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(+)-15β-Hydroxy-17-oxolupanine a lupin alkaloid from the seeds ofLupinus albus
0031-9422(94)00537-o
Pergamoo
Phytochemwtry, Vol. 37, No. 6, pp. 1751-1753, 1994 Copyright Q 1994 FAswier kiena Ltd Printed III Great Britain. AU r@ts rcsmml 0031~!4422/94 $7.00 + 0.00
(+)-15/3-HYDROXY-17-OXOLUPANINE, A LUPIN ALKALOID SEEDS OF LUPINUS ALBUS
FROM THE
MAHMOUD H. MOHAMED,* ZEDAN Z. IBRAHEIM,?OMAR M. ABDALLAH~ and ISAMU MURAKOSHI~ Department of Pharmacognosy, Faculty of Pharmacy, Al-Azhar University, 71524 Assiut, Egypt; TDepartmcnt of Pharmacqnosy, Faculty of Pharmacy, Assiut University, Assiut 71515, Egypt; SDepartment of Plant Chemistry and Pharmacognosy, Faculty of Pharmaceutical Sciences, I-33 Yayoi-cho, Chiba 263, Japan (Received in revised form 3 June 1994) Key Word Index-Lupinus
hydroxy-17-oxolupanine;
albus; Leguminosae; seeds; quinolizidine alkaloid, lupin alkaloid, (+)-lSfi( + )-tetrahydrorhombifoline.
Abstract--A new lupin alkaloid (+)-15/Shydroxy-17-oxolupanine was isolated from the ethanol extract of the seeds of Lupinus albus together with (+)-tetrahydrorhombifoline. 17-Oxolupanine was detected by GC-mass spectral analysis.
0
INTRODUCXON The various species of Lupinus have been a subject of interest to several investigators owing to their rich alkaloid content [ 11. Lupinus albus L. (Lupinus termis Forsk) is cultivated in the Mediterranean and Egypt for its edible seeds [2]. Some medicinal uses of this plant have been reported [3, 41. We have previously isolated ( - )-ASdehydromultiflorine, (-)-A’-dehydroalbine, ( f)-termisine and nine known lupin alkaloids from the seeds of
I. R=OH 2. R=H
L. albus [S-8].
In a further investigation of the less polar fraction of the basic components of viable seeds of the Egyptian lupine, ( + )-15&hydroxy-17-oxolupanine (l), was isolated together with (+)-tetrahydrorhombifoline. 17-Oxolupanine was detected as a very minor compound by GCmass spectral analysis. RlBULTGAND DISCUSSION ( + )-l SB-Hydroxy- 17-oxolupanine (1) accounted for 0.017% of fresh weight. The HR-mass spectrum of 1 afforded the molecular formula CISHzIN,O, ([Ml’, m/z 278.1638, talc. 278.1631) and [M-H,O]+ corresponding to CISH,,NaO, (m/z 260.1528, talc. 260.1526). The low resolution EI-mass spectrum showed the molecular ion peak at m/z 278 and the base peak at m/z 260 [M- 18]+. The presence of a hydroxyl group was indicated by the fragments at m/z 261 (20) and 260 (base peak) in the EI-mass spectrum, corresponding to [M -OH]’ and [M-H20]+, respectively [9]. The IR spectrum showed an intense broad band at 34203250 cm-’ indicative of the presence of hydroxyl groups [lo], two lactam carbonyl groups at 1640 and 1620 cm-‘,
*Author to whom correspondence should LXaddressed.
