发酵剂对牦牛乳硬质干酪成熟过程中生物胺的影响

乳酸菌产生物胺的能力具有菌株特异性,因此,为了探究不同种类发酵剂对牦牛乳硬质干酪中生物胺形成的影响,该试验利用高效液相色谱对3种不同发酵剂制作的硬质干酪成熟过程中生物胺进行了测定和分析。结果表明,嗜热和嗜温发酵剂牦牛乳硬质干酪中检测出2-苯乙胺、腐胺、尸胺、组胺和酪胺,混合发酵剂干酪中检测出腐胺、2-苯乙胺、尸胺和酪胺。各生物胺之间呈现正相关性。3种不同发酵剂干酪在1～6个月成熟过程中,其各生物胺整体呈现增加趋势,嗜热、嗜温和混合发酵剂干酪中总生物胺最高含量分别为(448.3±9.6)、(456.8±58.4)、(293±24.5)mg/kg。组胺和酪胺是2种毒性相对高的生物胺,嗜热发酵剂干酪中组胺和嗜温发酵剂干酪中酪胺最高,其最高含量分别为(20.8±7.9)、(92.9±6.7)mg/kg,混合发酵干酪中未检测出组胺,酪胺含量次之,3种不同发酵剂干酪中组胺、酪胺含量均低于推荐安全剂量50 mg/kg和100 mg/kg。这为合理选择发酵剂和控制干酪中生物胺形成提供了依据。     

Evaluation of biogenic amines and microbial counts throughout the ripening of goat cheeses from pasteurized and raw milk

The effect of the hygienic quality of milk on changes in microbial counts and biogenic amine content was evaluated during ripening of goat cheeses manufactured from pasteurized and raw milks at 1, 14, 30, 60 and 90 d. The original milk, rennet, curd and whey were also included in the study. The pH, salt content and extent of proteolysis in the cheese were also evaluated. Spermidine and spermine were the main amines in raw milk, while they were minor amines in cheeses. Other amines increased markedly during ripening, tyramine being the main amine in cheese made from raw milk and cadaverine and putrescine in those produced from pasteurized milk. Enterobacteriaceae counts decreased during ripening whereas those of lactic acid bacteria increased, especially lactobacilli and enterococci. Cheese made from raw milk showed higher microbial counts during ripening than those made from pasteurized milk, especially for Enterobacteriaceae and enterococci, counts being 2 or 3 log units higher. Raw milk cheese showed remarkably higher biogenic amines compared with pasteurized milk cheeses. Therefore, pasteurization of milk causes a decrease in final biogenic amine content of cheese as a result of the reduction of its microbial counts.     

Effect of added proteinases and level of starter culture on the formation of biogenic amines in raw milk Manchego cheese

The influence of two proteinases (Bacillus subtilis neutral proteinase and Micrococcus sp. cysteine proteinase) and two starter culture levels (0.1% and 1%) on biogenic amine formation has been studied in raw ewes' milk Manchego cheese. Amino acid decarboxylating micro-organisms were determined on tyrosine enriched selective media. Biogenic amines were analysed by capillary electrophoresis in citrate buffer at pH 3.6. Addition of proteinases and level of starter culture did not influence the population of micro-organisms with amino acid decarboxylating activity, which represented on average 1% of the bacterial population in 30-day-old cheeses. Tyramine and histamine were detected in all batches of cheese from day 30. Concentrations of tyramine and histamine were higher in cheeses made from milk with neutral proteinase (up to 356 and 284 mg kg(-1), respectively, after 90 days) than in cheeses made from milk with cysteine proteinase (up to 269 and 189 mg kg(-1), respectively) or with no proteinase added (up to 305 and 226 mg kg(-1), respectively). Formation of tyramine and histamine was also favoured in cheeses made with 1% starter culture with respect to cheeses made with only 0.1% starter culture, probably due to the higher pH values of the former cheeses. After 90 days of ripening, concentrations of 10-20 mg kg(-1) phenylethylamine were observed in 9 of the 12 batches, and levels < 10 mg kg(-1) tryptamine were only detected in 3 batches, with no significant relationship between the concentration of these amines and proteinase addition or level of starter culture.     

