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Levels of lantadenes, bioactive pentacyclic triterpenoids, in young and mature leaves of Lantana camara var. aculeata
Fitoterapia 71 Ž2000. 487᎐491
Levels of lantadenes, bioactive pentacyclic triterpenoids, in young and mature leaves of Lantana camara var. aculeata Om P. SharmaU , Anita Singh, Sarita Sharma Biochemistry Laboratory, Indian Veterinary Research Institute, Regional Station, Kangra Valley, Himachal Pradesh, Palampur 176 061, India
Received 16 November 1999; accepted in revised form 9 February 2000
Abstract Levels of the lantadene pentacyclic triterpenes were quantified in young and mature leaf samples of Lantana camara var. aculeata, by HPLC. The amount of different lantadenes Žmgr100 g dry wt.. in young and mature leaf samples, respectively, was: lantadene A, 491.5 " 6.3, 805.9 " 52.8; lantadene B, 347.0 " 3.0, 522.3 " 37.1; lantadene C, 191.3 " 10.3, 424.8 " 39.1; lantadene D, 49.7 " 5.3, 177.4 " 19.0; reduced lantadene A, 19.1 " 2.3, 28.7 " 4.5; reduced lantadene B, 13.0 " 1.3, 18.6 " 1.2; and 22 -hydroxyoleanonic acid, 82.5 " 11.4, 167.7 " 30.1. 䊚 2000 Elsevier Science B.V. All rights reserved. Keywords: Lantana camara var. aculeata; Lantadenes; Triterpenoids
1. Introduction Lantana camara L. ŽVerbenaceae ., lantana, is listed as one of the important medicinal plants of the world w1x. The lantadenes, pentacyclic triterpenes from the
U
Corresponding author. Fax: q91-1894-33063. E-mail address: [email protected] ŽO.P. Sharma.. 0367-326Xr00r$ - see front matter 䊚 2000 Elsevier Science B.V. All rights reserved. PII: S 0 3 6 7 - 3 2 6 X Ž 0 0 . 0 0 1 5 6 - 8
488
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
lantana plant ŽFig. 1. have a number of biological activities. Lantadenes inhibited the activity of the Epstein᎐Barr virus, lantadene B being the most active w2x. Lantadenes had an inhibitory effect on the two-stage carcinogenesis of mouse skin papilloma induced by 7,12-dimethylbenzŽa. anthracene and 12-O-tetradecanoyl phorbol-13-acetate w3x. Pentacyclic triterpenoids have a number of pharmacological properties such as antitumor, anti-AIDS, antibacterial and anti-inflammatory activities w4᎐11x. Oleanolic acid and ursolic acid, which contain the pentacyclic core part of the lantana triterpenoids w12,13x, inhibit tumor initiation and promotion w11,14x. Oleanolic acid has hepatoprotective activity as well w15x. Pentacyclic triterpenoids from L. camara and L. indica showed antibacterial activity w16,17x. Methanolic extracts of L. camara leaves inhibited human thrombin w18x. Thrombin inhibitory activity was associated with a series of 5,5-trans-fused cyclic lactone containing euphane triterpenes w18,19x. We report here the levels of different lantadenes in the young and mature leaf sample of L. camara var. aculeata. This information would be useful in choosing the plant sample for isolation of these compounds for drug research and related studies.
Fig. 1. Chemical structure of lantadenes.
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
489
2. Experimental 2.1. Plant material The samples of young and mature leaves of L. camara var. aculeata were collected from the vicinity of the Indian Veterinary Research Institute Palampur. The samples were dried at 70⬚C. Fine powder of 1-mm particle size was prepared using a Cyclotec grinder ŽTecator, Sweden.. 2.2. Reference substances Purified samples of lantadenes ŽFig. 1. previously isolated w20x from L. camara var. aculeata and authenticated by spectroscopic analysis were used. 2.3. Extraction and HPLC analysis Leaf powder Ž2 g. samples were extracted with 3 = 40-ml MeCN. The extracts were combined and the solvent was removed in vacuo. The residue was dissolved in 10 ml of dichloromethane. Aliquots Ž1 ml. were taken and the solvent was removed in vacuo. The residue was dissolved in 10 ml HPLC grade methanol. Aliquots Ž20 l. were taken for HPLC analysis w20x. The HPLC system used was from Waters ŽUSA. with 510 pump, Rheodyne injector with a 20-l loop, 490E multichannel detector and Millennium 2010 dataprocessor. The detector was set at 210 nm w20x.
