• Journal of environmental and Sanitary engineering
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• v.23 no.4
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• pp.37-44
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• 2008

In this study we developed the analytical methods for the determination of three neurotoxin; anatoxin-a, saxitoxin and neosaxitoxin using HPLC/FLD system and this analytical methods were applied to real sample; algae culture and algae extracts. For the HPLC/FLD analysis of anatoxin-a samples were concentrated on WCX(Weak Cation Exchanger) SPE and then anatoxin-a in concentrate was derivatized with NBD-F solution. Supernatant was injected on HPLC system. For the HPLC/FLD analysis of saxitoxin and neosaxitoxin samples were separated on the column and then derivatizied by post column reactor for fluorescen detection. For post column reaction of saxitoxin we feed two kinds of reaction solution; Oxidizing Reagent of which composition was periodic acid(7mM) in 50mM potassium phosphate buffer, pH 9 and acidifying reagent of which Composition was 0.5M acetic acid. The LOD value for anatoxin-a, saxitoxin and neosaxitoxin in HPLC/FLD method was 24.3 ng. $35{\mu}g/L$, $27{\mu}g/L$ respectively. We determined the anatoxin-a content of lyophilized anabaena flos-aquae and $20{\mu}g/g$ d.w. of anatoxin-a was detected. We analyzed saxitoxin and neosaxitoxin in algae culture media and extracts of lypopyllized algal cell cultured and that of Deachung reservior. Saxitoxin and neosaxitoxin in real sample were below the limit of detection. Although there are various water treatment processes for removing neurotoxins were suggested no process give simultaneous and complete removal of neurotoxins. It was cocluded that nanofiltration which reject material by size can be a process for removal of neurotoxins.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.25 no.2
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• pp.144-150
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• 1992

Paralytic shellfish toxins in mussels Mytilus edulis and dinoflagellate Alexandrium tamarene from Jinhae Bay, south coast of Korea were investigated. The mussels collected in March-April, 1989 showed toxicities of 7.5 MU/g of whole meat(31-88 MU/g of the digestive gland) , and those collected in 1990 showed toxicity level of 1.9-9.9 MU/g of whole meat by the standard mouse bioassay. Analysis of toxins by high performance liquid chromatography revealed the presence of gonyautoxin 1-4$(48-76\%)$ gonyautoxin 8 and epi-gonyautoxin $8(C1-C2,\;14-39\%)$, saxitoxin$(1-10\%)$, neosaxitoxin$(l-7\%)$ and trace amount of decarbamoylgonyautoxin 2 and 3(dcGTX2, dcGTX3) in the mussels of 1989. While, Mussels collected in 1990 contained a significantly larger proportion of neosaxitoxin $(44-50\%)$ than did those of 1989. A. tamarense isolated in April 1989 produced the same toxins in culture with slightly higher proportion of Cl, C2, dcGTX2 and dcGTX3 than in the mussels. The difference was within a range of toxin change during accumulation by shellfish and during sample preparation for analysis. It was thus concluded that the dinoflagellate was the cause of toxins in the mussels.

• Fisheries and Aquatic Sciences
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• v.2 no.2
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• pp.155-160
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• 1999

Studies on detoxification of Paralytic Shellfish Poison (PSP)-infested oyster, Crassostrea gigas were carried out using available processing resources. Changes of paralytic shellfish toxin components and specific toxicity during canning process were also investigated with high performance liquid chromatography (HPLC). Toxic oysters collected at Hachong in Koje Bay were used for experimental samples. The toxicity of oysters with range of 185-778 ug/100g was reduced below the quarantine limit of 80 ug/100g or not detected level by the mouse bioassay after canning process. The mole $\%$ of toxin components in the shucked oyster was in the order of 25.1 mole $\%$ of gonyautoxin 1, 19.2 mole $\%$ of gonyautoxin 3, 17.2 mole $\%$ of gonyautoxin 4 and 14.6 mole $\%$ of gonyautoxin 2. This sample had tracing amounts of Cl, C2, saxitoxin and neosaxitoxin. In the case of specific toxicity, the major toxins were consisted of gonyautoxin 1-4. The sum of gonyautoxin 1, 2, 3 and 4 was 80% of total toxicity of oyster. Saxitoxin and decarbamoylsaxitoxin were the more thermostable than any other toxin components.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.29 no.6
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• pp.900-908
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• 1996