R
and bands at 2950-2850 cm- ’ for C-H stretching. Thus, 1 was presumed to be a sparteine-type lupin alkaloid containing two lactams and hydroxyl groups in the molecule. The 13C NMR spectrum of 1 the presence of 15 carbons which were assigned as shown in Table 1. Determination of the multiplicity was carried out by DEPT experiments, which revealed that 1 contained five methine, eight methylene carbons, and two carbonyl groups. The substitution pattern was deduced as follows: the first carbonyl was placed at position 2 because C-2 had the same chemical shift 6171.1 as (+)-17oxolupanine [ll, 123 and the second carbonyl (6 168.9) was assigned to position C-17. The hydroxyl group was placed at position 15 because the signal corresponding to the carbinol carbon at 672.3 (d, C-15) [13] was in agreement with the calculated one [14]. The 13CNMR spectrum of 1 resembled that of (+)-17-oxolupanine (2) where the chemical shifts of the carbons of rings A and B were coincident with those of (+)-17-oxolupanine (2) which was also detected by GC-mass spectrometry. In the ‘HNMR spectrum, the downfield shifted carbinol proton resonated at 66.18 (lH, dd, J=6.2, 3.3 Hz, H-15,,). H-15 was coupled to both H-14 protons with
1751
1752
M, H. MOHAMED et al. Table 1. Comparison of the *3C NMR data of compounds 1 and 2 c
1
2*
2
171.1s
170.7s
3
32.6 t 18.2 t 32.7 t 58.8 d 43.6 d 26.7 t 33.2 d
33.4 t 19.4 t 32.8 t 58.9 d 43.6 d 27.0 f 33.8 d
47.9 E
48.0t
4 5 6 7 8 9 10
I1 x2 13 14 15 17
53.9 d 29.8 t 26.0 t 18.4 t 72.3 d 168.9 s
fif,f d 33.4 t 25.0 t 25.3 t 42.9 t
167.3s
*Taken from ref. [II].
relatively large coupling constants (3.3 and 6.2), diaxial coupling was observed and hence the H-15 proton must be axial. The other downfield proton of 1 which resonated at 64,81 (lH, dt, J = 13.4,2.4 Hz, H-IO,,) resembled that of [ + )-17-oxolupanine (2) fll, i23. These data provided further evidence that the sparteine skeleton of 1 was substituted by two carbonyl groups and one hydroxyl group at C-2, C-17 and C-15, respectively. The assignments including all the protons and carbons were canfirmed by fH-XHCOSY and 13C-lHCOSY. Camparing the ‘H and 13C NMR data of f with those uf f +>17-oxolupanine (2), the signal of the proton at C-l 1 of 1 was shifted downfield to 63.79 (53.20 in 2) and the signals of C-12 and C-14 of 1 were shifted upfield to 6 29,8 and 18.4, respectively. These results indicated that the cun~guratiun of rings C and D in 1 is chair-chair [S]. This is also supported by the absence of cross-peaks between C-11 and C-9 protons. As 17-oxolupanine is present in L. albus seeds, 1 is presumed to be biosynthesized from it by hydroxylation.
General. Mps: uncorr; fR: thin films of Kl3r or CHCI,; ‘H and 13CNMR: 500 and 125 MHz, respectively, CDCl, with TMS as int. standard; low resolution EI-MS and in-beam HREI-MS: 70 eV; TLC: silica gel (Kiesel gel F254) of 0.25 mm layer thickness in 60, CH,Cl,-MeOH-28% N&OH (90:9: 1). The chromatograms were visualized by spraying with Dragendroff and iodoplatinate reagents. Analytical GC was performed as described before [ 151. Extraction and isolation of alkaloids. The seeds 1;. albus were collected at the Medicinal Plant Experimental Station at Al-Azhar University, Assiut in April 1992. The voucher specimen was identified by Prof. A. Fayed
