红光响应眼镜蛇神经毒素PEG-PLGA纳米囊的制备与表征

以聚乙二醇-聚乳酸-聚乙醇酸嵌段共聚物(PEG-PLGA)为囊材,添加脱镁叶绿酸作为光敏剂,采用复乳法制备了光响应的眼镜蛇神经毒素纳米囊。以纳米囊的包封率、载药量和粒径为指标,采用单因素法对纳米囊的制备条件进行优化;以差示量热扫描分析其热流变性能,并以累积释药量研究其光控释行为。优化的PEGPLGA、眼镜蛇神经毒素及光敏剂脱镁叶绿酸质量配比为40∶12.5∶1,获得的纳米囊包封率为72.3%±3.6%,载药量为15.1%±1.3%,平均粒径为(862±23)nm,电位为(-46.5±3.8)m V,呈紧密球形,光敏剂分布在囊壳;在650 nm半导体激光照射30 min,体外释放明显加快。该纳米囊在不光照时具有增强药物稳定和缓释作用,而红光可促进药物释放,因而可实现光控靶向。     

Conjugated fluorescent polymer sensor for proteolytic activity detection with designed specificity

A portable fluorescence assay for direct endopeptidase activity detection has been developed with the use of a cyclophane‐based conductive conjugated fluorescent polymer and peptide substrates. The substrates, carrying internal quenching amino acid, were designed to be cleaved in a sequence‐specific manner by a protease of interest. Intact substrates were incapable of quenching the fluorescence of the polymer due to steric constraints. Upon specific cleavage, the quencher became exposed and could interact with the ring structure of the fluorescent polymer, disrupting the conjugation and quenching the fluorescence along the polymer chain. The approach was developed using a model Glu‐C endopeptidase from Staphylococcus aureus strain V8, detected in the picomolar to micromolar concentration range. The developed assay was tested for the detection of endopeptidase activity of botulinum toxin. The feasibility study showed that botulinum neurotoxin (BoNT)‐A could be detected down to picomolar concentration, with limit of detection of 5 pg, or 33 amol in 5 µL of sample, and a total assay time under 2 h. The assay exhibited high specificity and no cross‐reactivity with BoNT‐B was detected. Following this proof‐of‐concept work, the assay can be further optimized and expanded to differentiate between various botulinum toxin serotypes in their active proteolytic form, or modified for detection of proteases with other specificities. © 2015 Society of Chemical Industry     

Toxin Development by Clostridium botulinum in Modified Atmosphere-Packaged Fresh Tilapia Fillets During Storage

Potential for toxin development by Clostridium botulinum type E was investigated in retail-type packages of fresh tilapia fillets packaged in high barrier film under selected atmospheres [100% air, a modified atmosphere (MA) containing 75%CO₂:25%N₂, and vacuum] and stored under refrigeration (4°C) and abuse temperatures (8 and 16°C). Toxin development coincided with sensory spoilage at 16°C storage for fillets packaged in either MA, or vacuum. At 8°C, toxin development in fillets packaged in either of the atmospheres occurred 7-23 days after sensory spoilage. At 4°C, none of MA-packaged fillets became toxic until 10 days after onset of sensory spoilage. Toxin development did not precede sensory spoilage in any treatments or temperatures studied.     

石家庄市肉毒梭菌在环境中的分布研究

目的:了解石家庄市肉毒梭菌在环境中的分布情况,为预防、诊断、治疗肉毒梭菌食物中毒提供科学依据.方法:按照地貌,均匀抽取8个县(市、区),采集土壤、食品和水3类样品,检测肉毒梭菌污染情况.结果:采集土壤、水、食品3类样品共350份,肉毒梭菌检出15份,检出率为4.3%,其中,12份为8型肉毒毒素,3份为A型肉毒毒素.发生过肉毒中毒的县(市、区)肉毒梭菌检出率高于未发生过的县(市、区)(P<0.05).山区与平原肉毒梭菌检出率差异无统计学意义.结论:石家庄市外环境中肉毒梭菌污染较以前严重,并且存在A型肉毒毒素.     

肉毒梭菌与食物中毒

肉毒梭菌是严重食物中毒的病原菌之一，是一种典型的腐物寄生菌，侵入人及家畜肠道内不能生长繁殖，但在消毒不彻底或污染的罐头、香肠、豆制品等食物中，在厌氧的环境下生长繁殖，并产生强烈的外毒素。这种毒素毒性强烈，性质稳定，是目前已知化学毒物和生物毒素中毒性最强烈者，对人的最小致死量约为0．0000001g。     

The effect of growth temperature on the phospholipid and fatty acyl compositions of non-proteolytic Clostridium botulinum