不同NaCl添加量对牦牛乳硬质干酪成熟过程中生物胺产生的影响

以不同盐添加量（1.0%、1.3%、1.8%和2.3%）牦牛乳硬质干酪为研究对象,分析牦牛乳硬质干酪0～6 个月成熟过程中生物胺的动态变化,并对干酪中产胺微生物进行筛选。结果显示：不同盐添加量牦牛乳硬质干酪中生物胺主要为色胺、β-苯乙胺、尸胺、酪胺和腐胺,未检测到组胺,生物胺积累阶段主要集中在成熟后期。不同盐添加量干酪中色胺含量最低,且在成熟后期含量差异较小。当盐添加量从2.3%减少到1.0%时,干酪中β-苯乙胺含量减少。盐添加量分别为1.0%和1.3%时,干酪中尸胺含量较低,且未检测出腐胺。当盐添加量在1.8%～2.3%时,随着盐添加量的增加,干酪中尸胺和腐胺含量整体呈现增加趋势。不同盐添加量干酪成熟过程中,其酪胺含量范围为3.13～49.81 mg/kg,且盐添加量为1.3%干酪中酪胺含量较高。不同盐添加量干酪中生物胺总量最高为304.18 mg/kg。采用显色培养基筛选出一株产胺微生物,经分子生物学鉴定为Enterococcus durans。     

Formation of biogenic amines in a typical semihard Italian cheese

Given that the concentration of biogenic amines in cheeses depends on variety, age, and type of microflora, a study was undertaken to investigate the formation of these compounds during the ripening of a typical semihard Italian cheese. Tryptamine, phenylethylamine, putrescine, cadaverine, histamine, and tyramine contents were calculated in 30 samples of Montasio cheese characterized by different levels of proteolysis. Histamine and tyramine were the major amines. Tryptamine and phenylethylamine concentrations were very low at all ripening periods. Putrescine and cadaverine were present only in samples with anomalous fermentation processes. The relationship between the total amine content and the proteolytic maturation coefficient was calculated; however, even for higher levels of proteolysis, the biogenic amine content in Montasio cheese was below the level considered potentially toxic.     

Monitoring of biogenic amines in cheeses manufactured at small-scale farms and in fermented dairy products in the Czech Republic

The aim of the study was the monitoring of six biogenic amines (histamine, tyramine, phenylethylamine, tryptamine, putrescine, and cadaverine) and two polyamines (spermidine and spermine) in 112 samples of dairy products purchased in the Czech Republic, namely in 55 cheeses made in small-scale farms and in 57 fermented dairy products. The products were tested at the end of their shelf-life period. Neither tryptamine nor phenylethylamine was detected in the monitored samples; histamine was found only in four cheese samples containing up to 25 mg/kg. The contents of spermine and spermidine were low and did not exceed the values of 35 mg/kg. Significant amounts of tyramine, putrescine, and cadaverine occurred especially in cheeses produced from ewe’s milk or in long-term ripened cheeses. In about 10% of the tested cheeses, the total concentration of all the monitored biogenic amines and polyamines exceeded the level of 200 mg/kg, which can be considered toxicologically significant. In fermented dairy products, the tested biogenic amines occurred in relatively low amounts (generally up to 30 mg/kg) that are regarded safe for the consumer’s health.     

Influence of starter and nonstarter on the formation of biogenic amine in goat cheese during ripening

Two commercial starters were investigated for their potential ability to decarboxylate amino acids during goat cheese ripening. Two batches of goat cheese were produced with identical pasteurized milk but different starter cultures. One of them contained Lactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris and the other Lactococcus lactis subsp. lactis. The amine contents, microbial counts, proteolysis-related parameters, pH, total solids and salt content were studied in raw materials and cheeses. In raw materials, polyamines were the prevailing amines, whereas the main amines in cheeses were putrescine, tryptamine and, in particular, tyramine (94.59 mg/kg). Aerobic mesophilic microorganisms and Lactococcus counts increased throughout ripening, while Enterobacteriaceae were no longer detectable in cheese after 30 days of ripening. Amine concentration rose during cheese ripening in both batches. Moreover, the decarboxylase activity of microorganisms isolated from samples during cheese ripening was assayed and discussed.     

Influence of hygienic quality of raw materials on biogenic amine production during ripening and storage of dry fermented sausages