(Department of Systematic Botany, Faculty of Science, Assiut University, Assiut, Egypt) is deposited in the Department of Pharmacognosy, Faculty of Pharmacy, Al-Azhar University, Assiut, Egypt. The total alkaloid fraction (yield of 2.6% fr. wt) from the 75% EtOH extracts of the viable seeds was chromatographed on a silica gel column (Merck, type 60,230-400 mesh, 1 kg, 7 x 150 cm) with 1% MeOH in CH,Ct,-28% NH&W (500: 1). ( +)-Tetrahydrorhumb~oline was obtained as an oil (21 mg), [z]“’ 8 1 (CH,CI,; c 0.1). A mixture af 13-(2-methyibutyryl)-oxylupanine and 17-0x0lupanine was eluted by 1.5% MeOH in CH,Cl,. GCMS indicated the presence of these two alkaloids in a ratio of 1: 1 [ 16- 191, Pure I (17 mg) was obtained from the fraction eluted by 2% MeOH in CHZC12. ( + )-1 @-H@~xy- 17-ox&pan& (1). Fine needles, mp X5-87”; [ot32’ +4.5” fCH,Cl,; ~0.15); HR in-beam El-MS rnjz (rel. int.): 278,1638 [M] + (3), (talc, for C,,H,,N,O,, 278.1631); El-MS m/z (rel. int.): 278 (3), 261 (20), 260 (loo), 232 (8), 166 (X), 152 (ll), 150 (23), 148 (32), 133 (ll), 112 (32), 82 (15), 55 (30), 14 (13); 1R v=; cm- 1: 3420-3250 br {OH), 2950-2850 (CH), 164U {lactam C=O), 1620 (lactam C=O); lH NMR (CDCl,, 500 MHz): 66.18 (IH, dd, J =t:K&3.3 Hz, H-&,),4.81 (lH, dt, J= 13.4,2.4 Hz, H-lo,,), 3.79 (lH, dd, J= 12.3, 2.7 Hz, H-11), 3.46 (lH, m, H-6), 2.69 (lH, dd, J= 13.4, 2.4, H&,), 2SO flH, Br s, H-7), 2.37 flH, br 6,j = 13.2, H-5,,), 2.26 (IH, m, H-5,,), 2.19 (ZH, m, H-8,, H-13,,), 1.99-1.87 (ZH, m, H-8,,, H-14,), I+77(lH., br s, H-9X 1.72- 1.65 (3H, m, H-4,,, H-12,,, H-13,,), 1.59-1.52 (2H, m, H-12,,, H14,,), 1.4G1.39 (3H, m, 2 x H-3, H-4,,); 13CNMR fCDCl,, 125 MHz): Table 1. ( + )-Tetrahyd~orhomBifrrline.This compound was identified by spectral analysis. For full data see refs [20, 211. REFERENCES 1. Mears, J. A. and Mabry, T. J. (1971) in Chemotaxonomy of me Legwninusae (Harborne, J. B, BouIter, D. and Turner, B. L., eds), p. 73. Tackholm, V. (1974) Stti&nts F!ota of Egypt, 2nd Edn, p- 224. Cairo University Press, Cairo. Antaun, M. D. and Taha, 0. M. A. (1981) J. Nat. Prod. 44, 179. Mohamed, M. H., Kadry, H. A., Khalifa, T. I., Hassan, H. A., Fujii, F., Saito, K, and Murakoshi, f. f1991) 15th International Congress of Pharmaceutical Sciences of F.I.P. (Washington). Abstract 4P-008. 5. Mohamed, M. H., Saito, K., Murakoshi, I., Kadry, H. A., Khalifa, T. I. and Hassan, H. A. (1990) J. Nar. Prod. 53, 1578. 6. Mohamed, M. H., Saita, K., Kadry, H. A., Khalifa, T. I,, Ammar, H. H. and Murakoshi, I. (1991) Pkytuchemistry 30, 3 111. 7. Mohamed, M. H., Kadry, H. A., Khalifa, T. I., Ammar, H. H., Sekine, T., Saito, K., Ogata, K. and Murakoshi, I. (1992) Al-Azhm J. Nat. Prod. 8, 1. 8. Mohamed, M. H. and El-Shorbagi, A, A. H. (1993) 3. Na$. Pmd. 56, 1399.
An alkaloid from Lupinus albus
of Organic Compounds, 3rd Edn, p. 112. John Wiley, New York. 10. Herbert, S. A. (1985) in Topics in St~eochemistry (Allinger, N. L. and Ehei, E. L., eds), Vol. 11, p. 12. Academic Press, New York. 11. Takamatsu, S. (1987) Master Thesis, Faculty of Pharmaceutical Sciences, Chiba University. 12. Ghmiya, S., Otomasu, H., Haginiwa, J. and Murakoshi, I. (1984) Phytochem~try 23, 2665. 13. Arslanian, R. L., Harris, G. H. and Stem&z, F. R. (1990) J. Org. Chem. 55, 1204. 14. Breitmaier, E. and Voetter, W. (1987) in Carbon-13 NMR Spectroscopy, 3rd Edn, p. 270. VCH, New York.
1753
15. Sekine, T., Arai, K., Saito, K., Mohamed, M. H., Makboul, M. A. and Murakoshi, I. (1992) Thai J. Pharm. Sci. 16, 267. 16. Wink, M., Witte, L., Schiebel, H. M. and Hartmann, T. (1980) Planta Med. 38, 238. 17. Wink, M., Witte, L. and Hartmann, T. (1981) Planta Med. 43, 342. 18. Wink, M. and Hartmann, T. (1982) Pianta 1% 560. 19. Hussain, R. A., Kinghom, A. D. and Molyneux, R. J. (1988) J. Nat. Prod. 51, 809. 20. Balandrin, M. F. and Kinghorn, A. D. (1981) .I. Nat. Prod. 44,495. 21. McCoy, J. W. and Stermitz, F. R. (1983) J. Nat. Prod. 46,902.
