Analgesic Effects of Tetrodotoxin Combined with Acetylsalicylic Acid on Acetic Acid-induced Abdominal Constriction and Formalin Test in Mice

Aim To study the effects of tetrodotoxin （TTX） combined with acetylsalicylic acid （ASA） on nociceptive stimulus in mice. Methods To assess the antinociceptive effects of TTX, ASA or TTX plus ASA, the acetic acid-induced abdominal constriction test and formalin pain test were used. Results TTX （0.5 - 4.0 μg· kg^-1 ） or ASA （25 - 200 mg· kg^-1 ） im produced a significant inhibition of acetic acid-induced abdominal constriction. The median inhibitory doses （ID508） were 2.1 μg· kg^-1 for TTX（ and 64 mg· kg^-1 for ASA. TTX and ASA also showed a dose-dependent inhibition of the second phase response in the formalin pain model, the ID508, being 2.3μg·kg^-1 and 74.2 mg· kg^-1, respectively. The ihteraction between TTX and ASA was synergistic, as evidenced by the fact that （1） when ASA alone compared with the combination of TTX （0.79 μg · kg^-1 or 0.39μg· kg^-1 ） and ASA, the ID508, of ASA reduced from 64.0 mg· kg^-1 to 5.8 mg· kg^-1 or 12.6 mg· kg^-1, and from 74.2 mg· kg^-1 to 7.4 mg· kg^-1 or 13.0 mg· kg^-1 on tile two models of nociceptive tests, respectively; and that （2） synergism in the analgesic effects was shown by isobiolographic analysis. Conclusion TTX, ASA and the combination of the two drags produce analgesic effects in acetic acid-induced abdominal constriction test and formalin-induced pain test. The interactions between TTX and ASA may be useful in developing novel analgesic agents.

Simultaneous identification and quantification of tetrodotoxin in fresh pufferfish and pufferfish-based products using immunoaffinity columns and liquid chromatography/quadrupole-linear ion trap mass spectrometry

In this study, we established a comprehensive method for simultaneous identification and quantification of tetrodotoxin （TTX） in fresh pufferfish tissues and pufferfish-based products using liquid chromatography/quadrupole-linear ion trap mass spectrometry （LC-QqLIT-MS）. TTX was extracted by 1% acetic acid-methanol, and most of the lipids were then removed by freezing lipid precipitation, followed by purification and concentration using immunoaffinity columns （IACs）. Matrix effects were substantially reduced due to the high specificity of the IACs, and thus, background interference was avoided. Quantitation analysis was therefore performed using an external calibration curve with standards prepared in mobile phase. The method was evaluated by fortifying samples at 1, 10, and 100 ng/g, respectively, and the recoveries ranged from 75.8%--107%, with a relative standard deviation of less than 15%. The TTX calibration curves were linear over the range of 1-1 000 ~tg/L, with a detection limit of 0.3 ng/g and a quantification limit of 1 ng/g. Using this method, samples can be further analyzed using an information- dependent acquisition （IDA） experiment, in the positive mode, from a single liquid chromatography-tandem mass spectrometry injection, which can provide an extra level of confirmation by matching the full product ion spectra acquired for a standard sample with those from an enhanced product ion （EPI） library. The scheduled multiple reaction monitoring method enabled TTX to be screened for, and TTX was positively identified using the IDA and EPI spectra. This method was successfully applied to analyze a total of 206 samples of fresh pufferfish tissues and pufferfish-based products. The results from this study show that the proposed method can be used to quantify and identify TTX in a single run with excellent sensitivity and reproducibility, and is suitable for the analysis of complex matrix pufferfish samples.

Effects of continuous peripheral nerve block by tetrodotoxin on growth associated protein-43 expression during neuropathic pain development

