干酪乳杆菌SB27降低N-二甲基亚硝胺对大鼠肠黏膜细胞损伤作用研究

本研究选用分离自甘肃牦牛乳,且具有潜在益生功能的干酪乳杆菌SB27(Lactobacillus casei,SB27)进行研究,分析其降低N-二甲基亚硝胺(NDMA)对大鼠肠道黏膜细胞IEC-6毒性作用的能力。研究通过CCK-8实验、HE染色、TUNEL染色和透射电镜等进行全面分析。CCK-8实验结果显示,干酪乳杆菌SB27可改善NDMA致大鼠肠道黏膜细胞IEC-6毒性的影响,且该作用与干酪乳杆菌SB27的剂量有关。HE染色、TUNEL染色和透射电镜实验结果显示,NDMA(80 μg/mL)诱导了IEC-6细胞的损伤,在此过程中干酪乳杆菌SB27(10_⁵ CFU/mL、10_⁶ CFU/mL)可保护IEC-6细胞免受NDMA的毒性损伤。以上实验结果表明,干酪乳杆菌SB27降低NDMA对IEC-6细胞的毒性损伤,为该菌株降低NDMA毒性作用机理研究奠定了基础。     

副干酪乳杆菌X12抑制N-二甲基亚硝胺诱导IEC-6细胞损伤作用研究

目的 探究副干酪乳杆菌X12(Lactobacillus.paracaseisubsp.paracaseiX12)抑制N-二甲基亚硝胺(N-nltrosodimethylamine,NDMA)诱导IEC-6细胞(大鼠肠道黏膜细胞)毒性损伤作用。方法 通过胃液、肠液耐受性实验、扫描电镜法分析副干酪乳杆菌X12益生功能和菌株的形态;通过CCK-8实验对副干酪乳杆菌X12抑制NDMA诱导IEC-6细胞毒性作用进行分析。结果 胃肠道耐受性实验显示,副干酪乳杆菌X12可耐受胃肠液的消化,但对胆盐的耐受性略差;扫描电镜下副干酪乳杆菌X12菌体呈短而圆的杆状结构,无鞭毛;CCK-8实验结果显示,在MRS培养基中NDMA (0～80μg/mL)对副干酪乳杆菌X12的生长无显著影响(P>0.05);副干酪乳杆菌X12可抑制NDMA致大鼠肠道黏膜细胞IEC-6毒性的作用,且该作用与副干酪乳杆菌X12的剂量有关。结论 副干酪乳杆菌X12可抑制NDMA诱发的IEC-6细胞毒性损伤。     

不同培养介质中N-二甲基亚硝胺对干酪乳杆菌生长影响

目前研究已证实乳酸菌可降低食品中N-二甲基亚硝胺（NDMA）的含量,减小其对人体消化道的毒性作用。但乳酸菌抑制NDMA毒性作用的同时,NDMA对其生长影响鲜见相关报道。该研究选用具有潜在益生功能的干酪乳杆菌（Lactobacillus casei）SB27,分析其胃肠液耐受性及黏附性能,并检测不同培养介质中NDMA对干酪乳杆菌SB27生长的影响。结果表明,干酪乳杆菌SB27可耐受胃肠液的消化,具有良好的黏附性能,且在MRS培养基、低氮源MRS培养基和磷酸盐缓冲液中,NDMA对菌株SB27生长均无显著影响。该研究为进一步研究干酪乳杆菌SB27降解NDMA作用奠定了理论基础。     

多香果粉对肉丸中N-亚硝胺的抑制作用

为降低肉制品中N-亚硝胺的含量,探究不同浓度的多香果粉对肉丸中挥发性N-亚硝胺的影响,以及与亚硝酸盐残留量、脂肪氧化值（TBARS值）、挥发性盐基氮（TVB-N值）、p H 4个指标的相关性。结果表明,利用气相色谱串联质谱法（GC-MS/MS）测定了肉丸中8种挥发性N-亚硝胺,仅二甲基亚硝胺（NDMA）、亚硝基哌啶（NPIP）、亚硝基吡咯烷（NPYR）被检出,在5 d冷藏中0.02%、0.04%、0.08%多香果粉添加对肉丸中N-亚硝胺的含量具有降低作用,对NDMA、NPIP、NPYR抑制率最高分别为93.1%、80.3%、60%,多香果粉最高抑制的添加浓度为0.08%。添加多香果粉后,肉制品中NDMA、NPIP、NPYR与亚硝酸盐残留量、TBARS、TVB-N等指标相关性发生了改变,NDMA与TVB-N相关性上升、NPIP与TBARS相关性上升、NPYR与亚硝酸盐相关性上升、N-亚硝胺总量与TBARS和TVB-N相关性上升。      