Pergamoo
Phytochemwtry, Vol. 37, No. 6, pp. 1751-1753, 1994 Copyright Q 1994 FAswier kiena Ltd Printed III Great Britain. AU r@ts rcsmml 0031~!4422/94 $7.00 + 0.00
(+)-15/3-HYDROXY-17-OXOLUPANINE, A LUPIN ALKALOID SEEDS OF LUPINUS ALBUS
FROM THE
MAHMOUD H. MOHAMED,* ZEDAN Z. IBRAHEIM,?OMAR M. ABDALLAH~ and ISAMU MURAKOSHI~ Department of Pharmacognosy, Faculty of Pharmacy, Al-Azhar University, 71524 Assiut, Egypt; TDepartmcnt of Pharmacqnosy, Faculty of Pharmacy, Assiut University, Assiut 71515, Egypt; SDepartment of Plant Chemistry and Pharmacognosy, Faculty of Pharmaceutical Sciences, I-33 Yayoi-cho, Chiba 263, Japan (Received in revised form 3 June 1994) Key Word Index-Lupinus
hydroxy-17-oxolupanine;
albus; Leguminosae; seeds; quinolizidine alkaloid, lupin alkaloid, (+)-lSfi( + )-tetrahydrorhombifoline.
Abstract--A new lupin alkaloid (+)-15/Shydroxy-17-oxolupanine was isolated from the ethanol extract of the seeds of Lupinus albus together with (+)-tetrahydrorhombifoline. 17-Oxolupanine was detected by GC-mass spectral analysis.
0
INTRODUCXON The various species of Lupinus have been a subject of interest to several investigators owing to their rich alkaloid content [ 11. Lupinus albus L. (Lupinus termis Forsk) is cultivated in the Mediterranean and Egypt for its edible seeds [2]. Some medicinal uses of this plant have been reported [3, 41. We have previously isolated ( - )-ASdehydromultiflorine, (-)-A’-dehydroalbine, ( f)-termisine and nine known lupin alkaloids from the seeds of
I. R=OH 2. R=H
L. albus [S-8].
In a further investigation of the less polar fraction of the basic components of viable seeds of the Egyptian lupine, ( + )-15&hydroxy-17-oxolupanine (l), was isolated together with (+)-tetrahydrorhombifoline. 17-Oxolupanine was detected as a very minor compound by GCmass spectral analysis. RlBULTGAND DISCUSSION ( + )-l SB-Hydroxy- 17-oxolupanine (1) accounted for 0.017% of fresh weight. The HR-mass spectrum of 1 afforded the molecular formula CISHzIN,O, ([Ml’, m/z 278.1638, talc. 278.1631) and [M-H,O]+ corresponding to CISH,,NaO, (m/z 260.1528, talc. 260.1526). The low resolution EI-mass spectrum showed the molecular ion peak at m/z 278 and the base peak at m/z 260 [M- 18]+. The presence of a hydroxyl group was indicated by the fragments at m/z 261 (20) and 260 (base peak) in the EI-mass spectrum, corresponding to [M -OH]’ and [M-H20]+, respectively [9]. The IR spectrum showed an intense broad band at 34203250 cm-’ indicative of the presence of hydroxyl groups [lo], two lactam carbonyl groups at 1640 and 1620 cm-‘,
*Author to whom correspondence should LXaddressed.
R
and bands at 2950-2850 cm- ’ for C-H stretching. Thus, 1 was presumed to be a sparteine-type lupin alkaloid containing two lactams and hydroxyl groups in the molecule. The 13C NMR spectrum of 1 the presence of 15 carbons which were assigned as shown in Table 1. Determination of the multiplicity was carried out by DEPT experiments, which revealed that 1 contained five methine, eight methylene carbons, and two carbonyl groups. The substitution pattern was deduced as follows: the first carbonyl was placed at position 2 because C-2 had the same chemical shift 6171.1 as (+)-17oxolupanine [ll, 123 and the second carbonyl (6 168.9) was assigned to position C-17. The hydroxyl group was placed at position 15 because the signal corresponding to the carbinol carbon at 672.3 (d, C-15) [13] was in agreement with the calculated one [14]. The 13CNMR spectrum of 1 resembled that of (+)-17-oxolupanine (2) where the chemical shifts of the carbons of rings A and B were coincident with those of (+)-17-oxolupanine (2) which was also detected by GC-mass spectrometry. In the ‘HNMR spectrum, the downfield shifted carbinol proton resonated at 66.18 (lH, dd, J=6.2, 3.3 Hz, H-15,,). H-15 was coupled to both H-14 protons with
1751
1752
M, H. MOHAMED et al. Table 1. Comparison of the *3C NMR data of compounds 1 and 2 c
1
2*
2
171.1s
170.7s
3
32.6 t 18.2 t 32.7 t 58.8 d 43.6 d 26.7 t 33.2 d
33.4 t 19.4 t 32.8 t 58.9 d 43.6 d 27.0 f 33.8 d
47.9 E
48.0t
4 5 6 7 8 9 10
I1 x2 13 14 15 17
53.9 d 29.8 t 26.0 t 18.4 t 72.3 d 168.9 s
fif,f d 33.4 t 25.0 t 25.3 t 42.9 t
167.3s
*Taken from ref. [II].
