Biogenic amines in selected foods of animal origin obtained from the Croatian retail market

ABSTRACT

This study aimed to investigate the presence of eight biogenic amines (BAs): tryptamine (TRP), phenylethylamine (PHE), putrescine (PUT), cadaverine (CAD), histamine (HIS), tyramine (TYR), spermidine (SPD) and spermine (SPM) in cheese, fish & fishery products and meat & meat products obtained from the Croatian retail market. A selective and robust method of high-performance liquid chromatography (HPLC) with diode array detection (DAD) was applied for the determination of BAs in a total of 91 samples in accordance with the performance criteria outlined in the European legislation. A high inter- and intra-food group variability of the amounts of BAs was observed. In the analysed samples, the most represented amines were TYR, HIS, CAD and PUT. Based on the highest content of the most toxic BAs (HIS and TYR) and consequential food safety concerns, the studied food groups can be ranked in the following order: cheese (HIS up to 106.4 mg/kg; TYR up to 206.6 mg/kg), fish &fishery products (HIS up to 98.8 mg/kg; TYR up to 47.9 mg/kg), and meat & meat products (HIS up to 20.0 mg/kg; TYR up to 117.5 mg/kg). The total BA content was significantly higher (p < .05) in fermented in comparison with other food. The study aimed to contribute to the knowledge on BA toxicity and food quality, as well as to support the indispensable future studies of consumption data and exposure assessment, to the end of defining allowable BA concentrations in food.     

Formation of cadaverine, histamine, putrescine and tyramine by bacteria isolated from meat, fermented sausages and cheeses

Enterobacteriaceae (EB, n = 149), Lactobacillus (LB, n = 162) and Leuconostoc sp. (LC, n = 89) and enterococci (EC, n = 137), isolated from raw meat (n = 65), fermented sausages (n = 50) and cheese (n = 55) samples, were cultivated in a broth containing precursor amino acids (each 3 g/l). After incubation, the liquid culture was chemically analysed for cadaverine (CAD), putrescine (PUT), histamine (HIS) and tyramine (TYR) formation at pH 5.2 and at pH 6.7. The majority of EB isolates (147 of 149) was capable of forming >100 mg/l of either CAD or PUT. Among the most frequently isolated species Hafnia alvei and Serratia liquefaciens, formation of >100 mg/l HIS occurred, but with low prevalence (1.6 and 6.5%, respectively). Twelve of 149 isolates (8%) were able to produce more than 10 mg/l HIS. One hundred forty-two isolates (95.3%) produced less than 10 mg/l TYR, and 7 isolates (4.7%) 10 mg/l to a maximum of 35.3 mg/l TYR. For LB + LC, one isolate (Leuconostoc mesenteroides ssp. mesenteroides) formed >100 mg/l PUT and one >100 mg/l CAD (of all 251 LB + LC isolates 0.4% each). Formation of >100 mg/l HIS and TYR was detected in 3.6 and 19% of the LB + LC isolates, respectively. For the EC isolates, maximum levels for PUT, CAD and HIS were 25.4 mg/l, 6.0 mg/l and 15.7 mg/l, respectively. TYR was formed in quantities of 100–1000 mg/l by 47.9% of EC faecalis (n = 75), and 59.7% of EC faecium (n = 62) isolates. More than 1000 mg/l TYR were formed by 50.7 and 35.5% of the isolates, respectively. A low initial pH of 5.2 compared to the initial pH of 6.7 favoured tyramine production by lactic acid bacteria, but was associated with lower CAD yield by EB. A considerable intra-species variability in amine formation was observed.     

Determination of biogenic amines in selected Malaysian food

The biogenic amines tryptamine (TRP), putrescine (PUT), histamine (HIS), tyramine (TYR) and spermidine (SPD) were determined in 62 selected food items commonly consumed in Malaysia. This include the local appetisers “budu” and “cincalok”, canned fish, salt-cured fish, meat products, fruit juice, canned vegetables/fruits and soy bean products. After the aqueous extraction, the samples were derivatised with dansyl chloride before analysing using reversed phase HPLC with UV detection. Mean levels of TRP, PUT, HIS, TYR and SPD in eight budu samples were 82.7, 38.1, 187.7, 174.7 and 5.1 mg kg⁻¹, respectively. The main biogenic amines found in cincalok were PUT, HIS and TYR where the mean values were 330.7, 126.1 and 448.8 mg kg⁻¹, respectively. With the exception of “pekasam” and “belacan”, significantly lower levels of biogenic amines were found in canned fish and salt-cured fish samples. Non detectable or low levels of biogenic amines were found in meat, fruit juice and canned vegetables/fruit samples.     

