认识麻痹性贝类毒素

正织纹螺因含有麻痹性贝类毒素而被国家禁止食用。麻痹性贝类毒素是如何产生的?如何避免?既然称其为"贝类毒素",我们平时所吃的美味贝类都安全吗?麻痹性贝类毒素是一种生物毒素麻痹性贝类毒素并非来自贝类生物体本身,而是贝类摄食有毒藻类,并经其体内蓄积、放大、转化等过程形成的具有神经肌肉麻痹作用的赤潮生物毒素。人体若误食含有此类毒素的贝类     

连云港海州湾麻痹性贝类毒素中毒分析

为分析连云港地区麻痹性贝类毒素中毒特征,对连云港海州湾织纹螺进行形态鉴定,收集近年麻痹性贝类中毒资料,应用小白鼠生物测定法检测螺肉中麻痹性贝类毒素。结果连云港海州湾存有4种织纹螺,其中以半褶织纹螺含麻痹性贝类毒素量高(1.6×103μg100g螺肉),并导致连云港地区10起、银川1起麻痹性贝类中毒;1992年还发生一起由泥螺引起的麻痹性贝类中毒。中毒者均表现为下行性神经麻痹症状,重者致死。鉴于海洋污染的严重性,为保障海洋贝类的食用安全,应对贝类进行毒素等安全指标的监测。     

超高效液相色谱-串联质谱法测定闽东地区织纹螺中河豚毒素

建立了快速检测闽东地区织纹螺中河豚毒素含量的高效液相色谱-串联质谱方法。织纹螺均质样品用1%乙酸水溶液提取,0.5%乙酸甲醇溶液稀释,冷冻10min后离心除蛋白,亲水作用色谱柱分离,乙腈-0.1%甲酸水溶液作为流动相梯度洗脱,UPLC-MS/MS分析。基质匹配标准曲线外标法定量。方法的检出限为10μg/kg,定量限为25μg/kg,满足欧洲食品安全委员会对双壳贝类和腹足类中河豚毒素安全限量值44μg/kg的检测要求。在织纹螺中添加50、100、250μg/kg 河豚毒素进行加标回收试验,河豚毒素回收率为79.6%~118.1%,相对标准偏差为(RSD,n=6)7.7%~10.6%。采用该方法测定闽东地区14份织纹螺样品,最高检出河豚毒素 25718.2μg/kg。该研究方法准确性高、成本低、操作性强,能够满足当前快速定量检测织纹螺中河豚毒素含量的技术要求。     

南海海域养殖牡蛎中麻痹性贝类毒素分析

本研究在2013年3月采集湛江、阳江、钦州、北海、汕头、汕尾、深圳、台山等8个位于南海海域的养殖牡蛎,用小鼠生物(MBA)法和高效液相色谱(HPLC)法进行麻痹性贝类毒素检测。MBA毒性分析结果显示:8个区域养殖牡蛎的毒素含量为0MU/g-2.3MU/g,符合我国贝类食品安全限量要求;HPLC毒素成分分析结果显示:湛江、深圳、汕头、台山和阳江等5个海域的养殖牡蛎体内检测出痕量的麻痹性贝类毒素,湛江、深圳、汕头和阳江海域的养殖牡蛎中检出高毒性的STX和neo STX。本次麻痹性贝类毒素测试MBA法的检出率为25%,HPLC法的检出率为62.5%,HPLC法更适合于检测麻痹性贝类毒素含量低的样品。     

上海市售贝类产品中麻痹性贝类毒素污染状况调查及其评价

目的 调查上海市售贝类产品中麻痹性贝类毒素污染状况。方法 2010年8月~2011年7月间, 在上海水产品批发市场进行5种贝类样品采集, 每月抽取样品24份, 全年共288份。采用生物法(SC/T 3023-2004)对其进行了麻痹性贝类毒素的检测, 其中虾夷扇贝的肠腺和肌肉(扇贝柱)进行分开测定。结果 缢蛏、菲律宾蛤仔、牡蛎、文蛤、虾夷扇贝肠腺和肌肉中麻痹性贝类毒素的含量范围分别为ND~121.5 MU/100 g、ND~113.4 MU/100 g、ND~177.7 MU/100 g、ND~124.6 MU/100 g、261.7~3363.5 MU/100 g和ND。全年麻痹性贝类毒素的平均含量分别在98.5±10.5 MU/100 g、78.6±9.3 MU/100 g、50.4±10.1 MU/100 g、40.6±14.8 MU/100 g、1242.2±974.3 MU/100 g和0。按照目前我国贝类产品主要出口国家和国际组织对麻痹性贝类毒素的限量要求进行评价, 仅仅是虾夷扇贝肠腺中麻痹性贝类毒素超标, 超标率为98%, 因此在食用扇贝时应去除其肠腺; 而其余贝类产品中麻痹性贝类毒素均在限量规定范围内。结论 上海市售贝类产品对食用的安全性不产生负面影响。     

