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1.
J Chromatogr A ; 1608: 460409, 2019 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-31399210

RESUMO

In this study two different analytical approaches have been developed to determine the presence of several cyanotoxins in saline water samples from a continental salt marsh. A salting-out assisted liquid-liquid extraction (SALLE) has been used in combination with ultra-high performance liquid chromatography-tandem mass spectrometry and UV-diode array detection (UHPLC-MS/MS and UHPLC-DAD). The target analytes are eight microcystins named MC-RR, MC-YR, MC-LR, MC-WR, MC-LA, MC-LY, MC-LW, MC-LF and nodularin (NOD), covering a wide range of polarities. The separation was achieved using a Zorbax Eclipse Plus RRHD C18 column (50 × 2.1 mm, 1.8 µm) in less than 7.5 and 5.5 min for UV and MS/MS detection, respectively. The mobile phase used consisted of water (solvent A) and acetonitrile (MeCN) (solvent B), both containing 0.01% of formic acid for DAD and 0.4% of formic acid for MS/MS detection, at a flow rate of 0.4 mL min-1. The temperature of the column was set at 25 °C and 20 µL of sample were injected. The main parameters affecting the SALLE procedure were studied and the following optimum values were obtained: neutral pH, 2 mL of acetonitrile as extraction solvent and 1.2 g of ammonium sulfate as salting-out agent for 4 mL of water sample. The validation protocols for both methods were accomplished with real water samples obtaining LODs ranging from 1.0 to 3.4 µg L-1 and 0.02 to 0.11 µg L-1 for DAD and MS/MS respectively. Although the SALLE-UHPLC-DAD methodology is easier and cheaper than UHPLC-MS/MS significantly better detection limits were achieved with tandem mass spectrometry as well as allowing for unambiguous identification. Extraction recoveries were higher than 77.0% (except for MC-RR and NOD which were 53.2% and 54.3, respectively) with satisfactory inter-day and intra-day precisions (RSD below 13.3%).


Assuntos
Cromatografia Líquida de Alta Pressão , Monitoramento Ambiental/métodos , Extração Líquido-Líquido , Toxinas Marinhas , Microalgas/química , Água do Mar/química , Espectrometria de Massas em Tandem , Limite de Detecção , Toxinas Marinhas/análise , Toxinas Marinhas/isolamento & purificação , Microcistinas/análise , Microcistinas/isolamento & purificação , Reprodutibilidade dos Testes , Poluentes Químicos da Água/análise , Poluentes Químicos da Água/isolamento & purificação
2.
J Chromatogr A ; 1608: 460404, 2019 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-31378529

RESUMO

An easy-prepared adsorbent with high stable and good dispersibility is especially valuable for the development of magnetic solid-phase extraction (MSPE) techniques. In this study, magnetic nitrogen-doped carbon nanotube cages (N-CNTCs) were synthesized via direct carbonization of cobalt (II)-containing metal-organic frameworks. During carbonization, cobalt ions inside the MOFs were converted into magnetic functional nanoparticles of N-CNTCs. Simultaneously, large amounts of nitrogen, originating from the organic ligands, were doped into the carbon framework. This unique structure gave the N-CNTCs excellent chemical stability, high affinity, and good dispersibility. The synthesized magnetic N-CNTCs were then used for MSPE of okadaic acid (OA) from aquatic samples. A simple, efficient, and sensitive method for detecting and quantitating OA was developed by combining the above sample pretreatment technique with high-performance liquid chromatography -tandem mass spectrometry (HPLC-MS/MS). The resulting method boasts a linear dynamic range of 3.0-1000.0 pg mL-1 with good linearity (R2 ≥ 0.9994). The limit of detection (LOD) and limit of quantification (LOQ) were 1.3 pg mL-1 and 3.0 pg mL-1, respectively. Several shellfish and seafood samples were analyzed using the developed method, showing satisfactory recoveries (82.0-107.0%) and relative standard deviations (<4.5%). The developed method was also used to investigate the OA distribution in crab tissues. Our results demonstrate that magnetic N-CNTCs are promising adsorbents for providing reliable support for the early warning and tracing to the source of algae toxins.


