Two Kinds Of Toxic Red Tide Algae Growth And Production Conditions And Preparation Of Matrix Standard Materials

The outbreak of toxic red tide algae will destroy marine ecology,affect the development of marine fishery,and even cause public health events such as seafood poisoning.Alexandrium tamarense and Procentrum lima are the main toxogenic algae species of Paralytic Shellfish Poisoning and Diarrhetic Shellfish Poisoning.Paralytic Shellfish Poisoning and Diarrhetic Shellfish Poisoning have caused many cases of seafood poisoning.Because of the complex chemical structure of Paralytic Shellfish Poisoning and Diarrhetic Shellfish Poisoning,they can not be obtained by chemical synthesis.Therefore,It is of great significance to explore the conditions of the growth and toxin-producing of Alexandrium tamarense and Procentrum lima and prepare matrix standard materials of Paralytic Shellfish Poisoning and Diarrhetic Shellfish Poisoning.Firstly,we cultured Alexandrium tamarense and Procentrum lima under laboratory conditions.Then we studied the effect of nutrient concentration on the growth of two kinds of poisonous algae under different nitrogen and phosphorus concentration,followed the growth process and plotted the growth curve.Algal toxins were extracted and analyzed by LC-MS/MS.The experimental results can provide early warning and diagnosis index for the early warning of toxic red tide,as well as reference for the breeding of Alexandrium tamarense and Procentrum lima under laboratory conditions.The results showed that the growth conditions were more suitable when the nitrogen concentration was 80?mol/L and the phosphorus concentration was 4.2?mol/L;the nitrogen enrichment and phosphorus limitation both promoted the toxicity production of Alexandrium tamarense,which may be due to the imbalance of nitrogen-phosphorus ratio;and the nitrogen limitation promoted the toxicity production of Lima Prochaemia,but the phosphorus concentration had little effect on the toxicity production.Secondly,we optimized a liquid chromatography tandem mass spectrometry(LC-MS/MS)method to detect a total of 14 shellfish toxins(3 kinds of representative Diarrhetic Shellfish Poisoning toxins and 11 kinds of Paralytic Shellfish Poisoning toxins).Paralytic Shellfish Poisoning toxins include Saxitoxin(STX),G onyautoxin-1&4(GTX1&4),Neosaxitoxin(NEO),Gonyautoxin-5&6(GTX5&6),N-sul focarbamoyl toxins-1&2(C1&2),decarbamoylgonyautoxin-2&3(dcGTX2&3),decar-bamoylsaxitoxin(dcSTX)and Okadaic acid(OA),Dinophysistoxin-1&(DTX1&2).The optimization of detection methods not only provide a basis for the detectio nof toxin components in algae,but also provide a guarantee for the detection ofmatrix standard materials.Then,the scallops were fed with Alexandrium tamarense and Procentrum lima respectively.Homogenize the meat of scallop scallop,detect and analyze the accumulation of shellfish toxin and toxin types in shellfish by LC-MS/MS,study the accumulation and excretion of shellfish toxins in scallops and make a health risk assessment.The study found that vixcera was the main part for accumulation in shellfish;by the seventh day after feeding the low-toxicity Alexandrium tamarense and by the fourth day of feeding with Procentrum lima,the toxin in the shellfish exceeded the safty limit value.Finally,through the symbiosis of algae and shellfish,the bivalve is poisoned.The poisoned shellfish meat was homogenized and made into lyophilized powder type matrix reference material,and carry out the homogeneity and stability testing.Homogeneity testing,short time stability and 6-months stability results show that two kind of matrix reference material is homogeneous and stable.This experiment proved that it is feasible for shellfish to be poisoned naturally and made matrix reference material under laboratory conditions,and also provides the basis for the application of national secondary standard of Paralytic Shellfish Poisoning and Diarrhetic Shellfish Poisoning in the future.