Characterisation and application of Halomonas shantousis SWA25, a halotolerant bacterium with multiple biogenic amine degradation activity

Biogenic amines are identified as toxicological substances in foods and may have detrimental effects on consumers’ health. In recent years, the application of microorganisms that can degrade biogenic amines has become an emerging method for their reduction. The degradation characteristics and application potential of a salt-tolerant bacterium Halomonas shantousis SWA25 were investigated in this study. H. shantousis SWA25 exhibited degradation activity against eight biogenic amines at 10–40°C (optimum, 30–40°C) and pH 3.0–9.0 (optimum, 6.0–7.0) in the presence of 0–20% (w/v) NaCl (optimum, 0%). Specifically, H. shantousis SWA25 degraded all tryptamine (TRY) and tyramine (TYR) in 6 h, all phenethylamine (PHE) in 9 h, 66.7% of histamine (HIM), 52.4% of cadaverine (CAD), 48.0% of spermidine (SPD), 42.9% of putrescine (PUT) and 42.0% of spermine (SPM) in 20 h at 30°C and pH 7.0 with shaking at 120 r min^?1. The enzymes from H. shantousis SWA25 responsible for degradation of biogenic amines were mainly amine oxidases located on the cell membrane. Further studies showed that H. shantousis SWA25 effectively degraded TRY, PHE, PUT, CAD, HIM and TYR in commercial fish sauce and soy sauce samples. Nevertheless, significant SPD and SPM degradation were not observed due to low initial concentrations. Therefore, H. shantousis SWA25 can be applied as a potential biogenic amines degradation bacterium in foods.     

阿魏酸对风干鲈鱼中N-亚硝胺及生物胺的抑制作用

在风干鲈鱼腌制过程中分别添加0、50、100、150、200 mg/kg的抑制剂阿魏酸,15 ℃、相对湿度80%～90%条件下风干84 h,研究阿魏酸对风干鲈鱼中N-亚硝胺及生物胺含量等安全指标影响。结果显示,阿魏酸对风干鲈鱼中腐胺、尸胺、组胺、酪胺的生成具明显的抑制作用,但对样品中精胺和亚精胺的生成具有促进作用。当阿魏酸添加量不超过150 mg/kg时,随着阿魏酸添加量的提高,N-二甲基亚硝胺含量显著降低。贮藏27 d时对照组菌落总数为7.59 （lg（CFU/g））,添加200 mg/kg阿魏酸组菌落总数为5.45（lg（CFU/g））,相对于对照组降低了28.20%。结果表明：添加阿魏酸在不影响风干鲈鱼感官品质的基础上能显著降低风干鲈鱼中生物胺总量、微生物菌落总数、N-二甲基亚硝胺含量、亚硝酸盐残留量。     

Changes in biogenic amines in mature and germinating legume seeds and their behavior during cooking

Ungerminated legume seeds (broad bean, chick pea and lupine) were contained all tested biogenic amines. Tryptamine (TRY) was the main biogenic amine detected, and its concentration considerably increased during the germination. beta-Phenylethylamine (PHE) was detected in small amounts and its concentration slowly increased during germination. The concentration of tyramine (TYR) showed a fluctuation pattern of changes during germination in all tested legumes. The concentrations of cadaverine (CAD) and putrescine (PUT) increased during the germination period in all tested grains. However, histamine (HIS) showed a fluctuated pattern of changes in both broad bean and lupine, and a gradual increase in chick pea. Spermidine (SPD) and spermine (SPM) contents of broad bean and chick pea showed a fluctuation pattern of change, while, a decrement trend of change was recorded for lupine along the germination period. By cooking, legume samples became free of biogenic amines which appeared in the boiling water. Heat treatment seems to have little effect on the concentration of biogenic amines in legume sprouts. The amounts of biogenic amines detected in the boiling water are less than the initial amounts of the sprouts (expected amounts).     

