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At least 397 records · Page 22Linked to original sources

PubMed2003

Paralytic toxins in three new gastropod (Olividae) species implicated in food poisoning in southern Taiwan.

The toxins in the new gastropods Oliva miniacea, O. mustelina and O. nirasei implicated in a food paralytic poisoning incident in South Taiwan in February 2002 were studied. It was found that the three species of gastropods contained moderate amounts of toxin in edible portion only, and the highest toxicity score was 18 MU/g for O. miniacea, 10 MU/g for O. mustelina, and 27 MU/g for O. nirasei. The toxin was partially purified from the toxic specimens of each species by ultrafiltration using a YM-1 membrane, followed by chromatography on Bio-Gel P-2 column. Analyses by HPLC, GC-MS and LC-MS showed that the toxin from O. miniacea, O. nirasei and O. mustelina contained TTX, and related compounds 4-epi TTX and anhydro-TTX. The paralytic shellfish poisons were not found.

Animals↗

PubMed2001

A fluorimetric method based on changes in membrane potential for screening paralytic shellfish toxins in mussels.

To prevent the consumption of bivalves contaminated with paralytic shellfish poisoning (PSP), toxin levels in seafood products are estimated by using the official mouse bioassay. Because of the limitations of this bioassay other methods of monitoring toxins are clearly needed. We have developed a test to screen for PSP toxins based on its functional activity; the toxins bind to the voltage-gated Na+ channels and block their activity. The method is a fluorimetric assay that allows quantitation of the toxins by detecting changes in the membrane potential of human excitable cells. This assay gives an estimate of toxicity, since each toxin present in the sample binds to sodium channels with an affinity which is proportional to its intrinsic toxic potency. The detection limits for paralytic shellfish toxins were found to be 1 ng saxitoxin equivalents/ml compared to the regulatory limit threshold of 400 ng/ml (equivalent to 80 microg/100 g) used in most countries. Our results indicate that this fluorescent assay is a specific, very sensitive, rapid, and reliable method of monitoring PSP toxin levels in samples from seafood products and toxic algae.

Animals↗

PubMed1999

Production and characterization of a monoclonal antibody against domoic acid and its application to enzyme immunoassay.

For production of monoclonal antibodies against domoic acid, a causative agent of amnesic shellfish poisoning, three immunogens, domoic acid conjugated with bovine serum albumin (BSA), ovalbumin (OVA) and human gamma globulin (HGG), were prepared. The antiserum obtained from BALB/c mice immunized with domoic acid-BSA showed the highest affinity for domoic acid. The monoclonal antibody, DA-3, obtained from the mice was highly specific for domoic acid and showed a minor cross-reactivity with the isomers of domoic acid (isodomoic acids B, E, F, G and H), except for isodomoic acid A. Using DA-3 antibody, an indirect competitive enzyme immunoassay (idc-EIA) was developed for measurement of domoic acid. The working range for quantitative measurement of domoic acid and the quantification limit for domoic acid in shellfish were estimated to be 0.15-10 ng/ml and less than 0.04 microg/g, respectively. The mean recovery of domoic acid added to extracts of shellfish at toxin levels of 0.02 to 0.2 microg/ml was 103% with a coefficient of variation of 4.5%. The newly developed idc-EIA seems to be a useful method for monitoring domoic acid in shellfish.

Animals↗

PubMed2006

Detailed profiles of 7-O-acyl esters in plankton and shellfish from the Portuguese coast.

In bivalve mollusks from the Portuguese coast contaminated by diarrhetic shellfish poisoning (DSP), most of the parent toxins, okadaic acid (OA) or dinophysistoxin-2 (DTX2), are found esterified, and toxicity assessment is only performed after an alkaline hydrolysis step to recover the parent molecules in their free form. The presence of 7-O-acyl esters with fatty acids (FAs) has already been confirmed previously in Mytilus galloprovincialis and Donax trunculus samples. This paper reports the presence of acyl esters in a wider range of estuarine and offshore bivalve species found by direct analysis in LC-MS. The total of acyl esters found in each species represented the percentages commonly found by hydrolysis in those species in previous years, justifying the majority of the esters commonly found in shellfish. This implies that any diol esters remaining after digestion of toxic microalgae would represent only a minor contribution to the ester's contents. Esters with C14:0, C16:0, C16:1, C20:5 and C22:6 FAs were the most abundant, followed by esters with C18:0, C18:1, C18:2, C18:3 and C18:4. This is the first report of OA and DTX2 esters with odd FAs: C15:0, C17:0, C17:1, and probably a branched FA: iso-C16:0. Esters with iso-C16:0 where found in high percentages particularly in two species of estuarine clams, where they represented 13-34% of total esters found. Esters were also found in plankton, predominantly with C16:0. Total esters in plankton were not higher than 10%, not enough to justify per se the high levels found in bivalves.