原料及加工工艺对包馅鱼肉卷N-亚硝胺含量的影响

为评价原料及加工工艺对包馅鱼肉卷中N-亚硝胺含量的影响,分别测定包馅鱼肉卷馅和皮加工过程中9 种挥发性N-亚硝胺含量、亚硝酸盐含量、硫代巴比妥酸反应物（thiobarbituric acid reaction substances,TBARs）值以及挥发性盐基氮（total volatile nitrogen,TVB-N）含量的动态变化。结果表明：经过加工后鱼肉卷馅和皮中的TVB-N含量均下降,TBARs值和亚硝酸盐含量均上升;鱼肉卷馅的各种原料中含有不等量的N-二甲基亚硝胺（N-nitrosodimethylamine,NDMA）（0～27.51 μg/kg）、N-二乙基亚硝胺（N-nitrosodiethylamine,NDEA）（0～3.39 μg/kg）、N-甲基乙基亚硝胺（N-nitrosoethylmethylamine,NMEA）（0.11～4.33 μg/kg）以及N-亚硝基二苯胺（N-nitrosodiphenylamine,NDPheA）（0～0.82 μg/kg）;在鱼肉卷皮原料中检出了NDEA（0～12.56 μg/kg）、NMEA（0.08～15.26 μg/kg）以及N-亚硝基哌啶（N-nitrosopiperidine,NPIP）（0～8.13 μg/kg）;馅中NDMA、NMEA、NDEA及皮中NDMA、NMEA主要在盐擂期间形成,并在加工后期逐渐下降;NDPheA及NPIP含量在馅和皮的加工过程中始终低于检出限;鱼肉卷成品馅料中NDMA含量为（1.9±0.2） μg/kg,低于国家标准中的限量,而皮中未检出NDMA。     

亚硝化模拟体系中油脂对N-亚硝胺形成的影响

为了解油脂对N-亚硝胺形成的影响,采用体外模拟亚硝化反应体系,研究油脂种类（花生油、亚麻籽油和葵花籽油）和比例（0%、10%、20%、30%、40%）对N-二甲基亚硝胺（N-nitrosodimethylamine,NDMA）、N-二乙基亚硝胺（N-nitrosodiethylamine,NDEA）生成量的影响;同时,在油脂参与条件下（20%花生油）,以不含油脂的水相体系为对照,考察亚硝化反应条件（底物浓度比、pH值、反应温度及反应时间）对NDMA、NDEA、N-亚硝基吡咯烷（N-nitrosopyrrolidine,NPYR）生成量的影响。结果表明：3?种油脂对NDMA和NDEA的形成均有一定的促进作用,相比于葵花籽油,花生油和亚麻籽油的添加更易促进N-亚硝胺的形成,且随添加比例升高有增加的趋势。亚硝酸盐浓度对NDMA和NDEA形成的影响大于对NPYR形成的影响;在pH值为5.4～7.0范围内随着pH值的升高,NDMA和NDEA生成量有下降趋势,pH值对NPYR形成影响较小;当反应温度高于80?℃时,N-亚硝胺生成量随着温度上升有明显增加的趋势;NDMA和NDEA的生成量随着反应时间的延长呈现出先增加后略下降的趋势,NPYR的生成量随着反应时间的延长变化不大;含有油脂的乳化体系中NDMA、NDEA和NPYR的生成量均显著高于对照组水相体系,且在相同反应条件下,3?种N-亚硝胺的生成量依次为NDMA＞NDEA＞NPYR。结果表明,较高的温度、较低的pH值、较高的亚硝酸盐浓度以及一定量的油脂存在条件下,可以明显促进NDMA和NDEA的形成,而对NPYR形成影响最大的因素是反应温度,因此在肉制品实际生产中可以通过控制这些因素从而降低N-亚硝胺的生成。     