Simple,rapid and valid thin layer chromatographic method for determining biogenic amines in foods

A simple, rapid and valid thin layer chromatographic (TLC) method for determining biogenic amines in foods is established. Biogenic amines were extracted from foods with 5% TCA, the extract was washed by ethyl ether, the washed extract was dried and the dansyl derivatives were prepared. Separation of the dansylated amines was achieved on silica gel TLC plates by multiple development technique, and quantification was performed by densitometry at 254 nm. The response of the densitometer was linear and highly correlated with the amounts of dansylamines. The calculated determination coefficient (r²) ranged from 0.997 to 0.970. The sensitivity of the method was 45, 174, 703, 101, 132, 88, 515 and 61 for TRY, PUT, CAD, SPD, HIS, SPM, TYR and PHE, respectively. The relative standard deviation (RSD) of the proposed method (repeatability) ranged from 0.39 to 6.36% for TYR (80 ng) and PHE (20 ng), respectively. The accuracy of the proposed method ranged from 86.2 to 93.2% for PHE and HIS, respectively, and there was no significant differences (p<0.05) between the recoveries obtained for all kinds of food analyzed. The limit of detection of the proposed method was found to be 10ng for TRY, SPM and PHE, and 5 ng for the other biogenic amines tested. ©     

Lead,cadmium and organochlorine pesticide residues in hunted red deer and wild boar from northern Italy

The objectives of the present study were to assess heavy metal cadmium (Cd), lead (Pb) and organochlorine pesticide concentrations in tissues of red deer (Cervus elaphus) and wild boar (Sus scrofa) from nine hunting areas and to evaluate related risk factors for the host animal. Over a period of 2 years, a total of 1055 and 210 masseters, 424 and 201 livers, 642 and 152 kidneys were collected from wild boar and red deer, respectively, and concentrations of Cd, Pb and organochlorine pesticides were determined. Comparing the two species, Cd concentration in the kidney (3.72 mg/kg), liver (0.67 mg/kg) and muscle (0.02 mg/kg) of wild boar was found to be significantly higher than in the organs of red deer (1.02 mg/kg in the kidneys, 0.07 mg/kg in the liver and 0.006 mg/kg in muscle). Mean Pb concentrations were found to be similar in both animals, with 0.39, 0.52 and 2.60 mg/kg detected in the wild boar kidney, liver and muscle, respectively, and 0.24, 0.21 and 2.04 mg/kg in the respective organs of the red deer. No difference in concentrations were found based on age class, location of tissue sample or contaminant in the case of wild boar. By contrast, a significantly lower Cd concentration was found in the kidney of the young red deer. The search for organochlorine pesticides in both red deer and wild boar produced negative results with values below the limits of detection. Due to the high levels of renal Cd and muscle Pb detected in wild boar and red deer, further research needs to be carried out in an effort to identify the source of contamination and preserve the health of animals and humans.     

Concentration of biologically active polyamines in rabbit meat, liver and kidney after slaughter and their changes during meat storage and cooking

The concentration of putrescine (PUT), spermidine (SPD) and spermine (SPM) was determined in chilled meat and kidneys of 18 rabbits and in liver of 12 animals 24 h after slaughter. Very low PUT concentrations were detected only in kidneys. Mean SPD levels were 2.2, 2.2, 61.7 and 32.7 mg kg^− 1 in saddle, leg, liver and kidneys, respectively. The respective SPM concentrations were 14.7, 8.0, 115 and 88.4 mg kg^− 1. SPD and SPM losses of about one third of the initial levels were apparent in saddles stored at − 18 °C for 8 months. Losses of both polyamines of about 15-20% of the initial concentrations were found in saddles stored aerobically at + 2 °C for up to 9 days. Stewing of saddles caused significant SPD and SPM losses of about 20-25%, while upon roasting and pan-roasting without oil a decrease of about 50% of the initial concentration was observed.     

