Fungi associated with post-harvest rot of sweet orange (Citrus sinensis) and aflatoxin B1 production by isolates of Aspergillus flavus on plain and supplemented orange juice

Samples of rotting sweet orange (Citrus sinensis) were obtained from the depots, sales counters and waste baskets. Fungi associated with rotting fruits were isolated and identified. Out of 12 species of fungi isolated, 8 are known to be producers of toxins. The 7 isolates of Aspergillus flavus obtained were screened for aflatoxin production in a nutrient solution, and 4 were found to be aflatoxigenic, producing primarily aflatoxin B₁. Aflatoxin B₁ production of the toxigenic isolates were further studied on plain juice and juice separately supplemented with 2.0% yeast extract and 2.0% sucrose. The highest yield of aflatoxin B₁ was produced on juice supplemented with yeast extract by the 4 toxigenic A. flavus isolates, followed by sucrose supplementation while the lowest amount of aflatoxin B₁ was detected on plain juice. Optimum temperature for aflatoxin B₁ production by A. flavus isolate (IBA-O1) was 25 °C to 30 °C, for an incubation period of 7–11 days on plain and supplemented juice media.     

黄曲霉毒素B1的检测方法

随着农业市场的不断发展, 伴随产生了一系列的食品安全质量问题, 尤其是毒性最强的黄曲霉毒素B1所引起的安全问题。粮食安全问题是一个不容小觑的重大问题, 现在也越来越受到老百姓的重视。本文介绍了黄曲霉毒素B1(aflatoxin B1, AFB1)及其毒性危害; 综述了几种黄曲霉毒素B1的检测方法, 分别包括: 薄层层析法(thin–layer chromatography, TLC)、高效液相色谱法(high performance liquid chromatography, HPLC)、酶联免疫吸附法(enzyme linked immunosorbent assay, ELISA)、金标试纸法(gold labeled immunochromatographic assay, GICA)等, 并对这几种方法进行了分析和比较, 以期为黄曲霉毒素B1的监测提供参考。     

酶联免疫吸附法测定植物油中黄曲霉毒素B1

目的对酶联免疫吸附法(enzyme-linked immunosorbent assay,ELISA)快速检测植物油中黄曲霉毒素B_1进行方法验证,考查植物油中黄曲霉毒素B_1污染情况。方法对酶联免疫吸附法检测植物油中黄曲霉毒素B_1进行了准确性与回收率、重复性、复现性等方法学实验。用验证后的试剂盒检测156份植物油中黄曲霉毒素B_1含量。结果酶联免疫吸附法的回收率在102.8%~113.8%,相对标准偏差为2.0%~6.1%,重复性相对标准偏差为4.3%,复现性相对标准偏差为7.1%和7.6%。156份植物油中黄曲霉毒素B_1检出率为60.90%,其中山茶油的检出率为78.38%,高于平均水平。结论试剂盒检测植物油稳定性较好,定量准确。156份植物油中黄曲霉毒素B_1含量均符合国家标准,花生油中黄曲霉毒素B_1检出浓度最高,其次是玉米油。茶油的加工生产过程可能存在污染黄曲霉毒素B_1的情况,应注意加工过程中黄曲霉毒素B_1的污染。     

Detection of orange juice adulteration by tangelo juice using multivariate analysis of polymethoxylated flavones and carotenoids

Reverse phase HPLC has been applied to quantify levels of polymethoxylated flavones and carotenoids in orange and tangelo juices. Lower levels of sinensetin and tetramethyl‐o‐scutellarein and higher levels of heptamethoxyflavone and tangeretin relative to nobiletin indicated the addition of tangelo to orange juice. β‐Cryptoxanthin and its esters, identified by positive ion electrospray mass spectrometry, were present in larger amounts relative to β‐carotene in tangelo than in orange juice. Using canonical discriminant analysis, the addition of 100 g kg^?1 tangelo to orange juice can be detected. © 2002 Society of Chemical Industry     

酶联免疫法检测酱油中的黄曲霉毒素B1

目的建立酶联免疫法检测酱油中黄曲霉毒素B_1(Aflatoxin B_1,AFB_1)的分析方法。方法酱油样本经纯乙腈(料液比=1:2,V:V)提取,再做1:9稀释,通过酶联免疫吸附法(enzyme linked immunosorbent assay,ELISA)测定样本中AFB_1。结果当加标浓度为2μg/kg和5μg/kg时,纯乙腈对酱油中AFB_1的提取回收率结果分别为121.3%和106.5%;方法检出限为1μg/kg。与国标方法检测结果的相对标准偏差小于10%。结论本方法准确、灵敏度高,可适用于酱油中AFB_1的检测。     