Animals↗

PubMed2001

Structural elucidation of a new cytotoxin isolated from mussels of the Adriatic sea.

A detailed analysis of the toxic composition in the hepatopancreas of mussels from northern Adriatic sea has been performed. Along with some polyether toxins of DSP (diarrhetic shellfish poisoning) type, such as yessotoxin and its analogues, which are responsible for a variety of human seafood poisonings throughout the world, we have now isolated a new type of toxin, the chlorosulfolipid 1, which is completely different in structure from the polyether DSP-toxins isolated so far. The structural determination of the new toxin, including its absolute stereochemistry, has been performed by extensive NMR analysis and molecular mechanics and dynamics calculations.

Animals↗

PubMed2006

High amount of dinophysistoxin-3 in Mytilus chilensis collected in Seno de Reloncaví, chile, during massive human intoxication associated with outbreak of Vibrio parahaemolyticus.

This study describes the detection of high amount of 7-O-acyl-derivative dinophysistoxin-1 (Dinophysistoxin-3) in filter bivalves collected on February 2005 in the Seno de Reloncaví, Puerto Montt City, Southern Chile, in the same period of time where an intoxication episode was associated with the presence of Vibrio parahaemolyticus in shellfish. The Diarrhetic Shellfish Poisoning (DSP) mouse bioassay of mussel extract samples, performed as described for regulatory testing, were negative to DSP toxins. Therefore, the same mussel samples collected from 8 places of Seno de Reloncaví were then analyzed by the HPLC-FLD method with pre-column derivatization procedure for DSP toxins. The samples showed mainly 7-O-acyl-derivative dinophysistoxin-1 (Dinophysistoxin-3) in concentrations ranging from 190.3 +/- 6.8 to 311.1 +/- 4.8 ng of DSP toxin/g hepatopancreas and less amounts of Dinophysistoxin-1 ranging from 1.9 +/- 1.5 to 11.7 +/- 4.6 ng of DSP toxin/g hepatopancreas. After alkaline hydrolysis of the mussel extracts, 279.4 +/- 7.2 ng of Dinophysistoxin-1 /g hepatopancreas (mean +/- SEM, N=6) were found in mussel extracts (Zone 8). These data showed that these shellfish samples are contaminated with the ester form 7-O-acyl-derivatives of Dinophysisyoxin-1, far beyond the safe regulatory limit. This paper also shows a direct relation between lipid content in the mussel tissue extracts and the levels of Dinophysistoxin-3. The 7-O-acyl-derivative dinophysistoxin-1 ester was the only compound associated with DSP toxins detected in the shellfish samples, and in view of the fact that metabolic transformation of Dinophysistoxin-3 into Dinophysistoxin-1 in humans has recently been described in the literature, the consumption of shellfish contaminated with 7-O-acyl-derivatives dinophysistoxin-1 could be a major reason that explains the diarrhetic symptoms shown by the intoxicated patients.

Adult↗

PubMed1996

Interference of free fatty acids from the hepatopancreas of mussels with the mouse bioassay for shellfish toxins.

Determination of free fatty acids (FFA) in hepatopancreas of mussel, Mytilus coruscus, contaminated by diarrhetic shellfish poisoning (DSP) toxins, was carried out by high-performance liquid chromatography-fluorometry as their 9-anthryldiazomethane derivatives. The FFA in the hepatopancreas of mussels during May 30-June 14 were 443-666 micrograms/g. A marked increase in FFA content was observed on June 20 (2197 micrograms/g hepatopancreas) and June 27 (6322 micrograms/g hepatopancreas). The proportion of polyunsaturated fatty acids (PUFA), including 20:4n-6, 20:5n-3, and 22:6n-3, in mussel FFA increased during the experimental period. Diatoms almost completely dominated the phytoplankton community at the experimental site, suggesting that diatoms were the origin of the PUFA accumulated in mussels. The toxicity of FFA in mussel hepatopancreas on June 27 was sufficiently high as to interfere with the mouse bioassay by injection of an extract of shellfish, the official method for DSP toxin analysis.

Animals↗

PubMed1999

Confirmation of domoic acid production of Pseudo-nitzschia multiseries isolated from Ofunato Bay, Japan.