Concentration of biologically active polyamines in meat and liver of sheep and lambs after slaughter and their changes in mutton during storage and cooking

Putrescine (PUT), spermidine (SPD) and spermine (SPM) concentrations using a UPLC method, in chilled mutton, lamb and livers 24 h after slaughter were determined. PUT concentrations were quantifiable only in some samples. Mean SPD concentrations were 4–6, 13.5 and 16.8 mg kg⁻¹ in the meats, sheep and lamb livers, respectively. The respective SPM concentrations were 17–25, 128 and 79 mg kg⁻¹. SPD and SPM losses of about one fifth and half of the initial level, respectively, were apparent in mutton loins stored at −18 °C for 6 months. Significant losses of SPD and SPM were found in mutton loins stored aerobically, vacuum-packaged or in a modified atmosphere at + 2 °C. Boiling and stewing of mutton legs caused SPD and SPM losses of about 40% and roasting of about 60% of the initial content.     

BIOGENIC AMINE CONTENT OF SOME TURKISH CHEESES

Biogenic amines in ten samples of Kaşar (aged), five samples of Kaşar (fresh), four samples of Mihaliç, nine samples of Van Otlu (herbed), nine samples of Örgü, nine samples of Urfa and six samples of Civil cheeses were obtained from local supermarkets of Ankara. Tryptamine (TRY), phenylethylamine (PHA), putrescine (PUT), cadaverine (CAD), histamine (HIT), tyramine (TYA), spermine (SPM) and spermidine (SPD) contents of Turkish cheese samples were screened by high performance liquid chromatography (HPLC).
PUT, CAD, HIT, TYA and SPD were the predominant biogenic amines in maturated Kaşar (aged), vacuum-packed Kaşar (fresh), Mihaliç, Otlu (herbed) and Örgü cheeses. PHA, CAD, TYA and SPD were the predominant biogenic amines in Urfa cheese samples.
Civil cheeses were found to contain the highest HIT and TYA content with an average of 94.76 mg/100 g and 138.16 mg/100 g, respectively, much higher than the toxic dose. HIT level of Mihaliç cheeses were also higher than the accepted limit. The concentration of amines in all other cheeses was much lower than the toxic dose limits.     

Production of Biogenic Amines by Lactic Acid Bacteria Isolated from Uzicka Sausages

The aim of this study was to monitor production of seven biogenic amines (Cadaverine – CAD, Putrescine – PUT, Spermine – SPE, Spermidine – SPD, Histamine – HIS, Tyramine – TYR and Tryptamine – TRY) in selected 24 lactic acid bacteria (LAB) strains. The decarboxylase activity of the microorganisms was studied in growth medium after 24 h cultivation. The ability of 24 LAB isolates cultivated in MRS broth and M17 broth supplement with 0.5% glucose to produce biogenic amines was assessed using liquid chromatography tandem mass spectrometry (LC-MS/MS). The investigation showed that LAB isolated from Uzicka sausage are not significant producers of biogenic amines in vitro.     

Changes in the content of biologically active polyamines during storage and cooking of pig liver

Dietary polyamines putrescine (PUT), spermidine (SPD) and spermine (SPM) participate in numerous human physiological processes, including tumour growth. Eight experiments with pig liver were carried out. In two, livers were stored at -18°C for 168 days, in four, livers were stored aerobically (AE), vacuum-packaged (VP) and packaged in a modified atmosphere (MO; 70% N(2) and 30% CO(2), v/v) at +2°C for 9, 21 and 21 days, respectively, and in two, the effects of four cooking treatments were tested. Polyamines were determined as dansyl derivatives using an HPLC method. Distribution of both SPD and SPM in the four main liver lobes was homogenous. The initial SPD and SPM contents in 14 livers 24h after slaughter were 23.3±6.7 and 94.5±19.6mgkg(-1), respectively. The putrescine content was below the limit of detection. The content of SPD and SPM decreased during frozen-storage to about 70% of the initial values. On day-9 of storage, mean SPD and SPM contents decreased to about 85% of the initial values in livers stored in MO and to about 75-80% in AE and VP at 2°C. The decrease continued more extensively in VP than in MO. PUT was detected from day-15 of VP and MO storage. There was a significant decrease in SPD and SPM, to about 70-60% of the initial content during cooking.     