Assuntos
Técnicas de Química Analítica/métodos , Cobalto/química , Nanotubos de Carbono/química , Nitrogênio/química , Ácido Okadáico/isolamento & purificação , Extração em Fase Sólida/métodos , Técnicas de Química Analítica/normas , Cromatografia Líquida de Alta Pressão , Limite de Detecção , Magnetismo , Toxinas Marinhas/análise , Toxinas Marinhas/isolamento & purificação , Ácido Okadáico/análise , Frutos do Mar/análise , Espectrometria de Massas em Tandem
3.
Food Chem ; 294: 104-111, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31126442

RESUMO

Magnetic solid-phase extraction (MSPE), using a new reversed-phase/weak anion exchange mix-mode mesoporous magnetic SiO2 adsorbent, was assessed as an approach for reducing matrix effects in the analysis of six lipophilic marine biotoxins in shellfish using ultrahigh-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The adsorbent showed greater adsorption capacity and selectivity for the analytes and, thus, the MSPE microspheres reduced the matrix effects significantly in the subsequent analysis. In the UPLC-MS/MS analysis, precursor and product ions of the analytes were monitored quantitatively and qualitatively using multiple reaction monitoring and product ion confirmation modes. The proposed method exhibited a linear correlation of 0.9980-0.9991 in the working range for azaspiracids (2.0-200.0 ng/mL) and okadaic acid and its derivatives dinophysistoxins (4.0-200.0 ng/mL) with satisfactory recoveries (82.8-118.6%, RSD < 12%), lower LODs (0.4-1.0 µg/kg) and LOQs (1.0-4.0 µg/kg) than existing methods. In addition, consumption of the adsorbent was reduced, and the MSPE operation is simple and rapid relative to alternatives. These results suggest the proposed method has potential for use in the analysis of lipophilic marine biotoxins in shellfish samples.


Assuntos
Cromatografia Líquida de Alta Pressão , Magnetismo , Toxinas Marinhas/análise , Microesferas , Ácido Okadáico/análise , Compostos de Espiro/análise , Espectrometria de Massas em Tandem , Óxido Ferroso-Férrico/química , Limite de Detecção , Toxinas Marinhas/isolamento & purificação , Ácido Okadáico/isolamento & purificação , Porosidade , Frutos do Mar/análise , Dióxido de Silício/química , Extração em Fase Sólida , Compostos de Espiro/isolamento & purificação
4.
Mar Drugs ; 17(4)2019 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-30965587

RESUMO

A novel protein, soritesidine (SOR) with potent toxicity was isolated from the marine sponge Spongosorites sp. SOR exhibited wide range of toxicities over various organisms and cells including brine shrimp (Artemia salina) larvae, sea hare (Aplysia kurodai) eggs, mice, and cultured mammalian cells. Toxicities of SOR were extraordinary potent. It killed mice at 5 ng/mouse after intracerebroventricular (i.c.v.) injection, and brine shrimp and at 0.34 µg/mL. Cytotoxicity for cultured mammalian cancer cell lines against HeLa and L1210 cells were determined to be 0.062 and 12.11 ng/mL, respectively. The SOR-containing fraction cleaved plasmid DNA in a metal ion dependent manner showing genotoxicity of SOR. Purified SOR exhibited molecular weight of 108.7 kDa in MALDI-TOF MS data and isoelectric point of approximately 4.5. N-terminal amino acid sequence up to the 25th residue was determined by Edman degradation. Internal amino acid sequences for fifteen peptides isolated from the enzyme digest of SOR were also determined. None of those amino acid sequences showed similarity to existing proteins, suggesting that SOR is a new proteinous toxin.


Assuntos
Toxinas Marinhas/toxicidade , Poríferos , Sequência de Aminoácidos , Animais , Aplysia/efeitos dos fármacos , Artemia/efeitos dos fármacos , Comportamento Animal/efeitos dos fármacos , Bioensaio/métodos , Linhagem Celular Tumoral , Humanos , Japão , Larva/efeitos dos fármacos , Masculino , Toxinas Marinhas/administração & dosagem , Toxinas Marinhas/química , Toxinas Marinhas/isolamento & purificação , Camundongos , Peso Molecular , Testes de Mutagenicidade/métodos
5.
J Zhejiang Univ Sci B ; 20(1): 59-70, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30614230

