Toxicity Physiology And Food Safety Evaluation Of An Azaspiracid Toxin Producing Algae

The eutrophication of marine water bodies leads to frequent outbreaks of red tides.The toxins produced by harmful red tide algae are rich in shellfish toxins in shellfish,which seriously endangers the health of consumers and hinders the development of foreign trade in aquatic products,also has been one of the bottlenecks problem of sustainable development.In this paper,Q-Exactive high-resolution mass and spectrometry(GC-MS)for the preparation of various AZAs toxins was established by using the novel shellfish toxin tetracycline toxins as a research object.At the same time,Toxin-producing algae A.poporum(AZDY06)isolated from China was cultured and exposed to Chlamys farrerii,for the evaluation of azaspiracids(AZAs)accumulation,distribution and biotransformation in scallop.Components of AZA and responses of antioxidant enzymes were monitored in shellfishes fed with AZA-producing algae in laboratory.Meanwhile the mixtures of AZA and digestive gland tissues of shellfish were incubated in order to explore the metabolic pathways of AZA in shellfish and its influencing factors.Some original data and theoretical basis will be accumulated for developing rapid detoxification technology and antidote for AZA toxins.It is very significant to support the sustainable development of shellfish aquaculture and to protect consumer health.The main contents of the paper are as follows:(1)The Q-Exactive high-resolution mass spectrometry was used to establish a high-resolution non-directional qualitative screening and low-resolution target detection method for AZA toxin on the basis of liquid chromatography-tandem mass spectrometry.The samples were analyzed comprehensively with this method,there are four metabolites(AZA6,AZA12,AZA19 and AZA23)produced by the accumulationof Ampelite in the shellfish,and the real positive samples were analyzed by liquid chromatography-tandem mass spectrometry.The results showed that the method Quantitative test to achieve the same time accurate and accurate quantitative requirements for Azaspiracid toxins metabolic substances in the screening analysis,in order to further improve the aquatic products in China shellfish toxins monitoring system to provide reliable work Basic and technical support.(2)Chlamys farreri was exposed to three kinds of algae in different biomass models to simulate the early,middle and late marine environmental processes during the red tide burst.By comparing the toxin components,the accumulation of toxins in the tissues and organs and the specificity of metabolic transformation resistance,acid diazaspiro process of toxin(Azaspiracid,AZAs)in vivo scallop Chlamys hole formed hazards.The results showed that AZA2 is the dominant toxin of AZDY06 which distributed in the South China Sea,toxin production capacity up to 7.05±0.52 fg/cell;low ration toxic algae biomass production in vivo with a total of three groups scallop metabolites(AZA6,AZA12,AZA19)occurs,high biomass and biomass group of four metabolites(AZA6,AZA12,AZA19 and AZA23),indicating that the size of the biomass of algae-infected algae affects the transformation process of metabolite components.The changes of biomass and low biomass in the exposure stage are similar,The trend of rapid increase in the stage,the highest point followed by the overall decline,while the high biomass group in the accumulation stage showed a rapid upward trend in the early accumulation has reached a peak after a sharp downward trend,then slowly increased until after the end of exposure,in contrast,the experimental group of AZAs ability to accumulate toxins in descending order:biomass group> high biomass group> low biomass group,wherein the biomass The strongest ability to accumulate.Preliminary experimental data to explore the AZA2 metabolic conversion mechanisms in the body of the scallop,lay the foundation for subsequent experiments.(3)Previous experiments showed that the high biomass production of poisonous algae exposure experiment will increase the composition of the metabolites and the content of each component,but it will also increase the toxic effect on the scallop,will reduce the scallop of the snail The accumulation capacity of the cyclic toxins,the longer the exposure time,the weaker the accumulation and metabolism of the scallops to toxins.Chlamys farreri is exposed directly to higher biomass algae,while reducing exposure time.The results showed that Chlamys farreri had strong feeding ability and AZAs accumulation ability,After 12 h exposure experiments with5×10~7 cells AZDY06 ingested by every C.farrerii,AZAs in mussels reached highest concentration and exceeds EU regulatory limit,the concentration was 165.3 ?g AZA1eq/kg,while total accumulation efficiency reached 78.2%.The distribution of AZA toxins showed significant differences between organizations,gill>viscera>pallium>muscle.Four AZA analogues including AZA19,AZA12,AZA6 and AZA23 were found in all organizations.AZA19 was the main metabolite with a proportion of 40%,other metabolites content is rarely.This study proved the hazard of A.poporum(AZDY06)isolated from China is strong and underlined that AZA19 were present at significant levels already after 6 h and until the end of the study.Therefore,these analogue need to be considered as major metabolite which affect human health,It is highly recommended to speed up the formulation of AZA limits.(4)Exposed to oxidative reductase in the antioxidant defense system in viscera and gill tissues were activated at the beginning of the exposure experiment,MDA content increased,lipid peroxidation,corresponding GSH-PX and POD activities were enhanced,granulocyte secretion of ACP,POX activity increased,NADPH content decreased,some NADPH may be involved in the biotransformation of toxins.The effect of integrated AZAs on the ultrastructure of viscera and gill tissue of Chlamys farreri and the activation of oxidoreductase in tissue can provide a theoretical basis for the establishment of histological toxicological indexes of shellfish under AZAs stress.The results showed that the ultrastructures of viscera and gill tissues in the scallop of Chlamys farreri were all changed during the exposure experiment.The intestinal epithelial cells were vacuolated,the nucleus atrophy was deformed,and the necrosis was severe The mitochondria and lysosomes increased and aggregated in the columnar epithelium of the gills,and the mucosal granules were released from the mucous cells.The changes of oxidoreductase in the antioxidant defense system of viscera and gill tissues were examined by exposure test.The toxicological effects of AZAs on the ultrastructure of viscera and gill tissues of Chlamys farreri were also studied.The effects of AZAs on the internal organs of Chlamys farreri And the toxicological stress of gill tissue.