Micronucleus Test of Wild Ginseng Culture Extract Using the Marrow Cells in ICR Mice

Transcription

1 J. Fd Hyg. Safety 20(1), (2005) ICR s w n w x y*á **Á *** * $IFNPO** w w ƒœ l*** w w w yw Micronucleus Test of Wild Ginseng Culture Extract Using the Marrow Cells in ICR Mice Si-Whan Song*, Deok Chun Yang**, and Se Young Choung*** *$IFNPO*OD+FJMSJ:BOHKJNZVO:POHJOTJ(ZFOHHJEP,PSFB **0SJFOUBM.FEJDJOF.BUFSJBMBOE1SPDFTTJOH$FOUFS,ZVOH)FF6OJWFSTJUZ:POHJO,PSFB ***%FQBSUNFOUPG)ZHFOJD$IFNJTUSZ$PMMFHFPG1IBSNBDZ,ZVOH)FF6OJWFSTJUZ4FPVM,PSFB ABSTRACT To assess clastogenic effects of the wild ginseng culture extract (WGCE) in vivo micronucleus test was performed using 7 weeks old ICR mice. At 24 hours after 2nd treatment with wild ginseng culture extract at the doses of 0, 500, 1,000, and 2,000 mg/kg/day by peritoneal route mice were sacrified and marrow cells were prepared for smear slides. As a result of counting the micronucleated polychromatic erythrocyte (MNPCE) of 2,000 polychromatic erythrocyte(pce), all treatment groups did not show statistically significant increase than negative control group. And there was no clinical sign connected with injection of wild ginseng culture extract. It was concluded that wild ginseng culture extract did not induce micronucleus in the marrow cells of ICR mice. Key words: wild ginseng culture extract, micronucleus, polychromate erythrocyte, marrow cell w wì w š, w» w t w t ƒw t w y w š l w mw,, x,, y ƒ š w w» w g š w 1) 2) wì in vitro in vivo x mw y z ƒ, g 3) l w z 4) z 5)ƒ š ù z ƒ š l w š ù,» { p j w w t y ƒ w z 6 yw w» { š, ³ y (») ùkú w y w vw ª» t/ y Ýw w Author to whom correspondence should be addressed. s p š w y w p ginsenoside t,» t y» ù x ¾» t ƒ y y w t w w w ƒ s in vivo mw y wš ù ƒ» t œ» w x w w(micronucleus, MN) x x ü wš 24 z w w w in vivo x wù w ü erythroblastƒ w z kw x x (PCE, polychromatic erythrocyte)ƒ š PCE 10 z x ƒ x ƒ w erythrocyte k w ƒ Á yw wš qr û PCE ü 58

2 .JDSPOVDMFVT5FTUPG8JME(JOTFOH$VMUVSF&YUSBDU6TJOHUIF.BSSPX$FMMTJO*$3.JDF 59 w w w 6) w w e ICR s w t w w wš t t š y x» y t x» ww l w 4 w z w 3z wš 45 o C» 12 w 1 kg w L(1:10) ƒwš 85 C o 4 wš z œ» w brix 60 w w, þ w x w x w x x» ƒ t w x ICR m p ³ (SPF) kw œ w j g sww 7 y» w z, w x w x w, ww ü 23±3 C o w, 55±15%, y» z 10~20z/hr, 12, 150~300 Lux w x x w z e w ³ š ƒ ww w n w x w w x ³»w» w xk q w š x ³ x n w w (Table 1). x n 7 x 0, 500, 1,000 2,000 Table 1. s and route for treatment groups Treatment Groups Animals per dose Animal No. Volume (ml/kg) Route Vehicle I.P. WGCE (G1) I.P. WGCE (G2) , I.P. WGCE (G3) , I.P. CPA I.P. CPA : cyclophosphamide H 2 O (Positive control item) mg/kg 1 1z 2 ü n wš, cyclophosphamide monohydrate(cpa) x 2z n 1z n w n l 24 z s w w s sƒw ƒ l w n l 23 e, 3 ml FBS( k x, GIBCOBRL # ) ü xk k 100 rpm 5 w w z e s, w z p g 5 e w s š w š ƒ óù May- Grunwald 3, May-Grunwald 1:1 ( ) 2, Giemsa (5% in PBS, ph 6.8) w 1,000 w 7) w ƒ n l w kƒ yw 1 kw g yw z w x 2,000 PCE ùkù MNPCE w meanûs.d. t w w w s 1/5-1/20 j» w s w w ùkü x-k x w w, w w w z w w 500 PCE normochromatic erythrocyte(nce) w PCE/(PCE+NCE), s t x n w w n ƒ 1z w z w

