1/3 NSC B

Transcription

1 1/3 NSC B more effetive in orgni solvent (2 N hydrohlori id in (diosgenin) methnol or 2 N sulfuri id in 70% isopropnol) thn queous solution. The hydrolyste ws inuted t 80 C spirostnol for 30 min to evporte the orgni solvent nd to omplete glyoside the onvert, efore the hexne (liquid-liquid) extrtion. spirostnol ( 5 -furostnol) Four mong 19 tested ultivrs ontin diosgenin in the rnge 7 of 41 to 70 µg/g (dry sis) in flesh, nd the peel of these ultivrs lso ontin diosgenin with ontent 2 to 3 times 2 N - 2 N higher thn the ontent in the flesh. - (70 ) hromtogrphi onditions for the nlysis were: olumn, C 18 reverse phse olumn; eluent, etonitrile: wter = 95:5 (V/V); flow rte, 1 ml/min; detetion, UV 203 nm. 19 Keywords: Diosore, Chinese ym, diosgenin, steroidl sponin, HPLC 41 70µg Diosoree Diosore 2 3 (HPLC) Liu et l., 1995, nm C18 / 95/5 v/v 1 ml/min C 1-2 sponins diosgenin et l., 1995, 1999, 2000) (HPLC) ABSTRACT To estlish diosgenin ontent dtse of Chinese ym grown in Tiwn, the vlidtion of urrent nlytil methods ws re-evluted. The mjority of diosgenin omponents re in the form of glyosides (sponins). To urtely nlyzing the ontent of diosgenin, the glyosyl groups on the sponins, oth spirostnol nd furostnol glyosides, shll e removed y id hydrolysis nd onverted to diosgenin ( 5 -furostnol) efore the quntittive hromtogrphy. After evluting 7 different proedures, we suggested tht n exhustive extrtion with methnol ws required to seprte the sponins from rude fier of ym tht interfered the determintion. The id hydrolysis ws found The high-performne (Liu et l., 1994 Liu steroidl sponins (glyosides) spirostnol glyoside furostnol glyoside glyone (diosgenin, 5 - furostnol) steroidl spogenins steroidl hormones ortiosteroids Chen, 1985 Chen nd Wu, 1994 Chen Wu, 1994 Diosore olletti D. ltheoides D. olletti vr. hypoglu D.

2 grillim D. nipponi D. pnthi D. prviflor D. zingierensis (diosgenin) (HPLC)(Tl nd Golderg, ~3.6 (Chen nd Wu, 1994) (Liu et l., 1994 Liu et l., 1995) (Tindll, 1983 Liu et l., 1995, 1996) (diosgenin) HPLC UV 203nm ( Hithi-U3210 (Liu, et l.,1992) (Liu, et 1996) diosgenin (Sigm, D1634) C 18 (LUNA 1 (Diosore lt L.) 2 (Diosore lt L.) (Diosore lt L.) (Diosore lt L.) (Diosore lt L.) diosgenin (Diosore lt L.) (Diosore lt L. vr. purpure (Rox) M. Pouh) (Diosore lt L.) (Diosore lt L.) 1986 Chen nd Wu, (Diosore jponi Thum. Vr. pseudojponi (Hy.) Ymmoto) (Diosore ulifer L.) 3 (Diosore doryophor Hne) 12 Ly et l., mm Soxhlet (diosgenin) extrtor 150ml petroleum ether 65 Wng Lo (1968) 10 Chen (1985) 34 5ml HPLC Drpeu et l., 1986 Ortuno et 17 l., 1998 Svikin-Fodulovi et l., D. omposit ( ) 18.3% 80 reflux pprtus 8 (reflux) (1) 50ml (Wng nd Lo, 1968 Chen, 1985 Yeh et l., 1994 Tylor et l., 1997 Tylor et l., 2000) petroleum 3 ether Li nd Run, ml Mhto et l., 1981 Yng nd Chen, 1983 Drpeu et l., 1986 Chen nd Wu, ml isopropnol (hexne) 5ml HPLC (prtition) (Drpeu et l., 1986 Ortuno et l., 1998 Svikin-Fodulovi et l., 1998) 1994 Tylor et l., 1997 Tylor et l., ( mm) Liu et l., 1995, 1999 (spogenin) N (thin lyer hromtogrphy, TLC) ( Blunden et 1967 Blunden nd Hrdmn, 1968 Wng nd Lo, 1968 Chen, 1985) ( ) (GLC)(Knight, ml Liu et l., 1995 Pierre et l., 1995 Tylor et l., 1997 Svikin-Fodulovi et l., 1998, 1999 Tylor et l., 2000) Mhto et l., 1981 Chen, 1985 Yeh et l., 1994 Ortuno et l., 1998 Ortuno et l., 1999 Onin et l., 2000) (diosgenin) 201nm) C18, 4.6 x 250 mm, 5 µm (Phenomenox, CA) / 95/5 v/v 1ml/min diosgenin Li nd Run, 1980 Mhto et l., 1981 Yng nd Chen, 1983 Drpeu et l., 50ml 2N 80 reflux pprtus ml 2N 70 liquid-liquid extrtion 2 50 ml 50ml Wng nd Lo, 1968 Chen, 1985 Yeh et l., Soxhlet extrtor 150ml ml( ) 2 2.0N ( )20ml( ) 3 2N 70