Biologically active polyamines in beef, pork and meat products: A review

Dietary polyamines (PAs) putrescine (PUT), spermidine (SPD) and spermine (SPM) participate in an array of roles in human metabolism. Nevertheless, under some physiological conditions they can be undesirable. Meat and meat products are among important sources of PAs in human nutrition, mainly of SPM. The usual contents of PUT, SPD and SPM in fresh beef and pork are <2, <5 and 20-40mgkg(-1), respectively. Current information on changes of PAs during meat storage corresponds with PUT formation by bacterial activity mainly of pseudomonads and Enterobacteriaceae. However, data on SPD and SPM changes during meat chill-storage have been inconsistent. Culinary processing of meat probably does not change SPD and SPM levels. PUT can be formed in different meat products in relation to the microbial population of the raw materials used and the hygienic level of manufacturing process. SPD and SPM contents seem to remain stable during processing of non-fermented meat products or decrease during dry-cured ham ripening. PUT contents increase commonly to 60-140mgkg(-1) in dry spontaneously fermented sausages, however, contents up to several hundreds mgkg(-1) are not extraordinary. Starter cultures are usually able to decrease PUT formation considerably. SPD and SPM contents in dry fermented sausages are comparable with levels typical for fresh meat. Data on SPD and SPM changes during ripening and storage are inconsistent. A decrease of the both polyamines during a storage period has been usually reported.     

Effects of rosemary and sage tea extract on biogenic amines formation of sardine (Sardina pilchardus) fillets

Natural antimicrobials and antioxidants from rosemary (Rosmarinus officinalis) and sage tea (Salvia officinalis) were produced using solvent extraction method. The effect of two extracts on ammonia (AMN) and biogenic amines (BAs) formation in vacuum packed sardine (Sardina pilchardus) fillets stored at 3 ± 1 °C was investigated for 20 days. Although the effect of extracts was dependent on specific amine and storage time, phenolic compounds from rosemary and sage tea generally resulted in lower AMN and BAs accumulation in sardine muscle. Putrescine (PUT) and cadaverine (CAD) were the most abundant amines, while histamine (HIS) concentration ranged from 2.05 to 28.77 mg 100g^?1. Rosemary and sage tea extracts significantly reduced HIS, PUT, CAD and trimethylamine accumulation in the fish muscle (P < 0.05) while stimulating effect of extracts was observed on serotonin and agmatine formation. At the end of the storage period, PUT and CAD contents of control were 100‐fold higher than those of treated groups.     

Content of polyamines in beef and pork after animal slaughtering

Dietary polyamines putrescine (PUT), spermidine (SPD) and spermine (SPM) participate in an array of important human physiological roles, including tumour growth. Physicians and dieticians thus need reliable information on polyamine contents in foods. However, data for both fresh and processed beef and pork are limited. We therefore, determined the initial content of the polyamines, 24 h after slaughtering, in sirloin and rump of 63 young bulls and in loin and leg of 27 pigs of both genders. Polyamines were determined as N-benzamides by micellar electrokinetic capillary chromatography (MECC). PUT and SPD contents in most of the meat samples were negligible. Mean SPM contents were about 22±6 mg kg⁻¹ in sirloin and rump. No significant correlations at P<0.05 were found between the SPM contents in sirloin and rump, likewise between SPM contents and the age and between live weight and type of the efficiency of the bulls. Mean SPM contents in loin and legs of barrows were 26.1±7.0 and 28.4±8.5 mg kg⁻¹, respectively, while the corresponding values in gilts were 22.3±10.6 and 18.3±9.3 mg kg⁻¹.Between the genders the difference in SPM contents in the legs was significant. SPM contents relatively vary widely within the individual kinds of meat, which complicates application of the results for the controlled human nutrition.     

Elemental composition of game meat from Austria

Concentrations of 26 elements (B, Na, Mg, P, S, K, Ca, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Se, Rb, Sr, Mo, Cd, Sb, Ba, Hg, Pb, U) in wild game meat from Austria were analysed using an inductively coupled plasma mass spectrometer. All investigated animals were culled during the hunting season 2012/2013, including 10 chamois (Rupicapra rupicapra), 9 hare (Lepus europaeus), 10 pheasant (Phasianus colchicus), 10 red deer (Cervus elaphus), 12 roe deer (Capreolus capreolus) and 10 wild boar (Sus scrofa). In 19 out of 61 meat samples lead concentrations were higher than 0.1 mg/kg, the maximum limit in meat as set by the European Commission (Regulation EC No 1881/2006), which is most likely caused by ammunition residues. Especially, pellet shot animals and chamois show a high risk for lead contamination. Despite ammunition residues all investigated muscle samples show no further health risk with respect to metal contamination.     

