Precautions for seafood consumers: An updated review of toxicity, bioaccumulation, and rapid detection methods of marine biotoxins.

Seafood products are globally consumed, and there is an increasing demand for the quality and safety of these products among consumers. Some seafoods are easily contaminated by marine biotoxins in natural environments or cultured farming processes. When humans ingest different toxins accumulated in seafood, they may exhibit different poisoning symptoms. According to the investigations, marine toxins produced by harmful algal blooms and various other marine organisms mainly accumulate in the body organs such as liver and digestive tract of seafood animals. Several regions around the world have reported incidents of seafood poisoning by biotoxins, posing a threat to human health. Thus, most countries have legislated to specify the permissible levels of these biotoxins in seafood. Therefore, it is necessary for seafood producers and suppliers to conduct necessary testing of toxins in seafood before and after harvesting to prohibit excessive toxins containing seafood from entering the market, which therefore can reduce the occurrence of seafood poisoning incidents. In recent years, some technologies which can quickly, conveniently, and sensitively detect biological toxins in seafood, have been developed and validated, these technologies have the potential to help seafood producers, suppliers and regulatory authorities. This article reviews the seafood toxins sources and types, mechanism of action and bioaccumulation of marine toxins, as well as legislation and rapid detection technologies for biotoxins in seafood for official and fishermen supervision.

Foodborne Disease Outbreaks Caused by Biotoxins in Yantai City: A 10-Year Spatiotemporal Monitoring Study.

Unsafe food causes 600 million cases of foodborne diseases and 420,000 deaths every year. Meanwhile, biological toxins such as poisonous mushrooms, saponins, and aflatoxin can cause significant damage to humans. Therefore, it is particularly important to study foodborne disease outbreaks caused by biotoxins (FDOB). We collected FDOB in Yantai City from 2013 to 2022 and further established a corresponding database. Statistical analysis was carried out according to time, place, pathogen, and contamination of pathogenic factors. There were 128 FDOB, resulting in 417 patients and 6 deaths. The third quarter was a high season for foodborne disease outbreaks, the number of events, patients and deaths accounted for 65.63% (84/128), 55.88% (233/417), and 100% (6/6) of the total number, respectively. The highest number of outbreaks per 10,000 persons was Qixia (0.41), followed by Zhifu (0.36) and Laiyang (0.33). The top three causes of outbreaks were poisonous mushroom toxin, saponins and hemagglutinin, and Lagenaria siceraria (Molina) Standl. Sixty-five (50.78%) outbreaks were attributed to poisonous mushroom toxin, 18 (14.06%) outbreaks to saponin and hemagglutinin, and 12 (9.38%) outbreaks to L. siceraria (Molina) Standl. The largest number of outbreaks, patients and deaths all occurred in families, accounting for 82.81% (106/128) outbreaks, 66.19% (276/417) patients, and 100% (6/6) deaths, respectively. Followed by catering service establishments, accounting for 14.84% (19/128), 30.22% (126/417), and 0% (0/6), respectively. The main poisoning link of outbreaks was ingestion and misuse, accounting for 72.66% (93/128), followed by improper processing, accounting for 20.31% (26/128). It is necessary to carry out targeted family publicity and education, strengthen the integration of medical and prevention, explore innovative monitoring and early warning mechanisms for foodborne diseases, and reduce the occurrence of underreporting of foodborne disease outbreaks.

Recent advancements in LC-MS based analysis of biotoxins: Present and future challenges.

