高效液相色谱法检测牙鲆体内的生物胺

采用高效液相色谱(HPLC)法测定了牙鲆体内的多种生物胺.以邻苯二甲醛(OPA)为柱后荧光衍生剂,通过Capcell Pak MG C18色谱柱的梯度洗脱,将酪胺、腐胺、尸胺、组胺、胍丁胺、亚精胺和精胺等7种生物胺进行良好分离.测试结果表明,7种生物胺在0.05～30mg/L的范围内线性良好,相关系数都在0.999以上...     

不同加工对生物胺影响的研究

为破坏生物胺结构、抑制生物胺产生、有效去除生物胺,通过HPLC-柱后衍生-FLD检测技术分析了7种生物胺（酪胺、腐胺、尸胺、组胺、胍丁胺、亚精胺、精胺）在不同加工处理后的变化情况。结果表明,超声、微波、加热对生物胺没有破坏作用,超声和山梨酸钾前处理对鲅鱼生物胺的抑制率分别为45.89%和58.36%。醋酸、泡菜汁对生物胺的去除率为17.59%和27.76%。因此,生物胺不能通过超声、微波、加热技术来消除,可以通过控制细菌生长和酸液浸提等方式来抑制和去除。      

黄酒中9 种生物胺的高效液相色谱分析法

建立一种检测黄酒中生物胺的反相高效液相色谱法。用三氯乙酸提取黄酒中生物胺,丹磺酰氯作为衍生化试剂,1,7-二氨基庚烷作为内标定量。色谱条件为：Waters XBridge C18色谱柱（4.5 mm×250 mm,3.8 μm）作为分离柱,乙腈和水作为流动相进行梯度洗脱,流速1 mL/min,紫外检测波长254 nm。结果表明,9 种生物胺（盐酸吡哆胺、色胺、腐胺、尸胺、β-苯乙胺、组胺、酪胺、精胺和亚精胺）在40 min内能被很好分离。在1～50 mg/L质量浓度范围内,各种生物胺呈现良好的线性相关性（R2＞0.999）。各生物胺加标回收率为79.54%～89.55%,相对标准偏差为3.14%～6.35%,该法各组分检出限（RSN=3）为0.002～0.009 mg/L。应用柱前衍生化高效液相色谱-紫外检测法,实现了黄酒中生物胺的准确检测。     

不同加工对生物胺影响的研究

为破坏生物胺结构、抑制生物胺产生、有效去除生物胺,通过HPLC-柱后衍生-FLD检测技术分析了7种生物胺(酪胺、腐胺、尸胺、组胺、胍丁胺、亚精胺、精胺)在不同加工处理后的变化情况。结果表明,超声、微波、加热对生物胺没有破坏作用,超声和山梨酸钾前处理对鲅鱼生物胺的抑制率分别为45.89%和58.36%。醋酸、泡菜汁对生物胺的去除率为17.59%和27.76%。因此,生物胺不能通过超声、微波、加热技术来消除,可以通过控制细菌生长和酸液浸提等方式来抑制和去除。     

高效液相色谱法同时测定香肠中6种生物胺

采用丹磺酰氯为衍生试剂,建立反相高效液相色谱法同时测定香肠中腐胺、尸胺、酪胺、组胺、亚精胺和精胺6种生物胺的方法.色谱条件:C18色谱柱分离,以甲醇和0.1%乙酸溶液为流动相梯度洗脱,流速0.8mL/min,二极管阵列检测器,检测波长254 nm.以1,7-二氨基庚烷为内标,在设定的试验条件下,6种生物胺实现了良好的分离,并呈良好的线性相关性(r>0.999).方法的检测限:腐胺、亚精胺、尸胺和酪胺均为0.8μg/mL,组胺和精胺均为4 μg/mL.本检测方法具有良好的重现性和回收率.对发酵香肠样品中的生物胺含量进行了分析,其中部分香肠中含有一定量的生物胺,最高可达1.09mg/g.     

