Practice parameter: laryngeal electromyography (an evidence-based review)

This paper reports on an evidence-based review of laryngeal electromyography (EMG) as a technique for use in the diagnosis, prognosis, and treatment of laryngeal movement disorders, including the laryngeal dystonias, vocal fold paralysis, and other neurolaryngological disorders. The authors performed a systematic review of the medical literature from 1944 through 2001 on the clinical application of EMG to laryngeal disorders. Thirty-three of the 584 articles met the predefined inclusion criteria. The evidence demonstrated that in a double-blind treatment trial of botulinum toxin versus saline, laryngeal EMG used to guide injections into the thyroarytenoid muscle in persons with adductor spasmodic dysphonia was beneficial. A cross-over comparison between laryngeal EMG-guided injection and endoscopic injection of botulinum toxin into the posterior cricoarytenoid muscle in abductor spasmodic dysphonia found no significant difference between the two techniques and no significant treatment benefit. Based on the evidence, laryngeal EMG is possibly useful for the injection of botulinum toxin into the thyroarytenoid muscle in the treatment of adductor spasmodic dysphonia. There were no evidence-based data sufficient to support or refute the value of laryngeal EMG for the other uses investigated, although there is extensive anecdotal literature suggesting that it is useful for each of them. There is an urgent need for evidence-based research addressing other applications in the use of laryngeal EMG for other applications.     

Cloud-point extraction of nodularin-R from natural waters

A cloud-point extraction (CPE) technique for the determination of a cyanobacterial hepatotoxin, nodularin-R, in aqueous media using a cationic surfactant, tricaprylmethylammonium chloride (Aliquat-336), was developed. Cloud-point phase separation of Aliquat-336 at ambient temperature was induced by the addition of sodium sulfate. The Aliquat-336/Na₂SO₄ CPE system displayed large preconcentration factor, F_C, for nodularin-R. At the operational CPE conditions, F_C of 709.2 was achieved. Distribution constant, K_D, of the distribution of nodularin-R between the surfactant-rich and aqueous phases of the CPE system was estimated to be (4.94±0.8)×10³. Coupled to liquid chromatography with UV detection, the CPE method offered a detection limit of 330 pg ml⁻¹ (in freshwater)/1.3 ng ml⁻¹ (in seawater) and a repeatability of 6.4% (in freshwater) (n=7, P<0.05) for nodularin-R in a sample of 25 ml. The CPE is a rapid process and no cleanup step is required. Effects of pH, natural abundant anion (chloride) and dissolved organic matters (DOM, humic acid, HA) on the extraction efficiency were evaluated. A double CPE technique was developed to overcome interferences encountered in the analysis of environmental samples. Applicability of the new method to the determination of nodularin-R in real coastal marine water samples has also been demonstrated.     

Mass spectrometric identification of toxins of biological origin

The chemical/biological (CB) threat spectrum encompasses a wide range of potential agents including chemical warfare agents, biological warfare agents and toxins of biological origin that fall between these two main agent categories. These proteinaceous and non-proteinaceous toxins, commonly referred to as mid-spectrum agents, range in molecular mass from a few hundred to more than a hundred thousand daltons. The large number of potential candidates as well as the structural diversity of possible mid-spectrum agents makes identification of these compounds a challenge. The NATO defense community has recognized these challenges and has a working group that is developing identification protocols and evaluating methods through a series of international analytical exercises. Identification strategies rely heavily on recent advances that have been made in both mass spectrometry (MS) and liquid chromatography (LC), with LC-MS typically being employed as the primary method for separation/identification. While this paper focuses on the application of these and related instrumental analytical techniques for the identification of mid-spectrum agents, the approach described could be applied in the fields of toxicology, forensic science and environmental analysis. Areas for future research have been identified and application of developed mid-spectrum identification methods to the ongoing biological and toxin weapons convention (BTWC) are anticipated.     