There has been a rising concern regarding the harmful impact of biotoxins, source of origin, and the determination of the specific type of toxin. With numerous reports on their extensive spread, biotoxins pose a critical challenge to figure out their parent groups, metabolites, and concentration. In that aspect, liquid chromatography-mass spectrometry (LC-MS) based analysis paves the way for its accurate identification and quantification. The biotoxins are ideally categorized as phytotoxins, mycotoxins, shellfish-toxins, ciguatoxins, cyanotoxins, and bacterial toxins such as tetrodotoxins. Considering the diverse nature of biotoxins, both low-resolution mass spectrometry (LRMS) and high-resolution mass spectrometry (HRMS) methods have been implemented for their detection. The sample preparation strategy for complex matrix usually includes "QuEChERS" extraction or solid-phase extraction coupled with homogenization and centrifugation. For targeted analysis of biotoxins, the LRMS consisting of a tandem mass spectrometer operating in multiple reaction monitoring mode has been widely implemented. With the help of the reference standard, most of the toxins were accurately quantified. At the same time, the suspect screening and nontarget screening approach are facilitated by the HRMS platforms during the absence of reference standards. Significant progress has also been made in sampling device employment, utilizing novel sample preparation strategies, synthesizing toxin standards, employing hybrid MS platforms, and the associated data interpretation. This critical review attempts to elucidate the progress in LC-MS based analysis in the determination of biotoxins while pointing out major challenges and suggestions for future development.

Exploration of Vulcanodinium rugosum Toxins and their Metabolism Products in Mussels from the Ingril Lagoon Hotspot in France.

Over the year 2018, we assessed toxin contamination of shellfish collected on a monthly basis in Ingril Lagoon, France, a site known as a hotspot for Vulcanodinium rugosum growth. This short time-series study gave an overview of the presence and seasonal variability of pinnatoxins, pteriatoxins, portimines and kabirimine, all associated with V. rugosum, in shellfish. Suspect screening and targeted analysis approaches were implemented by means of liquid chromatography coupled to both low- and high-resolution mass spectrometry. We detected pinnatoxin-A and pinnatoxin-G throughout the year, with maximum levels for each one observed in June (6.7 µg/kg for pinnatoxin-A; 467.5 µg/kg for pinnatoxin-G), whereas portimine-A was detected between May and September (maximum level = 75.6 µg/kg). One of the main findings was the identification of a series of fatty acid esters of pinnatoxin-G (n = 13) although the levels detected were low. The profile was dominated by the palmitic acid conjugation of pinnatoxin-G. The other 12 fatty acid esters had not been reported in European shellfish to date. In addition, after thorough investigations, two compounds were detected, with one being probably identified as portimine-B, and the other one putatively attributed to pteriatoxins. If available, reference materials would have ensured full identification. Monitoring of these V. rugosum emerging toxins and their biotransformation products will contribute towards filling the data gaps pointed out in risk assessments and in particular the need for more contamination data for shellfish.

Assessing the Toxicity of Lagocephalus sceleratus Pufferfish from the Southeastern Aegean Sea and the Relationship of Tetrodotoxin with Gonadal Hormones.

Given the dramatic increase in the L. sceleratus population in the southeastern Aegean Sea, there is growing interest in assessing the toxicity of this pufferfish and the factors controlling its tetrodotoxin (TTX) content. In the present study, liver, gonads, muscle and skin of 37 L. sceleratus specimens collected during May and June 2021 from the island of Rhodes, Greece, were subjected to multi-analyte profiling using liquid chromatography-tandem mass spectrometry (LC-MS/MS) in order to quantitate TTX and evaluate whether this biotoxin interrelates with hormones. TTX and its analogues 4-epiTTX, 11-deoxyTTX, 11-norTTX-6-ol, 4,9-anhydroTTX and 5,11/6,11-dideoxyTTX were detected in all tissue types. Liver and gonads were the most toxic tissues, with the highest TTX concentrations being observed in the ovaries of female specimens. Only 22% of the analyzed muscle samples were non-toxic according to the Japanese toxicity threshold (2.2 µg TTX eq g-1), confirming the high poisoning risk from the inadvertent consumption of this species. Four steroid hormones (i.e., cortisol, testosterone, androstenedione and ß-estradiol) and the gonadotropin-releasing hormone (GnRH) were detected in the gonads. Androstenedione dominated in female specimens, while GnRH was more abundant in males. A positive correlation of TTX and its analogues with ß-estradiol was observed. However, a model incorporating sex rather than ß-estradiol as the independent variable proven to be more efficient in predicting TTX concentration, implying that other sex-related characteristics are more important than specific hormone-regulated processes.

Ciguatoxin in Hawai'i: Fisheries forecasting using geospatial and environmental analyses for the invasive Cephalopholis argus (Epinephelidae).