钦州湾7种经济贝类中麻痹性贝类毒素分析与安全评价

采用小鼠生物检测法和液相色谱-荧光检测法,分别对采自钦州湾的7种典型经济贝类中麻痹性贝类毒素(paralytic shellfish toxins,PSTs)组成成分与含量进行分析,同时参考我国渔政渔港监督管理局制订的贝类安全食用标准(400 MU/100 g或80μg/100 g STX_(eq))评价其食用安全性。结果如下,小鼠生物法分析表明小鼠在观察15 min内均不死亡,说明7种贝类中PSTs含量均小于400 MU/100 g或者不含有毒素;进一步对贝类样品进行液相色谱-荧光检测分析,表明7种贝类中可检测到麻痹性贝类毒素,毒素组成成分以高毒性的氨基甲酸酯类毒素为主如膝沟藻毒素4(gonyautoxin,GTX4)、膝沟藻毒素1(gonyautoxin,GTX1)、新石房哈毒素(neosaxitoxin,NEO)和石房蛤毒素(saxitxin,STX),其中异毛蚶中有最大毒素含量/毒性值,分别为0.27 nmol/g或13.1μg/100 g STX_(eq),低于安全标准80μg/100 g STX_(eq)。研究中有6种贝类可检测到毒素成分,检出率为86%。这说明,在钦州湾所采集的7种贝类中,尽管毒素含量低于食用安全标准规定的阈值,但多种贝类中仍可检测到麻痹性贝类毒素成分,因此其存在的安全性问题不容忽视,应加强该地区经济型贝类中麻痹性贝类毒素的监测,以防中毒事件的发生。     

东沙港沿岸贝类毒性监测与探讨

目的了解东沙港沿岸织纹螺体内带毒情况、消长规律及与其养殖在一起的鲜贝、毛贝、蛤蜊等经济贝类是否带毒,为经济贝类食用安全提供科学依据。方法用小鼠生物测试法测定织纹螺、鲜贝、毛贝、蛤蜊各8份样品的毒性。结果8份织纹螺普遍带毒,均大于400MU／100g,6月份毒性最强,达到1246MU／100g鲜肉重;24份共同养殖的其他经济贝类,毒素含量均小于175MU／100g。结论本调查结果显示应严格禁止食用该地区的织纹螺,与织纹螺养殖在一起的经济贝类没有带毒,食用安全。     

2018—2020年河北省市售贝类中麻痹性贝类毒素污染状况调查分析

目的了解2018—2020年河北省市售贝类中麻痹性贝类毒素(paralytic shellfish poison,PSP)污染状况。方法 2018年8月—2020年5月间,对河北省市售的7种双壳贝类,共508份进行检测分析。样品经0.5%乙酸水提取,石墨化碳黑固相萃取柱净化,采用高效液相色谱-串联质谱法进行检测。结果 508份样品,PSP阳性样品24份,检出率为4.7%, 15份样品超过世界卫生组织规定安全限量,超标率为3.0%。检出贝类为贻贝、毛蚶、杂色蛤、扇贝, PSP含量范围分别为217.0~13001.8μg石房蛤毒素当量(saxitoxin equivalent, STXeq/kg)、217.0~4893.2μg STXeq/kg、217.0~503.6μg STXeq/kg、217.0~11024.5μg STXeq/kg;超标贝类为贻贝、毛蚶、扇贝。贝类中检出的PSP类型有GTX1、GTX4、GTX2、GTX3、neoSTX、STX。结论河北省市售贝类麻痹性贝类毒素暴露风险整体较低,秦皇岛地区贻贝等贝类产品在4、5月份较易受到PSP污染,应持续关注,加强早期监测预警。     