Changes of paralytic shellfish toxin components and specific toxicity in blue mussel, Mytilus edu/is and oyster, Crassostrea gigas during canning process were investigated by high performance liquid chromatography (HPLC). The $mole\%$ of the frozen shucked blue mussel were in order of $27.5\;mole\%$ of gonyautoxin 1, $23.0\;mole\%$ of gonyautoxin 8 (C1) and $23.0\;mole\%$ of epi-gonyautoxin 8 (C2), while those of the frozen shucked oyster were in order of $29\;mole\%$ of C1, $22\;mole\%$ of C2, $16.7\;mole\%$ of gonyautoxin 2. Both samples had minor amounts of saxitoxin and neosaxitoxin. On the other hand, in case of specific toxicity, the major toxins were consisted of gonyautoxin $1\~4$ in both sample. The toxicity of gonyautoxin $1\~4$ were 88 and $84\%$ in blue mussel and oyster, respectively. According to the experimental results, C1, C2 and gonyautoxin 4 were very sensitive to heat treatment, while gonyautoxin 2 and saxitoxin were pretty heat resistant than any other toxin components.

• Journal of Food Hygiene and Safety
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• v.28 no.2
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• pp.188-193
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• 2013

Anatoxin-a, saxitoxin and neosaxitoxin are produced by Aphanizomenon flos-aquae that is a sort of the cyanobacteria phylum. Therefore, it is not permitted for food materials in Korea. Traditionally, the classification of cyanobacteria has been based on morphological characters such as trichome width, cell size, division planes, shape, and the presence of character such as gas vacuole. But, some diagnostic features, such as gas vacuole or akinetes, can show variation with different environmental or growth conditions and even be lost during cultivation. Therefore, we developed detection method for functional foods containing Aph. flos-aquae by PCR. To design the primer, 16S rRNA region of Aph. flos-aquae, Spirulina laxissima, and Spirulina spp. registered in the GeneBank (www.ncbi.nlm.nih.gov) have been used and for comparative analysis, BioEdit ver. 7.0.9.0. was used. As a result, we was design AFA-F1/AFA-R1 (363 bp) primer for the differentiation Aph. flos-aquae from chlorella, spirulina, green tea, and spinach. Also, it could be distinguished chlorella and spirulina products those are made to contain 1% Aph. flos-aquae.

• Journal of Korean Society on Water Environment
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• v.25 no.4
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• pp.597-605
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• 2009

Algae bloom occurred in reservoir in summer can cause taste and odor in water and disturb the flocculation and sedimentation processes in water treatment plant and cause sand filter plugging. It was also reported that microcystins, anatoxin and saxitoxin released from cyanobacteria had acute toxic effects on liver and nervous system. For these reasons, many advanced countries inclusive of WHO set the guideline for these toxins and cyanotoxins have been managed with regular monitoring in Korea as well. However, complex sample preparation steps such as a solid phase extraction (SPE) and derivatization are required with an existing analysis method with HPLC. We needed to improve an analysis method for low extraction efficiency and long sample preparation time. In this study, we have established a new LC/MS/MS method which can simultaneously determine 6 cyanotoxins (Microcystins-LR, Microcystins-RR, Microcystins-YR, Anatoxin-a, Saxitoxin, Neosaxitoxin) with only simple filtration step. When $75{\mu}L$ filterated sample was injected onto the LC-MS/MS, the recovery ranged from 86% to 112% and the MDL was $0.025{\sim}0.581{\mu}g/L$. We can make the MDL be lower than the guideline ($1{\sim}3{\mu}g/L$) of advanced countries with simple preparation.