不同温度储藏条件下鲅鱼生物胺变化的研究

为研究鲭鱼中毒的组胺来源以及温度对鲅鱼生物胺的影响,通过HPLC-柱后衍生-FLD检测技术分析了鲅鱼体内7种生物胺（酪胺、腐胺、尸胺、组胺、胍丁胺、亚精胺、精胺）的含量以及在不同储藏温度的变化情况。结果表明,新鲜鲅鱼肌肉中没有检出组胺。鲅鱼体内生物胺的变化随温度的升高而加快,0℃（冰藏）、4℃、20℃储藏8d的生物胺总量变化范围分别为3.09～6.77、16.28～274.65、154.96～1846.29mg/kg。因此,细菌是引起组胺中毒的主要来源,低温储藏是控制鲅鱼产生生物胺的有效措施。      

水产品中生物胺的两种衍生测定方法比较

分别采用邻苯二甲醛柱前衍生高效液相色谱(HPLC)荧光检测法(简称OPA法)和丹磺酰氯柱前衍生HPLC紫外检测法(简称DNS-Cl法)测定水产品中生物胺含量,比较这两种方法对水产品中多种生物胺检测的差异。结果表明,两种方法对生物胺的检测质量浓度范围在1~100 mg/L,OPA法的检出限(0.03~0.35 mg/kg)略优于DNS-Cl法(0.15~0.90 mg/kg)。方法的精确度试验表明:OPA法的加标回收率(76.2%~107%)略优于DNSCl法(62.2-118%),然而两种方法的测量精密度均小于10%(OPA法1.3%~6.7%,DNS-Cl法2.6%~9.9%,n=6)。OPA法操作简便、快速,然而其衍生产物不太稳定,对亚精胺和精胺的检测灵敏度低;而DNS-Cl法操作相对繁琐,其衍生产物稳定,可同时测定包括亚精胺和精胺在内的多种生物胺。用这两种衍生方法检测水产品中组胺、腐胺、酪胺和色胺存在显著性差异(P0.05)。     

不同温度储藏条件下鲅鱼生物胺变化的研究

为研究鲭鱼中毒的组胺来源以及温度对鲅鱼生物胺的影响,通过HPLC-柱后衍生-FLD检测技术分析了鲅鱼体内7种生物胺(酪胺、腐胺、尸胺、组胺、胍丁胺、亚精胺、精胺)的含量以及在不同储藏温度的变化情况。结果表明,新鲜鲅鱼肌肉中没有检出组胺。鲅鱼体内生物胺的变化随温度的升高而加快,0℃(冰藏)、4℃、20℃储藏8d的生物胺总量变化范围分别为3.09～6.77、16.28～274.65、154.96～1846.29mg/kg。因此,细菌是引起组胺中毒的主要来源,低温储藏是控制鲅鱼产生生物胺的有效措施。     

白酒和黄酒中生物胺的高效液相色谱分析法

本研究建立了1种检测某白酒与黄酒中常见的9种生物胺(尸胺、组胺、酪胺、色胺、盐酸吡哆胺、精胺、亚精胺、腐胺和2-苯乙胺)的柱前衍生-高效液相色谱法,并对其实际运用可行性进行验证与讨论。以丹磺酰氯(Dns-Cl)作为衍生试剂,采用Thermo Hypersil GOLD C18色谱柱(250 mm×4.6 mm, 5μm)分离,以乙腈和含0.1%甲酸的0.01 mol/L乙酸铵水溶液作为流动相A与B,流速0.8 mL/min,紫外检测器波长254 nm。分析结果显示,9种生物胺在质量浓度0.5～50 mg/L范围,生物胺浓度与峰面积间线性相关性良好(R20.998)。选取1.0,5.0,10.0 mg/L 3个水平加标,得到加标回收率83.30%～114.7%,相对标准偏差均低于4.56%,9种生物胺检出限为0.07～0.22 mg/L。采用柱前衍生-高效液相色谱法分离、检测白酒与发酵酒——黄酒中的生物胺,实现了白酒与黄酒中生物胺含量的准确检测。     

酿造过程及啤酒中生物胺的初步控制

啤酒中发现的最重要的生物胺主要有组胺、腐胺、尸胺、酪胺、色胺、β-苯乙胺、精胺和亚精胺。生物胺作为啤酒新鲜或腐败变质程度的指标,其分析显得非常重要。目前已经开发出几种测定生物胺的方法,其定量方法主要是依据色谱方法：薄层色谱（TLC）、气相色谱（GC）,毛细管电泳（CE）和高效液相色谱法（HPLC）,其中高效液相色谱是最常用的分析方法,大部分检测是采用预柱或者柱后衍生技术荧光检测。本文对发酵过程及成品啤酒中八种生物胺含量及总量的变化进行了研究。在发酵过程中,组胺、腐胺及生物胺总量有一定的增加,苯乙胺水平低于检测限;但是,在对应成品中,八种生物胺含量都有不同程度的下降;检测了分别添加组氨酸和酪氨酸的巴氏杀菌啤酒在存储12个星期内的生物胺变化情况。在接种了从麦芽表面筛选的混合乳酸菌培养直至混浊时发现,成品酒中组胺及酪胺含量变化不大。     