Changes in orange juice color by addition of mandarin juice

The most natural way of improving the color of orange juices is by adding other juices, which provide a more intense coloration. The US legislation allows the addition of up to 10% of mandarin juice to the orange juice to improve its color. The first objective of this study was to compare the color characteristics of juices from 11 mandarin cultivars, currently being grown in Spain. Experimental results proved that only the green-red coordinate, a*, of the orange juices can be improved by adding mandarin juice. The mandarin cultivar that provided a juice with the highest values of a* was Clemenules. Once this selection was made, the effects of adding mandarin juice at different ratios, up to 10%, on the color characteristics of the orange juice were studied. Values of the a* coordinate went from 5.50 for the pure orange juice up to 6.29 for the mixture of 90% orange juice plus 10% of mandarin juice. Finally, hedonic tests proved that regular juice consumers preferred the color of mandarin juice to that of orange juice and that they liked better the color of the juice mixture containing 10% mandarin juice than that containing 3% mandarin juice.     

Detection of target staphylococcal enterotoxin B antigen in orange juice and popular carbonated beverages using antibody-dependent antigen-capture assays

There is a critical need for qualitative and quantitative methodologies that provide the rapid and accurate detection of food contaminants in complex food matrices. However, the sensitivity of the assay can be affected when antigen-capture is applied to certain foods or beverages that are extremely acidic. This study was undertaken to assess the effects of orange juice and popular carbonated soft drink upon the fidelity of antibody-based antigen-capture assays and to develop simple approaches that could rescue assay performance without the introduction of additional or extensive extraction procedures. We examined the effects of orange juice and a variety of popular carbonated soft drink beverages upon a quantitative Interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) assay system and a lateral flow device (LFD) adapted for the detection of staphylococcal enterotoxin B (SEB) in foods. Alterations in the performance and sensitivity of the assay were directly attributable to the food matrix, and alterations in pH were especially critical. The results demonstrate that approaches such as an alteration of pH and the use of milk as a blocking agent, either singly or in combination, will partially rescue ELISA performance. The same approaches permit lateral flow to efficiently detect antigen. Practical Application: The authors present ways to rescue an ELISA assay compromised by acidity in beverages and show that either the alteration of pH, or the use of milk as a blocking agent are not always capable of restoring the assay to its intended efficiency. However, the same methods, when employed with lateral flow technology, are rapid and extremely successful.     

高效液相色谱法测定粮谷中黄曲霉毒素B1的研究

建立了用三氯甲烷提取、硅胶小柱净化、三氟乙酸衍生,再以荧光检测器来测定粮谷中黄曲霉毒素B1的高效液相色谱方法.试验证明,该方法安全限量标准最低可达0.4 μg/kg,线性在2～50 μg/kg良好,平均回收率较高,具有准确度高、精密度好、安全限量低的特点.     

2011~2015年花生和小麦制品中黄曲霉毒素B1的检测

目的 对2011~2015年花生和小麦制品中黄曲霉毒素B1进行检测分析和安全评价。方法 采用酶联免疫法对2011~2015年的1140个花生类和6475个小麦类制品的黄曲霉毒素B1的含量进行检测。结果 2011~2015年检测的花生类制品合格率分别为95.5%、98%、97.4%、98.3%和98.7%; 检测的各种小麦类制品合格率均为100%。结论 花生类制品合格率逐年上升, 但仍存在一定的质量隐患; 小麦类制品质量均符合国家规定限量。     

Binding of aflatoxin B1 by probiotic bacteria

The ability of six probiotic bacteria to bind a common food carcinogen, aflatoxin B₁, was assessed. The studied strains included Lactobacillus strains and one Bifidobacterium strain. The strains were incubated in vitro with alfatoxin B₁ and the toxin residue in the supernatant was measured using high‐performance liquid chromatography. The aflatoxin‐binding capacity of the strains was found to range from 5.8 to 31.3%. The results further support the observation that a number of probiotic bacteria are able to bind specific dietary contaminants. Although the extent of binding varies depending on the bacterial strain used, the data may explain some of the antimutagenic and anticarcinogenic effects of probiotic micro‐organisms. © 2000 Society of Chemical Industry     