RESUMO

Globally, peptide-based anticancer therapies have drawn much attention. Marine organisms are a reservoir of anticancer peptides that await discovery. In this study, we aimed to identify cytotoxic oligopeptides from Sarcophyton glaucum. Peptides were purified from among the S. glaucum hydrolysates produced by alcalase, chymotrypsin, papain, and trypsin, guided by a methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay on the human cervical cancer (HeLa) cell line for cytotoxicity evaluation. Purification techniques adopted were membrane ultrafiltration, gel filtration chromatography, solid phase extraction (SPE), and reversed-phase high-performance liquid chromatography (RP-HPLC). Purified peptides were identified by de novo peptide sequencing. From papain hydrolysate, three peptide sequences were identified: AGAPGG, AERQ, and RDTQ (428.45, 502.53, and 518.53 Da, respectively). Peptides synthesized from these sequences exhibited cytotoxicity on HeLa cells with median effect concentration (EC50) values of 8.6, 4.9, and 5.6 mmol/L, respectively, up to 5.8-fold stronger than the anticancer drug 5-fluorouracil. When tested at their respective EC50, AGAPGG, AERQ, and RDTQ showed only 16%, 25%, and 11% cytotoxicity to non-cancerous Hek293 cells, respectively. In conclusion, AERQ, AGAPGG, and RDTQ are promising candidates for future development as peptide-based anticancer drugs.


Assuntos
Antozoários/química , Citotoxinas/isolamento & purificação , Toxinas Marinhas/isolamento & purificação , Oligopeptídeos/isolamento & purificação , Sequência de Aminoácidos , Animais , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Cromatografia em Gel , Cromatografia Líquida de Alta Pressão , Cromatografia de Fase Reversa , Citotoxinas/farmacologia , Descoberta de Drogas , Células HEK293 , Células HeLa , Humanos , Hidrólise , Toxinas Marinhas/farmacologia , Oligopeptídeos/farmacologia , Extração em Fase Sólida , Espectrometria de Massas em Tandem
6.
Food Chem ; 272: 427-433, 2019 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-30309565

RESUMO

A simple QuEChERS method coupled with ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed to improve the extraction efficiency of lipophilic marine toxins (yessotoxins, dinophysistoxins, okadaic acid, azazspiracids, and spirolides) in fresh and processed shellfish products. The proposed method included freezing and dispersive solid-phase extraction with graphene oxide as the sorbent to clean complex matrices containing lipids (e.g., free fatty acids) and pigments. Quantification was performed using matrix-matched calibration curves. Recoveries were 85%-117.4% and the relative standard deviation for precision was less than 10% for marine toxins in fresh and processed shellfish products. The limits of detection (signal-to-noise = 3) and quantification (signal-to-noise = 10) were 0.10-1.47 and 0.32-4.92 µg/kg, respectively. The validated QuEChERS method, coupled with UPLC-MS/MS, was applied successfully to determine lipophilic marine toxins in fresh and processed shellfish samples.


Assuntos
Toxinas Marinhas/análise , Frutos do Mar/análise , Espectrometria de Massas em Tandem/métodos , Animais , Cromatografia Líquida de Alta Pressão , Congelamento , Grafite/química , Limite de Detecção , Toxinas Marinhas/isolamento & purificação , Ácido Okadáico , Oxocinas , Alimentos Marinhos/análise , Extração em Fase Sólida
8.
Mar Drugs ; 16(12)2018 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-30545093

RESUMO

Marine dinoflagellates are a valuable source of bioactive molecules. Many species produce cytotoxic compounds and some of these compounds have also been investigated for their anticancer potential. Here, we report the first investigation of the toxic dinoflagellate Alexandrium minutum as source of water-soluble compounds with antiproliferative activity against human lung cancer cells. A multi-step enrichment of the phenol⁻water extract yielded a bioactive fraction with specific antiproliferative effect (IC50 = 0.4 µg·mL-1) against the human lung adenocarcinoma cells (A549 cell line). Preliminary characterization of this material suggested the presence of glycoprotein with molecular weight above 20 kDa. Interestingly, this fraction did not exhibit any cytotoxicity against human normal lung fibroblasts (WI38). Differential gene expression analysis in A549 cancer cells suggested that the active fraction induces specific cell death, triggered by mitochondrial autophagy (mitophagy). In agreement with the cell viability results, gene expression data also showed that no mitophagic event was activated in normal cells WI38.