• Fisheries and Aquatic Sciences
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• v.16 no.3
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• pp.159-164
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• 2013

The mouse bioassay and high performance liquid chromatography (HPLC) post-column oxidation method are different methods of quantifying paralytic shellfish poisoning toxins. In this study, we compared their ability to accurately quantify the toxicity levels in two types of field sample (oysters and mussels) with different toxin profiles for routine regulatory monitoring. A total of 72 samples were analyzed by both methods, 44 of which gave negative results, with readings under the limit of detection of the mouse bioassay ($40{\mu}g/100g$ saxitoxin [STX] eq). In 14 oysters, the major toxin components were gonyautoxin (GTX) 1, -2, -3, -4, -5, decarbamoylgonyautoxin-2 (dcGTX2), and decarbamoylsaxitoxin (dcSTX), while 14 mussels tested positive for dcSTX, GTX2, -3, -4, -5, dcGTX2, neosaxitoxin (NEO), STX, and dcSTX. When the results obtained by both methods were compared in two matrices, a better correlation ($r^2=0.9478$) was obtained for mussels than for oysters ($r^2=0.8244$). Additional studies are therefore needed in oysters to investigate the differences in the results obtained by both methods. Importantly, some samples with toxin levels around the legal limit gave inconsistent results using HPLC-based techniques, which could have a strong economic impact due to enforced harvest area closure. It should therefore be determined if all paralytic shellfish poisoning toxins can be quantified accurately by HPLC, and if the uncertainties of the method lead to doubts regarding regulatory limits.

• Journal of Food Hygiene and Safety
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• v.15 no.4
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• pp.293-296
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• 2000

Paralytic shellfish poison (PSP) occurrence and variation in th\ulcorner different bivalve species including oyster,Crassostrea gigas and mussel, Mytilus edulis Jinhae bay Korea from January to December in 1997 were surveyed. And also compositional characteristics of PSP ingredients in the different bivalve species were investigated. PSP in shellfish was detected from late February and increased continuously until the middle of April in Jinhae bay. And after April PSP level had been decreased gradually and the toxicity was not detected by mouse bioassay in the early of June. Of the examined bivalve species, PSP content in the mussel exhibited the highest value and the PSP content in the mussel in the middle of April, PSP high season in Jinhae bay, was 6 times higher than that in the oyster. Gonyautokin (GTX) 1~4 group occupied 59.0~78.8% of whole PSP contents and identified as dominant ingredient in the examined bivalves except oyster. And it is also identified that the PSP toxicity in the tested species were derived from the GTX group. And the dominant ingredient of PSP in the oyster was carbamoyl-N-sulfo-11$\alpha$-hydroxysaxitoxin sulfate(Cl)(37.9%) and neosaxitoxin(neoSTX)(26.2%). But the toxicity of Cl in the tested oyster could be ignorable and most toxicity (80.0%) was derived from saxitoxin (STX) group.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.20 no.4
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• pp.293-299
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• 1987

At various times and places all over the world men have become ill and some have died after eating shellfish that were intoxicated with paralytic shellfish poison(PSP) caused by Protogonyaulax spp. In late March, 1986, two persons were dead by ingesting wild sea mussels, Mytilur edulis, grown at bottom of an anchored waste ship to be dismantled at Gamchun Bay, Pusan, Korea. The samples were collected from the bottom of the ship during April $1\~April$ 8 of the year to find the cause of the food poisoning accident. The toxicity was estimated by bioassay with ICR male mouse, while the toxins were extracted and characterized. The toxins were extracted with acidified $80\%$ ethanol. The extract was defatted three times with dichloromethane, treated with activated charcoal, and then purified by chromatography on Bio-Gel P-2 and Bio-Rex 70. The toxic fractions obtained were analysed by cellulose acetate membrane electrophoresis, thin layer chromatography and high performance liquid chromatogaphy. The range and the average of PSP-toxicity of the samples were $132\~295\;MU/g$, 203 MU/g respectively. The amount of PSP was $26.4\~58.9{\mu}g/g$ of whole meat in range and $40.6{\mu}g/g$ in average. The toxicity of the digestive gland of the samples was 9 times higher than that of edible meat (except digestive gland) as $439\~979MU/g$, and it was about $70\%$ in total toxin. The compositional analytical results of the paralytic shellfish toxin, Gonyautoxin $1\~4$ were the major part of the PSP and Saxitoxin and neosaxitoxin were detected as the minor component. It was concluded that the food poisoning accident was caused not by Saxitoxins but by Gonyautoxins.