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Traditional medicinal plants of Thailand XVII Biologically active constituents of Plumeria rubra
Journal of E!hnopharmucology,
33
( I99 I ) 289-292
289
Elsevier Scientific Publishers Ireland Ltd.
Short Communication
Traditional
Matthias
medicinal plants of Thailand XVII. * Biologically active constituents of Plumeria rutwa
0. Hamburgera’**,
Geoffrey
A. Cordell”
and Nijsiri Ruangrungsib
“Program for Collaborative Research in the Pharmaceutical Sciences. College of Pharmacy, University of Illinois at Chicago, Chicago, Illinois 60612 (U.S. A.) and ‘Department of Pharmacognosy. Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok (Thailand)
(Accepted January 22, 1991) The compounds l-6 were isolated from the heartwood of Plumeria rubra, following bioactivity-directed fractionation. Plumericin 1 and isoplumericin 2 displayed molluscicidal, cytotoxic and antibacterial activity, 4-hydroxyacetophenone 3 was weakly cytotoxic, whereas the remaining glycosidic isolates (plumieride, 4, l3-O-coumaroylplumieride, test systems.
5; protoplumericine
A, 6) were inactive in all
Key wora!s: Plumeria rubra; molluscicidal activity; iridoids: cytotoxicity.
Introduction As a part of our continuing search for biologically active compounds, in particular antimicrobial and cytotoxic agents from medicinal plants of Thailand (Hamburger et al., 1987), we have examined Plumeria rubra L. (syn. P. acutifoiiu Poir.) (Apocynaceae). There are numerous reports on the ethnomedical use of plants belonging to the pantropic genus Plumeria. In South America, decoctions of leaves, bark or heartwood are used as an anthelmintic, purgative, emmenagogue, and for the topical treatment of various afflictions of the skin (Peckolt, 1942). The fruit pulp,
Correspondeme IO: Professor G.A. Cordell. Program for Collaborative Research in the Pharmaceutical Sciences. College of Pharmacy, University of Illinois at Chicago. Chicago. Illinois 60612 (U.S.A.) *For the previous paper. see Topcu et al.. 1990. l*Presem address: lnstitut de Pharmacognosie et Phytochimie. Universitt de Lausanne, CH-1005 Lausanne. Switzerland.
0378-8741/$03.50 0 1991 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
latex and stem bark serve in eastern Asia as an abortifacient and purgative (Quisumbing, 1951; Wasuwat, 1967; Datta and Satta, 1976; Sawhney et al., 1978). Extracts of various Plumeriu species have been shown to exhibit significant anantifungal and antiviral activity tibacterial, (Sticher, 1977; Vanden Berghe et al., 1978; Jeven 1979). The iridoids plumericin, et al., isoplumericin and fulvoplumierin have been isolated as the antifungal and antiviral agents of P. rubra (Sticher, 1977). The same compounds, along with some iridoid glycosides, have been identified by co-TLC in some other Plumeria species (Coppen and Cobb, 1983). While the present investigation was in progress, a phytochemical study of the iridoids from the roots of P. rubra was published without reporting, however, on the activity of the isolates (Abe et al., 1988), and our colleagues have reported on the isolation of six cytotoxic constituents (Kardono et al., 1990a) and a new flavan-3-01 glycoside (Kardono et al., 1990b) from the bark of P. rubra of Indonesian origin.