如皋火腿中挥发性N-亚硝胺种类和含量分析

用带有氮磷检测器的气相色谱(GC-NPD)测定了从市场上买来的20条如皋火腿中的挥发性N-亚硝胺,并用GC-MS确证GC-NPD检测结果.结果表明:市售如皋火腿中含有3种挥发性N-亚硝胺,分别为N-二甲基亚硝胺(NDMA)、N-二乙基亚硝胺(NDEA)和N-吡咯烷亚硝铵(NPYR), 平均含量分别为0.20、0.81和3.85μg/kg,其中NPYR的含量最多;NPYR的检出率最高为100%,其次为NDEA,检出率为95%, NDMA最小为85%,NDHA,NPIP和NDBA未检测出.     

2种模拟体系下Vc和V_E联合作用对N-亚硝胺形成的影响

为研究肉制品加工中控制N-亚硝胺形成的有效亚硝化抑制剂,试验采用水相和乳相(油脂存在下)2种亚硝化模拟反应体系,探究不同浓度Vc、V_E或Vc+V_E对体系中N-二甲基亚硝胺(N-nitrosodimethylamine,NDMA)、N-二乙基亚硝胺(N-nitrosodiethylamine,NDEA)、N-亚硝基吡咯烷(N-nitrosopyrrolidine,NPYR)形成的影响;并研究Vc和V_E联合作用下,不同亚硝化条件(pH、反应温度、反应时间)对这2种反应体系中NDMA、NDEA、NPYR形成的抑制率变化。试验结果表明:在水相体系中抑制N-亚硝胺效果最好的是Vc,当Vc浓度为3mmol/L时,对NDMA、NDEA、NPYR的抑制率分别达到78.50%、59.97%和58.53%;而在乳化体系中,Vc和V_E联合作用(1:1,3mmol/L)对NDMA(69.02%)和NDEA(86.57%)抑制效果达到最佳。且在pH为6.2、反应温度为70℃、反应时间为1.5h的亚硝化反应条件下,Vc和V_E联合作用对乳化体系中NDMA和NDEA的抑制率可达到最高值,而对NPYR的抑制作用明显低于NDMA和NDEA。研究将为降低肉制品加工过程中N-亚硝胺的形成提供理论依据和数据支持。     

气相色谱-三重四极杆质谱测定广东地区市售腊肠中9种挥发性亚硝胺

采用QuEChERS技术(具有快速、简便、经济、高效、可操作性强和安全性)结合气相色谱-三重四极杆质谱联用法(gas chromatography tandem triple quadrupole mass spectrometry,GC-MS-MS)检测广东省6个市(县)的40份市售腊肠样品中常见挥发性亚硝胺的种类和含量,了解广东地区市售腊肠中的9种挥发性亚硝胺的污染现状,包括N-亚硝基二甲胺(N-nitrosodimethylamine,NDMA)、N-亚硝基二乙胺(N-nitrosodiethylamine,NDEA)、N-亚硝基甲乙胺(N-nitrosomethylethylamine,NMEA)、N-亚硝基二丙胺(N-nitrosodipropylamine,NDPA)、N-亚硝基吡咯烷(N-nitrosopyrrolidine,NPYR)、N-亚硝基哌啶烷(N-nitrosopiperidine,NPIP)、N-亚硝基二丁胺(N-nitrosodibutylamine,NDBA)、N-亚硝基吗啉(N-nitrosomorpholine,NMOR)、N-亚硝基二苯胺(N-nitrosodiphenylamine,NDPhA)。结果表明:本研究的检测方法灵敏度高,操作简单,适合批量样品的快速检测,其中线性范围为0.05~200 ng/mL,检出限为0.01~0.1μg/kg,加标回收率范围在90.7%~116.0%,相对标准差范围为1.8%~14.0%。广东地区市售腊肠中NDMA、NDBA、NMOR、NPYR、NPIP、NDPA检出率较高;15.0%的市售腊肠样品中NDMA超过我国肉质品的限量标准,最高值为7.37μg/kg,20.0%样品中总挥发性亚硝胺超过10.0μg/kg,含量最高的黑椒腊肠为15.32μg/kg;不同生产方式的市售腊肠中挥发性亚硝胺的种类不同,其中工业化生产的腊肠样品NDMA、NDPA、NDBA含量高于小作坊家庭自制腊肠样品,而小作坊、家庭自制腊肠样品中NMEA、NPYR、NMOR含量高于工业化生产。     