Invasive species can precede far-reaching environmental and economic consequences. In the Hawai'ian Archipelago Cephalopholis argus (family Serranidae) is an established invasive species, now recognized as the dominant local reef predator, negatively impacting the native ecosystem and local fishery. In this region, no official C. argus fishery exists, due to its association with Ciguatera seafood poisoning (CP); a severe intoxication in humans occurring after eating (primarily) fish contaminated with ciguatoxins (CTXs). Pre-harvest prediction of CP is currently not possible; partly due to the ubiquitous nature of the microalgae producing CTXs and the diverse bioaccumulation pathways of the toxins. This study investigated the perceived risk of CP in two geographically discrete regions (Leeward and Windward) around the main island of Hawai'i, guided by local fishers. C. argus was collected and investigated for CTXs using the U.S. Food and Drug Administration (FDA) CTX testing protocol (in vitro neuroblastoma N2a-assay and LC-MS/MS). Overall, 76% of fish (87/113) exceeded the FDA guidance value for CTX1B (0.01 ng g-1 tissue equivalents); determined by the N2a-assay. Maximum CTX levels were ≅2× higher at the Leeward vs Windward location and, respectively, 95% (64/67) and 54% (25/46) of fish were positive for CTX-like activity. Fisher persons and environmental understandings, regarding the existence of a geographic predictor (Leeward vs Windward) for harvest, were found to be (mostly) accurate as CTXs were detected in both locations and the local designation of C. argus as a risk for CP was confirmed. This study provides additional evidence that supports the previous conclusions that this species is a severe CP risk in the coastal food web of Hawai'i, and that ocean exposure (wave power) may be a prominent factor influencing the CTX content in fish within a hyperendemic region for CP.

Optimization and Validation of a High Throughput UHPLC-MS/MS Method for Determination of the EU Regulated Lipophilic Marine Toxins and Occurrence in Fresh and Processed Shellfish.

Under the name of lipophilic marine toxins, there are included more than 1000 toxic secondary metabolites, produced by phytoplankton, with the common chemical property of lipophilicity. Due to toxicological effects and geographical distribution, in European legislation relevant compounds are regulated, and their determination is accomplished with the reference liquid chromatography-tandem mass spectrometry method. In this study a modified ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method has been developed for the identification and quantification of EU-regulated lipophilic toxins. The method optimization included a refinement of SPE-C18 clean-up, in order to reduce matrix interferences. Improved LC conditions and upgraded chromatographic ammonia-based gradient ensured the best separation of all analytes and, in particular, of the two structural isomers (OA and DTX2). Also, different MS parameters were tested, and confirmation criteria finally established. The validation studies confirmed that all parameters were satisfactory. The requirements for precision (RSD% < 11.8% for each compound), trueness (recoveries from 73 to 101%) and sensitivity (limits of quantification in the range 3−8 µg kg−1) were fulfilled. The matrix effect, ranging from −9 to 19%, allowed the use of a calibration curve in solvent (3−320 µg kg−1 in matrix) for quantification of real samples. Method relative uncertainty ranged from 12 to 20.3%. Additionally, a total of 1000 shellfish samples was analysed, providing a first preliminary surveillance study that may contribute to the knowledge of lipophilic marine toxins contamination. Increase in algae proliferation events and intoxication cases, EFSA suggestions for modification of maximum permitted levels and toxicity equivalency factors, and new studies of important toxic effects underline that implementation of reference methods still represents an important task for health and food safety laboratories.

Investigation of the Genotoxic Potential of the Marine Toxin C17-SAMT Using the In Vivo Comet and Micronucleus Assays.