Production of Biogenic Amines and Their Potential Use as Quality Control Indices for Hake (Merluccius merluccius, L.) Stored in Ice

ABSTRACT: The production of tyramine, histamine, putrescine, cadaverine, agmatine, and spermidine was studied for use as quality indices in muscle of gutted hake stored in ice for 25 d. The changes in biogenic amine concentrations were correlated with trimethylamine nitrogen levels. All concentrations except for spermidine, increased progressively throughout storage. During storage, cadaverine reached 72.14 mg/kg while there were no significant increases in histamine and tyramine. The levels of free amino acids, which may be metabolized into biogenic amines, were not correlated to respective biogenic amines levels. The production of cadaverine and agmatine occurred prior to spoilage and these biogenic amines may indicate freshness of hake stored in ice.     

Formation of biogenic amines by Gram-negative bacteria isolated from poultry skin

The aim of this study is to explore production of biogenic amines (histamine, tyramine, putrescine, cadaverine, agmatine, spermine and spermidine) by 88 Gram-negative bacteria isolated from poultry skin (41 isolates of family Enterobacteriaceae, 21 isolates of genus Aeromonas, 16 isolates of genus Pseudomonas, and 10 isolates of other Gram-negative rods). Ion-exchange chromatography was employed to analyse the above mentioned amines. Enterobacteria were found to be the largest producers of amines with proved presence of tyramine, agmatine, putrescine, and cadaverine in cultivation broth after incubation of bacteria. Putrescine and cadaverine were the most abundant products. Presence of at least two biogenic amines, i.e. mainly concurrent presence of putrescine and cadaverine, was revealed in 19 enterobacteria strains. Eleven isolates classified into Aeromonas genus produced putrescine and five of them also formed cadaverine. The other observed biogenic amines (histamine, spermine and spermidine) were not found among tested isolates. Production of biogenic amines by any Pseudomonas family isolates was not proved.     

Application of the simplex method for optimization of chromatographic analysis of biogenic amines in fish

The aim of this study was to optimize the separation of biogenic amines with ion-exchange high-performance liquid chromatography, using the modified simplex method. A modified chromatographic response function was used, and two methodological parameters were selected as simplex variables: the proportion of the organic solvent 2-propanol in the mobile phases, added as a new mobile phase, and the variation of the gradient curve at one specific step in the elution program. In optimum conditions the correct resolution and separation of the biogenic amines (tyramine, β-phenylethylamine, histamine, tryptamine, putrescine, cadaverine, agmatine, spermidine and spermine) were completed in 16 min, reducing the analysis time by 9 min compared with the original method. The regression coefficients were higher than 0.99 in all the amines, except for spermine (0.989). The detection limits varied between 0.06 mg/L, for histamine and agmatine, and 0.22 mg/L, for tryptamine, while retention time repeatability, determined as a relative standard deviation, was between 4.64% for histamine and 11.95% for tryptamine. The repeatability variation for the amine concentrates was found between 0.71% in cadaverine and 3.23% for tryptamine. Applying the optimized method to samples of refrigerated sardine and sea bream demonstrated the effectiveness of the method in these fish species.     

六类鱼制品中生物胺的HPLC法测定

针对不同类型的鱼制品建立同时检测腐胺、尸胺、精胺、亚精胺、酪胺、苯乙胺、组胺和色胺8种生物胺含量的高效液相色谱(HPLC)法。应用该方法,测定了干燥制品、油炸制品、烘烤制品、发酵制品、混揉制品以及萃取制品6类鱼干制品中的生物胺含量。结果显示,萃取制品-鱼油中未检测出生物胺,其余5类产品中生物胺含量最高的为发酵制品(73.42~70.59 mg/kg)和混揉制品(55.48~49.88 mg/kg),其次为油炸制品(17.72~14.32 mg/kg),而鱼干制品(5.28~5.17 mg/kg)和烘烤制品(4.75~4.69 mg/kg)中的生物胺含量相对较少;另外,尸胺、组胺、精胺和亚精胺四种生物胺为除萃取制品之外的5类鱼制品中普遍存在的4种生物胺成分;其中尸胺是鱼制品中变化最大、影响最为明显的单体生物胺,检测产品中尸胺含量有利于监控、评判鱼干制品的质量。     