柑桔类果汁苦味物质的脱除研究

柠碱和柚苷是柑桔汁和柑桔其他制品中的主要苦味成分,采用吸附脱苦法、酶脱苦法、β-环糊精脱苦法、乙烯利代谢脱苦法及其他脱苦法可将它们除去,文中还提出了采用综合脱苦法应用于生产实际的观点。     

同步荧光光谱法测定橙汁中原果汁的含量

利用同步荧光光谱法对橙汁中原果汁含量进行定量分析。对样本的同步荧光光谱数据进行预处理然后选择不同的光谱波段结合偏最小二乘(partial least squares,PLS)法建立原果汁含量预测模型,并对预测集样本进行预测来验证模型的准确性以及此法的可行性。结果显示:平滑处理结合一阶导数处理后的光谱数据更加适用于模型建立;使用全波段光谱建立的模型性能优于区间波段所建立模型。最终得到的最优模型对预测集进行预测时所得预测均方根误差(Root mean square error of prediction,RMSEP)为0.035832,决定系数(coefficient of determination,R2)为0.972570。由此可以说明同步荧光光谱法结合PLS可实现原果汁含量的定量分析。      

时间分辨荧光免疫法检测食品中黄曲霉毒素B1的含量

目的建立高灵敏度的时间分辨免疫法检测黄曲霉毒素B_1的含量。方法通过双功能螯合剂异氰酸苄基二乙烯三胺四乙酸络合稀土元素Eu~(3+)标记抗AFB_1的单克隆抗体,以AFB_1-OVA为固相抗原,优化反应条件,建立直接竞争时间分辨免疫检测方法。结果优化条件后,方法灵敏度为0.02μg/L,IC_(50)为0.73μg/L,线性检测范围为0.01~30μg/L,大米、花生、黄豆和干果样品回收率在91%~104%之间,检测结果准确;与结构类似物AFB_2、AFG_1、AFG_2、AFM_1的交叉反应率分别为17.3%、4.49%、2.37%和0.73%。结论本方法灵敏度高、特异性强、检测快速,可以满足实际样品的检测需求。     

应用抗黄曲霉毒素单链抗体检测酱油中黄曲霉毒素B1

利用本实验室制备的抗黄曲霉毒素B1的单链抗体(ScFv),通过棋盘实验确定了抗原抗体的最适工作浓度,在此之上根据间接竞争酶联免疫法(ELISA)绘制标准曲线,检测酱油中AFB1的含量;通过改变样品的盐浓度及pH来确定其对ELISA检测结果的影响。研究结果表明,利用ScFv检测黄曲霉毒素的最小检测值为0.10ng/mL,平均加标回收率在84%～109%之间,本文建立的ELISA方法在pH5～8,盐浓度小于10%时较稳定。本文建立的利用抗AFB1的ScFv检测黄曲霉毒素含量的方法方便快捷,稳定性较好,并且成本较低,适合于食品中黄曲霉毒素的检测。     

花生油生物精炼法去除黄曲霉毒素的研究

目前已经筛选出能够固态发酵花生粕降解其中黄曲霉毒素B1(AFB1)的微生物菌株,因此可以利用发酵液提取粗酶来降解花生油中的AFB1,并将这种酶反应引入到花生油的精炼工艺中.通过对比精炼工艺,将酶法脱胶(采用磷脂酶A1)与去毒反应相结合.对脱胶去毒过程中反应温度、NaOH添加量、去毒粗酶添加量、磷脂酶A1添加量、反应时间等进行单因素实验,并通过正交实验确定脱胶去毒实验优化工艺条件为:反应温度40℃,反应时间4h,NaOH添加量0.025％,磷脂酶A1添加量600 mg/kg.以添加AFB1为100 μg/kg的花生毛油为实验对象,取样10 g,在去毒粗酶添加量100 μL及优化条件下,AFB1去除率高达81％,去毒反应后花生油的脱胶效果也达到精炼要求(磷含量降至13 mg/kg).     