SPME-GC-NPD法测定肉制品中挥发性N-亚硝胺

改进3种N-亚硝胺(N-亚硝基二乙胺,NDEA;N-亚硝基二甲胺,NDMA;N-亚硝基吡咯烷,NPYR)的测定方法。以固相微萃取(SPME)为样品萃取方式,配备氮磷检测器的气相色谱仪(GC-NPD)为检测工具,研究3种N-亚硝胺(NDEA、NDMA、NPYR)的测定方法。以PDMS/DVB/CAR为萃取头,萃取温度50℃、搅拌速度400 r/min、萃取时间30 min、盐离子浓度0.20 g/m L时NDEA能得到最大的萃取面积。对3种N-亚硝胺的线性分析得出,NDEA的测定在10 ng/m L~100 ng/m L时线性关系为0.994;NDMA在10 ng/m L~100 ng/m L时的线性关系为0.912;NPYR在0~10 ng/m L时没有表现出明显线性关系。采用SPME,结合GC-NPD测定N-亚硝胺的方法,对NDEA的测定灵敏度最高,RSD为6.7%,加标回收率为89.7%,最低检出限(LOD)0.67 ng/m L,可以满足肉制品中NDEA的定量测定。     

N-nitrosamines and residual nitrite in cured meats from the Dutch market

A total of 140 samples of 16 kinds of cured meats were analyzed for contents of residual nitrite and N-nitrosamines. Nitrite was determined by reaction with sulfanilamide/naphthylethylenediamine and colorimetric measurement. N-nitrosamines were isolated from the samples by vacuum distillation and determined by gas-chromatography with chemiluminescence detection (GC-TEA). In six samples no nitrite was detectable (less than 1 mg NaNO2/kg), the remaining samples contained 1-140 mg NaNO2/kg, median value 6.8 mg/kg. In 46 samples (33%) no N-nitrosamines were detected, i.e. less than 0.1-0.5 microgram/kg of the individual nitrosamines, depending upon their structure. N-nitrosodimethylamine (NDMA) was the nitrosamine present most frequently, in 75 samples, contents were 0.1-0.9 microgram/kg, mean 0.3 microgram/kg. Other N-nitrosamines found were: N-nitrosopiperidine (NPIP), 10 times, 0.3-25 micrograms/kg; N-nitrosodiethylamine (NDEA), three times, 0.2-0.9 microgram/kg; N-nitrosopyrrolidine (NPYR), three times, 1.3-4.2 micrograms/kg; N-nitrosomorpholine, once, 0.7 microgram/kg and N-nitrosothiazolidine (NTHZ), 36 times, 0.5-91 micrograms/kg, mean 5.7 micrograms/kg. NTHZ was found most often and with the highest contents in smoked products. Frying of bacon and cured, smoked pork bellies led to substantially increased levels of NPYR in both products, and for the pork bellies also of NTHZ. In five samples of cured, smoked pork bellies after frying NTHZ-contents of 3.6-490 micrograms/kg (mean 179) were found. No correlation between residual nitrite levels and N-nitrosamine contents could be established. Investigations during the nineteen seventies gave much higher levels for NDMA, NDEA, NPIP and NPYR in Dutch cured meats than now found; at that time NTHZ was not measured.     