3 60 4J8IBO4POHFUBM m q SAS m 8~10) w m wš, P<0.05 m w w q w w PCE/(PCE+NCE) l(x) arcsin[sqrt(x)] y mw ³ ƒ w. w ³ w, Kruskal-Wallis test wš w PCE/(PCE+NCE) ³, e w, w PCE/(PCE+NCE) 0.1 x Table 2. Results of dose-range finding study (Male) Chemical Treated Vehicle 0 x 2 WGCE 125 x 2 WGCE 250 x 2 WGCE 500 x 2 WGCE 1,000 x 2 WGCE 2,000 x 2 Animal No. k w wš11,12) MNPCE ƒ m w w ƒw ù wù x ùkú q w, x w ANOVA's test w š w w š y x w x ƒƒ 24 ICR f f x Body weights (g) at the time of 1st Admin. 2nd Admin. 3rd Admin. Sacrifice Mean ± S.D ± ± ± ± Mean ± S.D 33.01± ± ± ± Mean ± S.D 32.90± ± ± ± Mean ± S.D 33.69± ± ± ± Mean ± S.D 32.68± ± ± ± Mean ± S.D 32.54± ± ± ±1.686

4 .JDSPOVDMFVT5FTUPG8JME(JOTFOH$VMUVSF&YUSBDU6TJOHUIF.BSSPX$FMMTJO*$3.JDF 61 ü n w z y w, x ùkû (Table 2, 3). x n w z ¾ x n w q p w x n s d w ƒ w (Table 4), š m w w ùkü (P<0.05). w s x sƒwš s w w x w 7 x 1 1z 2 ü n wš, n l 24 z s w w s sƒw x w ü 2,000 PCE w w, 2,000 PCE w ƒ PCE 1z n», 500 mg/kg n (G1), 1,000 mg/kg n (G2), 2,000 mg/kg n (G3) Table 3. Results of dose-range finding study (Female) Vehicle 0 x 2 WGCE 125 x 2 WGCE 250 x 2 WGCE 500 x 2 WGCE 1,000 2 WGCE 2,000 2 Body weights(g) at the time of 1st Admin. 2nd Admin. 3rd Admin. Sacrifice Mean ± S.D ± ± ± ± Mean ± S.D 25.65± ± ± ± Mean ± S.D 25.24± ± ± ± Mean ± S.D 26.30± ± ± ± Mean ± S.D 26.15± ± ± ± Mean ± S.D 25.59± ± ± ±0.972