改进的QuEChERS结合RPLC-TMS测定水产品中生物胺的研究

采用改进的Qu ECh ERS方法,利用液相色谱-串联质谱（RPLC-TMS）检测,建立了几种水产品中生物胺的分析方法。样品用乙腈+甲醇（9+1）提取,无水硫酸钠除去水分,经乙二胺-N-丙基硅烷（PSA）和C18混合粉末净化,采用HILIC柱,以乙腈/1%甲酸水溶液为流动相梯度洗脱进行液相色谱分离。质谱分析采用电喷雾正离子电离,多反应监测模式,外标法定量。结果显示6种生物胺在10²00 ng/m L范围线性关系良好。方法的定量限（LOQ,信噪比为10）为0.10 mg/kg。加标水平在0.10 mg/kg时,鱼、虾、蟹及鱿鱼四种阴性基质中加标的回收率在74.18%⁹0.15%之间,相对标准偏差（RSD,n=6）在2.22%⁸.04%之间。结果表明本方法简便、快速、安全、适用于水产品中生物胺含量的定性和定量检测。      

Effect of controlled atmospheres enriched with O2 in formation of biogenic amines in chilled hake (Merluccius merluccius L.)

Changes in biogenic amines (tyramine, histamine, putrescine, cadaverine, agmatine and spermidine) contents during the storage of chilled hake in a controlled atmosphere with four gas mixtures of different CO₂ and O₂ concentrations, (CO₂/O₂/N₂ at 60/15/25, 40/40/20, 60/40/0 and 40/60/0 respectively) were studied. Biogenic amines were determined by high-pressure liquid chromatography with post-column derivatization and ortho-phtaldehyde. Histamine levels, potentially the most toxic amine in hake, were very low during storage. The amine in hake with the highest level was cadaverine. Spermidine and putrescine had very high levels at the beginning of the storage. However, agmatine and tyramine were not detected in the source raw material. An increase in biogenic amines was observed in the control lot from day 12 of storage except for agmatine, where levels began to increase earlier. Cadaverine and agmatine could be used as control indices due to their behaviour during the storage period studied. The varying O₂ and CO₂ concentrations in the different atmospheres did not seem to affect the production of spermidine. The high levels of CO₂ in thelot in M1 atmosphere were not sufficient to inhibit the production of biogenic amines. However, the high O₂ concentration had an inhibiting effect on the production of biogenic amines. Therefore, the formation of biogenic amines was inhibited by controlled atmospheres with oxygen-enriched binary gas mixtures.     

Determination of biogenic amines in cheese by ion exchange chromatography

A sensitive, fully automated method, yielding reproducible results, was developed for the determination of relevant biogenic amines in cheese. Histamine, tyramine, putrescine, cadaverine, tryptamine, agmatine, spermidine and spermine were separated in the ion exchanger of OSTION LG ANB column of the automatic amino acid analyser Mikrotechna 339 T. Elution was carried out at 60 degrees C using a system of two buffers. The samples were extracted and precipitated by trichloroacetic acid and purified, after removal of fat, by membrane filtration. The recovery for individual amines in cheese ranged between 86% and 108%. The detection limit was of 1-5 mg of the respective amine per 1 kg of cheese and the method was linear within the dose range of 0.1-10 micrograms (for tryptamine 0.5-10 micrograms).     

Biogenic amine index for freshness evaluation in iced Mediterranean hake (Merluccius merluccius)

Biogenic amine accumulation was studied during the ice storage of Mediterranean hake. Sensory analysis and counts of Shewanella, Pseudomonas, enterobacteria, psychrotrophic, and mesophilic bacteria provided complementary information on hake spoilage. Putrescine and cadaverine were the main amines accumulated, whereas histamine and tyramine were minor amines but had qualitative interest from the hygienic point of view. Although all biogenic amines were less abundant than in pelagic fish, they may also be used as indicators of freshness and/or spoilage in hake. Cadaverine was the amine best correlated with Shewanella, which was the specific spoilage organism. Therefore, cadaverine may be regarded as the specific spoilage biogenic amine for hake stored at chilling temperatures. However, the biogenic amine index, which considers cadaverine, putrescine, histamine, and tyramine, has several advantages as an indicator of hake quality. Taking into account sensory data, an acceptability limit of the biogenic amine index could be established in 15 to 20 microg/g.