Production of domoic acid (DA), the responsible toxin for amnesic shellfish poisoning, was examined for 44 strains of Pseudo-nitzschia spp. isolated from Ofunato Bay, Japan. Only one strain which was identified as Pseudo-nitzschia multiseries produced DA in a level comparable to Canadian strains. No significant DA was detected in the rest of the strains, indicating that toxic P. multiseries does not bloom in a high density in the bay.

Canada↗

PubMed2000

Development of a method for the identification of azaspiracid in shellfish by liquid chromatography-tandem mass spectrometry.

Azaspiracid is the main toxin responsible for a number of recent human intoxications in Europe resulting from shellfish consumption. The first micro liquid chromatography-tandem mass spectrometry (micro-LC-MS-MS) method was developed for the determination of this novel shellfish poisoning toxin in mussels. The analyte was extracted from whole mussel meat with acetone and chromatographed on a C18 reversed-phase column (1.0 mm I.D.) by isocratic elution at 30 microl/min with acetonitrile-water (85:15, v/v), containing 0.03% trifluoroacetic acid. The toxin was ionised in an ionspray interface operating in the positive ion mode, where only the intact protonated molecule, [M+H]+, was generated at m/z 842. This served as precursor ion for collision-induced dissociation and three product ions, [M+H-nH2O]- with n=1-3, were identified for the unambiguous toxin confirmation by selected reaction monitoring LC-MS-MS analysis. A detection limit of 20 pg, based on a 3:1 signal-to-noise ratio, was achieved for the analyte. This LC-MS-MS method was successfully applied to determine azaspiracid in toxic cultivated shellfish from two regions of Ireland.

Animals↗

PubMedDate not supplied

Determination of decarbamoyl saxitoxin and its analogues in shellfish by prechromatographic oxidation and liquid chromatography with fluorescence detection.

Oxidation and chromatographic conditions for detecting the decarbamoyl analogues of several paralytic shellfish poison (PSP) toxins were studied. Prechromatographic oxidation with periodate or hydrogen peroxide under slightly alkaline conditions was used as previously reported for the parent PSP toxins. Both periodate and hydrogen peroxide oxidations produced 2 fluorescent products separable by liquid chromatography for each decarbamoyl (dc) toxin (dc-saxitoxin, dc-neosaxatoxin and dc-gonyautoxins 2 and 3). Decarbamoyl saxitoxin produced the same 2 products as did dc-neosaxitoxin but in different ratios. One of these products was the same as the one obtained with neosaxitoxin after periodate oxidation. Decarbamoyl gonyautoxins 2 and 3 (together) produced 2 products, one of which was the same as the major product obtained with gonyautoxins 1 and 4 (together) after periodate oxidation. Decarbamoyl gonyautoxins 1 and 4 were not available for study. The method was used to detect dc-saxitoxin and dc-gonyautoxins 2 and 3 in shellfish extracts.

Animals↗

PubMed2000

Evaluation of 11-[3H]-tetrodotoxin use in a heterologous receptor binding assay for PSP toxins.

This report describes the preparative scale production of 11-[3H]-tetrodotoxin (TTX) and its evaluation as a substitute for [3H]-saxitoxin (STX) as the radioligand in a receptor binding assay for paralytic shellfish poisoning (PSP) toxins. Restrictions on the world-wide distribution of [3H]-STX imposed by the international Chemical Weapons Convention served as the primary impetus for this study. We have incorporated on a preparative scale, a nonexchangeable tritium label into the TTX molecule at a specific activity of 12.90 Ci/mmol and recovered material of high radiochemical purity (98%). The resulting 11-[3H]-TTX was found to exhibit site-specific binding characteristics in the receptor assay (dissociation constant(K(d))=4.77+/-1.54nM; maximum binding(B(max))=1. 62+/-0.24pmol/mg of synaptosomal protein). The inhibition constant (K(i)) for the assay was 1.46+/-0.28 nM STX equiv. (n=6), with an estimated detection limit of ca. 2-4 ng STX equiv./ml in a sample extract. Moreover, quantitative comparisons indicated that 11-[3H]-TTX could be used interchangeably with [3H]-STX in the receptor assay for determination of PSP toxicity in shellfish and algal extracts without compromising assay performance. We conclude that the 11-[3H]-TTX produced and evaluated herein exhibits physical, chemical and biological characteristics suitable not only for use in the PSP receptor binding assay, but likely for other applications employing [3H]-STX as the radioligand.