290
Material and Methods General procedures
These have been described elsewhere (Hamburger et al., 1987). Plant material
The heartwood of P. rubra was collected in 1987 in Phetchaburi Province, Thailand. The plant material was authenticated by one of us (N.R.), and a voucher specimen is deposited in the Botany Section, Technical Division, Department of Agriculture, Ministry of Agriculture and Cooperatives, Bangkok, Thailand. Extraction
and preliminary
biological
evaluation
The chopped heartwood (2.5 kg) was percolated at room temperature with EtOAc. Direct bioautography on precoated silica gel TLC plates (eluent: toluene/EtOAc (1: 1)) with B. subtilis ATCC 6633 as test organism (Hamburger and Cordell, 1987) revealed the presence of two antibacterial compounds at R/O.60 and 0.63. Preliminary small scale separation of the extract with toluene/EtOAc (1: 1) and monitoring cytotoxic activity against the P-388 cell line (Geran et al., 1972) revealed that the activity of the extract (EDSo 5 pg/ml) was located in the fraction containing the antibacterial compounds (EDSo < 0.5 &ml). Direct bioautography and the P-388 assay in vitro were used for monitoring the fractionation. Molluscicidal activity of the isolates was determined according to the procedure described by Hostettmann et al., 1982. l-6 A portion (44 g) of the EtOAc extract was subsequently separated by rapid column chromatography (CC) on silica gel using light petroleum/EtOAc (1: 1) as eluent, followed by MeOH. Four fractions (A-D) were collected. Recrystallization of fraction C (2.15 g) from toluene/EtOAc yielded 2 as colorless needles (1.22 g). Fraction B (6.3 g) was separated by CC on silica gel with toluene/EtOAc (9:l) as mobile phase. Six fractions were collected. Isoplumericin 2 (140 mg) was obtained through recrystallization of fraction 2 (400 mg). Fraction 4 (300 mg) was Isolation of compounds
submitted to reversed-phase chromatography on a Lobar RP-8 column (size B, Merck, Darmstadt, F.R.G.). Elution with MeOH/H,O (1:l) afforded 4-hydroxyacetophenone 3 (50 mg). A portion of fraction D (10.0 g) was further separated by CC on silica gel. Elution with CHCI,/MeOH/H@ (90:10:0.5 to 70:30:10) gave 7 fractions (l ’-7’). Fractions 3 ’ (2.0 g) and 7 ’ (1.6 g) consisted of 13-O-coumaroylplumieride 5 and protoplumericine A 6, respectively. A portion of each fraction was submitted to a final purification on Sephadex LH20 (MeOH). Plumieride 4 (120 mg) was isolated from fraction 5 ’ (500 mg) by CC on silica gel with CHClJ/MeOH/H20 (85:15:0.7) as mobile phase. All of the isolates were identified by MP, UV, IR, [&, EI- or D/(X-MS, ‘H and 13C-NMR (one- and two-dimensional) in comparison with published data (Yamauchi et al., 1981; Abe et al., 1984; Martin et al., 1985; Abe et al., 1988). In addition, glycosides 4-6 were submitted to acid (2N HCI, reflux for 1 h) and mild basic hydrolysis (NH3 10% in MeOH/H*O (2:1), RT). After the usual workup, the hydrolysates were analyzed by co-TLC with authentic samples of glucose, pcoumaric acid and plumieride 4. Results and Discussion In our preliminary screening, the EtOAc extract of the heartwood displayed cytotoxic activity against the P-388 cell line in vitro (ED50 5 pg/ml). Direct bioautography on TLC (Hamburger and
291
adds another facet to the broad range of biological activities of these iridoids, which have recently been the challenging target of various synthetic approaches (Trost et al., 1986; Parkes and Pattenden, 1988). Cordell, 1987), using B. subtilis as a test organism, revealed the presence of two antibacterial compounds in the same extract. Activity-guided fractionation led to the isolation of the cytotoxic and antibacterial iridoids plumericin 1 and isoplumericin 2, along with the moderately cytotoxic compound 4-hydroxyacetophenone 3 (EDSo 0.05, 0.06 and 3.2 &ml, respectively). In addition, the inactive iridoid glycosides plumieride 4, 13-Ocoumaroylplumieride 5 and protoplumericin A 6 were isolated from the polar fractions of the extract. Compounds 1 and 