Effects of organonitrogen, carbamate pesticides and others on beta-hexosaminidase release from rat basophilic leukemia cells (RBL-2H3)

There have been very few reports of the effect of pesticides on immediate allergy. In the previous report, the effect of pyrethroid pesticides and organophosphorus pesticides on immediate allergic reaction was investigated. Subsequently, 12 organonitrogen pesticides, 14 carbamate pesticides and 4 other pesticides were investigated for their effects on the enzyme activity of beta-hexosaminidase as an index of chemical mediator release from rat basophilic leukemia cells (RBL-2H3). Two organonitrogen pesticides, bitertanol and pyridaben, and two organotin pesticides, cyhexatin and fenbutatin oxide, were found to promote beta-hexosaminidase release. Bitertanol non-specifically caused the promotion of chemical mediator release, while the release-promotive action of pyridaben was related to IgE antibody and those of cyhexatin and fenbutatin oxide to cell injury. On the other hand, two organonitrogen pesticides, propiconazole and triadimenol, and imazalil showed release-inhibitory action. These data suggested that some pesticides can affect immediate allergy.     

Inhibitory effect of zinc on beta-hexosaminidase release from RBL-2H3 cells by synthetic chemicals

The effects of foods and chemicals related to food hygiene on degranulation were evaluated using a method for assaying the enzyme activity of beta-hexosaminidase as an index of chemical mediator release from RBL-2H3 cells in vitro. Using a previously developed assay system, we had found a large number of inhibitors and promoters of degranulation of RBL-2H3 cells. In the present study, we examined the inhibitory effect of zinc chloride on the degranulation (beta-hexosaminidase release) from RBL-2H3 cells with or without antigen in the presence of the degranulation-promotive chemicals, namely, 4 food additives, 7 pesticides and 2 veterinary drugs. These promotive chemicals were classified into two types on the basis of inhibitory profile by zinc chloride: 1) those which showed marked degranulation-inhibitory action when the cells were stimulated with antigen, such as butylhydroxyanisole, dibutylhydroxytoluene, EPN, cis- and trans-permethrin, prothiofos, pyridaben, terbufos, 2) those which showed marked degranulation-inhibitory action whether the cells were stimulated with antigen or not, such as butyl p-hydroxybenzoate, o-phenylphenol, bitertanol, salinomycin. In conclusion, zinc had a dramatic inhibitory effect on enhanced degranulation induced by synthetic chemicals in vitro.     

固相萃取-超高效液相色谱-串联质谱法同时测定动物源性食品中多种兽药残留

利用通过固相萃取法结合超高效液相色谱-串联质谱法建立同时检测猪肉、鱼、虾等动物源性食品中磺胺类、喹诺酮类、糖皮质激素类、大环内酯类、β-受体激动剂类、类固醇激素类、四环素类、青霉素类、氯霉素类、头孢菌素类10大类72种兽药残留的分析方法。样品经80%乙腈溶液提取,直接通过不经活化的Prime HLB固相萃取柱进行净化。使用C_(18)色谱柱分离,电喷雾离子源分段多反应监测扫描模式检测,外标法定量。72种兽药定量限为0.5~5μg/kg,各基质中回收率在75.6%~122.3%之间,相对标准偏差为0.6%~10.7%。本方法极大地提高了多残留兽药的检测效率,适用于食品安全突发事件应急处置中多残留兽药的快速分析。     

抗生素和其他兽药残留的危害

本文较为详细地介绍了动物性食品抗生素残留的危害，如细菌耐药性的普遍产生，易导致二重感染，扰乱机体微生态，药物对机体的直接不良反应、直接毒性作用，各种变态反应等等，本文也简要描述了其他兽药非抗生素兽药和激素的毒副作用，列举了我国违禁兽药清单。     