relatively large coupling constants (3.3 and 6.2), diaxial coupling was observed and hence the H-15 proton must be axial. The other downfield proton of 1 which resonated at 64,81 (lH, dt, J = 13.4,2.4 Hz, H-IO,,) resembled that of [ + )-17-oxolupanine (2) fll, i23. These data provided further evidence that the sparteine skeleton of 1 was substituted by two carbonyl groups and one hydroxyl group at C-2, C-17 and C-15, respectively. The assignments including all the protons and carbons were canfirmed by fH-XHCOSY and 13C-lHCOSY. Camparing the ‘H and 13C NMR data of f with those uf f +>17-oxolupanine (2), the signal of the proton at C-l 1 of 1 was shifted downfield to 63.79 (53.20 in 2) and the signals of C-12 and C-14 of 1 were shifted upfield to 6 29,8 and 18.4, respectively. These results indicated that the cun~guratiun of rings C and D in 1 is chair-chair [S]. This is also supported by the absence of cross-peaks between C-11 and C-9 protons. As 17-oxolupanine is present in L. albus seeds, 1 is presumed to be biosynthesized from it by hydroxylation.
General. Mps: uncorr; fR: thin films of Kl3r or CHCI,; ‘H and 13CNMR: 500 and 125 MHz, respectively, CDCl, with TMS as int. standard; low resolution EI-MS and in-beam HREI-MS: 70 eV; TLC: silica gel (Kiesel gel F254) of 0.25 mm layer thickness in 60, CH,Cl,-MeOH-28% N&OH (90:9: 1). The chromatograms were visualized by spraying with Dragendroff and iodoplatinate reagents. Analytical GC was performed as described before [ 151. Extraction and isolation of alkaloids. The seeds 1;. albus were collected at the Medicinal Plant Experimental Station at Al-Azhar University, Assiut in April 1992. The voucher specimen was identified by Prof. A. Fayed
(Department of Systematic Botany, Faculty of Science, Assiut University, Assiut, Egypt) is deposited in the Department of Pharmacognosy, Faculty of Pharmacy, Al-Azhar University, Assiut, Egypt. The total alkaloid fraction (yield of 2.6% fr. wt) from the 75% EtOH extracts of the viable seeds was chromatographed on a silica gel column (Merck, type 60,230-400 mesh, 1 kg, 7 x 150 cm) with 1% MeOH in CH,Ct,-28% NH&W (500: 1). ( +)-Tetrahydrorhumb~oline was obtained as an oil (21 mg), [z]“’ 8 1 (CH,CI,; c 0.1). A mixture af 13-(2-methyibutyryl)-oxylupanine and 17-0x0lupanine was eluted by 1.5% MeOH in CH,Cl,. GCMS indicated the presence of these two alkaloids in a ratio of 1: 1 [ 16- 191, Pure I (17 mg) was obtained from the fraction eluted by 2% MeOH in CHZC12. ( + )-1 @-H@~xy- 17-ox&pan& (1). Fine needles, mp X5-87”; [ot32’ +4.5” fCH,Cl,; ~0.15); HR in-beam El-MS rnjz (rel. int.): 278,1638 [M] + (3), (talc, for C,,H,,N,O,, 278.1631); El-MS m/z (rel. int.): 278 (3), 261 (20), 260 (loo), 232 (8), 166 (X), 152 (ll), 150 (23), 148 (32), 133 (ll), 112 (32), 82 (15), 55 (30), 14 (13); 1R v=; cm- 