重组培根加工过程中N-亚硝胺及其前体物质含量的动态变化

为了明确重组培根加工过程中N-亚硝胺含量的动态变化及其影响因素,监测原料肉、腌制、蒸煮、烟熏加工环节中pH值及亚硝酸盐、生物胺、N-亚硝胺含量的动态变化,同时考察重组培根的感官品质。结果表明：随着加工的进行,重组培根pH值和亚硝酸盐残留量均呈现先上升后下降的趋势;在监测的8 种生物胺中,原料肉中仅检出精胺,随着重组培根加工的进行,生物胺的种类不断丰富,含量逐渐升高,烟熏显著加速了生物胺的生成;原料肉中仅检测出N-二甲基亚硝胺（N-nitrosodimethylamine,NDMA）和N-亚硝基吡咯烷（N-nitrosopyrrolidin,NPYR）;腌制后可检出N-甲基乙基亚硝胺（N-nitrosomethylethylamine,NMEA）、N-二丙基亚硝胺（N-nitrosodipropylamine,NDPA）、N-二丁基亚硝胺（N-nitrosodibutylamide,NDBA）和N-亚硝基哌啶（N-nitrosopiperidine,NPIP）,NDMA含量超过了国标限量; 蒸煮后新增N - 二乙基亚硝胺（ N - n i t r o s o d i e t h y l a m i n e ,NDEA）和N-亚硝基吗啉（N- n i trosomorpholine,NMOR）,且NDMA、NDPA、NDBA、NPIP含量显著升高（P＜0.05）;烟熏过程中N-亚硝胺总量显著增加（P＜0.05）,烟熏6～9 h增幅最大;重组培根的感官评分随着烟熏时间的延长显著提高（P＜0.05）。综合食用安全性和感官评分,建议制作重组培根时选择腌制16 h,热熏法（55±2） ℃烟熏6h。     

Occurrence of N-nitrosodimethylamine in South Korean and imported alcoholic beverages.

Volatile N-nitrosamine (VNA) levels in South Korean and imported alcoholic beverages were determined between 1995 and 2002. A total of 147 alcoholic beverages, including lager beer, whiskey, liqueurs and traditional Korean alcoholic beverages (Chungju, Takju and Soju), were analysed for their VNA content by GC-TEA. Of eight VNAs (N-nitrosodimethylamine (NDMA), N-nitrosodiethylamine (NDEA), N-nitrosopyrrolidine (NPYR), N-nitrosomorphorine (NMOR), N-nitrosodibuthylamine (NDBA), N-nitrosopiperidine (NPIP), N-nitrosodiprophylamine (NDPA) and N-nitrosodiphenylamine (NDPhA)) only NDMA was detected. In 1995, NDMA was detected in 79.3% of domestic beers; the average was 0.8 microg kg-1. Seven years later, the average NDMA level for 18 domestic beers was 0.3 microg kg-1 and it was positive in 55.6% samples. In whisky and liqueurs, NDMA levels averaged <0.1 microg kg-1 in both 1995 and 2002. Average NDMA levels of Chungju in 1995 were <0.1 microg kg-1, but NDMA was not detected in 2002. Takju had undetectable levels of NDMA both times. In 1995, NDMA was found in four of six Soju samples, but in 2002, NDMA could not be detected.     

模拟肉制品加工条件下影响N-亚硝胺的形成因素

为探讨肉制品加工条件下影响N-亚硝胺的形成因素,本实验模拟肉品加工条件,研究温度、pH、底物浓度之比对N-亚硝基二甲胺（NDMA）和N-亚硝基二乙胺（NDEA）形成的影响,以及pH、底物浓度对N-亚硝基吡咯烷（NPYR）形成的影响。结果表明,NDMA和NDEA的形成量随着温度的升高而增加,随着pH的升高而降低:当温度大于80℃时,NDMA和NDEA的生成量显著增加（p<0.05）;在pH5.4～6.2之间,降低速度较快,在pH6.2～7.0时,降低速度缓慢。随着pH的升高,由吡咯烷（PYR）生成NPYR的量变化不大（p>0.05）,而由脯氨酸（Pro）生成NPYR的量则呈下降趋势（p<0.05）。二甲胺盐酸盐（DMA·HCl）与二乙胺盐酸盐（DEA·HCl）相比,DMA·HCl与NaNO2反应更容易形成N-亚硝胺,且均在低温肉制品加工温度下就可以形成;Pro与PYR相比,PYR与NaNO2反应更容易形成NPYR,且均在较高温度下才可以形成。上述四个反应中当NaNO2过量时更容易生成N-亚硝胺。实验结果表明,较高的温度、较低的pH、较高的亚硝酸钠可以促进N-亚硝胺的合成,因此在肉制品加工中应控制这些因素从而减少N-亚硝胺的生成。      

