The wormicidal substance of fresh water fishes on Clonorchis sinensis V. Purification and chemical characterization of <i>clonorchicidal substance</i> from epidermal mucus of <i>Cyprinus carpio</i>

, Rhee, Jae Ku; , Lee, Sang Bork; , Ahn, Byung Zun

doi:1984.22.1.127

Korean J Parasito > Volume 22(1):1984 > Article

Original Article


Korean J Parasitol. 1984 Jun;22(1):127-134. English. Published online Mar 20, 1994. http://dx.doi.org/10.3347/kjp.1984.22.1.127
Copyright © 1984 by The Korean Society for Parasitology


The wormicidal substance of fresh water fishes on Clonorchis sinensis V. Purification and chemical characterization of clonorchicidal substance from epidermal mucus of Cyprinus carpio
Jae Ku Rhee,Sang Bork Lee and Byung Zun Ahn
Department of Veterinary Medicine, Jeonbug National University, Jeonju 520, Korea.
Department of Pharmacy, Chungnam National University, Daejeon 300, Korea.


Abstract
As a series of studies to clarify clonorchicidal substances in body surface mucus of some fresh-water fishes, the substance in the epidermal mucus of Cyprinus carpio was isolated by silica gel column and thin layer chromatography and analyzed for its chemical nature. Wormicidal trial was done in vitro, and the results obtained are summarized as follows: The mucus was extracted by ethyl ether and separated into 4 fractions by column chromatography using benzene as solvent. The second fraction with yellowish red colour among them showed the strongest clonorchicidal effect. The yellowish red fraction obtained by column chromatography was then fractionated into 6 spots by thin layer chromatography with petrol. ether/chloroform(30/70, v/v), and the Rf. 0.714 spot among the 6 spots showed the strongest effect. The Rf. 0.714 spot was further fractionated into 6 spots by thin layer chromatography with benzene/acetone (90/10, v/v), and the Rf. 0.800 spot among the later 6 spots revealed the strongest effect. The Rf. 0.800 spot was chromatographed on column with benzene and 2 fractions were obtained. The second fraction of light brown colour represented the final purified fraction. By these purification procedures, clonorchicidal substance was purified 15-fold with 0.03 percent yield from the mucus of C. carpio, and 10mg of the final fraction killed the cercaria in 26 min, the metacercaria in 115 min, and the adult in 443 min. Infra red and nuclear magnetic resonance spectrometric analysis of the purified substance revealed that the substance belongs to an ethyl ester of unsaturated fatty acid with 2 double bonds, 15 methylene groups and l methyl group.

Figures


	Fig. 1 Fractionation method of clonorchicidal substance from Cyprinus carpio *; Clonorchicidal activity


	Fig. 2 IR-spectrum of the final colnorchicidal substance of Cyprimus crpio


	Fig. 3 NMR spectrum of the final clonorchicidal substance from Cyprimus crpio

Tables


	Table 1 Time required form killing cercaria in the second step fractions from epidermal mucus Solvent; Benzene


	Table 2 Time required form killing cercaria in the third step spots fromepidermal mucus Solvent; Pet. ether/Chloroform: 30/70


	Table 3 Time required form killing Clonorchis in the final fraction from epidermal mucus


	Table 4 Purification of clonorchicidal substance from Cyprinus carpio

References


1.	Rhee JK. [Experimental studies on the second intermediate hosts of Clonorchis sinensis: III. Observations on the relationship between clavate cells of epidermis and infectivity of metacercariae of Clonorchis sinensis in fresh-water fish]. Korean J Parasitol 1974;12(2):101–110.

2.	Rhee JK, Baek BK, Ahn BZ, Park YJ. [The Wormicidal Substances Of Fresh Water Fishes On Clonorchis Sinensis: I Preliminary Research On The Wormicidal Substance From Mucous Substance Of Carassius Carassius]. Korean J Parasitol 1979;17(2):121–126.

3.	Rhee JK, Baek BK, Ahn BZ, Park YJ. [The Wormicidal Substances Of Fresh Water Fishes On Clonorchis Sinensis: II. Preliminary Research On The Wormicidal Substances From Mucous Substances Of Various Fresh Water Fishes]. Korean J Parasitol 1980;18(1):98–104.

4.	Rhee JK, Baek BK, Ahn BZ, Park YJ. [The Wormicidal Substances Of Fresh Water Fishes On Clonorchis Sinensis: III. Seasonal variation on the wormicidal substances of mucous substances of Carassius carassius]. Korean J Parasitol 1980;18(2):179–184.

5.	Rhee JK, Kim PG, Baek BK, Lee SB, Ahn BZ. Clavate Cells Of Epidermis In Cyprinus Carpio Nudus With Reference To Its Defence Activity To Clonorchis Sinensis. Korean J Parasitol 1982;20(2):201–203.

6.	Rhee JK, Kim PG, Baek BK, Lee SB, Ahn BZ. [Experimental Infection Of Clonorchis Sinensis To Cyprinus Carpio Nudus]. Korean J Parasitol 1983;21(1):11–19.

7.	Rhee JK, Kim PG, Baek BK, Lee SB, Ahn BZ. [The Wormicidal Substance Of Fresh Water Fishes On Clonorchis Sinensis: Iv. Preliminary Research On The Wormicidal Substance From Mucus Of Cyprinus Carpio Nudus]. Korean J Parasitol 1983;21(1):21–26.

8.	Rhee JK, Baek BK, Lee SB, Koh HB. [Epidemiological Studies Of Clonorchis Sinensis In Mangyeong Riverside Areas In Korea]. Korean J Parasitol 1983;21(2):157–166.