Assuntos
Antineoplásicos/farmacologia , Organismos Aquáticos/química , Dinoflagelados/química , Toxinas Marinhas/farmacologia , /efeitos dos fármacos , Células A549 , Antineoplásicos/isolamento & purificação , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Toxinas Marinhas/isolamento & purificação , Toxinas Marinhas/uso terapêutico
9.
Mar Drugs ; 16(11)2018 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-30441860

RESUMO

Cyclic imine toxins are neurotoxic, macrocyclic compounds produced by marine dinoflagellates. Mass spectrometric screenings of extracts from natural plankton assemblages revealed a high chemical diversity among this toxin class, yet only few toxins are structurally known. Here we report the structural characterization of four novel cyclic-imine toxins (two gymnodimines (GYMs) and two spirolides (SPXs)) from cultures of Alexandrium ostenfeldii. A GYM with m/z 510 (1) was identified as 16-desmethylGYM D. A GYM with m/z 526 was identified as the hydroxylated degradation product of (1) with an exocyclic methylene at C-17 and an allylic hydroxyl group at C-18. This compound was named GYM E (2). We further identified a SPX with m/z 694 as 20-hydroxy-13,19-didesmethylSPX C (10) and a SPX with m/z 696 as 20-hydroxy-13,19-didesmethylSPX D (11). This is the first report of GYMs without a methyl group at ring D and SPXs with hydroxyl groups at position C-20. These compounds can be conceived as derivatives of the same nascent polyketide chain, supporting the hypothesis that GYMs and SPXs are produced through common biosynthetic genes. Both novel GYMs 1 and 2 were detected in significant amounts in extracts from natural plankton assemblages (1: 447 pg; 2: 1250 pg; 11: 40 pg per mL filtered seawater respectively).


Assuntos
Dinoflagelados/química , Compostos Heterocíclicos com 3 Anéis/química , Hidrocarbonetos Cíclicos/química , Iminas/química , Toxinas Marinhas/química , Fitoplâncton/química , Compostos de Espiro/química , Compostos Heterocíclicos com 3 Anéis/isolamento & purificação , Hidrocarbonetos Cíclicos/isolamento & purificação , Iminas/isolamento & purificação , Toxinas Marinhas/isolamento & purificação , Estrutura Molecular , Compostos de Espiro/isolamento & purificação
10.
Mar Drugs ; 16(11)2018 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-30366463

RESUMO

Blue biotechnologies implement marine bio-resources for addressing practical concerns. The isolation of biologically active molecules from marine animals is one of the main ways this field develops. Strikingly, cnidaria are considered as sustainable resources for this purpose, as they possess unique cells for attack and protection, producing an articulated cocktail of bioactive substances. The Mediterranean sea anemone Anemonia viridis has been studied extensively for years. In this short review, we summarize advances in bioprospecting of the A. viridis toxin arsenal. A. viridis RNA datasets and toxin data mining approaches are briefly described. Analysis reveals the major pool of neurotoxins of A. viridis, which are particularly active on sodium and potassium channels. This review therefore integrates progress in both RNA-Seq based and biochemical-based bioprospecting of A. viridis toxins for biotechnological exploitation.


Assuntos
Venenos de Cnidários/química , Venenos de Cnidários/genética , Toxinas Marinhas/química , Neurotoxinas/química , Neurotoxinas/genética , Anêmonas-do-Mar/química , Anêmonas-do-Mar/genética , Animais , Venenos de Cnidários/isolamento & purificação , Venenos de Cnidários/farmacologia , Mineração de Dados , Toxinas Marinhas/genética , Toxinas Marinhas/isolamento & purificação , Toxinas Marinhas/farmacologia , Neurotoxinas/isolamento & purificação , Neurotoxinas/farmacologia , RNA/química , RNA/genética , Análise de Sequência de RNA , Pesquisa Médica Translacional
11.
Toxicon ; 155: 27-31, 2018 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-30312694

RESUMO

Pseudo-nitzschia multiseries is a diatom species associated with the production of domoic acid (DA), a water soluble neurotoxin that is easily transferred up in the food web, causing devastating effects on top marine organisms and humans. Despite studies on Pseudo-nitzschia are relevant to human health safety, partitioning of marine toxins between intracellular and extracellular fractions are poorly documented. This study aimed to determine the growth rates and DA content, both intracellular and extracellular, of Pseudo-nitzschia multiseries cultures at three different light settings (15, 120 and 560 µmol m-2 s-1). The optimal conditions for cell growth were observed at 120 and 560 µmol m-2 s-1, whereas DA production was observed in P. multiseries at 15 and 120 µmol m-2 s-1, ranging between 0.18-2.56 and 0.16-3.5 pg DA cell-1, respectively. Higher intracellular DA concentrations were found during the senescence phase at low light intensity and during the exponential phase at medium light intensity, while higher concentrations of dissolved DA were found at low and medium light intensities in the senescence phase reaching 3 and 10 ng DA mL-1 respectively. The amount of toxin released into the culture medium represents the most important fraction ranging between 63 and 98% during the exponential phase and nearly 99% during the senescence phase. In contrast, under low light intensity, dissolved DA was detected in the culture medium only during the senescence phase. This study confirms the importance of light intensity on DA production and clearly shows that dissolved domoic acid is an important fraction in Pseudo-nitzschia cultures, suggesting with the careful assumptions of results from static cultures extrapolated to bloom situations that waterborne exposure of marine organism should be considered during blooms of Pseudo-nitzschia multiseries.