硇洲岛海域珊瑚礁螃蟹的麻痹性贝类毒素分析

本研究于2013年3月～5月间,采用小鼠生物(MBA)法检测在湛江硇洲岛海域捕获的花纹爱洁蟹、雷诺鳞斑蟹、善泳蟳和细纹爱洁蟹等珊瑚礁螃蟹体内麻痹性贝类毒素(PSP)的毒性值。毒性分析结果表明:珊瑚礁螃蟹体内检测出较高浓度的麻痹性贝类毒素;螃蟹的毒性大小为:花纹爱洁蟹细纹爱洁蟹善泳蟳雷诺鳞斑蟹;63.6%的珊瑚礁螃蟹毒性值超过了食用安全限量(4MU/g)要求,螃蟹的毒性值为2.69～54.8MU/g。采用高效液相色谱(HPLC)法检测花纹爱洁蟹和雷诺鳞斑蟹的毒素组成分析结果显示,蟹体内的PSP主要是低毒性的C_2毒素,以及少量的GTX_1毒素。推测硇洲岛海域分布有产毒的甲藻,该海域海产品有受到麻痹性贝类毒素污染的风险。     

基质分散固相萃取净化-亲水液相色谱-串联质谱法检测织纹螺与贝类中河鲀毒素

目的针对海产品常见中毒原因分析需求,建立基质分散固相萃取净化-亲水液相色谱-串联三重四极杆质谱(HILIC-MS/MS)快速定性定量检测织纹螺和贝类中河鲀毒素的新方法。方法 1.0 g样品经0.1%乙酸溶液沸水浴提取后,用50 mg亲水亲油平衡填料(HLB)和5 mg石墨化碳黑(GCB)吸附剂吸附净化,最后经乙腈蛋白沉淀后过0.22μm聚四氟乙烯(PTFE)滤膜,亲水液相色谱柱(150 mm×2.0 mm,3μm)分离,电喷雾离子化,选择反应监测(SRM)模式检测,基质匹配外标法定量。结果在2.0~40.0 ng/ml浓度范围内,河鲀毒素呈现良好的线性关系,相关系数r~2≥0.999;以3倍基线噪声所对应的浓度为检出限时,河鲀毒素的方法检出限可达10.0μg/kg;在25、100和200μg/kg的加标水平时,方法回收率为74.2%~87.9%,相对标准偏差为2.3%~9.1%。应用本方法对浙江沿海地区市售织纹螺和贝类样品进行检测,15份织纹螺中有14份检出河鲀毒素,检出率为93.3%,含量范围为0.04~15.75 mg/kg,60份贝类样品均未检出河鲀毒素。结论该检测方法准确、快速、易操作,能满足典型海产品中河鲀毒素的公共卫生应急检测或日常监测要求。     

秦皇岛海域食用贝类重金属污染情况分析

为了解秦皇岛海域食用贝类中重金属的污染水平及其存在的健康风险。采用电感耦合等离子体发射质谱法(ICP-MS)和原子荧光光谱法测定了该地区15种常见贝类肌肉和内脏中As(无机砷和总砷)、Pb(铅)、Cr(铬)、Cd(镉)、Hg(甲基汞)的含量,并采用污染指数法、系统聚类法评价了各重金属的污染特征和分类。结果表明:海虹(贻贝,Mytilus edulis)、栉孔贝(Chlamys farreri)、生蚝(Ostrea gigas thunberg)和毛蚶子(Scapharca subcrenata)等的重金属综合污染(MPI)较高。根据NY 5073-2006 无公害食品水产品中有毒有害物质限量标准进行重金属评价。秦皇岛海域食用贝类六种重金属含量最高为Cd,均值为1.129 mg/kg,超出NY 5073-2006 无公害食品水产品中有毒有害物质限量标准,污染较严重,总As和无机As均值分别为0.161、0.021 mg/kg。Cr和Hg均值分别为0.107和0.013 mg/kg,均未受污染,Pb均值为0.188 mg/kg,污染较轻。但部分贝类Cd污染比较严重。其中内脏污染比肌肉部分严重,尤其是大小海螺(Busycon canaliculatu)。因此,需进一步研究秦皇岛海域可食用贝类中Pb和Cd的污染来源并进行有效防治。     

Identification of tetrodotoxin in a marine gastropod (Nassarius glans) responsible for human morbidity and mortality in Taiwan