5 62 4J8IBO4POHFUBM Table 4. Body weights of ICR mice. Chemical Treatment Animals per dose Body weights (kg)* at the time of 1st Admin. 2nd Admin. Sacrifice Vehicle ± ± ±0.864 WGCE (G1) ± ± ±1.085 WGCE (G2) 1, ± ± ±0.825 WGCE (G3) 2, ± ± ±0.841** CPA ± ± ±1.353 Cyclophosphamide?H2O (positive control) administered once on the day of 2nd Admin. *Mean±S.D. g **Significantly different from the vehicle control group at P<0.05. Table 5. Micronucleus test of WGCE in male ICR mice. Chemical Treatment Animals per dose MNPCE/ 2000 PCE (Mean±S.D.) PCE/ (PCE+NCE) (Mean±S.D.) Vehicle ± ± 0.05 WGCE (G1) ± ± 0.07 WGCE (G2) 1, ± ± 0.06 WGCE (G3) 2, ± ± 0.05 CPA ± 11.64* 0.38 ± 0.06 MNPCE : PCE with one or more micronuclei PCE : polychromatic erythrocyte NCE : normochromatic erythrocyte Cyclophosphamide?H2O (Positive control item) administered once on the day of 2nd Admin. *Significantly different from the vehicle control group at P<0.01. s³ 5.17, 5.50, (Table 5)., x n w n w m w w ƒ ùkü wr w w m w w x w ƒƒ ùkû (P<0.01). s t PCE/(PCE+NCE), 500 mg/kg n (G1), 1,000 mg/kg n (G2), 2,000 mg/kg n (G3) ƒƒ s³ 0.36, 0.32, 0.33, ùkü (Table 5). x n w s³ 0.25 š, x n w s eƒ w w ùkû ù x m w w ùkù x w s w w x sƒ w f ICR s w w x w 1 z n š x w 7 f x 0, 500, 1,000 2,000 mg/kg 1 1z 2 ü n wš, n l 24 z s w w s sƒw 2,000 x (michromatic erythrocyte, PCE) ùkù w ƒ x (micronucleated polychromatic erythrocyte, MNPCE) w, x n w m w w ƒ ùkù x n w q x š w ƒ x, x w s w w

6 .JDSPOVDMFVT5FTUPG8JME(JOTFOH$VMUVSF&YUSBDU6TJOHUIF.BSSPX$FMMTJO*$3.JDF 63 š x 1. û» : š ( z r)., pp (1996). 2. Yamaguchi, H,. Matsuura, H., Kasai, R., Tanaka, O., Satake, M., Kodha, H., Izumi, H., Nuno, M., Katsuki, S. and Isoda, S.: Analysis of saponins of wild ginseng. Chem. Pharm. Bull., 36(10), (1988). 3. Mizuno, M., Yamada, J., Terai, H., Kozukue, N., Lee, Y.S. and Tsuchida, H.: Differences in immunomodualating effects between wild and cultured Panax ginseng. Biochem. Biophys. Res. Commun., 200(3), (1994). 4. Lee, E.J., Jhao, H.L., Li, D.W., Jung, C.S., Kim, J.H. and Kim, Y.S.: Effect of MeOH extract of adventious root culture of Panax ginseng on hyperlipidemic rat induced by high fat-rich diet. Korean J. Pharmacogn,. 34(2), (2003). 5. : w. wy t wz, 27(2), (2001). 6. Alder, I.D.: Cytogenetic tests in mammals. In mutagenecity testing a practical approach. Venitt, S. Parry, JM (eds.), IRL press, Oxford, pp (1984). 7. Schmid, W.: The micronucleus test. Mutat. Res., 31:9-15 (1975). 8. SAS Intitute: SAS/STAT User's guide, Version 6, 4th ed., Vol.1, Cary, NC., USA (1989a). 9. SAS Intitute: SAS/STAT User's guide, Version 6, 4th ed., Vol.2, Cary, NC., USA (1989b). 10. Zar JH: Bioststistical Analysis, 2nd Edition, Prentice-Hall International Editions (1990). 11. Heddle, J.A., Stuart, E., Salamone, M.F.: The bone marrow micronucleus test. In Handbook of mutagenicity test procedures, 2nd edition Elsevier Science Publishiers BV. pp (1984). 12. Lovell, D.P., Anderson, D., Albanese, R., Amphlett, G.E., Clare, G., Ferguson, R., Richold, M., Papworth, D.G. and Savage, J.R.K.: Statistical analysis on in vivo cytogenetic assays, In Statistical evaluation of mutagenicity test data (Kirkland, D.J. ed), Chambridge University Press, Cambridge, U.K., pp (1998).