) is HIM. HIM was not detected in 78 samples. In 19 tested bodies, HIM concentrations below 10 mg/kg were recorded. In other 9 cases, HIM concentrations of were below 100 mg/kg (P < 0.05) which is still considered as the safe limit for a healthy human (Halász et al., 1994; Silla Santos, 1996; Ten Brink et al. 1990). On the other hand, in 6 bodies (two Greenland halibuts, and a sample of Atlantic mackerel, Atlantic cod, striped catfish and Atlantic pomfret) the limit of 100 mg/kg was significantly exceeded (P < 0.05). Although the all latter mentioned fish samples were stored at freezing temperatures (the fish bodies were obtained from six different restaurants), the concentrations reached even the level above 1000 mg/kg (P < 0.05), which should be considered as dangerous (Halász et al., 1994; Silla Santos, 1996; Ten Brink et al. 1990). These results emphasize the necessity of BAs monitoring and evaluation of health risks.
Fig. 1. Separation of mixtures of biogenic amines as dansylamides. Part A: a standard mixture (tryptamine, phenylethylamine, putrescine, cadaverine, histamine, ISTD e internal standard e 1,7eheptandiamine, tyramine, spermidine and spermine) in concentration of 50 mg/l in acetonitrile. Part B: a derivatized sample extracted from the body of Atlantic salmon.
PUT was the most abundant biogenic amine observed in our samples (Table 1). Only in 14 cases, PUT was not detected. PUT concentrations up to 10 mg/kg and between 10 and 100 mg/kg were recorded in 26 and 69 samples, respectively. Three bodies (Atlantic salmon, Wels catfish and scallop) possessed PUT concentration >100 mg/kg (P < 0.05), which should be pointed out as serious. In 92 bodies, CAD was not detected or the concentrations were up to 10 mg/kg. More significant CAD concentrations were observed in 20 cases, where only once the amount exceeded the level 50 mg/kg (P < 0.05). PUT and CAD could act as potentiators of toxicological effects of HIM and TYM (Arnold & Brown, 1978; Halász
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et al., 1994; Prester, 2011; Silla Santos, 1996; Ten Brink et al. 1990). The occurrence of PUT and CAD is usually related to the hygiene conditions in the process of meat treatment and could point out to the presence of contaminating microflora, especially gramnegative strains of family Enterobacteriaceae and genus Pseudomonas (Halász et al., 1994; Hwang et al., 2012; Rawles et al., 1996; Zhai et al., 2012). In nearly 80% of samples, SPD and SPN concentrations were very low (up to 10 mg/kg) or these polyamines were not detected. SPD concentration ranging between 50 and 100 mg/kg was determined in the only case of Atlantic salmon. In two fish samples (striped catfish and escolar) SPN concentration above 50 mg/kg were observed. Apart from the concentrations of individual BAs, the sum of determined BAs amounts was also evaluated. In almost 85% of tested samples, the sum of BAs was below 100 mg/kg. On the other hand, in 7% of observed bodies, the BAs concentrations between 100 and 200 mg/kg were recorded. Surprisingly, in 8% of samples, the BAs concentrations above 200 mg/kg and in 5% of bodies even the amounts exceeded 1000 mg/kg. The latter mentioned concentrations represent amounts which could cause serious health problems even for healthy consumers. Unfortunately, potentially dangerous concentrations of BAs were also observed in fish bodies which are widely served in restaurants in the Middle Europe (e.g. Atlantic mackerel, Atlantic salmon and striped catfish). Analysis of variance showed no significant difference between the samples stored frozen and “on ice” (P 0.05). Similarly, the BAs concentration in freshwater and sea fish did not differ significantly (P 0.05). The BAs concentrations of the other seafood samples were not significantly higher in comparison with the bodies of freshwater and sea fish (P 0.05). 4. Conclusion Monitoring of the occurrence of 8 biogenic amines in the bodies of 112 fish and other seafood obtained from restaurants in the Czech Republic has been performed. In more than three quarters of samples, the sum of concentrations of biogenic amines was under 100 mg/kg, which could be considered to be a safe limit for healthy consumers. On the other hand, in 8% of tested samples, dangerous amounts of BAs were found out and in 6 cases even legislatively set limits were exceeded. Although fish and fish products should be a part of human diet, it is important to deal with their short shelf-life which could be determined also by biogenic amines concentrations.
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