The effect of the hygienic status of raw materials on biogenic amine production during ripening and storage (at 4 and 15 degrees C) of fermented sausages was studied. Two portions of fresh lean and back fat pork were stored for 5 days at -20 degrees C (treatment A) and at 4 degrees C (treatment B), respectively. Raw materials of treatment A maintained their hygienic quality high and low amine content. Raw materials of treatment B showed from 1 to 3 log (CFU/g) higher microbial counts and a biogenic amine index near 50 mg/kg, indicating poorer hygienic quality. The quality of raw materials influenced the composition and the concentration of biogenic amines produced during the ripening sausages. Sausages of treatment A (A-sausages) showed a large accumulation of tyramine (up to 100 mg/kg dm) followed by putrescine and cadaverine (<15 mg/kg). In contrast, B-sausages resulted in earlier and much greater amine production, and cadaverine, tyramine, and putrescine levels were 50-, 2.6-, and 6.5-fold higher than those of A-sausages. Other biogenic amines, such as octopamine, tryptamine, phenylethylamine, and histamine were also produced in B-sausages. The higher proteolysis and the lower pH of B-sausages might have favored the decarboxylase activity of microorganisms. Biogenic amine contents of sausages during storage depended on the raw materials used and storage temperature. No significant modification on the amine contents was observed during the storage of A-sausages at either temperature. Greater changes occurred in B-sausages stored at 15 degrees C than in those stored at 4 degrees C. Higher temperatures favored proteolytic and decarboxylase reactions, resulting in increased amine concentrations after storage.     

Content and distribution of biogenic amines in Dutch-type hard cheese

Content of biogenic amines (BA; histamine, tyramine, tryptamine, putrescine, 2-phenylethylamine, cadaverine, spermidine, spermine) and counts of microorganisms (total aerobic and facultative anaerobic mesophilic microorganisms, TAC; lactic acid bacteria, LAB; coliforms; total anaerobic microorganisms, TAN; yeasts + moulds), were evaluated within the ripening interval of 1–6 months in Dutch-type hard cheese, produced by two different producers (the cheeses designated as R30 and R45) using pasteurized milk and different starter cultures (designated as R30-FD and R30-CH in the case of the R30 cheeses). The cheeses from the two producers differed in their fat content (30% and 45% of fat in dry matter, respectively).     

牦牛乳硬质干酪成熟过程中生物胺与细菌群落结构分析

成熟干酪易产生生物胺,生物胺对人体健康具有影响,因此,为了探究牦牛乳硬质干酪的质量安全性,本实验利用高效液相色谱和高通量测序技术对干酪成熟过程中生物胺和细菌群落结构进行测定和分析。结果表明：牦牛乳硬质干酪中主要生物胺为腐胺、2-苯乙胺、酪胺、组胺和尸胺,在1～6 个月成熟过程中,干酪中各生物胺含量呈现增加趋势。干酪中生物胺含量最高阶段均出现在成熟期5～6 个月。干酪中组胺、酪胺和总生物胺含量分别低于推荐安全剂量50、100 mg/kg和1 000 mg/kg。干酪中游离氨基酸含量与除组胺之外的各生物胺含量、总生物胺含量、成熟时间之间具有显著正相关性（P＜0.01,P＜0.05）,各生物胺含量之间也存在正相关性。不同成熟期干酪包含相同属的微生物,其中链球菌属（Streptococcus）为优势菌属,平均相对丰度为84.63%,明串珠菌属（Leuconostoc）次之,平均相对丰度为6.91%,其他分别为乳杆菌属（Lactobacillus）、拉乌尔菌属（Raoultella）、不动杆菌属（Acinetobacter）、假单胞菌属（Pseudomonas）、肠杆菌属（Enterobacter）和无分类的肠杆菌科（unclassified_Eunnterobacteriaceae）。不同成熟期干酪中各菌属的相对丰度具有差异。本研究可为评估牦牛乳硬质干酪的质量安全性和生物胺形成机理提供理论依据。     

Changes in biogenic amine and polyamine contents in smear-ripened cheeses during storage

The content of eight biologically active amines in two groups of smear-ripened cheeses (unwashed-rind and washed-rind) was analysed. The biogenic amine levels were determined after storage at 5 °C in three periods of shelf lifetime: at the minimum durability date; 1 and 2 weeks after this date. The analytical procedure consisted of amine extraction from sample matrix, derivatisation with dansyl chloride and ultra-performance liquid chromatography quantification. The content of biogenic amines and polyamines significantly differed according to the technology of ripening. The cheeses unwashed during ripening had much higher contents of all observed amines and polyamines in comparison with the washed-rind cheeses. The mean content of putrescine, cadaverine and tyramine exceeded 100 mg/kg in unwashed-rind cheeses, while the other amines occurred at lower levels. The content of all detected amines was very low in washed-rind cheeses; no tryptamine, phenylethylamine and histamine were found. The effect of storage on the amine formation was not confirmed.     