A non-proteolytic strain of Clostridium botulinum (NCIB 4270) was found to have a complex lipid composition, comprising five major phosphorus-containing lipids: phosphatidylethanolamine (PE), phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), phosphatidylserine (PS) and a glycophospholipid of unknown structure (GPL), in order of abundance. Changing the growth temperature did not alter the lipid composition either qualitatively or quantitatively. The main fatty acyl components of the lipids are 14:0, 16:0 and 16:1. When the growth temperature was lowered from 37 to 8°C, there was an increase in 14:0 from 16.4 to 37.5%, an increase in 16:1 from 10.5 to 22.5%, and a decrease in the proportion of 16:0 from 40.3 to 19.1%. There was also a decrease in the proportion of cyclopropane fatty acids (15:0cyc and 17:0cyc) from 7.3 to 0.5%, and in the equivalent chain length of the total fatty acids from 15.9 to 15.3 as the temperature was lowered. The same temperature-dependent changes occurred in the five major lipid classes examined. Despite reports of the presence of plasmalogenic forms of phospholipids (i.e. those lipids which have the acyl chain in the sn-1 position replaced by an alk-1-enyl group) in some Clostridium spp., none were detected in C. botulinum NCIB 4270 using either commercially available spray reagents or by gas-liquid chromatographic analysis of the products or acid methanolysis of total lipid extracts. It is concluded that non-proteolytic C. botulinum lacks plasmalogens, typical of other clostridia, in its membranes and instead modulates its fatty acid composition in response to temperature changes in a manner that is typical of other (non-clostridial) bacteria.     

不同浓度芽孢萌发剂对肉毒梭菌芽孢萌发的影响

肉毒梭菌（Clostridium botulinum）及芽孢是真空包装、冷冻食品及罐装食品中的主要杀菌对象,其抗逆性强,生长速度快,而成为食品中毒的主要原因之一。本试验采用单因素试验和响应面分析法研究芽孢萌发剂L-丙氨酸和KCl添加浓度,处理温度和处理时间对肉毒梭菌芽孢萌发条件的影响,探究影响肉毒梭菌芽孢萌最适条件,并结合常压100 ℃,20 min杀菌,研究最终肉毒梭菌的杀灭效果。结果显示,当处理温度为75.5 ℃,处理时间为25.0 min,添加L-丙氨酸浓度为15.4 mmol/L,KCl浓度为0.53%时,芽孢萌发率最大,可达89.91% ± 1.31%,经过两次100 ℃杀菌20 min,肉毒梭菌杀灭率可达98%。研究结果可为食品工业中罐头的常温杀菌提供一定的技术指导。     

不同碳源对传统酸肉中微生物菌群和生物胺的影响

为了研究添加小米饭、糯米饭、籼米饭、再接种籼米饭以及加入葡萄糖酸内酯对传统酸肉制品中肉毒梭状芽孢杆菌、生物胺含量及产品品质的影响,对预腌和发酵期间产品的水分含量、水分活性、pH及菌相的变化进行了分析。结果表明,传统酸肉制品的水分含量介于65.21%～70.44%之间,添加不同的米饭并未促进初期pH的下降;各组均未检出肉毒梭状芽孢杆菌,且大肠菌群经发酵2d后便低于4个对数单位,经发酵5d后低于1个对数单位;除了微球菌科和真菌类增殖较慢外,耐盐性细菌、乳酸菌和乳酸菌群等微生物均生长良好,发酵5d后菌数均维持在7～8个对数单位。添加GDL组在发酵初期有降低pH和水分含量,且能抑制微球菌生长的效果,添加GDL组挥发性盐基氮和生物胺含量较低,能提高传统酸肉的贮藏品质。     

乳制品中肉毒杆菌安全事件警示

自恒天然乳粉事件发生后,对我国乳品行业产生了一定的不良影响,由此引发了一系列的问题。本文重点阐述了国际上一些国家在乳制品或食品标准中对肉毒杆菌及其毒素的限量标准现状及检测方法,以期为我国乳制品或食品中肉毒杆菌及其毒素标准的制订提供基础参考,为我国乳品加工行业提供相关技术指导。     

An Integrated Pharmacophore/Docking/3D-QSAR Approach to Screening a Large Library of Products in Search of Future Botulinum Neurotoxin A Inhibitors

Botulinum toxins are neurotoxins produced by Clostridium botulinum. This toxin can be lethal for humans as a cause of botulism; however, in small doses, the same toxin is used to treat different conditions. Even if the therapeutic doses are effective and safe, the adverse reactions could be local and could unmask a subclinical impairment of neuromuscular transmissions. There are not many cases of adverse events in the literature; however, it is possible that sometimes they do not occur as they are transient and, if they do occur, there is no possibility of a cure other than to wait for the pharmacological effect to end. Inhibition of botulinum neurotoxin type A (BoNT/A) effects is a strategy for treating botulism as it can provide an effective post-exposure remedy. In this paper, 13,592,287 compounds were screened through a pharmacophore filter, a 3D-QSAR model, and a virtual screening; then, the compounds with the best affinity were selected. Molecular dynamics simulation studies on the first four compounds predicted to be the most active were conducted to verify that the poses foreseen by the docking were stable. This approach allowed us to identify compounds with a calculated inhibitory activity in the range of 316–500 nM.