Evaluation Studies as Topic↗

PubMed1990

Analysis of domoic acid and related compounds by mass spectrometry and gas chromatography/mass spectrometry as N-trifluoroacetyl-O-silyl derivatives.

A method is presented for the analysis of shellfish tissue for domoic acid, a neurotoxic amino acid responsible for cases of amnesic shellfish poisoning. Tissue extracts are first taken through a two-stage solid-phase extraction clean-up, using reversed-phase and strong cation exchange cartridges. A two-stage derivatization, using N-methyl-bis-trifluoroacetamide followed by either N-methyl-tert-butyldimethylsilyltrifluoroacetamide or N, O-bis-trimethylsilyltrifluoroacetamide, is then used to produce an N-trifluoroacetyl-O-silyl derivative which can be analyzed by mass spectrometry with introduction via direct inlet probe, moving-belt liquid chromatograph/mass spectrometer interface, or capillary column gas chromatography. The N-trifluoroacetyl-O-tert-butyldimethylsilyl derivative, which has good stability towards hydrolysis, provides a spectrum well suited to gas chromatography/mass spectrometry (GC/MS) using selected ion recording. GC/MS data for two related compounds, kainic acid and dihydrokainic acid, are also reported. The latter is used as an internal standard for quantification of domoic acid, although the method reported is intended primarily for confirmation of the toxin and related compounds in shellfish tissue.

Chromatography, Gas↗

PubMed2004

Rapid determination of polyether marine toxins using liquid chromatography-multiple tandem mass spectrometry.

The diarrhetic shellfish poisoning (DSP) toxins, okadaic acid (OA), dinophysistoxins (DTX); pectenotoxin-2 (PTX2) and pectenotoxin-2 seco acids, were determined in marine phytoplankton, Dinophysis acuta, and mussels (Mytilus edulis) collected along the southwest coast of Ireland. Liquid chromatography-multiple tandem mass spectrometry (LC-MS/MS) was employed for the simultaneous determination of a series of marine toxins with large polarity differences. Separation of five DSP toxins was achieved on a C18 column (Luna-2, 150 mm x 2.1 mm, 5 microm) using an acetonitrile-water gradient with ammonium acetate as an eluent modifier. Electrospray ionisation (ESI) in negative mode, was used to generate the molecule related ion, [M-H]-, for each toxin. To develop a multiple reaction monitoring (MRM) method, fragmentation studies were performed to determine the optimum precursor-product ion combinations: OA (803/255), DTX2 (803/255), DTX1 (817/255), PTX2SAs (875/137) and PTX2 (857/137). This highly sensitive method had detection limits better than 1 pg (on-column). Linear calibrations were obtained for shellfish extracts that were spiked with toxins, OA, 0.007-1.00 microg/ml (r2 = 0.9993, N = 3) and DTX2, 0.054-8.5 microg/ml (r2 = 0.9992, N = 3). Good reproducibility data were also achieved with %RSD values (N = 3) ranging from 3.15% (0.56 microg DTX2/ml) to 5.71% (0.14 microg DTX2/ml), for shellfish extracts. The method was sufficiently sensitive to permit the determination of DSP toxins in small numbers of picked phytoplankton cells (N = 12-40). In one sample of D. acuta the average toxin composition per cell was: OA (7.0 pg), DTX2 (11 pg) and PTX2 (7.2 pg).

Chromatography, High Pressure Liquid↗

PubMedDate not supplied

Determination of brevetoxins in shellfish by the neuroblastoma assay.

A neuroblastoma assay for determination of brevetoxins in shellfish was developed together with a method for sample cleanup that allows separation of brevetoxins from most of the components that cause matrix interference in the assay. This improved assay method was applied to a range of shellfish samples with different characteristics. Extracts of naturally contaminated and nontoxic shellfish together with extracts spiked with known amounts of toxin were tested. The results demonstrated that brevetoxins could be reliably detected in shellfish extracts at concentrations below the regulatory limit. Brevetoxin activity was detected in 15 of 23 samples from 5 separate toxicity incidents in which shellfish tested positive in the neurotoxic shellfish poisoning (NSP) mouse bioassay. Twelve of these positive NSP results came from 2 toxicity incidents. Yessotoxin was the major contributor to toxicity in 2 other incidents, although some samples contained both yessotoxin and brevetoxin. The sample from the remaining incident contained an unidentified toxin bioactivity, together with gymnodimine. In contrast to earlier versions of the neuroblastoma assay, gymnodimine was not detected by this modified method.