The contaminant responsible for the atypical toxicity reported in mussels from Bizerte Lagoon (Northern Tunisia) during the last decade has been characterized as C17-sphinganine analog mycotoxin (C17-SAMT). This neurotoxin showed common mouse toxic symptoms, including flaccid paralysis and severe dyspnea, followed by rapid death. For hazard assessment on human health, in this work we aimed to evaluate the in vivo genotoxic effects of this marine biotoxin using the classical alkaline and modified Fpg comet assays performed to detect DNA breaks and alkali-labile sites as well as oxidized bases. The micronucleus assay was used on bone marrow to detect chromosome and genome damage. C17-SAMT induces a statistically insignificant increase in DNA tail intensity at all doses in the duodenum, and in the spleen contrary to the liver, the percentage of tail DNA increased significantly at the mid dose of 300 µg/kg b.w/d. C17-SAMT did not affect the number of micronuclei in the bone marrow. Microscopic observations of the liver showed an increase in the number of mitosis and hepatocytes' cytoplasm clarification. At this level of study, we confirm that C17-SAMT induced DNA damage in the liver but there was no evidence of effects causing DNA oxidation or chromosome and genome damage.

LC-HRMS Profiling of Paralytic Shellfish Toxins in Mytilus galloprovincialis after a Gymnodinium catenatum Bloom.

Saxitoxin and its more than 50 analogues are a group of naturally occurring neurotoxins collectively designated as paralytic shellfish toxins (PSTs). PSTs are toxic to humans and maximum legal limits in seafood have been implemented by regulatory authorities worldwide. In the European Union, monitoring of PSTs is performed using the AOAC Official Method 2005.06, based on liquid chromatography coupled with fluorescence detection (LC- FLD). However, this method has been suggested to not effectively detect the emerging C-11 hydroxyl (M-toxins) and benzoate (GC-toxins) analogues, with these analogues currently not being surveyed in monitoring programs. In this study, a liquid chromatography-high resolution mass spectrometry (LC-HRMS) method was used to search for these emerging PSTs in mussels, Mytilus galloprovincialis, contaminated following an intense Gymnodinium catenatum bloom in the Tagus estuary (Lisbon, Portugal). Five M-toxins (M1, M2, M6, dcM6, and dcM10), but no GC-toxins, were detected in the mussels' whole-soft body tissue. Moreover, the classical PSTs (C1 to C4, GTX 4 to GTX6, dcGTX1 to dcGTX4, dcSTX, dcNEO, and STX) were also found and comprised the largest fraction of the PSTs' profile. The presence of unregulated PSTs in edible mussel samples suggests potential seafood safety risks and urges further research to determine the frequency of these analogues in seafood and their contribution to toxicity.

A High Throughput Screening HPLC-FLD Method for Paralytic Shellfish Toxins (PSTs) Enabling Effective Official Control.

Paralytic Shellfish Toxins (PSTs) are marine biotoxins, primarily produced by dinoflagellates of the genera Gymnodinium spp., Alexandrium spp. They can accumulate in shellfish and, through the food chain, be assimilated by humans, giving rise to Paralytic Shellfish Poisoning. The maximum permitted level for PSTs in bivalves is 800 µg STX·2HCl eqv/kg (Reg. EC N° 853/2004). Until recently, the reference analytical method was the Mouse Bioassay, but Reg. EU N° 1709/2021 entered into force on 13 October 2021 and identified in the Standard EN14526:2017 or in any other internationally recognized validated method not entailing the use of live animals as official methods. Then the official control laboratories had urgently to fulfill the new requests, face out the Mouse Bioassay and implement instrumental analytical methods. The "EURLMB SOP for the analysis of PSTs by pre-column HPLC-FLD according to OMA AOAC 2005.06" also introduced a simplified semiquantitative approach to discriminate samples above and below the regulatory limit. The aim of the present paper is to present a new presence/absence test with a cut-off at 600 µg STX·2HCl eqv/kg enabling the fast discrimination of samples with very low PSTs levels from those to be submitted to the full quantitative confirmatory EN14526:2017 method. The method was implemented, avoiding the use of a large number of certified reference standards and long quantification procedures, resulting in an efficient, economical screening instrument available for official control laboratories. The protocol was fully validated, obtaining good performances in terms of repeatability (<11%) and recovery (53−106%) and accredited according to ISO/IEC 17025. The method was applied to mollusks collected from March 2021 to February 2022 along the Marche region in the frame of marine toxins official control.

Possible food safety hazards of ready-to-eat raw fish containing product (sushi, sashimi).