原料乳中药物残留和掺假物质检测方法的研究进展

原料乳中药物残留或掺假物质的存在造成的安全事件屡屡发生,这使我国乳品行业面临着巨大的挑战。本文主要综述了常规检测法、免疫分析法、仪器检测分析法、生物传感器和蛋白质芯片技术等对原料乳中常见聚醚类抗生素、类固醇激素、苯二氮卓类药物、苯基脲类除草剂、四环素类抗生素和青霉素类抗生素等药物残留、物理性质与原料乳相近的物质、常见电解质、非电解质、防腐剂等掺假物质的检测方法,以及对单一成分掺假物和多种成分掺假物的检测方法,并比较了不同检测方法的准确度、灵敏度、成本等优缺点,以期为相关检测部门进行质量监测或生产厂家牛乳收购提供技术参考。     

Direct induction of CCK and GLP-1 release from murine endocrine cells by intact dietary proteins

Scope: Consumption of high‐protein diets cause elevated levels of CCK and GLP‐1. Although unknown, this might be due to protein breakdown by various proteases that originate from the gastrointestinal tract. This study investigated which dietary proteins, hydrolysates, or synthetic‐peptides are most potent to affect secretion of CCK and GLP‐1 in STC‐1 cells known for satiety hormone release. Methods and results: Addition of intact proteins to STC‐1 cells exerted strong effects on secretion of satiety hormones. Casein, whey, and pea showed strongest effects on CCK release, whereas casein, codfish, egg, and wheat showed most pronounced effects on GLP‐1 release. Egg‐hydrolysate stimulated release of CCK and GLP‐1, whereas all other tested hydrolysates and synthetic‐peptides showed no significant effects on hormone release. Addition of a combination of trypsin and casein‐hydrolysate, codfish, egg, egg‐hydrolysate, sodium‐casein, wheat‐hydrolysate, or wheat resulted in additional stimulation of CCK release, compared to only the protein. Addition of a combination of DPP‐IV and egg‐hydrolysate, ovomucoid, or sodium‐casein decreased GLP‐1 levels. Conclusion: This study showed that specific intact, or partially digested proteins, in contrast to protein‐hydrolysates and synthetic‐peptides, stimulated hormone release. We conclude that intact proteins exert strong effects on satiety hormone release, and may therefore provide potent dietary supplements for prevention or treatment of obesity.     

Simultaneous determination of multi-class antibiotics and steroid hormones drugs in livestock and poultry faeces using liquid chromatography–quadrupole time-of-flight mass spectrometry

ABSTRACT

A method for simultaneous determination of multi-class antibiotics and steroid hormone analysis in faeces of livestock and poultry was developed using liquid chromatography–quadrupole time-of-flight mass spectrometry (LC-QTOF MS). An in-house database was built for 156 detected drugs using Personal Compound Database Library software (PCDL) including compound name, monoisotopic mass, chemical formula, RT, chemical structure and three CID MS/MS spectra. The linearity result showed that all the drugs exhibited good linearity with determination coefficients (R²) higher than 0.99. The drug recoveries and their RSDs for all three faeces samples (pig, cattle and chicken) were tested and 81, 96 and 92 drugs were chosen for analysis in pig, cattle and chicken faeces, respectively. Further validation showed that 73 veterinary drugs in all three kinds of faeces samples can be quantified in one analytical run. This work shows that qualitative and quantitative analysis using LC-QTOF MS represents a simple, sensitive, low-cost and high-throughput methodology in routine laboratory analyses.     