Assuntos
Diatomáceas/crescimento & desenvolvimento , Diatomáceas/efeitos da radiação , Ácido Caínico/análogos & derivados , Toxinas Marinhas/análise , Células Cultivadas , Diatomáceas/metabolismo , Relação Dose-Resposta à Radiação , Ácido Caínico/análise , Ácido Caínico/isolamento & purificação , Ácido Caínico/metabolismo , Luz , Toxinas Marinhas/isolamento & purificação
12.
J Agric Food Chem ; 66(37): 9801-9809, 2018 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-30153406

RESUMO

In this study, a novel magnetic separation-based multiple systematic evolution of ligands by exponential enrichment (SELEX) was applied to select aptamers simultaneously against three kinds of marine biotoxins, including domoic acid (DA), saxitoxin (STX), and tetrodotoxin (TTX). Magnetic reduced graphene oxide (MRGO) was prepared to adsorb unbound ssDNAs and simplify the separation step. In the multiple SELEX, after the initial twelve rounds of selection against mixed targets and the subsequent four respective rounds of selection against each single target, the three resulting ssDNA pools were cloned, sequenced, and analyzed. Several aptamer candidates were selected and subjected to the binding affinity and specificity test. Finally, DA-06 ( Kd = 62.07 ± 19.97 nM), TTX-07 ( Kd = 44.12 ± 15.38 nM), and STX-41 ( Kd = 61.44 ± 23.18 nM) showed high affinity and good specificity for DA, TTX, and STX, respectively. They were also applied to detect and quantify DA, TTX, and STX successfully. The other two multitarget aptamers, DA-01 and TTX-27, were also obtained, which can bind with either DA or TTX. These aptamers provide alternative recognition molecules to antibodies for biosensor applications.


Assuntos
Ácido Caínico/análogos & derivados , Magnetismo/métodos , Toxinas Marinhas/isolamento & purificação , Técnica de Seleção de Aptâmeros/métodos , Saxitoxina/isolamento & purificação , Tetrodotoxina/isolamento & purificação , Aptâmeros de Nucleotídeos/química , Grafite/química , Ácido Caínico/química , Ácido Caínico/isolamento & purificação , Cinética , Magnetismo/instrumentação , Toxinas Marinhas/química , Óxidos/química , Técnica de Seleção de Aptâmeros/instrumentação , Saxitoxina/química , Tetrodotoxina/química
13.
J Agric Food Chem ; 66(11): 2962-2969, 2018 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-29502403

RESUMO

Azaspiracids belong to a family of more than 50 polyether toxins originating from marine dinoflagellates such as Azadinium spinosum. All of the azaspiracids reported thus far contain a 21,22-dihydroxy group. Boric acid gel can bind selectively to compounds containing vic-diols or α-hydroxycarboxylic acids via formation of reversible boronate complexes. Here we report use of the gel to selectively capture and release azaspiracids from extracts of blue mussels. Analysis of the extracts and fractions by liquid chromatography-tandem mass spectrometry (LC-MS) showed that this procedure resulted in an excellent cleanup of the azaspiracids in the extract. Analysis by enzyme-linked immunoasorbent assay (ELISA) and LC-MS indicated that most azaspiracid analogues were recovered in good yield by this procedure. The capacity of boric acid gel for azaspiracids was at least 50 µg/g, making this procedure suitable for use in the early stages of preparative purification of azaspiracids. In addition to its potential for concentration of dilute samples, the extensive cleanup provided by boric acid gel fractionation of azaspiracids in mussel samples almost eliminated matrix effects during subsequent LC-MS and could be expected to reduce matrix effects during ELISA analysis. The method may therefore prove useful for quantitative analysis of azaspiracids as part of monitoring programs. Although LC-MS data showed that okadaic acid analogues also bound to the gel, this was much less efficient than for azaspiracids under the conditions used. The boric acid gel methodology is potentially applicable to other important groups of natural toxins containing diols including ciguatoxins, palytoxins, pectenotoxins, tetrodotoxin, trichothecenes, and toxin glycosides.


