Comparative Analysis of a Neurotoxin from Calliostoma canaliculatum by On-Line Capillary Isotachophoresis/H NMR and Diffusion ¹H NMR

Abstract

NMR spectroscopy has been coupled on-line to capillary isotachophoresis (cITP) to enhance structural analyses of dilute charged species through separation and sample concentration. Microcoils, the most mass-sensitive NMR probes available, provide optimal detection for cITP/NMR. To evaluate the utility of cITP/NMR for natural product analysis, a homogenate of the hypobranchial gland from the marine snail Calliostoma canaliculatum containing a cationic neurotoxin (1, a disulfide-bonded dimer of 6-bromo-2-mercaptotryptamine) was studied. For comparison, hypobranchial gland homogenate was also examined by diffusion-NMR, an alternative approach for NMR mixture analysis. cITP/NMR concentrated the neurotoxin by almost 20-fold and isolated it from some of the other components present in the matrix. However, a minor component, likely a precursor or degradant, co-migrated with compound 1. Diffusion-NMR also did not resolve the two, indicating that the compounds possessed similar diffusion coefficients and electrophoretic mobilities. The strengths and limitations of the two approaches for NMR mixture analysis are discussed.

Capillary isotachophoresis from the student point of view – images and the reality

“…The state of ITP in many fields was reviewed by many authors [97 -118] [190,191] Natural neurotoxins [192,193] Aromatic and aliphatic sulfonates [194,195] Pharmaceuticals [85, 86, 196 -205] Coupling of ITP with NMR [190,192,196,[232][233][234][235][236][237] [260] Gebauer and Bocek [119] was published in 2002. From this date, over a hundred novel applications and methods were published; a short survey of these works is given in Table 2.…”

Efficient identification of flavones, flavanones and their glycosides in routine analysis via off-line combination of sensitive NMR and HPLC experiments

“…Recent trends in coupling NMR with chromatographic techniques, like e.g. HPLC (Christophoridou, Dais, Tseng, & Spraul, 2005;Dai et al, 2009;Lambert, Hansen, Sairafianpour, & Jaroszewski, 2005;Tatsis et al, 2007) or capillary isotachophoresis (cITP) (Kautz et al, 2001;Wolters, Jayawickrama, & Sweedler, 2005) led to a significant improvement in identification of natural products. Comprehensive reviews on on-line and off-line hyphenated NMR methods are given by Jaroszewski (Jaroszewski, 2005a(Jaroszewski, , 2005b and Seger & Sturm (Seger & Sturm, 2007).…”

Suppressive subtractive hybridisation transcriptomics provides a novel insight into the functional role of the hypobranchial gland in a marine mollusc

“…isolated from the hypobranchial gland of the vetigastropod Calliostoma 60 canaliculatum, has been characterized as a potent neurotoxin, targeting potassium 61 channels (Kelley et al, 2003;Wolters et al, 2005). These secondary metabolites 62 within the hypobranchial gland of distinct gastropod lineages suggests that one of the 63 roles of the gland is the production of bioactive secondary metabolites, which may 64 have key roles in the chemical defence of molluscs.…”