我国与欧盟、美国、国际食品法典委员会的禽蛋抗生素类兽药残留限量标准对比分析

近年来,鸡蛋中兽药残留成为食品安全问题的焦点。本文对国际食品法典委员会、欧盟、美国、中国关于禽蛋中抗生素类兽药的最大残留限量和限制种类进行总结比较。结果表明,我国关于动物源性食品中兽药最大残留限量的法规更新速度慢,禽蛋抗生素类兽药限制种类不全面,限量值有待进一步研究。2020年4月1日实施的食品安全国家标准(GB31650-2019)基本覆盖了我国兽药种类和动物组织,进一步完善了我国的兽药管理制度。结合我国实际,需加强兽药监管能力建设,加快相关标准的修订与更新,为我国养殖业的科学用药提供法规依据,从源头上减少和控制兽药的使用。     

免疫分析技术在乳及乳制品抗生素类兽药残留检测中的应用

为了预防与治疗乳牛疾病,抗生素类兽药被广泛应用于乳牛的饲养过程中。而滥用抗生素导致牛奶中抗生素残留超出最大允许残留量,不仅会对牛奶品质造成影响,而且可能会危害到牛奶饮用者的身体健康。因此,如何快速、准确地检测乳与乳制品中抗生素类兽药残留是乳品检测行业和企业亟需解决的问题。本文主要介绍了牛乳中兽药残留的来源、危害及其现状,阐述了酶联免疫分析技术、胶体金免疫层析技术的检测原理及其在应用过程中所具有的不同特点,最后对2种方法的优势与不足进行简要总结,并对免疫分析法在乳与乳制品抗生素类兽药残留检测中的应用前景做出展望。     

我国与国际食品法典委员会（CAC）食品中兽药最大残留限量标准的对比分析

对比分析我国与国际食品法典委员会(CAC)食品中兽药最大残留限量标准的异同。从标准涉及的兽药种类、具体食品种类、残留限量值三个层面逐级对比。结果表明我国标准涉及的兽药种类为217种,CAC标准为57种,二者相同的兽药种类42种;我国标准在残留具体食品动物的种类与数量上与CAC标准基本相同;我国兽药残留限量指标的总数为1035个,远远多于CAC的477个;在可比指标范围内,我国有302个残留限量指标值与CAC相同,26个残留限量指标值严于CAC,仅8个残留限量指标值宽于CAC。因此我国兽药残留限量标准中有98%的可比指标值已达到或超过CAC标准的要求。     

Inhibitory effects of water extracts of fermented rice bran on allergic response

In this study, the effects of water extracts of fermented rice bran (FRBE) on immediate-type allergic reactions in vitro (RBL-2H3 cell line) and in vivo [ovalbumin (OVA) sensitized mice model] were investigated. FRBE inhibited β-hexosaminidase release from RBL-2H3 cells activated by anti-dinitrophenyl (DNP) IgE while FRBE pretreatment profoundly inhibited the systemic allergic reaction induced by compound 48/80. Moreover, FRBE had a significant inhibitory effect on the mRNA expression of anti-DNP IgE-induced tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-4, IL-6, and IL-10, all of which mediate inflammatory reactions triggered by allergic reaction. To determine the mRNA levels of cytokines in an OVA-sensitized mice model after FRBE treatment, spleen cells were harvested with several concentrations of FRBE. The FRBE-treated group showed decreased levels of TNF-α, IFN-γ, IL-6, and IL-10. These results suggest that FRBE suppressed allergic and inflammatory reactions through inhibition of degranulation, histamine release, and pro-inflammatory cytokine production from mast cells.     

Investigation of transformation products of selected veterinary drugs generated by electrochemistry,microsomal assays,hydrolysis & photolysis