2 were found to exhibit significant molluscicidal activity against the schistosomiasis transmitting snail Biomphaluriu glabrata (LDloo 6.25 ppm). The other isolates 3-6 were inactive, when tested at 50 ppm. The bioactivity-guided fractionation of the extract yielded the two known compounds, plumericin 1 and isoplumericin 2, as the major antibacterial and cytotoxic principles in the heartThe weakly active of P. rubra. wood 4-hydroxyacetophenone 3 contributes only marginally to the cytotoxic activity of the extract. The three iridoid glycosides 4-6 are also known compounds, which have been reported from some Plumeria or Allamanda species. The strong molluscicidal activity of the tetracyclic iridoids 1 and 2 can be explained by the presence of an c~susceptible to methylene y-lactone moiety, undergo a Michael-type addition with biological nucleophiles (Kupchan et al., 1970). Other molluscicidal compounds with the similar structural features include the sesquiterpene lactones helenalin, pyrethrosin, 7-hydroxy-3-deoxyzaluzanin and the iridoid oruwacin (Hostettmann and Marston, 1987). In contrast, the a-substituted endocyclic o&unsaturated y-lactone ring has been found to be much less reactive towards nucleophiles (Kupchan et al., 1970). This may explain the lack of molluscicidal activity of glycosides 4-6. The discovery of the molluscicidal properties of plumericin 1 and isoplumericin 2
Acknowledgements
This work was supported by a grant from the Division of Cancer Treatment, National Cancer Institute, Bethesda, MD. We thank the Research Resources Center of the University of Illinois at Chicago for the provision of NMR and mass spectrometric facilities, and Prof. K. Hostettmann, University of Lausanne, Switzerland, for the molluscicidal tests. References Abe, F., Mori, T. and Yamauchi, T. (1984) Iridoids of Apocynaceae. III. Minor iridoids from Aliamanda neritfolia. Chemical and Pharmaceutical Bulletin 32, 2947-2956. Abe, F., Chen, R.F. and Yamauchi, T. (1988) Minor iridoids from the roots of Plumeria acuttfolia. Chemical and Pharmaceutical Bulletin 36, 278G2789. Coppen, J.J.W. and Cobb, A.L. (1983) The occurrence of iridoids in Plumeria and Allamandn. Phytochemistry 22, 125-128. Datta, S. and Satta, P.C. (1976) Bark drugs of Plumeria. Quarterly Journal of Crude Drug Research 14, 129-142. Geran, R.I., Greenberg, N.H., MacDonald, M.M., Schumacher, A.M. and Abbott, B.J. (1972) Protocols for screening chemical agents and natural products against animal tumors and other biological systems (Third Edition). Cancer Chemotherapy Reports 3, l-68. Hamburger, M.O. and Cordell, G.A. (1987) A direct bioautographic TLC assay for compounds possessing antibacterial activity. Journal of Natural Products 50, 19-22. Hamburger, M.O., Cordell, G.A., Tantivatana, P. and Ruangrungsi, N. (1987) Traditional medicinal plants of Thailand. VIII. Isoflavonoids of Dalbergia candenatensis. Journal of Natural Products 50, 696-699. Hostettmann. K.. Kizu, H. and Tomimori. T. (1982) Molluscicidal properties of various saponins. P/on/o Mcchcw 44. 3+35. Hostettmann, K. and Marston, A. research An update. In: Mollwcicides. J. Wiley and Sons, Jeven, M., Vanden Berghe, D.A.,
(1987) Plant molluscicide K.E. Mott (Ed.), Plant Chichester, pp. 299-320. Mertens, F., Vlietinck, A.
and Lammens, E. (1979) Screening of higher plants for biological activities. 1. Antimicrobial activity. Planta Medica 36, 311-321. Kardono, L.B.S., Tsauri, S., Padmawinata, K., Pezzuto, J.M. and Kinghom, A.D. (1990a) Cytotoxic Constituents of the
292
bark of flunrrriu ruhrlr collected in Indonesia. Journcd e/ Ninurul Products 53. 1447-1455.
Kardono. L.B.S.. Tsauri. S., Padmawinata, K.. Pezzuto. J.M. and Kinghorn. A.D. (1990b) Plumerubroside. a novel flavan-3-01 glycoside from the bark of Plumeriu ruhra Phytochemisrry
29. 2995--2997.
Kupchan, S.M., Giacobbe, T.J., Krull, J.S., Thomas, A.M., Eakin, M.A. and Fessler, D.C. (1970) Reaction of endocyciic c&unsaturated y-lactones with thiols. Journal of Orgunic Chemistry
35, 3539-3543.