3. Results and discussion The content of different lantadenes in young and mature leaf samples of L. camara var. aculeata is shown in Table 1. An HPLC profile of an extract is given in Fig. 2. The content of all the triterpene acids was lower in young leaves than in mature leaves. For lantadene C and lantadene D, the amount in mature leaves was more than double that in young leaves ŽTable 1.. Earlier, quantitation of lan-
Table 1 Lantadenes content Žmgr100 g dry wt.. of young and mature leaves of Lantana camara var. aculeataa Constituent
Young leaves
Mature leaves
Lantadene A Lantadene B Lantadene C Lantadene D Reduced lantadene A Reduced lantadene B 22 -Hydroxyoleanonic acid
491.5 " 6.3 347.0 " 3.0 191.3 " 10.3 49.7 " 5.3 19.1 " 2.3 13.0 " 1.3 82.5 " 11.4
805.9 " 52.8 522.3 " 37.1 424.8 " 39.1 177.4 " 19.0 28.7 " 4.5 18.6 " 1.2 167.7 " 30.1
a
Values are mean " S.D., n s 3.
490
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
Fig. 2. HPLC profile of lantadenes in Ža. young leaves and Žb. mature leaves of Lantana camara var. aculeata. LA, lantadene A; LB, lantadene B; LC, lantadene C; LD, lantadene D; RLA, reduced lantadene A; RLB, reduced lantadene B; OA, 22 -Hydroxyoleanonic acid.
tadenes has been done by batch extractions, crystallization and gravimetry w12,21x. Hart et al. w12x analyzed lantadenes in different taxa of Lantana camara in Australia. L. camara Common Pink-edged Red contained lantadene A and B as the major lantadenes. This taxon contained small amounts of reduced lantadene A and reduced lantadene B as well w12x. In general, the taxa toxic to livestock were rich in lantadene A and B. L. camara Common-Pink which is non-toxic did not contain lantadene A or B w12x. Earlier, lantadene C has been reported as a minor constituent in leaves and stem samples of L. camara Southern-Red w12x. The biological activity of 22 -hydroxyoleanonic acid, which is present in lantana leaves ŽTable 1. and can be prepared by hydrolysis of lantadenes w22x, is not known. Pentacyclic triterpenes are attracting increasing attention in the development of
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
491
novel anti-inflammatory drugs w23x. The core structure of the lantadenes is similar to that of pentacyclic triterpenoids, which show inhibition of protein kinase w5x.
References w1x Ross IA. Medicinal plants of the world. Chemical constituents, traditional and modern medical uses. New Jersey: Humana Press Inc, 1999. w2x Inada A, Nakanish T, Tokuda H, Nishino H, Iwasima A, Sharma OP. Planta Med 1995;61:558. w3x Inada A, Nakanish T, Tokuda H, Nishino H, Sharma OP. Planta Med 1997;63:272. w4x Kapoor VK, Chawla AS. J Sci Ind Res 1986;45:503. w5x Pengsuparp T, Cai L, Fong HHS, Kinghorn AD, Pezzuto JM. J Nat Prod 1994;57:415. w6x Fujioka T, Kashiwada Y, Kilkuskie RE et al. J Nat Prod 1994;57:243. w7x Li HY, Sun NJ, Kashiwada Y et al. J Nat Prod 1993;56:1130. w8x Xu HX, Zeng FQ, Wan M, Sim KW. J Nat Prod 1996;59:643. w9x Nick A, Wright AD, Sticher O. J Nat Prod 1994;57:1245. w10x Loggia RD, Tubaro A, Sosa S, Becker H, Saar St, Isaac O. Planta Med 1994;516. w11x Liu J. J Ethanopharmacol 1995;49:57. w12x Hart NK, Lamberton JA, Sioumis AA, Suares H. Aust J Chem 1976;29:655. w13x Sharma OP, Sharma PD. J Sci Ind Res 1989;48:471. w14x Lee KH, Lin YM, Wu TS et al. Planta Med 1988;54:308. w15x Misra LN, Dixit AK, Sharma RP. Planta Med 1997;63:582. w16x Verma DK, Singh SK, Nath G, Tripathy V. Indian Drugs 1997;34:390. w17x Barre JT, Bowden BF, Coll JC et al. Phytochemistry 1997;45:321. w18x O’Neill MJ, Lewis JA, Noble HM et al. J Nat Prod 1998;61:1328. w19x Weir MP, Bethell SS, Cleasby A et al. Biochemistry 1998;37:6645. w20x Sharma OP, Sharma S, Dawra RK. J Chromatogr 1997;786:181. w21x Johns SR, Lamberton JA, Morton TC, Suares H, Willing RI. Aust J. Chem 1983;36:1895. w22x Beeby PJ. Aust J Chem 1978;31:1313. w23x Safayhi H, Sailer ER. Planta Med 1997;63:487.
