Cylindrospermopsin determination using 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid (HEPES) as the internal standard

Cylindrospermopsin (CYN) was determined by liquid chromatography/electrospray ionization-mass spectrometry (LC/ESI-MS) using 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid (HEPES) as the internal standard. In the selected ion monitoring of LC/ESI-MS, m/z 414 for CYN and 237 for HEPES were monitored using the negative mode; the retention times of CYN and HEPES were 12.41 and 14.21 min, respectively. CYN was determined from peak area ratios of m/z 414/237. By the treatment of an anion exchange cartridge using a buffer at pH 10.5, CYN was isolated and condensed. No interfering peak was observed. Linearity of this method was observed at the range of 0.10-31.12 ng. Total coefficients of variation were 5.1 and 2.9% at 104 and 1038 μg CYN L⁻¹. The quantitative limit at a signal-to-noise (S/N) ratio of 10 was 0.16 ng.CYN concentration in natural waters is low. CYN in waters should be condensed for determination. This method including the treatment for isolation and condensation of CYN is useful for determination of CYN in environmental and/or drinking waters.     

Fit-for-purpose shellfish reference materials for internal and external quality control in the analysis of phycotoxins

The need for reference materials for quality control of analysis of foodstuffs has been stressed frequently. This has been particularly true in the phycotoxins field, where there is a great shortage of both pure calibration standards and reference materials. Worldwide there are very few independent bodies that produce certified reference materials for phycotoxins, the main producers currently being the National Research Council Canada and the Japanese Food Research Laboratory. Limited availability of contaminated shellfish and algae, as well as the time and knowledge necessary for the production of adequate reference materials, continuously lead to limited editions of certified reference materials and even more limited production of in-house reference materials. The restricted availability of in-house quality control materials promotes the rapid use of the limited certified reference materials, which in turn hampers the production of the suite of materials required globally for complete protection of public health. This paper outlines the various options that analysts can pursue in the use of reference materials for internal and external quality control, with a view to optimising the efforts of both reference materials users and reference materials producers. For this purpose, the logical sequence is reviewed from the discovery of a new bioactive compound in shellfish, through initial method development up to regulation for food safety purposes including accepted reference methods. Subsequently, the requirements for and efforts typically spent in the production and characterisation of laboratory reference materials, certified reference materials and other test materials used in inter-laboratory studies or proficiency testing, in the area of marine biotoxins are evaluated. Particular emphasis is put on practical advice for the preparation of in-house reference materials. The intricate link between reference material characterisation and method performance is outlined to give guidance on the appropriate in-house method validation in the rapidly developing field of phycotoxins.      

Paradoxical vocal fold motion: Presentation and treatment options

Paradoxical vocal fold motion is a rare disorder in which adduction of the folds occurs on inspiration. The disorder presents with signs of airway obstruction and often airway distress, so proper diagnosis by the otorhinolaryngologist is critical to subsequent management. We present a retrospective review of 10 patients with the diagnosis of paradoxical vocal fold motion seen over a 6-year period. Eight patients were females, and 6 required an acute airway intervention at presentation; 3 patients eventually underwent tracheotomy for respiratory decompensation. Six patients had a prior diagnosis of asthma, and this was determined to contribute to their respiratory status. Five patients were treated with botulinum toxin and 2 with flexible nasolaryngoscopic biofeedback, which improved the outcome. A review of the literature confirms a female predominance of patients presenting with paradoxical adduction and airway distress, often with a history of asthma and psychopathology. Our experience with botulinum toxin and biofeedback suggests that these procedures are viable treatment options in the management of patients with this disorder.     

农产品与饲料中T-2毒素免疫亲和柱净化-液相色谱串联质谱检测技术研究

利用T-2毒素单克隆抗体2G7和CNBr活化的Sepharose 4B研制出T-2免疫亲和柱,考察了T-2免疫亲和柱的最佳应用条件。利用免疫亲和柱,建立了农产品与饲料中T-2毒素免疫亲和柱净化/液相色谱-串联质谱(IAC/LC-MS/MS)确证性检测技术,并考察该方法的准确度和精密度。结果表明,T-2毒素在0.5~500.0 ng/g范围内线性关系良好,相关系数为0.999 7,检出限为0.05 ng/g,定量下限为0.17 ng/g。大米、玉米、饲料样品在T-2毒素10,50,100 ng/g的加标水平下,回收率为92.9%~109.7%,相对标准偏差为2.1%~8.3%。对市售36份农产品与饲料样品进行测定,检测结果的相对标准偏差均小于10.0%。     