It is undeniable that with the popularity of sushi and sashimi over the last decade the consumption of raw fish has extremely increased. Raw fish is very appreciated worldwide and has become a major component of human diet because of its fine taste and nutritional properties. Possible hazards concerning fish safety and quality are classified as biological and chemical hazards. They are contaminants that often accumulate in edible tissue of fish and transmit to humans via the food chain affecting the consumer's health. Although their concentration in fish and fishery products are found at non-alarming level of a daily basis period, they induce hazardous outcome on human health due to long and continuous consumption of raw fish. Regular sushi and sashimi eaters have to be aware of the contaminants found in the other components of their dish that often add up to acceptable residue limits found in fish. Hence, there is the urge for effective analytical methods to be developed as well as stricter regulations to be put in force between countries to monitor the safety and quality of fish for the interest of public health.

Chemodiversity of Brevetoxins and Other Potentially Toxic Metabolites Produced by Karenia spp. and Their Metabolic Products in Marine Organisms.

In recent decades, more than 130 potentially toxic metabolites originating from dinoflagellate species belonging to the genus Karenia or metabolized by marine organisms have been described. These metabolites include the well-known and large group of brevetoxins (BTXs), responsible for foodborne neurotoxic shellfish poisoning (NSP) and airborne respiratory symptoms in humans. Karenia spp. also produce brevenal, brevisamide and metabolites belonging to the hemi-brevetoxin, brevisin, tamulamide, gymnocin, gymnodimine, brevisulcenal and brevisulcatic acid groups. In this review, we summarize the available knowledge in the literature since 1977 on these various identified metabolites, whether they are produced directly by the producer organisms or biotransformed in marine organisms. Their structures and physicochemical properties are presented and discussed. Among future avenues of research, we highlight the need for more toxin occurrence data with analytical techniques, which can specifically determine the analogs present in samples. New metabolites have yet to be fully described, especially the groups of metabolites discovered in the last two decades (e.g tamulamides). Lastly, this work clarifies the different nomenclatures used in the literature and should help to harmonize practices in the future.

Guidance Level for Brevetoxins in French Shellfish.

Brevetoxins (BTXs) are marine biotoxins responsible for neurotoxic shellfish poisoning (NSP) after ingestion of contaminated shellfish. NSP is characterized by neurological, gastrointestinal and/or cardiovascular symptoms. The main known producer of BTXs is the dinoflagellate Karenia brevis, but other microalgae are also suspected to synthesize BTX-like compounds. BTXs are currently not regulated in France and in Europe. In November 2018, they have been detected for the first time in France in mussels from a lagoon in the Corsica Island (Mediterranean Sea), as part of the network for monitoring the emergence of marine biotoxins in shellfish. To prevent health risks associated with the consumption of shellfish contaminated with BTXs in France, a working group was set up by the French Agency for Food, Environmental and Occupational Health & Safety (Anses). One of the aims of this working group was to propose a guidance level for the presence of BTXs in shellfish. Toxicological data were too limited to derive an acute oral reference dose (ARfD). Based on human case reports, we identified two lowest-observed-adverse-effect levels (LOAELs). A guidance level of 180 µg BTX-3 eq./kg shellfish meat is proposed, considering a protective default portion size of 400 g shellfish meat.

Emerging Optical Materials in Sensing and Discovery of Bioactive Compounds.

Optical biosensors are used in numerous applications and analytical fields. Advances in these sensor platforms offer high sensitivity, selectivity, miniaturization, and real-time analysis, among many other advantages. Research into bioactive natural products serves both to protect against potentially dangerous toxic compounds and to promote pharmacological innovation in drug discovery, as these compounds have unique chemical compositions that may be characterized by greater safety and efficacy. However, conventional methods for detecting these biomolecules have drawbacks, as they are time-consuming and expensive. As an alternative, optical biosensors offer a faster, simpler, and less expensive means of detecting various biomolecules of clinical interest. In this review, an overview of recent developments in optical biosensors for the detection and monitoring of aquatic biotoxins to prevent public health risks is first provided. In addition, the advantages and applicability of these biosensors in the field of drug discovery, including high-throughput screening, are discussed. The contribution of the investigated technological advances in the timely and sensitive detection of biotoxins while deciphering the pathways to discover bioactive compounds with great health-promoting prospects is envisaged to meet the increasing demands of healthcare systems.