BACKGROUND： Peripheral nerve injury may lead to neuropathic pain and cause a markedly increase expression of growth associated protein-43 （GAP-43） in the spinal cord and dorsal root ganglion, local anesthetics blocking electrical impulse propagation of nerve fibers may also affect the expression of GAP-43 in the spinal cord and dorsal root ganglion. OBJECTIVE： To determine the effects of continuous peripheral nerve block by tetrodotoxin before and after nerve injury on GAP-43 expression in the dorsal root ganglion during the development of neuropathic pain. DESIGN： A randomized controlled animal experiment. SETTINGS： Department of Anesthesiology, the Second Hospital of Xiamen City; Department of Anesthesiology, the Second Affiliated Hospital of Shantou University Medical College. MATERIALS： Thirty-five Spragne Dawley （SD） rats, weighing 200 - 250 g, were randomly divided into four groups： control group （n =5）, simple sciatic nerve transection group （n =10）, peripheral nerve block before and after sciatic nerve transection groups （n =10）. All the sciatic nerve transection groups were divided into two subgroups according to the different postoperative survival periods： 3 and 7 days （n =5） respectively. Mouse anti-GAP-43 monoclonal antibody （Sigma Co., Ltd.）, supervision TM anti-mouse reagent （HRP, Changdao antibody diagnosis reagent Co., Ltd., Shanghai）, and HMIAS-100 image analysis system （Qianping Image Engineering Company, Tongji Medical University） were employed in this study. METHODS： This experiment was carried out in the Department of Surgery and Pathological Laboratory, the Second Affiliated Hospital of Shantou University Medical College from April 2005 to April 2006. ①The animals were anesthetized and the right sciatic nerve was exposed and transected at 1 cm distal to sciatic notch. ② Tetrodotoxin 10 μg/kg was injected percutaneously between the greater trochanter and the posterior superior iliac spine of fight hind limb to block the sciatic nerve proximally at 1 hour before or 4 hours after nerve injury respectively, the injection was repeated in all the rats every 12 hours.③ At 3 or 7 days after nerve injury, immunohistochemistry and image analysis were used to evaluate the expression of GAP-43 in the dorsal root ganglions of L5 to the transected sciatic nerve, and quantitative analysis was also performed. ④ Statistical analysis was performed using one way analysis of variance followed by t test. MAIN OUTCOME MEASURE： Expression of GAP-43 in the fight dorsal root ganglions of L5. RESULTS： All the 35 SD rats were involved in the final analysis of results. In normal rats, there were very low expressions of GAP-43 in the dorsal root ganglions. In simple sciatic nerve transection rats 3 and 7 days after sciatic nerve transection, the average absorbance value of GAP-43 immunopositive neurons were significantly different from that in normal rats （t =8.806, 6.771, P 〈 0.01）. Whereas 3 and 7 days after sciatic nerve transection in rats with peripheral nerve block before and after nerve injury, the average absorbance value of GAP-43 immunopositive neurons were not significantly different from that in normal rats （P 〉 0.05）. CONCLUSION： Local anesthetic continuous peripheral nerve block before or after nerve injury can suppress nerve injury induced high expression of GAP-43 during the development of neuropathic pain.

The development and optimization of ELISA for the determination of tetrodotoxin

Objective： To optimize the ELISA for the determination of tetrodotoxin. Methods： A competitive enzyme-linked immunosorbent assay （ELISA） was used. In the ELISA, 100 μl antigen （1. 0 μg/ml） was coated on the microtiter plate for 60 min at 37 C or over night at 4 C. The plate was then washed 3 times with PBS-T for 3-5 s each time. The optimal incubation time for monoclonal antibody （mAb）, goat anti-mice IgG peroxidase conjugate and OPD were 30 min, 20 min and 10 min at 37 C, re- spectively. Results.. The detection limit is 0. 05 ng in each well. The curve was linear for TTX doses be- tween 5-5 000 ng/ml （0. 25-250 ng for every assay）. The linear regress equation was Y = 0. 30 88X-0.17 41 （R=0.99 01）. The average callback for TTX of muscles and gonads were 99.74% and 100.30%, respectively. The sensitivity of optimization ELISA was 5 times than traditional method and the time of 1.8 h were saved. Conclusion： The optimized ELISA is an ideal method for the determination of tetrodotoxin.

特异性Tetrodotoxin亲和吸附微珠的制备及其提取性能

将横纹东方豚卵巢与醋酸乙醇溶液混合匀浆,经离心,加热,沉淀,脱脂等处理,制得粗提液,活性炭吸附粗提液得到粗毒液;以壳聚糖为载体合成亲和微珠分别来分离纯化粗提液和粗毒液,用传统的小鼠生物法检测TTX毒素量.潮湿活性炭对TTX的吸附量约为2μg/g,潮湿亲和微珠吸附量约为10μg/g;亲和微珠吸附提取粗毒液,回收率达51%,直接吸附粗提液,回收率达到80%,同时蛋白也得到有效的去除.