A capacitive immunosensor for detection of cholera toxin

Contamination of food with biological toxins as well as their potential use as weapons of mass destruction has created an urge for rapid and cost effective analytical techniques capable of detecting trace amounts of these toxins. This paper describes the development of a sensitive method for detection of cholera toxin (CT) using a flow-injection capacitive immunosensor based on self-assembled monolayers. The sensing surface consists of monoclonal antibodies against the B subunit of CT (anti-CT), immobilized on a gold transducer. Experimental results show that the immunosensor responded linearly to CT concentrations in the range from 1.0 × 10⁻¹³ to 1.0 × 10⁻¹⁰ M under optimized conditions. The limit of detection (LOD) was 1.0 × 10⁻¹⁴ M. Two more analytical methods were employed for detection of CT using the same antibody namely, sandwich ELISA and surface plasmon resonance (SPR)-based immunosensor. The former had an LOD of 1.2 × 10⁻¹² M and a working range from 3.7 × 10⁻¹¹ to 2.9 × 10⁻¹⁰ M whereas, the later had an LOD of 1.0 × 10⁻¹¹ M and a linearity ranging from 1.0 × 10⁻⁹ to 1.0 × 10⁻⁶ M. These results demonstrate that the developed capacitive immunosensor system has a higher sensitivity than the other two techniques. The binding affinity of CT to the immobilized anti-CT was determined using the SPR-based immunosensor and an association constant (K_A) of 1.4 × 10⁹ M⁻¹ was estimated.     

高效液相色谱分析鱼毒性藻类海洋卡盾藻溶血毒素

采用高效液相色谱技术(HPLC)分离和分析了海洋卡盾藻(香港株)溶血毒素的甲醇粗提物,通过对比不同流动相、检测波长、等度和梯度洗脱方式,初步建立了分析其溶血毒素的HPLC方法.结果表明,采用梯度洗脱的方法,海洋卡盾藻溶血毒素的粗提物在50 min内取得了较好地分离.色谱条件为C8硅胶柱(4.6 mm× 150 mm,5 μm),流动相为乙腈和水,梯度洗脱(0～10 min,60％→80％乙腈;10～40 min,80％ →90％乙腈;40～50 min,90％乙腈);紫外检测器检测波长为205和448nm;流速为0.8 mL/min;柱温为25℃.用兔血红细胞法对各个主要色谱峰进行溶血活性检测显示,海洋卡盾藻(香港株)的溶血毒素至少含有5种成分.光谱分析显示一种成分的紫外吸收高峰在448 nm,一种未完全分离的混合组分的紫外吸收高峰为440和446nm,另外3种成分的紫外吸收高峰为205 nm.     

藻类肝毒素的富集提取与分离

采用固相萃取（SPE）和高效液相色谱法（HPLC）对蓝绿藻肝毒素进行富集与分离。以悦目颤藻为研究对象,选择最佳方式破坏其细胞,使其释放出细胞内肝毒素,对肝毒素进行SPE富集提取。用反相HPLC技术,以C18柱作分离术,32%乙腈-水（含0.01mol/L乙酸铵）为流动相,达到了对藻类肝毒素的良好分离。     

白喉毒素活性中心的量子化学计算与149位突变体的酶学动力学

通过量子化学计算确定白喉毒素分子催化区活性中心的关键氨基酸残基, 评价其取代后的酶活性的改变, 为导向性抗癌药物研究提供高效杀伤细胞工具. 结合目前关于白喉毒素结构与功能的研究状况和量子化学计算结果, 将白喉毒素催化区的第149位酪氨酸突变为苯丙氨酸, 对其酶活性和与底物的结合能力进行评价. Y149位酪氨酸位于正电中心, 起受电子作用, 与野生白喉毒素相比, 苯丙氨酸突变体的酶催化活性增加约一倍, 而与底物结合能力没有变化. Y149是酶活性中心的关键氨基酸残基, 对其取代能够影响蛋白质的生物活性.     

A comparison of injection techniques using botulinum toxin injection for treatment of the spasmodic dysphonias

The benefit duration and side effects following effective treatment of spasmodic dysphonia or voice tremor using botulinum toxin injection were studied following 477 open trial unilateral or bilateral injections provided to 178 patients. A comparison of unilateral and bilateral injections in male and female patients with adductor spasmodic dysphonia demonstrated an increased benefit duration following unilateral injections in women, longer periods of breathiness in men than in women following bilateral or unilateral injections, and a trend for reduced benefit durations following repeated treatments using bilateral injections. Outcomes were similar in the adductor spasmodic dysphonia and vocal tremor groups following thyroarytenoid injection while benefit durations were reduced and side effects lasted longer in patients with abductor spasmodic dysphonia following cricothyroid injections. Long-term benefit > 1 year occurred in 12.3% of patients following injection.