Levels of lantadenes, bioactive pentacyclic triterpenoids, in young and mature leaves of Lantana camara var. aculeata Om P. SharmaU , Anita Singh, Sarita Sharma Biochemistry Laboratory, Indian Veterinary Research Institute, Regional Station, Kangra Valley, Himachal Pradesh, Palampur 176 061, India
Received 16 November 1999; accepted in revised form 9 February 2000
Abstract Levels of the lantadene pentacyclic triterpenes were quantified in young and mature leaf samples of Lantana camara var. aculeata, by HPLC. The amount of different lantadenes Žmgr100 g dry wt.. in young and mature leaf samples, respectively, was: lantadene A, 491.5 " 6.3, 805.9 " 52.8; lantadene B, 347.0 " 3.0, 522.3 " 37.1; lantadene C, 191.3 " 10.3, 424.8 " 39.1; lantadene D, 49.7 " 5.3, 177.4 " 19.0; reduced lantadene A, 19.1 " 2.3, 28.7 " 4.5; reduced lantadene B, 13.0 " 1.3, 18.6 " 1.2; and 22 -hydroxyoleanonic acid, 82.5 " 11.4, 167.7 " 30.1. 䊚 2000 Elsevier Science B.V. All rights reserved. Keywords: Lantana camara var. aculeata; Lantadenes; Triterpenoids
1. Introduction Lantana camara L. ŽVerbenaceae ., lantana, is listed as one of the important medicinal plants of the world w1x. The lantadenes, pentacyclic triterpenes from the
U
Corresponding author. Fax: q91-1894-33063. E-mail address: [email protected] ŽO.P. Sharma.. 0367-326Xr00r$ - see front matter 䊚 2000 Elsevier Science B.V. All rights reserved. PII: S 0 3 6 7 - 3 2 6 X Ž 0 0 . 0 0 1 5 6 - 8
488
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
lantana plant ŽFig. 1. have a number of biological activities. Lantadenes inhibited the activity of the Epstein᎐Barr virus, lantadene B being the most active w2x. Lantadenes had an inhibitory effect on the two-stage carcinogenesis of mouse skin papilloma induced by 7,12-dimethylbenzŽa. anthracene and 12-O-tetradecanoyl phorbol-13-acetate w3x. Pentacyclic triterpenoids have a number of pharmacological properties such as antitumor, anti-AIDS, antibacterial and anti-inflammatory activities w4᎐11x. Oleanolic acid and ursolic acid, which contain the pentacyclic core part of the lantana triterpenoids w12,13x, inhibit tumor initiation and promotion w11,14x. Oleanolic acid has hepatoprotective activity as well w15x. Pentacyclic triterpenoids from L. camara and L. indica showed antibacterial activity w16,17x. Methanolic extracts of L. camara leaves inhibited human thrombin w18x. Thrombin inhibitory activity was associated with a series of 5,5-trans-fused cyclic lactone containing euphane triterpenes w18,19x. We report here the levels of different lantadenes in the young and mature leaf sample of L. camara var. aculeata. This information would be useful in choosing the plant sample for isolation of these compounds for drug research and related studies.
Fig. 1. Chemical structure of lantadenes.
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
489
2. Experimental 2.1. Plant material The samples of young and mature leaves of L. camara var. aculeata were collected from the vicinity of the Indian Veterinary Research Institute Palampur. The samples were dried at 70⬚C. Fine powder of 1-mm particle size was prepared using a Cyclotec grinder ŽTecator, Sweden.. 2.2. Reference substances Purified samples of lantadenes ŽFig. 1. previously isolated w20x from L. camara var. aculeata and authenticated by spectroscopic analysis were used. 2.3. Extraction and HPLC analysis Leaf powder Ž2 g. samples were extracted with 3 = 40-ml MeCN. The extracts were combined and the solvent was removed in vacuo. The residue was dissolved in 10 ml of dichloromethane. Aliquots Ž1 ml. were taken and the solvent was removed in vacuo. The residue was dissolved in 10 ml HPLC grade methanol. Aliquots Ž20 l. were taken for HPLC analysis w20x. The HPLC system used was from Waters ŽUSA. with 510 pump, Rheodyne injector with a 20-l loop, 490E multichannel detector and Millennium 2010 dataprocessor. The detector was set at 210 nm w20x.