The knowledge of transformation pathways and transformation products of veterinary drugs is important for health, food and environmental matters. Residues (original veterinary drug and transformation products) are found in food products of animal origin and also in the environment (e.g., soil or surface water). Several transformation processes can alter the original veterinary drug, ranging from biotransformation in living organism to environmental degradation processes like photolysis, hydrolysis, or microbial processes. In this thesis, four veterinary drugs were investigated, three ionophore antibiotics Monensin, Salinomycin and Lasalocid and the macrocyclic lactone Moxidectin. Ionophore antibiotics are mainly used to cure and prevent coccidiosis in poultry especially prophylactic in broiler farming. Moxidectin is an antiparasitic drug and is used for the treatment of internal and external parasites in food- producing and companion animals. The main objective of this work was the usage of different laboratory approaches to generate and identify transformation products. The identification was conducted using high-resolution mass spectrometry (HR-MS). A major focus was put on the application of electrochemistry for simulation of transformation processes. The electrochemical reactor, equipped with a three-electrode flow-through cell, enabled the oxidation or reduction by applying a potential. Derived transformation products were analyzed by online coupling of the electrochemical reactor and a HR-MS and offline by liquid chromatography (LC) combined with HR-MS. The main modification reaction of the identified transformation products was different for each investigated veterinary drug. Monensin was showing decarboxylation and demethylation as main modification reactions, for Salinomycin mostly decarbonylation was occurring and for Lasalocid methylation was prevalent. For Moxidectin I observed oxidation (hydroxylation) reaction and adduct formation with solvent. In general, for salinomycin and Lasalocid more transient transformation products (online measurement) than stable transformation products (offline measurements) were detected. In contrast, the number of transformation products using online and offline measurements were identical for monensin and moxidectin. As a complementary approach, metabolism tests with rat or human liver microsomes were made for the ionophore antibiotics. Monensin was investigated by using rat liver microsomes and identified transformation products were based on decarboxylation and demethylation. Salinomycin and Lasalocid were converted by human and rat liver microsomes. For both substances were more transformation products found by using human liver microsomes. The transformation products of the rat liver microsome conversion were redundant, the transformation products were also found at the human liver microsome assay. Oxidation (hydroxylation) was found to be the main modification reaction for both. In addition, a frequent ion-exchange between sodium and potassium was identified. The last two experiments were performed for one substance each, the hydrolysis of monensin and the photolysis of moxidectin was investigated. The transformation products of the pH- dependent hydrolysis were based on ring-opening and dehydration. Moxidectin formed several transformation products by irradiation with UV-C light and main modification reactions were isomeric changes, (de-)hydration and changes of the methoxime moiety. In summary, transformation products of the four investigated veterinary drugs were generated by the different laboratory approaches. Most of the identified transformation products were identified for the first time. The resulting findings provide an understanding for clarifying the transformation behavior.     

核酸适配体在食品危害物多靶标检测中的应用进展

文章总结了已报道筛选获得的致病菌、生物毒素、抗生素、兽药、农药、重金属等食品危害物核酸适配体现状,重点综述了近年来利用核酸适配体技术实现食品危害物多靶标检测研究进展,并展望了其未来发展方向。     

Untersuchungen zum Transfer pharmakologisch wirksamer Substanzen aus der Nutztierhaltung in Porree und Weißkohl

Investigation on the transfer of pharmacologically active substances used in animal husbandry into leek and cabbage. The potential of leek and cabbage for uptake of highly prescribed veterinary drugs (antibiotics) was tested in hydroponically grown plants. For this purpose the antibiotics sulfadiazine (SFD), enrofloxacine (ENR), tetracycline (TC), chlortetracycline (CTC) and monensine (MON) were chosen. A further aim was to gain data on the situation of vegetables grown in agricultural practise with regard to antibiotic residues. The evident effects of the antibiotics on plants grown hydroponically (each antibiotic was administered at 5 μmol/l nutrient solution) were greatly different: With regard to leek there were no visible effects (MON, SFD), a weak bleaching of the younger leaf sections (CTC), and strong effects of ENR. The (phytotoxic) effects of antibiotics on cabbage were much more distinct. CTC caused a yellowing of the plant vasculature in cabbage. MON induced lesions on some leaves and finally led to leaf wilting. With administration of ENR a nearly complete bleaching of young leaves was observed. Using LC-MS/MS-methods (low-resolution and high-resolution MS) the administered antibiotics, as well as conversion products and metabolites, were separately identified and quantified in various organs of leek (roots, young and old sections of leaves) and cabbage (roots, stalks, young and old leaves). Depending on the type of antibiotic, vegetable species, and plant organ, the detected concentrations of antibiotic residues comprised several orders of magnitude ranging from μg/kg to mg/kg of fresh weight (fw). The highest concentrations of antibiotics were found in roots of both vegetable species: CTC and TC were detected at approximately 10 mg/kg fw in cabbage roots and at approximately 20 mg/kg fw in leek roots and ENR was determined at approximately 12 mg/kg fw in cabbage roots. Low amounts of ENR were metabolised to ciprofloxacine (CIP). ENR occurred at similar concentrations of approximately 7 mg/kg fw in roots and old leaves of cabbage, indicating a high transport rate of this antibiotic in the cabbage plant. In stalks, young and old leaves of cabbage and in young and old leaf sections of leek all administered antibiotics were detected. Within these antibiotics, ENR and CTC and their conversion products, e. g. demeclocycline (DMC) and TC, occurred at the highest concentrations. SFD and MON were found in considerably lower concentrations (<100 μg/kg fw). The results of our experiments in hydroponic cultures, using defined concentrations of antibiotics in the nutrient solution, evidently demonstrate that cabbage and leek have a very high potential for uptake of a number of veterinary antibiotic drugs, especially for tetracycline and ENR.     