Effects of tetrodotoxin monoclonal antibody on the blocking action of tetrodotoxin on sodium channels

The effect of tetrodotoxin(TTX) monoclonal antibody (McAb) 8A5 on the blocking action of TTX on sodium channels was studied by using the electrophysiological technique of whole cell recording.We found the specific TTX McAb have the following characterizations: TTX sensitivity to NG108-15 cell was high , with sodium ion current of NG108-15 cell completely blocked by only 10-6 mol/L level of TTX; when the cell was treated with TTX McAb 8A5 for 1 min and 5min, after the sodium current was completely abolished by TTX, the sodium ion current was restored to 79. 44%?. 20% and 73. 89%?. 74% (n=5) of the control values respectively; when the cell was treated for 1 min with 8A5 and TTX which had been mixed for 1 h before added,the sodium ion current was maintained at 89. 21%?. 41% (n=4) of the control. These results indicated that TTX-induced blockage on the sodium ion current could be powerfully antagonized by TTX McAb 8A5 with two distinct administering ways.

Effects of dragon’s blood resin and its component loureirin B on tetrodotoxin-sensitive voltage-gated sodium currents in rat dorsal root ganglion neurons

Using whole-cell patch clamp technique on the membrane of freshly isolated dorsal root ganglion (DRG) neurons, the effects of dragons blood resin and its important component loureirin B on tetrodotoxin-sensitive (TTX-S) voltage-gated sodium currents were observed. The results show that both blood resin and loureirin B could suppress TTX-S voltage-gated sodium currents in a dose-dependent way. The peak current amplitudes and the steady-state activation and inactivation curves are also made to shift by 0.05% blood resin and 0.2 mmol/L loureirin B. These results demonstrate that the effects of blood resin on TTX-S sodium current may contrib-ute to loureirin B in blood resin. Perhaps the analgesic effect of blood resin is caused partly by loureirin B directly interfering with the nociceptive transmission of primary sensory neurons.

Modulation of dragon's blood on tetrodotoxin-resistant sodium currents in dorsal root ganglion neurons and identification of its material basis for efficacy

To clarify the modulation of dragon's blood on the tetrodotoxin-resistant (TTX-R) sodium currents in dorsal root ganglion (DRG) neurons and explore its corresponding material basis for the efficacy, using whole-cell patch clamp technique, the effects of dragon's blood and the combined effects of three components (cochinchinenin A, cochinchinenin B, and loureirin B) extracted from dragon's blood on the TTX-R sodium currents in acute-isolated DRG neurons of rats were observed. According to the operational definition of material basis for the efficacy of TCM established, the material basis of the modulation on the TTX-R sodium currents in DRG neurons of dragon's blood was judged from the experimental results. The drug interaction equation of Greco et al. was used to assess the interaction of the three components extracted from dragon's blood. This investigation demonstrated that dragon's blood suppressed the peak TTX-R sodium currents in a dose-dependent way and affected the activations of TTX-R sodium currents. The effects of the combination of cochinchinenin A, cochinchinenin B, and loureirin B were in good agreement with those of dragon's blood. Although the three components used alone could modulate TTX-R sodium currents, the concentrations of the three components used alone were respectively higher than those used in combination when the inhibition rates on the TTX-R sodium currents of them used alone and in combination were the same. The combined effects of the three components were synergistic. These results suggested that the interference with pain messages caused by the modulation of dragon's blood on TTX-R sodium currents in DRG neurons may explain some of the analgesic effect of dragon's blood and the corresponding material basis for the efficacy is the combination of cochinchinenin A, cochinchinenin B, and loureirin B.

2022年浙江省沿海海水贝类中河豚毒素膳食暴露及健康风险评价

目的了解2022年浙江省沿海城市野生、养殖贝类中河豚毒素暴露情况。方法采用分层随机抽样的方法,液相色谱-串联质谱法对样品中河豚毒素测定,结合《中国居民营养与慢性病状况报告(2015年)》和《2014年国民体质监测公报》对居民膳食摄入量调查。采用世界卫生组织/联合国粮食及农业组织推荐的《食品中化学物质膳食暴露评估》方法点评估法,对浙江省几个沿海城市中居民膳食中河豚毒素水平评估。结果贝类中河豚毒素含量差异较大,主要集中在彩虹明樱蛤和织纹螺,检出率为14.7%,含量最高达到5220.00μg/kg,河豚毒素的平均含量167.30μg/kg。一次性摄入织纹螺时,小于10岁的儿童低于15 g,10~20岁青少年低于40 g,20岁以上成年人低于60 g的时,其急性膳食暴露水平处于可接受的安全状态;摄入其他贝类的急性风险指数远小于100,人群暴露水平处于安全状态。结论贝类整体急性膳食暴露水平在安全状态,织纹螺河豚毒素含量极高,食用织纹螺极不安全,应加以监管。