3 50ml 5ml HPLC diosgenin ( ) diosgenin ( C 18 / 95/5 v/v ) ( diosgenin S/N=10/1 200ng Wng Lo (1968) diosgenin 10% ( ) diosgenin diosgenin diosgenin diosgenin diosgenin 18 diosgenin diosgenin 5mg 2N -70 diosgenin diosgenin ( ) 2N -70 diosgenin ( ) ± ± µ diosgenin diosgenin diosgenin ( ) 4 diosgenin diosgenin ± µ ( ) Drpeu (1986) Pseshnihenko (1973) spirostnol furostnol diosgenin 5mg 10g diosgenin 85-92% 60-70% diosgenin 2N 70 diosgenin diosgenin diosgenin furostnol F diosgenin ( ) spirostnol furostnol diosgenin 70 spirostnol furostnol diosgenin diosgenin diosgenin ( ) diosgenin ( -70% ) 2N 70 diosgenin diosgenin diosgenin 50ml diosgenin diosgenin TLC / =97/3(v/v) diosgenin furostnol diosgenin 2N diosgenin diosgenin 2N 2N Tlyor (2000) 70% diosgenin -70 (fenugreek 2N (70% ) diosgenin seed) (steroidl spogenin)

4 (steroidl pp sponin) 1999 diosgenin pp % pp5-15 diosgenin : 1-3. diosgenin 5. Blunden, G. nd Hrdmn, R n Improved method diosgenin of densitometri thin lyer hromtogrphy s pplised to the determintion of spogenin in Diosore tuers. J. 70 Chromtogr. 38: Blunden, G., Hrdmn, R. nd Morrison, J diosgenin Quntittive elimintion of diosgenin in Diosore tuers y densitometri thin lyer hromtogrphy. J. Phrm. Si (8): spirostnol furostnol 7. Chen, A. H Applied studies on the sponins of Diosore I. Studies on the sponins of Diosore in diosgenin diosgenin Tiwn. Chin. Chem. So. J. 43: diosgenin 8. Chen, Y. nd Wu, Y Progress in reserh nd ( ) mnufturing of steroidl spogenins in Chin. J. Her diosgenin Spi. Med. Plnt. 2: diosgenin 9. Drpeu, D., Suvire, Y., Blnh, H. W. nd Wilke, C. R Improvement of diosgenin yield form Diosore diosgenin 5mg 10g deltoide plnt ell ultures y use of non-trditionl diosgenin hydrolysis method. Plnt Med. 6: Ly, H. L., Denq, S. M., Liu, S. Y. nd Sheu, B. W Studies on the identifition of Chinese drugs mteril in yms (Diosore spp.). J. Food Drug Anl. 7: diosgenin 11. Li, D. G. nd Run, Y. C Diosgenin nd ymogenin diosgenin form four speies of Diosoee L. in Yunnn. At Botn. Yunnni 2: Liu, B., Lokwood, G. B. nd Gifford, L. A Superritil fluid extrtion of diosgenin from tuers of Diosore nipponi. J. Chromtogr. A. 690 : diosgenin 13. Liu, S. Y., Wng, J. Y., Shyu, Y. T., Hu, M. F., Yng, H. R. diosgenin ( ) nd Ho, C. C The development of ym new vriety Tinung No. 2. J. Agri. Res. Chin. 45: diosgenin Liu, S. Y., Wng, J. Y., Shyu, Y. T. nd Song, L. M Studies on yms (Diosore spp.) in Tiwn. J. Chin. Med. 30 6: Liu, S. Y., Wng, J. Y., Song, L. M. nd Shyu, Y. T HPLC Studies on the prodution nd qulity of ym (Diosore 203nm C18 spp.). J. Chin. Med. 5: Liu, S. Y., Wng, J. Y., Shyu, Y. T., Dun, C. H The / 95/5 v/v 1 ml/min development nd dptility of Diosore lt L. v. 2.0N ( ) Tinung No. 1. J. Agri. Res. Chin. 41(2): N Mhto, S. B., Shu, N. P. nd Roy, S. K Highperformne liquid hromtogrphi for the determintion of diosgenin in plnts. J. Chromtogr. 121: diosgenin 18. Onin, R., Boti,,J. M., Dio Rio, J. A. nd Ortuno, A Bioprodution of diosgenin in Cllus ultures of Trigonell foenum-greum L. Food Chem. 70: diosgenin 19. Ortuno, A., Onin, R., Boti,J. M. nd Dio Rio, J. A Regultion of the diosgenin expression in Trigonell foenum-greum plntsy different plnt growth regultors. Food Chem. 65: Ortuno, A., Onin, R., Boti,J. M. nd Dio Rio, J. A.