Martin, G.A., Sanduja, R. and Alam, M. (1985) Twodimensional NMR studies of marine natural products. 2. Utilization of two-dimensional proton double quantum coherence NMR spectroscopy in natural products structure elucidation - Determination of long-range couplings in plumericin. Journal of Organic Chemistry 50, 2383-2386. Parkes, K.E.B. and Pattenden, G. (1988) A total formal synthesis of allamcin. An approach to antileukaemic iridoid lactones and formal syntheses of plumericin and allamandin. Journal of the Chemical Society, Perkin Transactions I, 1119-1134. Peckolt, G. (1942) Brazilian anthehnintic plants. Revista de la Flora Medicinal 9, 333
Quisumbing, E. (1951) Medicinal planrs of the Philippines.
Technical Bulletin 16, Department of Agricultural and Natural Resources, Manila, Republic of the Philippines. Sawhney, A.N., Khan, MR., Ndaalio, G., Nkunya, M.H.H. and Wevers, H. (1978) Studies on the rationale of African traditional medicine, Part III. Preliminary screening of medicinal plants for antifungal activity. Pakistan Journal of Scientific and Industrial Research 21, 193-196.
Sticher, 0. (1977) Plant mono-, di- and sesquiterpenoids with pharmacological or therapeutical activity. In: H. Wagner and P. Wolff (Eds.), New Natural Products with Pharmacological, Biological or Therapeutical Activity. Springer Verlang, Berlin, pp. 137-176. Topcu. R.. Che. C.-T., Cordell. G.A. and Ruangrungsi (1990) lridolactones from Alysb reinwrrrdti. Phytochemistry 29. 3 197. Trost, B.M., Balkovec, J.M. and Mao, M.K.-T. (1986) A total synthesis of plumericin, allamcin and allamandin. 2. A biomimedic strategy. Journal of rhe American Chemical Society 108, 4914-4983.
Vanden Berghe, D.A., Jeven, M., Mertens, F., Vlietink, A. and Lammens, E. (1978) Screening of higher plants for biological activities. II. Antiviral activity. Journal of Natural Products 41, 463467.
Wasuwat, S. (1967) A List of Thai Medicinal Plt~ms. Report No. 1 on Research project 17 ASRCT, Bangkok.
33
( I99 I ) 289-292
289
Elsevier Scientific Publishers Ireland Ltd.
Short Communication
Traditional
Matthias
medicinal plants of Thailand XVII. * Biologically active constituents of Plumeria rutwa
0. Hamburgera’**,
Geoffrey
A. Cordell”
and Nijsiri Ruangrungsib
“Program for Collaborative Research in the Pharmaceutical Sciences. College of Pharmacy, University of Illinois at Chicago, Chicago, Illinois 60612 (U.S. A.) and ‘Department of Pharmacognosy. Faculty of Pharmaceutical Sciences, Chulalongkorn University, Bangkok (Thailand)
(Accepted January 22, 1991) The compounds l-6 were isolated from the heartwood of Plumeria rubra, following bioactivity-directed fractionation. Plumericin 1 and isoplumericin 2 displayed molluscicidal, cytotoxic and antibacterial activity, 4-hydroxyacetophenone 3 was weakly cytotoxic, whereas the remaining glycosidic isolates (plumieride, 4, l3-O-coumaroylplumieride, test systems.
5; protoplumericine
A, 6) were inactive in all
Key wora!s: Plumeria rubra; molluscicidal activity; iridoids: cytotoxicity.
Introduction As a part of our continuing search for biologically active compounds, in particular antimicrobial and cytotoxic agents from medicinal plants of Thailand (Hamburger et al., 1987), we have examined Plumeria rubra L. (syn. P. acutifoiiu Poir.) (Apocynaceae). There are numerous reports on the ethnomedical use of plants belonging to the pantropic genus Plumeria. In South America, decoctions of leaves, bark or heartwood are used as an anthelmintic, purgative, emmenagogue, and for the topical treatment of various afflictions of the skin (Peckolt, 1942). The fruit pulp,
Correspondeme IO: Professor G.A. Cordell. Program for Collaborative Research in the Pharmaceutical Sciences. College of Pharmacy, University of Illinois at Chicago. Chicago. Illinois 60612 (U.S.A.) *For the previous paper. see Topcu et al.. 1990. l*Presem address: lnstitut de Pharmacognosie et Phytochimie. Universitt de Lausanne, CH-1005 Lausanne. Switzerland.