3. Results and discussion The content of different lantadenes in young and mature leaf samples of L. camara var. aculeata is shown in Table 1. An HPLC profile of an extract is given in Fig. 2. The content of all the triterpene acids was lower in young leaves than in mature leaves. For lantadene C and lantadene D, the amount in mature leaves was more than double that in young leaves ŽTable 1.. Earlier, quantitation of lan-
Table 1 Lantadenes content Žmgr100 g dry wt.. of young and mature leaves of Lantana camara var. aculeataa Constituent
Young leaves
Mature leaves
Lantadene A Lantadene B Lantadene C Lantadene D Reduced lantadene A Reduced lantadene B 22 -Hydroxyoleanonic acid
491.5 " 6.3 347.0 " 3.0 191.3 " 10.3 49.7 " 5.3 19.1 " 2.3 13.0 " 1.3 82.5 " 11.4
805.9 " 52.8 522.3 " 37.1 424.8 " 39.1 177.4 " 19.0 28.7 " 4.5 18.6 " 1.2 167.7 " 30.1
a
Values are mean " S.D., n s 3.
490
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
Fig. 2. HPLC profile of lantadenes in Ža. young leaves and Žb. mature leaves of Lantana camara var. aculeata. LA, lantadene A; LB, lantadene B; LC, lantadene C; LD, lantadene D; RLA, reduced lantadene A; RLB, reduced lantadene B; OA, 22 -Hydroxyoleanonic acid.
tadenes has been done by batch extractions, crystallization and gravimetry w12,21x. Hart et al. w12x analyzed lantadenes in different taxa of Lantana camara in Australia. L. camara Common Pink-edged Red contained lantadene A and B as the major lantadenes. This taxon contained small amounts of reduced lantadene A and reduced lantadene B as well w12x. In general, the taxa toxic to livestock were rich in lantadene A and B. L. camara Common-Pink which is non-toxic did not contain lantadene A or B w12x. Earlier, lantadene C has been reported as a minor constituent in leaves and stem samples of L. camara Southern-Red w12x. The biological activity of 22 -hydroxyoleanonic acid, which is present in lantana leaves ŽTable 1. and can be prepared by hydrolysis of lantadenes w22x, is not known. Pentacyclic triterpenes are attracting increasing attention in the development of
O.P. Sharma et al. r Fitoterapia 71 (2000) 487᎐491
491
novel anti-inflammatory drugs w23x. The core structure of the lantadenes is similar to that of pentacyclic triterpenoids, which show inhibition of protein kinase w5x.
References w1x Ross IA. Medicinal plants of the world. Chemical constituents, traditional and modern medical uses. New Jersey: Humana Press Inc, 1999. w2x Inada A, Nakanish T, Tokuda H, Nishino H, Iwasima A, Sharma OP. Planta Med 1995;61:558. w3x Inada A, Nakanish T, Tokuda H, Nishino H, Sharma OP. Planta Med 1997;63:272. w4x Kapoor VK, Chawla AS. J Sci Ind Res 1986;45:503. w5x Pengsuparp T, Cai L, Fong HHS, Kinghorn AD, Pezzuto JM. J Nat Prod 1994;57:415. w6x Fujioka T, Kashiwada Y, Kilkuskie RE et al. J Nat Prod 1994;57:243. w7x Li HY, Sun NJ, Kashiwada Y et al. J Nat Prod 1993;56:1130. w8x Xu HX, Zeng FQ, Wan M, Sim KW. J Nat Prod 1996;59:643. w9x Nick A, Wright AD, Sticher O. J Nat Prod 1994;57:1245. w10x Loggia RD, Tubaro A, Sosa S, Becker H, Saar St, Isaac O. Planta Med 1994;516. w11x Liu J. J Ethanopharmacol 1995;49:57. w12x Hart NK, Lamberton JA, Sioumis AA, Suares H. Aust J Chem 1976;29:655. w13x Sharma OP, Sharma PD. J Sci Ind Res 1989;48:471. w14x Lee KH, Lin YM, Wu TS et al. Planta Med 1988;54:308. w15x Misra LN, Dixit AK, Sharma RP. Planta Med 1997;63:582. w16x Verma DK, Singh SK, Nath G, Tripathy V. Indian Drugs 1997;34:390. w17x Barre JT, Bowden BF, Coll JC et al. Phytochemistry 1997;45:321. w18x O’Neill MJ, Lewis JA, Noble HM et al. J Nat Prod 1998;61:1328. w19x Weir MP, Bethell SS, Cleasby A et al. Biochemistry 1998;37:6645. w20x Sharma OP, Sharma S, Dawra RK. J Chromatogr 1997;786:181. w21x Johns SR, Lamberton JA, Morton TC, Suares H, Willing RI. Aust J. Chem 1983;36:1895. w22x Beeby PJ. Aust J Chem 1978;31:1313. w23x Safayhi H, Sailer ER. Planta Med 1997;63:487.