Statistical evaluation of residue data for the assessment of predicted values (half-life, withdrawal time) as an example of toltrazuril and enrofloxacin in trout

The half-lives and withdrawal times of the veterinary drugs Toltrazuril and Enrofloxacin in trout have been assessed by statistical analysis. Confidence intervals were computed using a normal distribution of residual data and an empirical distribution by the Bootstrap method. Both methods produced similar statistics for the two drugs. Simulation of the residue data according to the regression lines of the decay curves has shown that the Bootstrap method is better for use when the residue patterns are not distributed normally. Using confidence intervals, a statistical mean of withdrawal times can be assessed. Taking into account the decays of the individual antibiotics in all treated trout, tolerance intervals for the regression lines are obtained: the calculated 10-20% longer withdrawal time includes values for which the antibiotic concentration in 95% of the treated trout is decreased below the tolerance level of Toltrazuril or Enrofloxacin.     

Novel in vitro systems for prediction of veterinary drug residues in ovine milk and dairy products

A new in vitro tool was developed for the identification of veterinary substrates of the main drug transporter in the mammary gland. These drugs have a much higher chance of being concentrated into ovine milk and thus should be detectable in dairy products. Complementarily, a cell model for the identification of compounds that can inhibit the secretion of drugs into ovine milk, and thus reduce milk residues, was also generated. The ATP-binding cassette transporter G2 (ABCG2) is responsible for the concentration of its substrates into milk. The need to predict potential drug residues in ruminant milk has prompted the development of in vitro cell models over-expressing ABCG2 for these species to detect veterinary drugs that interact with this transporter. Using these models, several substrates for bovine and caprine ABCG2 have been found, and differences in activity between species have been reported. However, despite being of great toxicological relevance, no suitable in vitro model to predict substrates of ovine ABCG2 was available. New MDCKII and MEF3.8 cell models over-expressing ovine ABCG2 were generated for the identification of substrates and inhibitors of ovine ABCG2. Five widely used veterinary antibiotics (marbofloxacin, orbifloxacin, sarafloxacin, danofloxacin and difloxacin) were discovered as new substrates of ovine ABCG2. These results were confirmed for the bovine transporter and its Y581S variant using previously generated cell models. In addition, the avermectin doramectin was described as a new inhibitor of ruminant ABCG2. This new rapid assay to identify veterinary drugs that can be concentrated into ovine milk will potentially improve detection and monitoring of veterinary drug residues in ovine milk and dairy products.     