The effects of gender,age and region on macro- and micronutrient contents and fatty acid profiles in the muscles of roe deer and wild boar in Mecklenburg-Western Pomerania (Germany)

Samples of M. longissimus were collected from a total of 203 feral roe deer (n = 118) and wild boar (n = 85) in two regions of Mecklenburg-Western Pomerania (Germany). The muscle lipid saturated fatty acid proportions of roe deer and wild boar ranged between 33 and 49 g/100 g total fatty acids and 31 and 35 g/100 g total fatty acids, respectively. The total n − 3 PUFA proportions in roe deer muscle varied between 8.0 and 14 g/100 g fatty acids, and in wild boar muscle between 2.6 and 6.0 g/100 g fatty acids. The major vitamin E homologue, α-tocopherol, was determined to be between 5.8 and 13.1 mg/kg in roe deer muscles. Lower levels between 1.2 and 4.7 mg/kg were measured in wild boar muscles. The iron and zinc concentrations in roe deer and wild boar muscle ranged from 26.3 to 33.9 mg/kg and from 17.0 to 21.7 mg/kg, and from 13.6 to 39.3 mg/kg and 18.1 to 31.9 mg/kg, respectively.     

Changes in the content of biologically active polyamines during pork loin storage and culinary treatments

Dietary polyamines putrescine (PUT), spermidine (SPD) and spermine (SPM) participate in numerous human physiological processes, including tumour growth. Physicians and dieticians thus need reliable information on their contents in foods. However, data for processed meat are rather limited. Eight experiments with pork loin were therefore carried out. Loins were stored at −18 °C for 168 days in three experiments, in next three ones pork was stored aerobically, vacuum-packaged and packaged in a modified atmosphere (70% N₂ and 30% CO₂, v/v) at +2 °C for 9, 21 and 21 days, respectively. The effects of five usual culinary treatments were tested in two experiments. The polyamines were determined as dansyl derivatives using a HPLC method. Only SPM was determined at levels 20.3–25.2 mg kg⁻¹ in fresh loins; PUT and SPD contents were below the detection limits. SPM content slightly, but significantly (P < 0.05) increased during frozen storage, while it mildly and insignificantly (P > 0.05) decreased during all three variants of cold storage. SPM losses of about 40% of the initial content were observed during boiling, stewing, pan-roasting without oil and breaded pork frying, while roasting caused the loss of about 55%. Similar losses were found for loins processed 24 h and 7 days after slaughter. No SPM content was detectable in broth and grease produced during the processing.     

Content of biologically active polyamines in livers of cattle, pigs and chickens after animal slaughter

Dietary polyamines putrescine (PUT), spermidine (SPD) and spermine (SPM) participate in an array of important human physiological roles, including tumour growth. Physicians and dieticians thus need reliable information on polyamine contents in foods. However, data for livers are lacking. We determined therefore the content of these polyamines 24 h after slaughter in livers of young bulls, cows, pigs and chicken in 58, 19, 36 and 38 samples, respectively. Polyamines were determined as N-benzamides by micellar electrokinetic capillary chromatography. Mean PUT contents about 25 mg kg⁻¹ were found in cattle livers, while very low or negligible contents were determined in livers of the other animals. Extremely high mean SPD contents of 122 and 161 mg kg⁻¹ were found in livers of bulls and cows, respectively and mean levels of 32 and 57 mg kg⁻¹ in livers of pigs and chicken. An opposite relation was observed for SPM. Its mean contents were 43, 35, 115 and 120 mg kg⁻¹ for bulls, cows, pigs and chicken livers, respectively. Thus, livers of the tested animal species belong among foods with the highest polyamine contents. However, the contents ranged very widely, that makes application of the results for the control of human nutrition rather difficult. Polyamine contents in bovine blood were found to be below the detection limits of 2.1, 1.0 and 1.4 mg kg⁻¹ for PUT, SPD and SPM, respectively. Thus, the blood content did not contribute to the substantial polyamine contents in bovine liver found in this study.