Occurence and Variability of Domoic Acid in Mussel (Mytilus galloprovincialis) Samples from the Golden Horn Estuary, Sea of Marmara (Turkey).

The occurrence and variability of domoic acid (DA) levels in wild Mytilus galloprovincialis samples, compared with the Pseudo-nitzschia spp. abundance and particulate DA (pDA) concentrations in relation to the environmental changes in the Golden Horn Estuary, Turkey from October 2018 to September 2019. Biotoxin analysis were performed by high-performance liquid chromatography with diode-array detection (HPLC-DAD). DA concentrations in particulate matter (pDA) and mussel samples were found between 0.090-0.685 µg L-1 and 0.905-2.413 µg g-1, respectively. Accumulation of DA in wild mussel samples could be the result of the increasing tendency of P.nitzschia spp. abundances between April and May. Maximum DA levels were detected in particulate matter when the salinity was measured as the lowest in May. Thus, it can be said that the DA production was driven by the significant salinity decrease in the GHE. This is the first attempt regarding the presence of DA in M. galloprovincialis samples collected from Turkish coasts.

Natural Products as Potential Antiviral Drugs: The Specific Case of Marine Biotoxins.

To fight against various viral infections researchers turned to new chemical structures resulting from natural medicinal plants and more recently from "marine origin" as sources of active molecules against viral infections. The present manuscript describes complex marine origin drugs, their chemical complex structure, their therapeutic use, and their antiviral properties. Emphasis is placed more particularly on the properties of ionic channels (Na+, K+, Ca2+) blockers compounds from marine origin, named Dinotoxins, derived from "dinoflagellates microalgae". These compounds are of particular pharmaceutical interest since ionic channels blockers could be used to fight against a wide diversity of viruses, including SARS-CoV2 virus.

[Contamination status and the evaluation of dietary exposure of marine biotoxins in seafood in Ningbo City in 2017-2019].

OBJECTIVE: Ultra-fast liquid chromatography-tandem mass spectrometry method was developed for the analysis of contamination degree of biotoxins in seafood in Ningbo City from 2017 to 2019 and the assessment of dietary exposure was conducted. METHODS: Samples were extracted and purified with optimized pretreatment process and then injected for analysis. According to the result of the measurements, an international point estimate model was used to evaluate the dietary exposure of the population. RESULTS: For tetrodotoxin and 16 shellfish toxins monitored routinely, gonyautoxin5(GTX5), tetrodotoxin and homo-yessotoxin(hYTX) had higher detection rate, other toxins including okadaic acid(OA), dinophysistoxin1(DTX1), decarbamoyl gonyautoxin2(dcGTX2) and decarbamoyl gonyautoxin3(dcGTX3) were detected sporadically. The detection rates of TTX、GTX and hYTX were 27%, 52% and 12%, respectively. The concentration ranges of TTX, GTX and hYTX in polluted samples were 0. 003-0. 535, 0. 008-0. 189 and 0. 032-0. 110 mg/kg. The exposure risk indices(ERI) of TTX, GTX5, hYTX, dcGTX2 and dcGTX3 were 2. 5, 0. 026, 0. 0080, 0. 79 and 0. 32, respectively. CONCLUSION: Marine biotoxins have a lower dietary health risk to the population. It is must be given great attention that in the season of toxic red tide, the detection rates of higher toxic toxins, dcGTX2 and dcGTX3 increased significantly with high risks to human. Moreover, the dietary health risk of tetrodotoxin in routine surveillance in 2019 was higher.

[Determination of lipophilic marine biotoxins in shellfish foods by ultra performance liquid chromatography-hybrid triple quadrupole linear ion trap-mass spectrometry].