处理发酵剂对促熟干酪中生物胺和游离氨基酸的影响

以0、30、60、90 d促熟干酪中生物胺（组胺、色胺、苯乙胺、尸胺、酪胺）和游离氨基酸含量为指标,研究了添加复合处理发酵剂对干酪产生游离氨基酸和生物胺量的影响。结果表明：不同处理发酵剂添加量对各种生物胺产生的影响不同;添加处理发酵剂干酪中生物胺和游离氨基酸含量增加,其含量随着复合处理发酵剂添加量增加而增加。     

Effect of proteolytic starter cultures of Staphylococcus spp. on biogenic amine formation during the ripening of dry fermented sausages

The effect of proteolytic starter cultures of Staphylococcus carnosus and Staphylococcus xylosus on biogenic amine production was examined during the fermentation process of dry sausages. Microbial counts (lactic acid bacteria, Micrococcaceae and Enterobactenaceae), pH, moisture and proteolysis-related parameters were also studied. The polyamines spermine and spermidine were the main amines found in the raw material and they only showed slight fluctuations during the fermentation. The four elaborated batches presented a significant (P < 0.001) formation of tyramine and putrescine. The main rate of amine production was during the first three days, when a sharp pH decrease and the development of lactic acid bacteria occurred. Sausages fermented with starters had lower amounts of tyramine than naturally fermented sausages (control), but differences in the Micrococcaceae counts were only significant during the first week of the ripening process. A slight formation of diaminopropane, cadaverine, agmatine, tryptamine and phenylethylamine was observed. The amounts of histamine were constant and remained below 0.5 mg/kg of dry matter, while serotonin, octopamine and dopamine were not detected. The sausages with Staphylococcus as starter culture showed strong proteolysis that was correlated with higher pH values than those of the control sausages. However, no positive correlation was found between the proteolysis index and biogenic amine production. Since proteolysis was stronger during the second half of the ripening process, the release of free amino acids as amine precursors occurred later than the early amine production.     

Interrelationships among microbiological, physicochemical, and biochemical properties of Terrincho cheese, with emphasis on biogenic amines

Changes in the microbiological, physicochemical, and biochemical characteristics of Terrincho cheese as represented by native microflora, pH, water activity, soluble nitrogen fractions, free amino acids, and biogenic amines (e.g., ethylamine, dimethylamine, tryptamine, phenylethylamine, putrescine, cadaverine, histamine, tyramine, cystamine, and spermine) during ripening were monitored. Terrincho is a traditional Portuguese cheese manufactured from raw ewe's milk. The main groups of microorganisms (lactococci, lactobacilli, enterococci, pseudomonads, staphylococci, coliforms, yeasts, and molds) were determined following conventional microbiological procedures. Free amino acids and biogenic amines were determined by reverse-phase high-performance liquid chromatography, following extraction from the cheese matrix and derivatization with dabsyl chloride. The total content of free amino acids ranged from 1,730 mg/kg of dry matter at the beginning of the ripening stage to 5,180 mg/kg of dry matter by day 60 of ripening; such an increase was highly correlated with the increase of water-soluble nitrogen in total nitrogen, 12% trichloroacetic acid-soluble nitrogen in total nitrogen, and 5% phosphotungstic acid-soluble nitrogen in total nitrogen throughout ripening. Histamine was consistently present at very low levels, whereas putrescine, cadaverine, and tryptamine were the dominant biogenic amines and increased in concentration during ripening. Ethylamine, tryptamine, phenylethylamine, and cystamine reached maxima by 30 days of ripening and decreased thereafter. Significant correlations between amino acid precursors and corresponding biogenic amines, as well as between biogenic amines and microbial viable numbers, were observed.     

Profile of Biogenic Amines in Goat Cheese Made from Pasteurized and Pressurized Milks

ABSTRACT: The formation of biogenic amines in goat cheese can be prevented through ensuring hygiene in raw materials and during manufacture, thereby avoiding potential decarboxylating microorganisms. High-pressure treatment may be used to inactivate microorganisms and is a potential alternative to pasteurization. This treatment could provoke higher proteolysis than pasteurization, leading to a higher availability of biogenic amine precursors. We compared the biogenic amine profile throughout the ripening of goat cheese made from pressurized milk with that obtained from pasteurized milk. Results indicate that the profile of biogenic amines is very similar for both cheeses. Tyramine was the prevailing amine in both cheeses, followed by cadaverine, putrescine, and histamine.     