Assuntos
Ácidos Bóricos/química , Toxinas Marinhas/isolamento & purificação , Mytilus edulis/química , Frutos do Mar/análise , Extração em Fase Sólida/métodos , Compostos de Espiro/isolamento & purificação , Adsorção , Animais , Cromatografia Líquida , Dinoflagelados/química , Géis/química , Toxinas Marinhas/química , Extração em Fase Sólida/instrumentação , Compostos de Espiro/química , Espectrometria de Massas em Tandem
14.
Ned Tijdschr Geneeskd ; 162: D2155, 2018.
Artigo em Holandês | MEDLINE | ID: mdl-29424330

RESUMO

BACKGROUND: When two or more people fall ill after eating the same food, this is called an outbreak of food poisoning or food-related infection. In the Netherlands, physicians have to notify the Municipal Health Services (GGD) of such outbreaks. The GGD informs the Netherlands Food and Consumer Product Safety Authority (NVWA). CASE DESCRIPTION: Two clusters of scombroid poisoning (histamine poisoning in this case after eating tuna) occurred in one month. Due to the quick reporting of patients by physicians, the GGD and the NVWA immediately tracked down the source. In both clusters the NVWA confirmed high histamine levels in tuna. In reaction to these findings and the first cluster, the supplier recalled the tuna batch. This may have prevented other cases of food poisoning. In the second cluster, the implicated tuna batch had already been sold. CONCLUSION: Timely notification by physicians of food poisoning or food-related outbreaks to the GGD enables swift tracing of the source and appropriate measures by the GGD and the NVWA..


Assuntos
Notificação de Doenças , Surtos de Doenças/prevenção & controle , Doenças Transmitidas por Alimentos , Toxinas Marinhas/isolamento & purificação , Papel do Médico , Atum , Animais , Notificação de Doenças/legislação & jurisprudência , Notificação de Doenças/normas , Doenças Transmitidas por Alimentos/diagnóstico , Doenças Transmitidas por Alimentos/epidemiologia , Doenças Transmitidas por Alimentos/etiologia , Doenças Transmitidas por Alimentos/prevenção & controle , Humanos , Países Baixos/epidemiologia
15.
Mar Drugs ; 16(1)2018 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-29301247

RESUMO

Ciguatera Fish Poisoning (CFP) is a human illness caused by the consumption of marine fish contaminated with ciguatoxins (CTX) and possibly maitotoxins (MTX), produced by species from the benthic dinoflagellate genus Gambierdiscus. Here, we describe the identity and toxicology of Gambierdiscus spp. isolated from the tropical and temperate waters of eastern Australia. Based on newly cultured strains, we found that four Gambierdiscus species were present at the tropical location, including G. carpenteri, G. lapillus and two others which were not genetically identical to other currently described species within the genus, and may represent new species. Only G. carpenteri was identified from the temperate location. Using LC-MS/MS analysis we did not find any characterized microalgal CTXs (P-CTX-3B, P-CTX-3C, P-CTX-4A and P-CTX-4B) or MTX-1; however, putative maitotoxin-3 (MTX-3) was detected in all species except for the temperate population of G. carpenteri. Using the Ca2+ influx SH-SY5Y cell Fluorescent Imaging Plate Reader (FLIPR) bioassay we found CTX-like activity in extracts of the unidentified Gambierdiscus strains and trace level activity in strains of G. lapillus. While no detectable CTX-like activity was observed in tropical or temperate strains of G. carpenteri, all species showed strong maitotoxin-like activity. This study, which represents the most comprehensive analyses of the toxicology of Gambierdiscus strains isolated from Australia to date, suggests that CFP in this region may be caused by currently undescribed ciguatoxins and maitotoxins.


Assuntos
Ciguatoxinas/isolamento & purificação , Dinoflagelados/classificação , Toxinas Marinhas/isolamento & purificação , Oxocinas/isolamento & purificação , Animais , Austrália , Linhagem Celular Tumoral , Cromatografia Líquida/métodos , Intoxicação por Ciguatera , Ciguatoxinas/toxicidade , Dinoflagelados/química , Humanos , Toxinas Marinhas/toxicidade , Oxocinas/toxicidade , Espectrometria de Massas em Tandem , Clima Tropical
16.
Chemosphere ; 190: 166-173, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28987405