低温贮藏下生肉丸中6种氨基酸与N-亚硝胺形成的关系探讨

为初步确定氨基酸与N-亚硝胺之间的转化关系,本研究以精瘦肉为原料,在肉丸中分别添加1 mg/kg赖氨酸、精氨酸、丙氨酸、脯氨酸、酪氨酸、缬氨酸,低温(4 ℃)贮藏9 d,利用气相色谱-串联质谱联用(GC-MS/MS)方法定期测定肉丸中N-亚硝胺含量,探究氨基酸与N-亚硝胺间的转化关系,并分析五味子对N-亚硝胺的产生以及外源氨基酸与N-亚硝胺之间关系的影响。结果表明,肉丸中检出N-亚硝基二甲胺(NDMA)、N-亚硝基哌啶(NPIP)、N-亚硝基吡咯烷(NPYR)、N-亚硝基二正丁胺(NDBA)、N-亚硝基二苯胺(NDPHA)五种N-亚硝胺。精氨酸、脯氨酸可促进N-二甲基亚硝胺(NDMA)、N-亚硝基二正丁胺(NDBA)的生成,其中精氨酸的添加使NDMA含量最高增加了73.18%,脯氨酸的添加使NDBA含量最高增加了156.18%;同时发现五味子使添加精氨酸的NDMA含量最高降低了28.37%,使添加脯氨酸的NDBA含量最大降幅达到29.47%,NPIP含量最大降幅达到48.62%。综上,精氨酸可提高肉制品中NDMA的含量,脯氨酸可提高NDBA的含量;五味子的添加总体上降低了NPIP含量,但差异不显著;五味子对添加精氨酸的NDMA含量有降低作用,同时对添加脯氨酸的NDBA含量也有降低作用。     

Occurrence and source of nitrosamines and secondary amines in groundwater and its adjacent Jialu River basin, China

The presence of mutagenic and carcinogenic nitrosamines in groundwater is of great concern. In this study, eight nitrosamines including N-nitrosodimethylamine (NDMA), N-nitrosodiethylamine (NDEA), N-nitrosomethylethylamine (NMEA), N-nitrosopyrrolidine (NPYR), N-nitrosomorpholine (NMOR), N-nitrosopiperidine (NPIP), N-nitrosodi-n-propylamine (NDPA), and N-nitrosodi-n-butylamine (NDBA) and corresponding secondary amines were investigated in shallow groundwater, river water, and wastewater samples collected from the Jialu River basin. The total concentrations of nitrosamines and secondary amines in groundwater were ND-101.1 ng/L and 0.36-4.38 μg/L, respectively. NDMA and its secondary amine DMA (44.7%/40.1%) were the predominant compounds in groundwater, followed by NDEA/DEA (21.7%/29.3%) and NDBA/DBA (26.4%/27.4%). Relatively high concentrations of these six compounds were also observed in river water that was influenced by the direct discharge of industrial and domestic wastewater. Using acesulfame as a quantitative population marker, the contribution of domestic sources to the concentrations of nitrosamines and secondary amines was 39-85% in downstream reaches of the Jialu River, and that of industrial sources was estimated to be 65-98% in other sites of the area. Both on-site leakage of domestic and industrial wastewater and leaching from river water would contribute to the occurrence of target pollutants in groundwater. The target pollutants posed a cancer risk of 4.12 × 10(-5) to the local populations due to the direct usage of groundwater as potable water.     

Volatile nitrosamines in microwave-cooked bacon

Bacon was analysed for volatile nitrosamines after microwave cooking, and the results were compared with those obtained after frying bacon in a pan. Microwave cooking gave statistically significantly lower levels of all three volatile nitrosamines detected in the bacon. NPYR was found in all 20 samples fried in a pan, but in only five microwave-cooked samples. The average volatile nitrosamine levels (micrograms/kg) in microwave-cooked bacon and pan-fried bacon were: NDMA, 0.3 and 1.2; NPIP, 0.04 and 0.2; NPYR, 0.1 and 4.5.