LC-MS-MS aboard ship: tandem mass spectrometry in the search for phycotoxins and novel toxigenic plankton from the North Sea

Phycotoxins produced by various species of toxigenic microalgae occurring in the plankton are a global threat to the security of seafood resources and the health of humans and coastal marine ecosystems. This has necessitated the development and application of advanced methods in liquid chromatography coupled to mass spectrometry (LC-MS) for monitoring of these compounds, particularly in plankton and shellfish. Most such chemical analyses are conducted in land-based laboratories on stored samples, and thus much information on the near real-time biogeographical distribution and dynamics of phycotoxins in the plankton is unavailable. To resolve this problem, we conducted ship-board analysis of a broad spectrum of phycotoxins collected directly from the water column on an oceanographic cruise along the North Sea coast of Scotland, Norway, and Denmark. We equipped the ship with a triple-quadrupole linear ion-trap hybrid LC-MS-MS system for detection and quantitative analysis of toxins, such as domoic acid, gymnodimine, spirolides, dinophysistoxins, okadaic acid, pectenotoxins, yessotoxins, and azaspiracids (AZAs). We focused particular attention on the detection of AZAs, a group of potent nitrogenous polyether toxins, because the culprit species associated with the occurrence of these toxins in shellfish has been controversial. Marine toxins were analyzed directly from size-fractionated plankton net tows (20 μm mesh size) and Niskin bottle samples from discrete depths, after rapid methanolic extraction but without any further clean-up. Almost all expected phycotoxins were detected in North Sea plankton samples, with domoic acid and 20-methylspirolide G being most abundant. Although AZA was the least abundant of these toxins, the high sensitivity of the LC-MS-MS enabled detailed quantification, indicating that the highest amounts of AZA-1 were present in the southern Skagerrak in the 3–20 μm size-fraction. The direct on-board toxin measurements enabled isolation of plankton from stations with high AZA-1 levels and from the most suspicious size-fraction, i.e. most likely to contain the AZA-producer. A large number (>100) of crude cultures were established by serial dilution and later screened for the presence of AZAs after several weeks growth. From one crude culture containing AZA, a small dinoflagellate was subsequently isolated and brought into pure culture. We have thus proved that even sophisticated mass spectrometers can be operated in ship laboratories without any limitation caused by vibrations of the ship’s engine or by wave movement during heavy seas at wind forces up to nine Beaufort. On-board LC–MS–MS is a valuable method for near real-time analysis of phycotoxins in plankton for studies on bloom dynamics and the fate of toxins in the food web, and for characterization and isolation of putatively toxigenic organisms.     

Development and validation of a sensitive enzyme immunoassay for surveillance of Cry1Ab toxin in bovine blood plasma of cows fed Bt-maize (MON810)

The increasing global adoption of genetically modified (GM) plant derivatives in animal feed has provoked a strong demand for an appropriate detection method to evaluate the existence of transgenic protein in animal tissues and animal by-products derived from GM plant fed animals. A highly specific and sensitive sandwich enzyme immunoassay for the surveillance of transgenic Cry1Ab protein from Bt-maize in the blood plasma of cows fed on Bt-maize was developed and validated according to the criteria of EU-Decision 2002/657/EC. The sandwich assay is based on immuno-affinity purified polyclonal antibody raised against Cry1Ab protein in rabbits. Native and biotinylated forms of this antibody served as capture antibody and detection antibody for the ELISA, respectively. Streptavidin-horseradish peroxidase conjugate and TMB substrate provided the means for enzymatic colour development.The immunoassay allowed Cry1Ab protein determination in bovine blood plasma in an analytical range of 0.4-100 ng mL⁻¹ with a decision limit (CCα) of 1.5 ng mL⁻¹ and detection capability (CCβ) of 2.3 ng mL⁻¹. Recoveries ranged from 89 to 106% (mean value of 98%) in spiked plasma.In total, 20 plasma samples from cows (n = 7) fed non-transgenic maize and 24 samples from cows (n = 8) fed transgenic maize (collected before and, after 1 and 2 months of feeding) were investigated for the presence of the Cry1Ab protein. There was no difference amongst both groups (all the samples were below 1.5 ng mL⁻¹; CCα). No plasma sample was positive for the presence of the Cry1Ab protein at CCα and CCβ of the assay.     

Solid-phase extraction-fluorimetric high performance liquid chromatographic determination of domoic acid in natural seawater mediated by an amorphous titania sorbent