RESUMO

The influence of nutritional regime and water temperature on depuration rates of OA-group toxins in the wedge shell Donax trunculus was examined by exposing naturally contaminated specimens to three nutritional regimes (microalgae, commercial paste of microalgae, and starvation) for 14 days at 16 °C and 20 °C. Total OA was quantified in the whole soft tissues of the individuals collected in days 2, 4, 6, 8, 10, 12 and 14. Mortality, dry weight, condition index, gross biochemical composition and gametogenic stages were surveyed. Low variation of glycogen and carbohydrates during the experiments suggest that wedge shells were under non-dramatic stress conditions. Wedge shells fed with non-toxic diets showed similar depuration rates being 15 and 38% higher than in starvation, at 16 and 20 °C, respectively. Depuration rates under non-toxic diets at 20 °C were 71% higher than at 16 °C. These results highlight the influence of water temperature on the depuration rate of total OA accumulated by D. trunculus, even when the increase is of only 4 °C, as commonly observed in week time scales in the southern Portuguese coastal waters. These results open the possibility of a faster release of OA in harvested wedge shells translocated to depuration systems when under a slight increase of water temperature.


Assuntos
Bivalves/química , Recuperação e Remediação Ambiental/métodos , Avaliação Nutricional , Ácido Okadáico/isolamento & purificação , Temperatura , Animais , Carboidratos/análise , Dieta/efeitos adversos , Glicogênio/análise , Toxinas Marinhas/isolamento & purificação
17.
J AOAC Int ; 101(2): 468-479, 2018 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-28851479

RESUMO

Paralytic shellfish toxins (PSTs) in bivalve molluscs represent a public health risk and are controlled via compliance with a regulatory limit of 0.8 mg saxitoxin (STX)⋅2HCl equivalents per kilogram of shellfish meat (eq/kg). Shellfish industries would benefit from the use of rapid immunological screening tests for PSTs to be used for regulation, but to date none have been fully validated. An interlaboratory study involving 16 laboratories was performed to determine the suitability of the Neogen test to detect PSTs in mussels and oysters. Participants performed the standard protocol recommended by the manufacturer and a modified protocol with a conversion step to improve detection of gonyautoxin 1&4. The statistical analysis showed that the protocols had good homogeneity across all laboratories, with satisfactory repeatability, laboratory, and reproducibility variation near the regulatory level. The mean probability of detection (POD) at 0.8 mg STX⋅2HCl eq/kg using the standard protocol in mussels and oysters was 0.966 and 0.997, respectively, and 0.968 and 0.966 using the modified protocol. The estimated LOD in mussels was 0.316 mg STX⋅2HCl eq/kg with the standard and 0.682 mg STX⋅2HCl eq/kg with the modified protocol, and 0.710 and 0.734 mg STX⋅2HCl eq/kg for oysters, respectively. The Neogen test may be acceptable for regulatory purposes for oysters in accordance with European Commission directives in which the standard protocol provides, at the regulatory level, a probability of a negative response of 0.033 on 95% of occasions. Its use for mussels is less consistent at the regulatory level due to the wide prediction interval around the POD.


Assuntos
Toxinas Marinhas/análise , Saxitoxina/análogos & derivados , Animais , Crassostrea/química , Dinoflagelados , Imunoensaio/métodos , Limite de Detecção , Toxinas Marinhas/imunologia , Toxinas Marinhas/isolamento & purificação , Mytilus/química , Kit de Reagentes para Diagnóstico , Saxitoxina/análise , Saxitoxina/imunologia , Saxitoxina/isolamento & purificação
18.
J AOAC Int ; 101(2): 480-489, 2018 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-28797318

RESUMO

Detection of paralytic shellfish toxins (PSTs) in bivalve shellfish by analytical methods is complicated and costly, requiring specific expertise and equipment. Following extensive blooms of Alexandrium tamarense Group 1 in Tasmania, Australia, an investigation was made into commercially available screening test kits suitable for use with the toxin profiles found in affected bivalves. The qualitative Neogen rapid test kit, with a modified protocol to convert gonyautoxins GTX1&4 and GTX2&3 into neosaxitoxin and saxitoxin (STX), respectively, with higher cross-reactivities, was the best fit-for-purpose. This validation study of the test kit and the modified protocol was undertaken following AOAC INTERNATIONAL guidelines for the validation of qualitative binary chemistry methods. The validation used four different PST profiles representing natural profiles found in Australia and in Europe: two in a mussel matrix and two in an oyster matrix. The test kit was shown to have appropriate selectivity of the toxin analogs commonly found in bivalve shellfish. The matrix and probability of detection (POD) study showed that the rapid test kit used with the modified protocol was able to consistently detect PST at the bivalve regulatory level of 0.8 mg STX⋅2HCl eq/kg, with a POD estimated via the binomial logistic regression of 1.0 at 0.8 mg STX⋅2HCl eq/kg in all tested profiles in both matrixes. The POD at 0.4 mg STX⋅2HCl eq/kg was 0.75 and 0.46 for the two toxin profiles in an oyster matrix and 0.96 and 1.0 for the two toxin profiles in a mussel matrix. No significant differences in the PODs of the PSTs at the regulatory level were found between production lots of the test kits. The results suggest the method is suitable to undergo a collaborative validation study.