超高效液相色谱−串联质谱法测定织纹螺中河豚毒素的方法优化

【背景】近年来,因食用织纹螺引发的河豚毒素(TTX)中毒事件频发,然而现行国家标准(GB 5009.206-2016)中的液相色谱−串联质谱法(LC-MS/MS)仅适用于河鲀中TTX含量的检测,因此亟需建立一种适用于检测织纹螺中TTX含量的高效、准确的方法。【目的】基于超高效液相色谱−串联质谱法(UPLC-MS/MS)建立一种高效且能准确测定织纹螺中TTX含量的方法。【方法】通过比较不同前处理方法(提取溶液、提取方式、冷冻时间)下TTX回收率的大小,以及不同色谱条件(流动相、进样量)对色谱峰的影响,获得最佳实验条件。此外,采用优化后的方法和现行国家标准方法对采自福建省莆田市和宁德市的织纹螺样品进行测定,以比较该检测方法的准确性和可靠性。【结果】最佳前处理条件:样品首先采用0.1%乙酸溶液60℃超声提取20 min,随后使用80%(体积分数)0.1%乙酸甲醇溶液稀释并在−18℃下冷冻30 min后离心,取上清液过膜后,上机测定;UPLC-MS/MS条件:以含5 mmol·L^(−1)乙酸铵的0.1%甲酸溶液和乙腈为流动相,经Amide-80色谱柱梯度洗脱后,采用电喷雾正离子选择反应监测模式,以基质标准曲线外标法定量。TTX在织纹螺中的基质效应为68.9%,在1~100 ng·mL^(−1)基质标准曲线范围内线性关系良好,线性回归方程相关系数R^(2)>0.99。本方法的检出限和定量限分别为10、25μg·kg^(−1),能同时满足中国和欧洲食品安全委员会对水产品中TTX安全限量值(分别为2200、44μg·kg^(−1))的要求。此外,在3个TTX加标浓度(50、500、4000μg·kg^(−1))下,平均回收率为74.8%~104%,相对标准偏差(RSD)均小于15%(n=6)。分别采用优化后的实验方法和现行国家标准方法对9份织纹螺样品进行检测,2种方法检测的TTX含量范围分别为<0.0250~22.8、0.0120~23.5 mg·kg^(−1),相对偏差为1.51%~14.6%,表明本检测方法准确可靠。【结论】本研究优化建立了一种适用于测定织纹螺中TTX含量的UPLC-MS/MS法,具有前处理简单、快速准确和低成本等特点,为实现织纹螺中TTX含量的快速批量检测奠定良好基础。

Inhibitive effect of loureirin B plus capsaicin on tetrodotoxin-resistant sodium channel

OBJECTIVE: To investigate whether the effect of loureirin B plus capsaicin on tetrodotoxin-resistant(TTX-R) sodium channel.METHODS: By using whole-cell patch-clamp recordings, in acutely isolated dorsal root ganglion(DRG) neurons, the combined effects of loureirin B and capsaicin on TTX-R sodium channel were observed. Based on the data, the interaction between loureirin B and capsaicin in their modulation on TTX-R sodium channel was assessed.RESULTS: Loureirin B could not induce transient inward TRPV1 current. Capsazepine, a transient receptor potential vanilloid l(TRPV1) antagonist, could not attenuate the block of 0.64 mmol/L loureirin B on TTX-R sodium channel. There was no significant difference(P > 0.05) between IC_(50) of loureirin B(0.37 mmol/L) on TTX-R sodium channel in capsaicin-sensitive DRG neurons and that(0.38 mmol/L)in capsaicin-insensitive DRG neurons. However,there was a significant difference(P < 0.05) between the IC_(50) of capsaicin(0.28 μmol/L) on TTX-R sodium channel in capsaicin-sensitive DRG neurons and that(52.24 μmol/L) in capsaicin-insensitive DRG neurons. Four combinations composed of various concentrations of loureirin B and capsaicin could all inhibit TTX-R sodium currents but have different interactions between loureirin B and capsaicin.CONCLUSION: Loureirin B plus capsaicin could produce double blockage on TRPV1 and modulation on TTX-R sodium channel. The action of loureirin B onTTX-R sodium channel was independent ofTRPV1 but similar with that of capsaicin on TTX-R sodium channel in capsaicin-insensitive DRG neurons.