The toxicity of the gastropod Nassarius glans was investigated. This gastropod was implicated in an incident of food paralytic poisoning on Tungsa Island, Taiwan, in April 2004. Six victims consumed both digestive glands and muscle. These tissues contained high concentrations of toxin; their highest toxicity scores were 2,048 and 2,992 MU/g, respectively, based on the tetrodotoxin (TTX) bioassay. The toxin was purified from these gastropods and analyzed by high-performance liquid chromatography, which revealed TTX and related compounds 4-epi TTX and anhydro-TTX; paralytic shellfish poisons were not found. The urine and blood samples from patients were cleansed using a C18 Sep-Pak cartridge column and 3,000 molecular weight cutoff Ultrafree microcentrifuge filters, and the eluate was filtered and analyzed by liquid chromatography and mass spectrometry. The detection limit for TTX was 1 ng/ml. The standard curves were linear in the range 30 to 600 ng/ml for urine and 1 to 30 ng/ml for blood. TTX was detected in all urine samples but in only three of four blood samples tested. Thus, the causative agent of gastropod food poisoning was identified as TTX.     

Monitoring phytoplankton and marine biotoxins in production waters of the Netherlands: results after one decade

Shellfish products may be contaminated with marine biotoxins which, after consumption, may lead to human illness. The Netherlands has a regular monitoring programme for marine biotoxins and the possible toxic phytoplankton in shellfish production waters. The aim of the current study was to evaluate the presence of potential toxic phytoplankton species and marine biotoxins in Dutch production waters over the last decade, and to analyse the relationship between toxin levels and abundance of possible causative phytoplankton species. The results of the monitoring programme of the period 1999-2009 were used. The presence of Alexandrium spp. were negligible, but Pseudo-nitzschia spp. and phytoplankton causing diarrhetic shellfish poisoning (DSP toxin-producing phytoplankton) were present in nearly all three main production areas and years. The main DSP toxin-producing species was Dinophysis acuminata followed by D. rotundata and Prorocentrum lima. Toxins causing paralytic shellfish poisoning (PSP) and amnesic shellfish poisoning (ASP) were present in only a few individual shellfish samples, all at low levels. At the end of 2002, an episode of DSP toxicity was recorded, based on the rat bioassay results. Of the samples that were chemically analysed for DSP toxins in 2007 and 2008, about half of the samples in 2007 contained these toxins, although levels were low and no positive results were obtained using the rat bioassay. There was a slight positive correlation between concentrations of DSP toxin-producing phytoplankton and levels of DSP toxins in 2007. Increased DSP toxin levels were found up to 5 weeks after the peak in DSP toxin-producing phytoplankton. This positive, but weak, relationship needs to be confirmed in future research using more samples and chemical methods to quantify the presence of DSP toxins. If this relationship is further substantiated and quantified, it could be used within the current monitoring programme in the Netherlands to predict the risk areas regarding DSP toxicity in shellfish.     

Tetrodotoxin-associated snail poisoning in Zhoushan: a 25-year retrospective analysis

Outbreaks of paralytic snail poisoning have recently occurred in Asia, especially in China. The epidemiological characteristics of this disease from an outbreak in Zhoushan City, China, were recorded. Forty-two outbreaks of paralytic snail poisoning, involving 309 cases of illness, occurred from 1977 to 2001. Sixteen people (5.2%) died, 48 people (15.5%) required intubations, and 140 people (45.3%) required emergency hospital treatment as a result of these outbreaks. Outbreaks involved multiple marine snail species and occurred primarily during the summer (from June to August) on 11 islands with high population densities. Peak numbers of outbreaks and amounts of snail toxicity occurred from 1978 to 1979, from 1985 to 1987, and from 1992 to 1994. Toxicity varied depending on specimen, region, and season. The toxin involved was identified as tetrodotoxin. The data obtained in this study suggest that snails should not be eaten unless they are certified to be nontoxic.     

The Gastropods Possessing TTX and/or PSP

Paralytic gastropod poisoning incidents have frequently occurred in the world. In the outbreaks, the symptoms of victims exhibited quite different patterns depending on the specific outbreak and most of all showed parasthesis with rare fatal cases. The toxin identified was mainly tetrodotoxin (TTX), sometimes with minor paralytic shellfish poison (PSP) and other toxins. Toxic gastropods included Family Nassariidae, Naticidae, Olividae, Muricidae, Buccinidae, Ranellidae, Harpidae, Trochidae, Turbinidae, Burdidae, and Melongenidae. The sources of toxins are from bacteria, dinoflagellate, or biosynthesis. The physiological function of toxin in toxic gastropods acts as defensive and/or attacking agent. The more toxic gastropod has higher preference or palatability preference to TTX and/or PSP.     