基于金标抗体的电化学免疫传感器检测米糠油中黄曲霉毒素B1

目的 为了快速检测食品中黄曲霉毒素B1的含量,本文提出一种以胶体金标记抗体的电化学免疫传感器用于米糠油中AFB1的检测。方法 首先在金电极(AuE)表面修饰抗原,再进一步修饰制备好的金标抗体,最后带有辣根过氧化物酶的二抗(IgG-HRP)通过与抗体反应结合到电极表面。当目标物AFB1存在时,与抗原竞争结合金标抗体,HRP催化过氧化氢(H2O2>)将溶液中对苯二酚(HQ)氧化成苯醌(BQ)的电流信号会下降,间接检测AFB1。结果 该免疫传感器在抗原包被量和金标抗体用量为3 μL且金标抗体与AFB1孵育80 min条件下具有最佳性能。该法线性范围为0.02 ng/mL~80 ng/mL,检出限为0.027 ng/mL。将该传感器应用于米糠油中进行加标回收实验,回收率在88.7%～108.0%。结论 该传感器具有良好的特异性、重现性和稳定性,可用于植物油中AFB1的快速检测。     

郫县豆瓣原料蚕豆中黄曲霉毒素B1的提取

郫县豆瓣中所用蚕豆是产自四川省和云南省的优质干蚕豆,经筛选、浸泡、接种、制曲等一系列加工后形成调味豆瓣酱。采用酶联免疫吸附法(ELISA),根据Box-Behnken Design软件的设计原理,检测原料蚕豆中黄曲霉毒素B1。探讨了甲醇浓度、料液比、超声波提取时间等因素对样品中黄曲霉毒素B1提取的影响,以确定检测原料蚕豆中黄曲霉毒素B1含量的最优方案。以黄曲霉毒素B1含量为响应值,利用响应面分析法对提取参数进行优化。结果表明:甲醇浓度43.41%,料液比1∶5.35,超声波提取时间39.89 min是检测原料蚕豆中黄曲霉毒素B1含量的最优条件,以该优化条件检测原料蚕豆中黄曲霉毒素B1其值达到最大,为1.50μg/kg。     

Ultraviolet treatment of orange juice

Ultraviolet (UV) with a wavelength of 254 nm tends to inactivate most types of microorganisms. Most juices are opaque to UV due to the high-suspended solids in them and hence the conventional UV treatment, usually used for water treatment, cannot be used for treating juices. In order to make the process efficient, a thin film reactor was designed and constructed from glass with the juice flowing along the inner surface of a vertical glass tube as a thin film. The decimal reduction doses required for the reconstitute orange juices (OJ; 10.5° Brix) were 87±7 and 119±17 mJ/cm² for the standard aerobic plate count (APC) and yeast and moulds, respectively. The shelf life of fresh squeezed orange juice was extended to 5 days with a limited exposure of UV (73.8 mJ/cm²). The effect of UV on the concentration of Vitamin C was investigated using both HPLC and titration methods of measurements. The degradation of Vitamin C was 17% under high UV exposure of 100 mJ/cm², which was similar to that usually found in thermal sterilization. Enzyme pectin methylesterase (PME) activity, which is the major cause of cloud loss of juices, was also measured. In contrast to the heat treatment, UV processing does not inactivate enzyme pectin methylesterase. The energy required for UV treatment of orange juice (2.0 kW h/m³) was much smaller than that required in thermal treatment (82 kW h/m³). The color and pH of the juice were not significantly influenced by the treatment.

Industrial relevance

This paper is of interest since it suggest—despite the low UV transmittance in orange juices—the use of a thin film UV reactor. The data suggest that shelf life extension from 2 to more than 5 days could be achieved with less energy requirements than for thermal processing. However, it seems unlikely that such thin film reactors could provide a real alternative to current conventional or new (e.g. high pressure pulsed electric field) processing.     

Quantitative determination of carbohydrates in orange juice by gas chromatography

 A gas chromatographic method using a capillary column is described for the quantitative analysis of fructose, glucose, sucrose and myo-inositol in orange juice. The method is evaluated for precision and recovery using phenyl-β-glucoside as an internal standard. The results support the suitability of the method. Carbohydrates (fructose, glucose, sucrose and myo-inositol) were determined in different kinds of orange juice. The determination of carbohydrate composition and ratios of the carbohydrate constituents provide a method to assess orange juice quality, especially the myo-inositol content and myo-inositol/fructose ratio. These new indices, which were found to be lower in samples made from concentrates, provide information on the quality and genuineness of orange juice. Received: 10 June 1997