Adsorption of tetrodotoxin by flexible shape-memory polymers synthesized from silica-stabilized Pickering high internal phase emulsion

Flexible shape-memory polymers were synthesized by Pickering high internal phase emulsion(HIPE)polymerization and used to adsorb and separate tetrodotoxin(TTX)from an aqueous solution.SiO_(2)nanoparticles were used to stabilize the Pickering oil-in-water(O/W)HIPEs.We introduced imidazolium-modified bromobutyl rubber(IBR)with excellent mechanical properties and high viscosity into the emulsion system as the shape-memory monomer.The properties,such as shape memory and morphology,were characterized by various methods,and batches of static adsorption experiments were conducted to analyze the adsorption performance of SiO_(2)@IBR on TTX.The characterization revealed that the SiO_(2)@IBR had a porous structure and good shape memory.Thus,the combination of SiO_(2)particles and IBR prevented shedding of SiO_(2)and enhanced the mechanical and adsorption properties of SiO_(2)@IBR.The results of the adsorption experiments indicated that the SiO_(2)@IBR had good adsorption of TTX.Both the Langmuir and Freundlich models fitted the isothermal adsorption experiment process.The TTX adsorption capacity of SiO_(2)@IBR was about 290.44 mg/g at 308 K.The fitting results of the pseudo-first-order and pseudosecond-order kinetic models showed that the adsorption process involved both chemical bonding and physical adsorption.After 10 adsorption and desorption experiments,the adsorption capacity of SiO_(2)@IBR decreased less than 0.03%,indicating that it had good adsorption and regeneration performance.

Rapid Determination of Tetrodotoxin in Human Plasma by Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry

A sensitive analytical method was developed to determine tetrodotoxin（TTX） in human plasma samples using protein precipitation, followed by ultra performance liquid chromatography（UPLC） analysis coupled with tandem mass spectrometry（MS/MS） using 11-deoxytetrodotoxin（11-deoxyTTX） as an internal standard. The plasma samples were prepared using protein precipitation prior to being analyzed by UPLC-MSfMS to identify TTX over a zwitterionic-hydrophilic interaction liquid chromatography column. The retention time values of TTX and 11-deoxyTTX were 4.12 and 3.67 min, respectively. TTX and 11-deoxyTTX were monitored and quantitated on the basis of their ion transitions for their respective precursor ions to their product ions（i.e., m/z 320.0→162. l for TTX and m/z 304.0→176.0 for 11-deoxyTTX） in the multiple reaction-monitoring mode. The lower limit of quantification of this method was determined to be 0.0199 ug/mL. This method showed good linearity for plasma samples that contained TTX concentrations in the range of 0.0199--1.99 ng/mL. The specificity, precision, accuracy, matrix effect, and stability characteristics of this method were also examined. The intra-assay precision and accuracy ranged from 1.89% to 6.00% and from 92.21% to 100.00%, whereas the inter-assay precision and accuracy ranged from 0.64% to 7.75% and from 99.38% to 101.26%, respectively. This new method therefore represents a rapid, accurate, reliable, and highly sensitive method for the qualitative and quantitative analyses of a trace amount of TTX in human plasma samples.

产河豚毒素微生物的研究进展

【背景】河豚毒素(TTX)是已知毒性较强的海洋毒素之一,广泛分布于河鲀等海洋生物体内。作为典型的钠离子通道阻断剂,TTX在镇痛、戒毒和抗心律失常等医疗领域展现出巨大的应用价值。然而,因获取困难和产量低下,TTX在医药领域的应用受到制约。因此,利用微生物发酵生产TTX成为了一个备受关注的研究方向。【目的】了解TTX的概况、产TTX微生物的分布和分类学多样性、TTX的发酵生产研究及TTX在微生物和环境中的迁移机制。【进展】目前,学界对TTX的理化性质已有较为清晰的认识,但其来源仍无明确定论。自1986年以来,科研人员陆续从河鲀、蓝环章鱼(Octopus maculosus)等生物及其栖息环境中分离出约150株产TTX菌株,其中以弧菌属、芽胞杆菌属为主,这些发现不仅为TTX的微生物起源说提供了强有力的理论支撑,也为后续利用微生物发酵生产TTX的研究奠定了坚实的基础。许多学者对TTX菌株发酵产毒中关键影响因素以及TTX迁移机制进行了探究,并取得了一定的成果。【展望】未来研究可重点考虑定向改造产TTX菌株、完善菌株大规模培养条件、探索影响菌株产TTX能力的关键因素以及分析微生物群落的相互作用对其合成TTX的影响,从而为TTX生物合成机制的阐明提供借鉴。【意义】通过对产TTX微生物的综述,阐明TTX的生物合成途径,为实现TTX规模化生产,解决TTX获取困难、成本高的难题,推动其在医药领域的应用提供理论参考。