The feasibility of using sol-gel amorphous titania (TiO₂) as a solid-phase sorbent for the pre-concentration of domoic acid (DA), a potent amnesic shellfish poisoning (ASP) toxin, directly from seawater was explored. The sol-gel titania material is able to adsorb DA from seawater, via the formation of ester-linkage between the carboxylic moieties of DA and the Ti-OH groups on the sorbent surface, at low pH and desorb it at high pH. The chemisorption process is not significantly interfered by the seawater matrix. The optimum pH values for the adsorption and desorption of DA were found to be pH 4 and 11, respectively. The optimal sorbent loading for the batch-type solid-phase extraction of DA was 0.67 mg-TiO₂ ng-DA⁻¹ and adsorption equilibrium was achieved in 2 h at room temperature. The desorbed DA in 500 μL of 0.1 M alkaline borate buffer can be directly derviatized by 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) in aqueous media for fluorimetric HPLC quantification. Analyte recovery, repeatability and detection limit of this titania SPE-fluorimetric HPLC determination are 89%, 6.2% and 120 pg-DA mL⁻¹ (n = 7, P < 0.05), respectively, for a sample volume of 30 mL. This titania SPE technique should also be applicable to the pre-concentration of other polar carboxylate- and phosphonate-containing biomolecules and pharmaceuticals in complex and interfering environmental sample matrices.     

新型蛋白结合类毒素血液灌流吸附剂的制备及吸附性能

蛋白结合类毒素(PBUT)在尿毒症并发症的发生发展中起着重要作用, 现有血液净化模式对其清除效果较差, 开发用于高效清除尿毒症患者体内PBUT的血液灌流吸附材料已成为迫切的临床需求. 本文首先采用悬浮聚合法制备了咪唑基改性低交联聚苯乙烯微球P(St-DVB-VMZ); 然后通过小分子外交联剂的一步法傅克烷基化后交联反应, 制备出血液灌流用含咪唑基超高交联聚苯乙烯多孔树脂吸附剂HCP(St-DVB-VMZ). 利用傅里叶变换红外光谱(FTIR)、 X射线光电子能谱(XPS)、 扫描电子显微镜(SEM)及N₂吸附-脱附分析等表征了吸附树脂的化学结构和微观孔结构. 结果表明, HCP(St-DVB-VMZ)具有丰富的孔结构, 比表面积达到709 m²/g. 尿毒症毒素吸附实验结果表明, HCP(St-DVB-VMZ)对蛋白结合类毒素[对硫酸吲哚酚(IS)、 对甲酚硫酸盐(PCS)和吲哚乙酸(IAA)]和中大分子毒素[甲状旁腺激素(PTH)、 β₂-微球蛋白(β₂M)及白细胞介素6(IL-6)]均具有优异的吸附性能并展示出较好的血液相容性, 有望实现全血灌流临床应用.     

Identification and quantification of domoic acid by UHPLC/QTOF tandem mass spectrometry,with simultaneous identification of non-target photodegradation products

Amnesic shellfish poisoning is a potentially lethal human toxic syndrome which is caused by domoic acid (DA), a neurotoxin produced by marine phytoplankton, principally from Pseudonitzschia genus. In this report, a method to identify and quantify the DA toxin, with simultaneous identification of its photodegradation products, has been developed. It uses an ultra high performance liquid chromatography coupled to a quadrupole-time-of-flight tandem mass spectrometer (UHPLC–QTOF) after solid-phase extraction (SPE). An unambiguous identification of DA was carried out by considering both the retention time of DA in UHPLC and the exact mass of protonated DA molecule ([M + H]⁺ = 312.1447 m/z) and of the most intense fragment ion (m/z 266.1391). The quantification was conducted using protonated DA molecule with protonated Glafenin as internal standard, obtaining a limits of detection of 0.75 µg L^?1. Large screening with UHPLC–QTOF could also give structural information about degradation products of DA present in samples after UV-irradiation. This method was applied for the determination of DA in complex liquid samples after SPE and is applicable for environmental monitoring of this toxic substance in the aquatic environment.     

Characterization of (<Superscript>13</Superscript>C<Subscript>24</Subscript>) T-2 toxin and its use as an internal standard for the quantification of T-2 toxin in cereals with HPLC–MS/MS

In this paper, the structure and the identity of fully ¹³C-substituted T-2 toxin were confirmed using high-resolution mass spectrometry, ¹H-NMR, ¹³C-NMR, tandem mass spectrometry and HPLC–DAD. The purity of this compound was estimated to be at least 98.8% according to UV data. The isotopic distribution of (¹³C₂₄) T-2 toxin indicated a total isotopic enrichment of 98.2 ± 1.0 atom% ¹³C, and the application of different MS measurement modes revealed the MS/MS fragmentation pattern of T-2 toxin. Furthermore, a stable isotope dilution mass spectrometry method for the quantification of T-2 toxin was developed using (¹³C₂₄) T-2 toxin as internal standard. The method was evaluated with and without conventional clean-up and validated for maize and oats. Both cereals showed strong matrix enhancement effects, which could be compensated for through the application of the isotope-substituted internal standard.