Occurrence of tetrodotoxin and paralytic shellfish poisons in a gastropod implicated in food poisoning in southern Taiwan

The toxicity of the gastropod Nassarius papillosus implicated in a food paralytic poisoning incident in Liuchiu Island, Taiwan, in October 2005 is reported. The symptoms of a victim (67 years old) were featured by general paresthesia, paralysis of phalanges and extremities, paralysis, coma, and aphasia. The remaining specimens of shell were assayed for toxicity. The range of specimen toxicity was found to be 63-474 mouse units (MU) per specimen for N. papillosus by a tetrodotoxin (TTX) bioassay. The mean (SD) toxicity of the digestive gland and other portions were 296 ± 120 and 382 ± 156 MU in N. papillosus. The toxin was partially purified from the acidic methanol extract of the gastropod by using a C18 solid-phase extraction column. The eluate was then filtered through a 3000 MW cut-off ultrafree microcentrifuge filter. It was shown that the toxin purified from gastropods analysed by high-performance liquid chromatography and liquid chromatography/mass spectrometry contained TTX 42-60 µg g^-1 (about 90%), whereas along with minor paralytic shellfish poisons (PSP) it was 3-6 µg g^-1 (about 10%).     

Toxin composition and toxicity dynamics of marine gastropod Nassarius spp. collected from Lianyungang, China

Consumption of nassariid gastropods often leads to poisoning incidents in some coastal provinces in China. To elucidate the pattern of toxicity dynamics and origin of toxins, samples of gastropod Nassarius spp. were collected from late May to early August 2007 from Lianyungang, Jiangsu province, where the poisoning incidents have been frequently reported. Toxicity was first screened with the mouse bioassay method, and tetrodotoxin and its analogues (TTXs) were analysed with high-performance liquid chromatography coupled with an ion-trap mass spectrometer (HPLC-MS(n)). The toxicity of nassariid N. semiplicatus showed an 'M'-shaped pattern of fluctuation during the sampling season. Two peaks of toxicity appeared in late May and late July. The maximum toxicity was recorded on 24 May, with the value of 846 mouse unit (MU) g(-1) of tissue (wet weight). TTX and its analogues trideoxyTTX, 4-epiTTX, anhydroTTX and oxoTTX were detected in the nassariid samples. TrideoxyTTX but not TTX was the major toxin in all the samples. No paralytic shellfish poison (PSP) was detected in the sample with the maximum toxicity by HPLC-FLD analysis. Variation of TTX content in the tissue of nassariid gastropods correlates well with the dynamics of toxicity. It is suggested that TTXs are the major toxins corresponding to the toxicity of the nassariids, and May and July are the high-risk seasons for consumption of nassariids, which is critical for the management of poisoning incidents.     

Tetrodotoxin-binding proteins isolated from five species of toxic gastropods

Toxic gastropods Polinices didamy, Natica lineata, Oliva miniacea, O. mustelina and O. hirasei are tetrodotoxin (TTX) bearing animals, which accumulate TTX in their muscle and digestive gland. Analysis by gel filtration on Sepharose CL-6B revealed that 0.05 M NaCl extracts of the muscle of five species of toxic gastropods contained TTX-binding high molecular weight substances (HMWS) (1500–2000 kDa). The TTX-binding capacities of those HMWS were 0.12, 0.62, 0.45, 0.28 and 0.35 MU/mg protein, respectively, but those HMWS had no neutralising effect on TTX or paralytic shellfish poison. The HMWS of the five toxic gastropods could be hydrolyzed with HCl and protease at 37 °C, pH 7.4, but not with ribonuclease T2, deoxyribonuclease I or α-amylase. After purifying the TTX-binding protein of N. lineata by Q Fast-Flow strong anion exchanger and then BioSep-SEC-S 2000, the TTX-binding capacity increased to 3.5 MU/mg and 4.2 MU/mg protein, respectively. The TTX-binding capacity of N. lineata HMWS had no obvious seasonal variation. The molecular weight of TTX-binding protein of N. lineata was estimated to be about 434 kDa, while it comprised two subunits with molecular weights of about 272 kDa and 47 kDa, respectively, under SDS–PAGE.     

海产品中麻痹性贝类毒素快速检测技术研究进展

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