0378-8741/$03.50 0 1991 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
latex and stem bark serve in eastern Asia as an abortifacient and purgative (Quisumbing, 1951; Wasuwat, 1967; Datta and Satta, 1976; Sawhney et al., 1978). Extracts of various Plumeriu species have been shown to exhibit significant anantifungal and antiviral activity tibacterial, (Sticher, 1977; Vanden Berghe et al., 1978; Jeven 1979). The iridoids plumericin, et al., isoplumericin and fulvoplumierin have been isolated as the antifungal and antiviral agents of P. rubra (Sticher, 1977). The same compounds, along with some iridoid glycosides, have been identified by co-TLC in some other Plumeria species (Coppen and Cobb, 1983). While the present investigation was in progress, a phytochemical study of the iridoids from the roots of P. rubra was published without reporting, however, on the activity of the isolates (Abe et al., 1988), and our colleagues have reported on the isolation of six cytotoxic constituents (Kardono et al., 1990a) and a new flavan-3-01 glycoside (Kardono et al., 1990b) from the bark of P. rubra of Indonesian origin.
290
Material and Methods General procedures
These have been described elsewhere (Hamburger et al., 1987). Plant material
The heartwood of P. rubra was collected in 1987 in Phetchaburi Province, Thailand. The plant material was authenticated by one of us (N.R.), and a voucher specimen is deposited in the Botany Section, Technical Division, Department of Agriculture, Ministry of Agriculture and Cooperatives, Bangkok, Thailand. Extraction
and preliminary
biological
evaluation
The chopped heartwood (2.5 kg) was percolated at room temperature with EtOAc. Direct bioautography on precoated silica gel TLC plates (eluent: toluene/EtOAc (1: 1)) with B. subtilis ATCC 6633 as test organism (Hamburger and Cordell, 1987) revealed the presence of two antibacterial compounds at R/O.60 and 0.63. Preliminary small scale separation of the extract with toluene/EtOAc (1: 1) and monitoring cytotoxic activity against the P-388 cell line (Geran et al., 1972) revealed that the activity of the extract (EDSo 5 pg/ml) was located in the fraction containing the antibacterial compounds (EDSo < 0.5 &ml). Direct bioautography and the P-388 assay in vitro were used for monitoring the fractionation. Molluscicidal activity of the isolates was determined according to the procedure described by Hostettmann et al., 1982. l-6 A portion (44 g) of the EtOAc extract was subsequently separated by rapid column chromatography (CC) on silica gel using light petroleum/EtOAc (1: 1) as eluent, followed by MeOH. Four fractions (A-D) were collected. Recrystallization of fraction C (2.15 g) from toluene/EtOAc yielded 2 as colorless needles (1.22 g). Fraction B (6.3 g) was separated by CC on silica gel with toluene/EtOAc (9:l) as mobile phase. Six fractions were collected. Isoplumericin 2 (140 mg) was obtained through recrystallization of fraction 2 (400 mg). Fraction 4 (300 mg) was Isolation of compounds
submitted to reversed-phase chromatography on a Lobar RP-8 column (size B, Merck, Darmstadt, F.R.G.). Elution with MeOH/H,O (1:l) afforded 4-hydroxyacetophenone 3 (50 mg). A portion of fraction D (10.0 g) was further separated by CC on silica gel. Elution with CHCI,/MeOH/H@ (90:10:0.5 to 70:30:10) gave 7 fractions (l ’-7’). Fractions 3 ’ (2.0 g) and 7 ’ (1.6 g) consisted of 13-O-coumaroylplumieride 5 and protoplumericine A 6, respectively. A portion of each fraction was submitted to a final purification on Sephadex LH20 (MeOH). Plumieride 4 (120 mg) was isolated from fraction 5 ’ (500 mg) by CC on silica gel with CHClJ/MeOH/H20 (85:15:0.7) as mobile phase. All of the isolates were identified by MP, UV, IR, [&, EI- or D/(X-MS, ‘H and 13C-NMR (one- and two-dimensional) in comparison with published data (Yamauchi et al., 1981; Abe et al., 1984; Martin et al., 1985; Abe et al., 1988). In addition, glycosides 4-6 were submitted to acid (2N HCI, reflux for 1 h) and mild basic hydrolysis (NH3 10% in MeOH/H*O (2:1), RT). After the usual workup, the hydrolysates were analyzed by co-TLC with authentic samples of glucose, pcoumaric acid and plumieride 4. Results and Discussion In our preliminary screening, the EtOAc extract of the heartwood displayed cytotoxic activity against the P-388 cell line in vitro (ED50 5 pg/ml). Direct bioautography on TLC (Hamburger and
