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Journal of the Korean Chemical Society (JKCS)

ISSN 1017-2548(Print)
ISSN 2234-8530(Online)
Volume 45, Number 2
JKCSEZ 45(2)
April 20, 2001 

Study on New Extraction Method of Microcystins from Cyanobacteria

남조류로부터 마이크로시스틴을 추출하는 새로운 추출법 연구
Dong-Jin Pyo, Hyun-Du Shin

표동진, 신현두
남조류독소, 마이크로시스틴을 초임계유체추출을 이용하여 추출하는 새로운 분석법이 개발되었다. 이 연구에서 사용된 마이크로시스틴들은 초임계유체 CO2에는 거의 녹지 않지만 90% CO2, 9.0% 메탄올, 1.0%물로 된 혼합유체를 사용한 결과 성공적으로 추출할 수 있었다, 이 연구에서 사용된 초임계유체 추출법은 고체상 추출법보다 많은 장점들을 가지고 있다. 우선 시료처리 단계가 줄어들고, 시료의 손실이 적으로, 분석시간을 단축시킬수 있다는 장점이 있다, 또한 전처리 단계가 생략된다는 장점도 있다. ODS 카트리지를 사용하는 고체상 추출법에서 ODS 카트리지가 마이크로시스틴에 대한 흡착력이 떨어지기 대문에 좀 더 극성이 있는 CN 카트리지를 대신 사용하였다, 동결건조된 세포(100mg)을 5% 아세트산으로 추출하였고, 추출물을 원심분리하여 CN 카트리지에 주입 시켰다. CN 카트리지는 물 5mL, 0.5M 아세트산 5mL, 5% 아세트니트릴 5mL로 빗은 후 70% 아세트니트릴로 최종용리 시킨후 HPLC로 분리하였다.CN 카트리지를 사용하였을 때 ODS 카트리지 보다 더 좋은 회수율과 크로마토그램을 보여 주었다.

A new analytical method of cyanobacterial toxins, i.e. microcystins was developed using supercritical fluid extraction(SFE). The microcystins included in this study are sparsely soluble in neat supercritical fluid CO2. However, the microcystins were successfully extracted with a ternary mixture(90% CO2, 9.0% methanol, 1.0% water) at 40 ℃ and 250 atm. The SFE method developed in this study has several advantages over solid-phase extraction(SPE) sample preparation for the analysis of microcystins. Sample handling steps are minimized, thus reducing possible losses of analytes and saving analysis time. No clean-up steps are employed in this SFE method. Although many methods have been described for microcystins RR and LR, the method using solid-phase extraction with ODS cartridges is the most commonly used. However, the adsorbing power of ODS cartridges for microcystins is weak, so we have attempted to use a more polar CN cartridge, to increase the adsorbing power for microcystins. Lyophilized cells(100 mg) were extracted with 5%(v/v) acetic acid. The extract was centrifuged and then the supernatant was applied to a CN cartridge. The cartridge which contained microcystins was rinsed with 5 ml of water and 5 ml of 0.5 M acetic acid, followed by 5 ml of 5% acetonitrile in water. Microcystins were finally eluted from the CN cartridge with 70% acetonitrile in water, and were determined by HPLC. Better recoveries and chromatograms were observed than with ODS cartridge.

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