Simultaneous determination of veterinary drugs in livestock foods and seafoods using liquid chromatography/tandem mass spectrometry

A simultaneous determination of veterinary drugs in livestock food and seafood using liquid chromatography with tandem mass spectrometry (LC/MS/MS) was developed. Veterinary drugs were extracted with 95% acetonitrile. The solution was passed through a Florisil column, and the solvent was replaced with phosphate buffer. The extract was charged on a Sep-Pak Plus C(18) mini-column and divided into 40% methanol eluate fraction and 70% acetonitrile eluate fraction. Test solutions were analyzed by LC/MS/MS with gradient elution. By using this method, 37 kinds of veterinary drugs were obtained with over 60% recovery, and quantitation was possible in cattle muscle, egg and fish. This method was inapplicable to 28 kinds of veterinary drugs. Although quantitation was not achieved, 42 other kinds of veterinary drugs can be screened. Since the limit of quantitation for this method is less than the provisional limit in general, it is useful as a screening method in residual analysis of veterinary drugs.     

农作物中抗生素检测技术的研究进展

随着养殖业的规模化和集约化发展,抗生素作为疾病预防与治疗药物、生长促进剂、饲料添加剂等被广泛用于畜禽养殖和水产养殖业中,进而进入水环境和土壤环境,被农作物根系吸收进入茎、叶中并在植物体内富集,从而通过食物链进入人体内。近年来研究者对农作物中抗生素的研究逐渐增多。目前对农作物中兽用抗生素残留的前处理技术主要包括Qu ECh ERs方法和固相萃取法,检测技术主要包括高效液相色谱法、高效液相色谱-串联质谱法和酶联免疫法。高效液相色谱-串联质谱法具有高灵敏度、高选择性和高准确度,可同时快速测定多个目标物,更适合农作物中兽用抗生素残留的检测。本文对环境中抗生素的来源、农作物中抗生素污染的现状、抗生素残留检测的提取、净化方法以及检测方法的研究进展进行了综述。     

Efficient removal of estrogenic activity during oxidative treatment of waters containing steroid estrogens

The discharge of synthetic and natural steroid estrogens from municipal wastewaters to the aquatic environment has received increased attention because of their potential reproductive effects on fish. Using 17alpha-ethinylestradiol (EE2) as a representative steroid estrogen, several oxidants applied in wastewater treatment (chlorine, bromine, ozone, hydroxyl radical, chlorine dioxide, and ferrate) were shown to selectively and rapidly transform EE2. For typically applied oxidant doses, these transformations occur in the time range of seconds to minutes. The resulting initial transformation products of EE2 exhibit a substantially lower in vitro estrogenic activity (< 13% of EE2). For selected structural derivatives of EE2, a quantitative structure-activity relationship was established between substituents changed on the phenolic moiety and the relative in vitro estrogenic activity. In addition, the initial EE2 transformation products that still exhibit residual estrogenic activity are quickly further transformed by most of the tested oxidants. Therefore, oxidative wastewater treatment may serve as a powerful tool to remove estrogenic activity induced by steroid estrogens.     

4种不同净化方法在猪肉兽药残留检测中的应用比较研究

目的 通过对猪肉中194种兽药残留的回收率和基质效应进行分析, 评价ProElut、PRiME、EMR、LipoNo 4种方式的净化效果。方法 样品采用0.1%甲酸-90%乙腈混合溶液提取后, 取上清液分别经ProElut、PRiME、EMR、LipoNo 4种不同的方式净化, 氮吹浓缩复溶后经超高效液相色谱-串联质谱法分析, 考察兽药的回收率和基质效应。结果 经4种方式净化后, 分别有84.5%、82.0%、80.0%、91.2%兽药的回收率在60%~120%之间, 回收率低于60%或高于120%的兽药数量分别是30、36、38、17种; 分别有62.9%、55.7%、83.0%、83.0%兽药的基质效应为弱基质效应, 6.2%、4.6%、3.1%、3.1%的兽药为强基质效应。结论 LipoNo处理后各类兽药平均回收率在60%~120%间比例最多, 基质效应干扰相对较小, 适用于猪肉中194种兽药残留量的测定。