1: 3420-3250 br {OH), 2950-2850 (CH), 164U {lactam C=O), 1620 (lactam C=O); lH NMR (CDCl,, 500 MHz): 66.18 (IH, dd, J =t:K&3.3 Hz, H-&,),4.81 (lH, dt, J= 13.4,2.4 Hz, H-lo,,), 3.79 (lH, dd, J= 12.3, 2.7 Hz, H-11), 3.46 (lH, m, H-6), 2.69 (lH, dd, J= 13.4, 2.4, H&,), 2SO flH, Br s, H-7), 2.37 flH, br 6,j = 13.2, H-5,,), 2.26 (IH, m, H-5,,), 2.19 (ZH, m, H-8,, H-13,,), 1.99-1.87 (ZH, m, H-8,,, H-14,), I+77(lH., br s, H-9X 1.72- 1.65 (3H, m, H-4,,, H-12,,, H-13,,), 1.59-1.52 (2H, m, H-12,,, H14,,), 1.4G1.39 (3H, m, 2 x H-3, H-4,,); 13CNMR fCDCl,, 125 MHz): Table 1. ( + )-Tetrahyd~orhomBifrrline.This compound was identified by spectral analysis. For full data see refs [20, 211. REFERENCES 1. Mears, J. A. and Mabry, T. J. (1971) in Chemotaxonomy of me Legwninusae (Harborne, J. B, BouIter, D. and Turner, B. L., eds), p. 73. Tackholm, V. (1974) Stti&nts F!ota of Egypt, 2nd Edn, p- 224. Cairo University Press, Cairo. Antaun, M. D. and Taha, 0. M. A. (1981) J. Nat. Prod. 44, 179. Mohamed, M. H., Kadry, H. A., Khalifa, T. I., Hassan, H. A., Fujii, F., Saito, K, and Murakoshi, f. f1991) 15th International Congress of Pharmaceutical Sciences of F.I.P. (Washington). Abstract 4P-008. 5. Mohamed, M. H., Saito, K., Murakoshi, I., Kadry, H. A., Khalifa, T. I. and Hassan, H. A. (1990) J. Nar. Prod. 53, 1578. 6. Mohamed, M. H., Saita, K., Kadry, H. A., Khalifa, T. I,, Ammar, H. H. and Murakoshi, I. (1991) Pkytuchemistry 30, 3 111. 7. Mohamed, M. H., Kadry, H. A., Khalifa, T. I., Ammar, H. H., Sekine, T., Saito, K., Ogata, K. and Murakoshi, I. (1992) Al-Azhm J. Nat. Prod. 8, 1. 8. Mohamed, M. H. and El-Shorbagi, A, A. H. (1993) 3. Na$. Pmd. 56, 1399.
An alkaloid from Lupinus albus
of Organic Compounds, 3rd Edn, p. 112. John Wiley, New York. 10. Herbert, S. A. (1985) in Topics in St~eochemistry (Allinger, N. L. and Ehei, E. L., eds), Vol. 11, p. 12. Academic Press, New York. 11. Takamatsu, S. (1987) Master Thesis, Faculty of Pharmaceutical Sciences, Chiba University. 12. Ghmiya, S., Otomasu, H., Haginiwa, J. and Murakoshi, I. (1984) Phytochem~try 23, 2665. 13. Arslanian, R. L., Harris, G. H. and Stem&z, F. R. (1990) J. Org. Chem. 55, 1204. 14. Breitmaier, E. and Voetter, W. (1987) in Carbon-13 NMR Spectroscopy, 3rd Edn, p. 270. VCH, New York.
1753
15. Sekine, T., Arai, K., Saito, K., Mohamed, M. H., Makboul, M. A. and Murakoshi, I. (1992) Thai J. Pharm. Sci. 16, 267. 16. Wink, M., Witte, L., Schiebel, H. M. and Hartmann, T. (1980) Planta Med. 38, 238. 17. Wink, M., Witte, L. and Hartmann, T. (1981) Planta Med. 43, 342. 18. Wink, M. and Hartmann, T. (1982) Pianta 1% 560. 19. Hussain, R. A., Kinghom, A. D. and Molyneux, R. J. (1988) J. Nat. Prod. 51, 809. 20. Balandrin, M. F. and Kinghorn, A. D. (1981) .I. Nat. Prod. 44,495. 21. McCoy, J. W. and Stermitz, F. R. (1983) J. Nat. Prod. 46,902.