291
adds another facet to the broad range of biological activities of these iridoids, which have recently been the challenging target of various synthetic approaches (Trost et al., 1986; Parkes and Pattenden, 1988). Cordell, 1987), using B. subtilis as a test organism, revealed the presence of two antibacterial compounds in the same extract. Activity-guided fractionation led to the isolation of the cytotoxic and antibacterial iridoids plumericin 1 and isoplumericin 2, along with the moderately cytotoxic compound 4-hydroxyacetophenone 3 (EDSo 0.05, 0.06 and 3.2 &ml, respectively). In addition, the inactive iridoid glycosides plumieride 4, 13-Ocoumaroylplumieride 5 and protoplumericin A 6 were isolated from the polar fractions of the extract. Compounds 1 and 2 were found to exhibit significant molluscicidal activity against the schistosomiasis transmitting snail Biomphaluriu glabrata (LDloo 6.25 ppm). The other isolates 3-6 were inactive, when tested at 50 ppm. The bioactivity-guided fractionation of the extract yielded the two known compounds, plumericin 1 and isoplumericin 2, as the major antibacterial and cytotoxic principles in the heartThe weakly active of P. rubra. wood 4-hydroxyacetophenone 3 contributes only marginally to the cytotoxic activity of the extract. The three iridoid glycosides 4-6 are also known compounds, which have been reported from some Plumeria or Allamanda species. The strong molluscicidal activity of the tetracyclic iridoids 1 and 2 can be explained by the presence of an c~susceptible to methylene y-lactone moiety, undergo a Michael-type addition with biological nucleophiles (Kupchan et al., 1970). Other molluscicidal compounds with the similar structural features include the sesquiterpene lactones helenalin, pyrethrosin, 7-hydroxy-3-deoxyzaluzanin and the iridoid oruwacin (Hostettmann and Marston, 1987). In contrast, the a-substituted endocyclic o&unsaturated y-lactone ring has been found to be much less reactive towards nucleophiles (Kupchan et al., 1970). This may explain the lack of molluscicidal activity of glycosides 4-6. The discovery of the molluscicidal properties of plumericin 1 and isoplumericin 2
Acknowledgements
This work was supported by a grant from the Division of Cancer Treatment, National Cancer Institute, Bethesda, MD. We thank the Research Resources Center of the University of Illinois at Chicago for the provision of NMR and mass spectrometric facilities, and Prof. K. Hostettmann, University of Lausanne, Switzerland, for the molluscicidal tests. References Abe, F., Mori, T. and Yamauchi, T. (1984) Iridoids of Apocynaceae. III. Minor iridoids from Aliamanda neritfolia. Chemical and Pharmaceutical Bulletin 32, 2947-2956. Abe, F., Chen, R.F. and Yamauchi, T. (1988) Minor iridoids from the roots of Plumeria acuttfolia. Chemical and Pharmaceutical Bulletin 36, 278G2789. Coppen, J.J.W. and Cobb, A.L. (1983) The occurrence of iridoids in Plumeria and Allamandn. Phytochemistry 22, 125-128. Datta, S. and Satta, P.C. (1976) Bark drugs of Plumeria. Quarterly Journal of Crude Drug Research 14, 129-142. Geran, R.I., Greenberg, N.H., MacDonald, M.M., Schumacher, A.M. and Abbott, B.J. (1972) Protocols for screening chemical agents and natural products against animal tumors and other biological systems (Third Edition). Cancer Chemotherapy Reports 3, l-68. Hamburger, M.O. and Cordell, G.A. (1987) A direct bioautographic TLC assay for compounds possessing antibacterial activity. Journal of Natural Products 50, 19-22. Hamburger, M.O., Cordell, G.A., Tantivatana, P. and Ruangrungsi, N. (1987) Traditional medicinal plants of Thailand. VIII. Isoflavonoids of Dalbergia candenatensis. Journal of Natural Products 50, 696-699. Hostettmann. K.. Kizu, H. and Tomimori. T. (1982) Molluscicidal properties of various saponins. P/on/o Mcchcw 44. 3+35. Hostettmann, K. and Marston, A. research An update. In: Mollwcicides. J. Wiley and Sons, Jeven, M., Vanden Berghe, D.A.,
(1987) Plant molluscicide K.E. Mott (Ed.), Plant Chichester, pp. 299-320. Mertens, F., Vlietinck, A.