OBJECTIVE: An ultra performance liquid chromatography-hybrid triple quadrupole linear ion trap-mass spectrometry(UPLC-QqLIT-MS) was established for determination of lipophilic marine biotoxins in shellfish. And the 12 lipophilic marine biotoxins in shellfish were surveyed. METHODS: The lipophilic marine biotoxins in homogenized shellfish were ultrasonically extracted by methanol in super-sonic instrument, and cleaned up by solid phase extraction of Strata-X column, and eluted with methanol(containing 0.3% ammonia water). The elution was diluted with water, and cleaned by 0.22 µm millipore filter. The filtrate was separated on a Waters ACQUITY UPLC BEH C_(18) column(150 mm×2.1 mm, 1.7 µm)by gradient elution in 12 minutes with acetronitrile-water(containing 0.01%(V/V) ammonia and 2 mmol/L ammonium formate) as mobile phase, and detected by ultra performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS), identified by electrospray ionization(ESI) in simultaneous scanning mode of positive and negative ions using multiple reaction monitoring, and quantified with external standards. Information dependent acquisition scan function(IDA) combined with enhanced production scan(EPI) was used to confirm the 12 lipophilic marine biotoxins. RESULTS: The calibration curves of 12 lipophilic marine biotoxins showed good linearity in the range of 0.5-50 µg/L with correlation coefficients were 0.9984-0.9999.The detection limits of the method were 0.15-0.29 µg/kg. The recoveries of three spiking levels ranged from 80.0% to 116.0%, and the relative standard deviation(RSD) were 0.6%-6.4%(n=7). CONCLUSION: The method for determination of 12 lipophilic marine biotoxins in shellfish by UPLC-QqLIT-MS was of operation convenience, less interference from impurities and good accuracy, which could meet the requirements for the determination of 12 lipophilic marine biotoxins residues in sea foods.

l-Cysteine-modified magnetic microspheres for extraction and quantification of saxitoxin in rat plasma with liquid chromatography and tandem mass spectrometry.

Saxitoxin, which is one of the most typical paralytic shellfish poisoning toxins, ranks the highest intoxication rate of marine biological poisoning cases globally. Efficient clean-up and extraction of saxitoxin from complex biological matrices are imperative for the analysis and concentration monitoring of the toxin when correlative poisoning cases happen. Herein, l-cysteine-modified magnetic microspheres based on metal-organic coordination were synthesized by a facile approach and applied for magnetic solid-phase extraction of saxitoxin from rat plasma samples before liquid chromatography-tandem mass spectrometry detection. Parameters, including adsorbent amount, extraction time, desorption solution, and desorption time that could affect the extraction efficiency, were respectively investigated. The developed method demonstrated good linearity in the range of 5-300 ng/mL (R2  = 0.9985) with a limit of quantification of 5 ng/mL and a limit of detection of 0.5 ng/mL, acceptable accuracy. and precision of within-run and between-run.

Improved Isolation Procedures for Okadaic Acid Group Toxins from Shellfish (Mytilus edulis) and Microalgae (Prorocentrum lima).

Okadaic acid (OA) group toxins may accumulate in shellfish and can result in diarrhetic shellfish poisoning when consumed by humans, and are therefore regulated. Purified toxins are required for the production of certified reference materials used to accurately quantitate toxin levels in shellfish and water samples, and for other research purposes. An improved procedure was developed for the isolation of dinophysistoxin-2 (DTX2) from shellfish (M. edulis), reducing the number of purification steps from eight to five, thereby increasing recoveries to ~68%, compared to ~40% in a previously reported method, and a purity of >95%. Cell densities and toxin production were monitored in cultures of Prorocentrum lima, that produced OA, DTX1, and their esters, over ~1.5 years with maximum cell densities of ~70,000 cells mL-1 observed. Toxin accumulation progressively increased over the study period, to ~0.7 and 2.1 mg L-1 of OA and DTX1 (including their esters), respectively, providing information on appropriate harvesting times. A procedure for the purification of OA and DTX1 from the harvested biomass was developed employing four purification steps, with recoveries of ~76% and purities of >95% being achieved. Purities were confirmed by LC-HRMS, LC-UV, and NMR spectroscopy. Additional stability observations led to a better understanding of the chemistry of these toxins.