Animals↗

PubMed2001

PSP toxins from Aphanizomenon flos-aquae (cyanobacteria) collected in the Crestuma-Lever reservoir (Douro river, northern Portugal).

The presence of paralytic shellfish poisoning (PSP) toxins in cultures of Aphanizomenon flos-aquae, isolated from the Crestuma-Lever reservoir, was found by reversed phase high performance liquid chromatography employing two isocratic elution systems for the separation of PSP toxins. With the first isocratic elution protocol, the presence of apolar toxins as saxitoxin, decarbamoyl saxitoxin and neosaxitoxin not detected. On the other hand, GTX4, GTX1 and GTX3 as well as Cs toxins were present either in the Aphanizomenon flos-aquae cells collected directly from the bloom or in the other toxic isolates priorly cultivated in laboratory conditions.

Bacterial Toxins↗

PubMed2006

Analysis of paralytic shellfish toxins in Aphanizomenon DC-1 from Lake Dianchi, China.

Lake Dianchi is in Yunnan Province in southwestern China. In recent years, significant cyanobacterial blooms have occurred in this lake nearly every year because of eutrophication. Monitoring data for the past 5 years acquired by our research group showed that phytoplankton composition alternated between species of Microcystis sp. during warm seasons and those of Aphanizomenon sp. during cool seasons. In March 2003, when phytoplankton composition was highly dominated by Aphanizomenon sp., samples were taken from the lake for toxin detection and immediate strain isolation. A mouse bioassay with extracts from the lyophilized field material showed obvious intoxication from paralytic shellfish poisons (PSPs), and all mice died within 30 min. Further analysis of both field and isolated algal strain Aphanizomenon DC-1 by the postcolumn HPLC-FLD method confirmed its PSP-producing ability. The analogues found in the extracts from the field material were neoSTX, dcSTX, and dcGTX3, with contents of 2.279, 1.135, and 0.547 ng/mg DW, respectively. Under laboratory culture condition, toxin content in the Aphanizomenon strain DC-1 varied greatly during different growth phases, with two peaks: in the early-exponential and late-stationary growth phases. When the culture grew at a relatively high rate during the mid- to late-exponential growth phase, toxin content declined gradually. Moreover, the types of toxin in the DC-1 strain varied greatly during a single culture cycle. The HPLC results showed that dcSTX was the only toxin isomer detected throughout the culture period, and its level remained stable. On the other hand, dcGTX2 and GTX4 were the major toxins during the early-exponential and stationary phases, respectively. This article presents the first data on the identification and detection of paralytic shellfish toxins from cyanobacteria in Lake Dianchi. As far as we know, this is also the first report of this type of toxin in inland water bodies in China. Our study indicates the threat associated with PSP toxins in Lake Dianchi and suggests that necessary measures and programs for control are urgently needed to prevent the spread of toxic cyanobacterial blooms.

Animals↗

PubMed1996

Liquid chromatographic determination of okadaic acid and dinophysistoxin-1 in shellfish after derivatization with 9-chloromethylanthracene.

The reagent 9-chloromethylanthracene was evaluated for derivatization of the diarrhetic shellfish poisons, okadaic acid and dinophysistoxin-1 (DTX-1), to form fluorescent products separable by liquid chromatography. The toxins were reacted with the reagent in acetonitrile in the presence of tetramethylammonium hydroxide for 1 h at 90 degrees C. The products were purified by using two silica solid-phase extraction cartridges before being determined by reversed-phase liquid chromatography with fluorescence detection. The results are comparable to those obtained using 9-anthryldiazomethane (ADAM) for okadaic acid and DTX-1 in mussel tissue. Detection limits were estimated to be about 70-100 ng/g hepatopancreas (equivalent to 12-20 ng/g whole tissue) for each toxin.

Animals↗

PubMedDate not supplied

The use of Daphnia magna for detection of okadaic acid in mussel extracts.

Okadaic acid (OA), the main toxin responsible for diarrheic shellfish poisoning (DSP) has toxicity for Daphnia magna (EC50 = 15 +/- 1.8 micrograms/l). A Daphnia bioassay was developed and used to analyse okadaic acid in mussel extracts. A linear correlation was found between OA concentration determined by the Daphnia bioassay and by HPLC assay (r = 0.85; p < 0.001). The Daphnia bioassay can measure OA levels 10 times below the threshold of the mouse bioassay method. It is an inexpensive sensitive tool which can be used in replacement of mouse bioassay for the screening of OA and some co-extracting toxins in mussel extracts.

Animals↗