and Lammens, E. (1979) Screening of higher plants for biological activities. 1. Antimicrobial activity. Planta Medica 36, 311-321. Kardono, L.B.S., Tsauri, S., Padmawinata, K., Pezzuto, J.M. and Kinghom, A.D. (1990a) Cytotoxic Constituents of the
292
bark of flunrrriu ruhrlr collected in Indonesia. Journcd e/ Ninurul Products 53. 1447-1455.
Kardono. L.B.S.. Tsauri. S., Padmawinata, K.. Pezzuto. J.M. and Kinghorn. A.D. (1990b) Plumerubroside. a novel flavan-3-01 glycoside from the bark of Plumeriu ruhra Phytochemisrry
29. 2995--2997.
Kupchan, S.M., Giacobbe, T.J., Krull, J.S., Thomas, A.M., Eakin, M.A. and Fessler, D.C. (1970) Reaction of endocyciic c&unsaturated y-lactones with thiols. Journal of Orgunic Chemistry
35, 3539-3543.
Martin, G.A., Sanduja, R. and Alam, M. (1985) Twodimensional NMR studies of marine natural products. 2. Utilization of two-dimensional proton double quantum coherence NMR spectroscopy in natural products structure elucidation - Determination of long-range couplings in plumericin. Journal of Organic Chemistry 50, 2383-2386. Parkes, K.E.B. and Pattenden, G. (1988) A total formal synthesis of allamcin. An approach to antileukaemic iridoid lactones and formal syntheses of plumericin and allamandin. Journal of the Chemical Society, Perkin Transactions I, 1119-1134. Peckolt, G. (1942) Brazilian anthehnintic plants. Revista de la Flora Medicinal 9, 333
Quisumbing, E. (1951) Medicinal planrs of the Philippines.
Technical Bulletin 16, Department of Agricultural and Natural Resources, Manila, Republic of the Philippines. Sawhney, A.N., Khan, MR., Ndaalio, G., Nkunya, M.H.H. and Wevers, H. (1978) Studies on the rationale of African traditional medicine, Part III. Preliminary screening of medicinal plants for antifungal activity. Pakistan Journal of Scientific and Industrial Research 21, 193-196.
Sticher, 0. (1977) Plant mono-, di- and sesquiterpenoids with pharmacological or therapeutical activity. In: H. Wagner and P. Wolff (Eds.), New Natural Products with Pharmacological, Biological or Therapeutical Activity. Springer Verlang, Berlin, pp. 137-176. Topcu. R.. Che. C.-T., Cordell. G.A. and Ruangrungsi (1990) lridolactones from Alysb reinwrrrdti. Phytochemistry 29. 3 197. Trost, B.M., Balkovec, J.M. and Mao, M.K.-T. (1986) A total synthesis of plumericin, allamcin and allamandin. 2. A biomimedic strategy. Journal of rhe American Chemical Society 108, 4914-4983.
Vanden Berghe, D.A., Jeven, M., Mertens, F., Vlietink, A. and Lammens, E. (1978) Screening of higher plants for biological activities. II. Antiviral activity. Journal of Natural Products 41, 463467.
Wasuwat, S. (1967) A List of Thai Medicinal Plt~ms. Report No. 1 on Research project 17 ASRCT, Bangkok.