Effectiveness of a Lactobacillus sakei starter culture in the reduction of biogenic amine accumulation as a function of the raw material quality

The effectiveness of an amine-negative starter culture (Lactobacillus sakei CTC494) in the reduction of biogenic amine production during the ripening of fermented sausages was examined. Four batches were manufactured in parallel: spontaneously fermented and starter-mediated sausages were manufactured from two lots of raw materials of different hygienic quality. Besides the biogenic amine contents, changes in the microbial counts, nitrogenous fractions, pH, and water content were measured at several sampling points during the ripening process. In sausages manufactured from good quality meat, the starter strain of L. sakei reduced and even inhibited biogenic amine accumulation during sausage fermentation, the end products showing extremely low biogenic amine contents (tyramine levels less than 15 mg/kg of dry matter and putrescine and cadaverine levels less than 5 mg/kg of dry matter). Nevertheless, starter-mediated sausages made from poorer-quality raw materials showed much higher amine contents (308, 223, and 36 mg/kg of dry matter of cadaverine, tyramine, and putrescine, respectively), which were only slightly lower than those of the spontaneously fermented sausages made from the same raw materials. The relatively high bacterial numbers of raw materials of poorer-hygienic quality diminished the beneficial effect of the starter strain. Therefore, the effectiveness of the starter was strongly dependent on the hygienic quality of the raw materials used.     

Comparison of biogenic amine profile in cheeses manufactured from fresh and stored (4 degrees C, 48 hours) raw goat's milk

In this study, the evolution of microbial counts, biogenic amine contents, and related parameters (pH, moisture, and proteolysis) in goat cheese made from fresh raw milk or raw milk stored for 48 h at 4 degrees C was examined. In both cases the milk was nonpasteurized. This study was designed to evaluate the effect of milk quality on the profile of biogenic amines in relation to the evolution of the microbial population during cheese making. Cheese made from raw milk stored for 48 h at 4 degrees C showed the highest microbial counts and biogenic amine levels. The storage of milk under refrigeration caused significant increases in the levels of some microbial and biogenic amines during ripening, but not initially. Tyramine was the main biogenic amine in the two cheeses tested, followed by cadaverine. However, the main differences in amine contents between batches were found for putrescine, histamine, and beta-phenylethylamine, whose levels were more than twofold higher in samples from raw milk refrigerated for 48 h than in samples from fresh milk.     

大蒜精油和发酵剂对熏马肠中生物胺及微生物分布的影响

为研究大蒜精油和发酵剂对熏马肠中生物胺的影响,采用高效液相色谱法测定熏马肠中生物胺含量,利用 宏基因组学技术分析熏马肠中微生物多样性和生物胺含量的变化情况。结果表明：将大蒜精油和发酵剂联合使用能 够很好地减少熏马肠中尸胺、腐胺、组胺、酪胺和苯乙胺的产生,对控制生物胺在熏马肠中的含量起到非常显著的 作用;熏马肠中含有丰富的微生物物种,通过样品科分类水平物种分析,得出乳酸菌在熏马肠整个加工过程都是优 势菌种,而葡萄球菌在成熟后期数量较多。     

Biogenic amines and their producers in Akawi white cheese

The aim of this work was to study the biogenic amine content of brine‐ripened cheeses after one year of storage and then to investigate possible contaminating micro‐organisms with decarboxylase activity. The biogenic amine production of isolates was tested in vitro. The most frequent biogenic amines were putrescine, histamine and tyramine. The biogenic amine content detected in one cheese sample was above 120 mg/kg; this can be considered toxicologically relevant. Decarboxylase activity was found for 33 contaminating micro‐organisms. Isolates belonging to Bacillus licheniformis, Debaryomyces hansenii, Staphylococcus equorum and Serratia marcescens produced significant amounts of putrescine and cadaverine.     

FORMATIONS OF BIOGENIC AMINES IN TURKISH STYLE SAUSAGE DURING RIPENING AND STORAGE PERIODS

Changes of biogenic amine (histamine, β‐phenylethylamine, serotonin, spermine, spermidine, cadaverine, putresine, 1,7‐diamino heptane, tryptamine and tyramine) concentrations and pH values of Turkish style sausages were investigated during ripening and storage. Cadaverine and β‐phenylethylamine were not detected during the ripening and storage periods for all sausage types. Storage time had a significant (P<0.05) effect on the formation of all biogenic amines. Starter culture usage decreased (P<0.05) biogenic amine formation. Histamine levels were higher in sausages prepared without additives and lower in the sausages made with starter culture than the sausages made without starter culture. There was no effect of starter culture and additives on formation of tryptamine, 1,7‐diamino heptane, serotonin or spermidine. Starter culture and additives affected (P<0.05) the formation histamine, putrescine and spermine. pH values of sausages sharply decreased (P< 0.05) from 5.98 to about 4.53 during 3 days of ripening. After that, pH values increased slowly and were nearly constant during the storage period.