title: Comparative analyses by HPLC and the sodium channel and saxiphilin 3H-saxitoxin receptor assays for paralytic shellfish toxins in crustaceans and molluscs from tropical North West Australia.
abstract: The increased frequency and distribution of red tides requires the development of high-throughput detection methods for paralytic shellfish toxins (PST). Community ethics also requires that there be a reduced reliance upon the standard mouse bioassay. A biomolecular assay such as the sodium channel 3H-saxitoxin binding assay can satisfy both of these requirements but may be compromised by cross-reactivity with the structurally unrelated tetrodotoxins (TTX). This study utilised the sodium channel assay but also an alternative 3H-saxitoxin binding assay based upon a saxiphilin isoform from the centipede Ethmostigmus rubripes to screen for PSTs. Saxiphilin is a novel transferrin which binds saxitoxin (STX) but differs from the sodium channel in not having any measurable affinity for TTX. A detailed analysis of toxin composition was achieved by high performance liquid chromatography (HPLC). Various crustaceans and molluscs accumulate PSTs and TTX, thus proving useful biomarkers for these toxins in their immediate environment and an ideal challenge to the detection and analysis of PSTs in this presumptive screening program. Also, there has been little investigation of PSTs in invertebrates from the Indian Ocean so this region was selected to extend our knowledge of the distribution of these toxins. 190 crabs and shellfish encompassing 31 species were collected from reefs along the North-West Australian coast and tested for PSTs and TTX by sodium channel and saxiphilin bioassays as well as HPLC. PSTs were detected in 18 species of crabs and shellfish of the 31 species tested. Eight of these species have not been previously described as toxic, these being the crabs Euzanthus exsculptus, Lophozozymus octodentatus, Metopograpsus frontalis, Pilumnus pulcher, Platypodia pseudogranulosa and Portunus pelagicus, and the molluscs Tectus fenestratus and Trochus hanleyanus. By HPLC, only one or both of STX and decarbamoyl-STX was detected in any extract. Some extracts markedly inhibited 3H-saxitoxin binding by the sodium channel but not by saxiphilin. The close agreement between toxin quantification by the PST specific methods of HPLC and the saxiphilin bioassay is indicative that the additional toxicity detected by the sodium channel assay is TTX.
authors.affiliation: Australian Institute of Marine Science, Dampier.
authors.forename: A
authors.initials: A
authors.lastname: Negri
authors.forename: L
authors.initials: L
authors.lastname: Llewellyn
mesh_terms: D029845:Amphibian Proteins; D000818:Animals; D001315:Australia; D002352:Carrier Proteins / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003445:Crustacea / Q000737:chemistry*; D019156:Indian Ocean; D008974:Mollusca / Q000737:chemistry*; D051381:Rats; D012530:Saxitoxin / Q000032:analysis* / Q000145:classification / Q000302:isolation & purification / Q000378:metabolism / Q000633:toxicity; D015222:Sodium Channels / Q000378:metabolism
chemical_list: D029845:Amphibian Proteins; D002352:Carrier Proteins; D015222:Sodium Channels; C027525:saxitoxin-binding protein, Rana catesbeiana; D012530:Saxitoxin