Assuntos
Toxinas Marinhas/análise , Saxitoxina/análogos & derivados , Animais , Cromatografia Líquida , Crassostrea/química , Dinoflagelados , Imunoensaio/métodos , Toxinas Marinhas/isolamento & purificação , Mytilus/química , Saxitoxina/análise , Saxitoxina/isolamento & purificação , Espectrometria de Massas em Tandem
19.
Mar Drugs ; 15(12)2017 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-29261163

RESUMO

The long-lasting interest in bioactive molecules (namely toxins) produced by (microalga) dinoflagellates has risen in recent years. Exhibiting wide diversity and complexity, said compounds are well-recognized for their biological features, with great potential for use as pharmaceutical therapies and biological research probes. Unfortunately, provision of those compounds is still far from sufficient, especially in view of an increasing demand for preclinical testing. Despite the difficulties to establish dinoflagellate cultures and obtain reasonable productivities of such compounds, intensive research has permitted a number of advances in the field. This paper accordingly reviews the characteristics of some of the most important biotoxins (and other bioactive substances) produced by dinoflagellates. It also presents and discusses (to some length) the main advances pertaining to dinoflagellate production, from bench to large scale-with an emphasis on material published since the latest review available on the subject. Such advances encompass improvements in nutrient formulation and light supply as major operational conditions; they have permitted adaptation of classical designs, and aided the development of novel configurations for dinoflagellate growth-even though shearing-related issues remain a major challenge.


Assuntos
Organismos Aquáticos/química , Biotecnologia/métodos , Dinoflagelados/química , Toxinas Marinhas/farmacologia , Organismos Aquáticos/crescimento & desenvolvimento , Biotecnologia/tendências , Dinoflagelados/crescimento & desenvolvimento , Toxinas Marinhas/química , Toxinas Marinhas/isolamento & purificação
20.
Environ Monit Assess ; 189(12): 621, 2017 Nov 09.
Artigo em Inglês | MEDLINE | ID: mdl-29124474

RESUMO

Pseudo-nitzschia is a diatom genus capable of producing the neurotoxin domoic acid (DA), which has been related to mortalities of marine vertebrates, and the amnesic shellfish poisoning (ASP) in human consumers of contaminated bivalves. This study reports DA production among Pseudo-nitzschia strains isolated from shellfish farming areas in southern Brazil. Twenty-seven cultures of potentially toxigenic Pseudo-nitzschia species were established. Growth, stepped-chain formation, and DA production were evaluated in static, intermittently illuminated (12:12 photoperiod) batch cultures for 12 selected strains, and under continuous light and/or turbulence for a single Pseudo-nitzschia calliantha strain. Growth rate ranged from 0.16 to 0.39 day-1 among the 12 strains. Only P. calliantha and P. cf. multiseries yielded detectable levels of intracellular DA, reaching up to 0.054 fg cell-1 in late exponential and 0.15 pg cell-1 in early stationary phase, respectively. Continuous light impaired cell growth, and turbulence enhanced step-chain formation by threefold during exponential growth phase, but no DA production was detected under both conditions. The effect of turbulence on chain formation should be further evaluated in the field, once particle size is expected to affect the ingestion of toxic cells and thus toxin accumulation by certain DA vectors. The low cell toxicity revealed herein under laboratory conditions is in accordance with the low frequency of DA contamination episodes in south Brazilian waters. Nevertheless, monitoring of Pseudo-nitzschia abundance and DA presence in farming areas is continuously required to assure the quality of local shellfish products.


Assuntos
Monitoramento Ambiental , Toxinas Marinhas/análise , Animais , Bivalves , Brasil , Diatomáceas , Ácido Caínico/análogos & derivados , Laboratórios , Toxinas Marinhas/isolamento & purificação , Neurotoxinas/toxicidade , Frutos do Mar , Intoxicação por Frutos do Mar
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