5 1998. Distriution nd hnges of diosgenin during development of Trigonell foenum-greum plnts. Modultion y enzylminopurine. Food Chem. 63: Pierre, P. R., Suvire, Y. D., Hillire-Buys, D. M., Leonte, O. M., Biss, Y. G., Ponsin, G. R. nd Ries, G. R Steroid sponins form fenugreek seeds: extrtion, purifition,nd phrmologil investigtion on feeding ehvior nd plsm holesterol. Steroid 60: Svikin-Fodulovi, K., Gruisi, D., Culfi, L., Menkovi, N. nd Risti, M Diosgenin nd phytosterols ontent in five llus lines of Diosore lni. Plnt Si. 135: Tl, B. nd Golderg, I High-performne liquid hromtogrphi seprtion of steroidl spogenins. J. Nt. Prod. 44: Tylor, W. G., Zmn, M. S., Mir, Z., Mir, P. S., Ahry, S. N., Mers, G. J. nd Elder, J. L Anlysis of steroidl spogenins from Amer fenugreek (Trigonell foenum-greum) y pillry gs hromtogrphy nd omined gs hromtogrphy/mss spetrometry. J. Agri. Food Chem. 45: Tylor, W. G., Elder, J. L., Chng, P. R. nd Rihrds, K. W Mirodetermintion of diosgenin from fenugreek (Trigonell foenum-greum) seeds. J. Agri. Food Chem. 48: Tindll, H. D Deosoreee, Vegetle in the Tropis, Vol. 1, Mmilln Press, London, pp Wng, N. Y nd Lo,. T. B Studies on thin lyer hromtogrphy of steroidl spogenins nd preliminry report of spogenin frtion of Formosn Diosore. Chin. Chem. So. J. 1: Yng, M. H. nd Chen, Y. Y Steroidl spogenins in Diosore ollettii [isolted from the id treted rhizome]. Plnt Med. 49 (1): Yeh, F. T., Hung, W. W., Cheng, C. C., N, C., nd Tsy, H. S Tissue ulture of Diosore doryophor Hne II. Estlishment of suspension ulture nd the mesurement of diosgenin ontent. Chin. Agron. J. 4:

6 Tle 1. Diosgenin ontent in fresh of yms ( Diosore spp.) determined y using HPLC with different smple prepring proedures. Proedures d ( ) e d ( ) d ( ) ( ) ( ) d d ± ( ) 1. Diosgenin ontent = µg of diosgenin / g of dried ym ( Diosore spp.) 2 Smple prepring proedures re desried in the text 3. Vlues re men ± S. D. 4. Mens of three replites 5. ND = not deteted 6. Figures ering different letters in the sme row re signifintly different ( p<0.05)

7 Tle 2. Diosgenin ontent in peels of yms ( Diosore spp.) determined y using HPLC with different smple prepring proedures. ( ) Proedures ± ( ) ± ( ) ( ) ( ) ± ± 5 ± ( ) 1. Diosgenin ontent =µg of diosgenin / g of dried peel of ym ( Diosore spp.) 2. Smple prepring proedures re desried in the text 3. Vlues re men ± S. D. 4. Mens of three replites 5. ND = not deteted 6. Figures ering different letters in the sme row re signifintly different ( p<0.05)

8 2N -70% Tle 3. The yield disrepny of diosgenin form seleted hexne extrting methods fter hydrolyzed with 2N H 2 SO 4 in 70% isopropnol. Smples Proedures mg diosgenin 79.07(0.12) 90.64(0.11) 73.36(0.24) 5mg diosgenin (5.96) 10mg 30.74(10.72) 24.19(8.41) 1. Mens of three replites 2. Smple prepring proedures re desried in the text 3. Vlues in prentheses re oeffiient of vrition (%) 4. Figures ering different letters in the sme row re signifintly different ( p<0.05)

9 Tle 4.The yield disrepny of diosgenin form seleted hydrolysis methods fter extrted with methnol. Proedures mg diosgenin + 10mg ( ) 5mg diosgenin + 10mg ( ) 1. Mens of three replites 2. Smple prepring proedures re desried in the text 3. Vlues in prentheses re oeffiient of vrition (%) 4. Figures ering different letters in the tle re signifintly different ( p <0.05)