LC-MS-MS测定血浆中河豚毒素

河豚毒素(tetrodotoxin,TTX)是一种小分子非蛋白类海洋生物毒素,具有极好的镇痛作用,临床用于戒毒治疗.由于TTX药效强而毒性大(肌注:LD50=19μg/kg),给药剂量很小,注射液每次给药仅为30μg,因此对检测方法的要求很高[1].本文初步建立了测定人血浆中TTX的LC-MS-MS方法,最低检测浓度为1 ng/mL,灵敏度比文献报道的方法提高了10倍[2].……

HPLC-柱后衍生荧光法测定水产品中河豚毒素含量结果的不确定度评定

通过建立HPLC-柱后衍生荧光法测定水产品中河豚毒素含量的不确定度评定方法,量化分析测定结果的不确定度分量,并对各分量相对贡献进行比较。结果表明:测定结果的不确定度主要来源于标准品的校准过程、样品称量、实验过程移取及定容、样品重复实验误差、前处理过程中的回收率不同等。其中校准不确定度中标准溶液配制的不确定度贡献最多,称样量不确定度最小;当水产品中河豚毒素的含量为90.6μg/kg时,其扩展不确定度为0.69μg/kg(包含因子k=2)。

抗河豚毒素单克隆抗体的制备及其特性的初步研究

将用合成的匙孔形虫戚血蓝蛋白－甲醛－河豚毒素（ＫＬＨ－ＨＣＨＯ－ＴＴＸ）连接物免疫的ＢＡＬＢ／ｃ小鼠的脾细胞与小鼠骨髓瘤细胞系Ｓｐ２／０融合，经３～４次亚克隆，建立了３个稳定分泌抗河豚毒素的杂交瘤细胞株，分别命名为１Ｇ３、４Ｇ３、６Ｄ９。其中１Ｇ３和４Ｇ３分属ＩｇＧ２ｂ亚类，６Ｄ９属ＩｇＧ１亚类。腹水的抗体稀释度为１∶１０５～１：５×１０６。用竞争抑制性酶联免疫吸附试验（ＥＬＩＳＡ）测定三种抗体的最低反应浓度为１×１０－３ｍｇ／Ｌ。

河豚毒素对小鼠和家兔的毒性研究

为研究抗河豚毒素 (TTX)的实验疫苗。测得昆明小鼠经ip ,sc ,ig三种TTX中毒途径的LD50 分别为 1 0 7、1 2 5、532 μg kg ;LD99分别为 1 2 5、1 4 6、62 2 μg kg。毒性反应具性别差异 ,♂性比♀性更敏感 ;无明显蓄积毒性。报告了大耳白家兔经im、ivTTX中毒毒性 ,最小致死剂量 (MLD)分别为 5 3、3 1 μg kg ;全死剂量(LD)分别为 5 8、3 8μg kg。描述了两种动物及各种途径中毒的症状特征。小鼠毒性剂量反应关系曲线均非常陡峭 ,家兔的MLD与LD非常接近 ,提示在致死量中毒时免疫抗毒较易实现 ,而在高倍致死量时要困难得多。TTX经口服中毒死亡潜伏期长 。

柱后衍生高效液相色谱法测定水产品中河豚毒素含量

采用柱后衍生高效液相色谱法检测水产品中的河豚毒素含量，选用河豚（Takifugu）和织纹螺（Nassarius）为主要研究对象，样品先由0．1％乙酸提取，经过C18固相萃取柱净化。以庚烷磺酸钠为离子对试剂，应用反相离子对液相色谱分离河豚毒素，采用在线柱后衍生系统，在110℃高温和4mol·L^-1 NaOH强碱条件下，将河豚毒素衍生化成具有荧光特性的C9碱，荧光检测器定量检测。河豚毒素在5～1600ng范围内呈良好的线性相关，相关系数r=0．9997；1、2、8μg·g^-1，3个浓度水平添加样，日内和日间的回收率为80．9％～91．2％、相对标准偏差为1．93％～5．57％；方法定量限为1μg·g^-1；从添加样色谱图上看，河豚毒素目标峰附近没有干扰峰，定性准确性好。采用柱后衍生高效液相色谱法与小鼠生物法检测同一阳性河豚样，检测结果一致性好。实验结果表明，柱后衍生高效液相色谱法适用于水产品中河豚毒素的检测。