Engineering

A New Larvicidal Lignan from Piper fimbriulatum

Description
A New Larvicidal Lignan from Piper fimbriulatum
Categories
Published
of 4
All materials on our website are shared by users. If you have any questions about copyright issues, please report us to resolve them. We are always happy to assist you.
Related Documents
Share
Transcript
  A New Larvicidal Lignan from  Piper fimbriulatum Pablo N. Solı´s 1 , Dionisio Olmedo 1 , Norio Nakamura 2 , A´ngela I. Calder  oon 1 , Masao Hattori 2 , and Mahabir P. Gupta 11 Centro de Investigaciones Farmacogn  oosticas de la Flora Paname ~ nna (CIFLORPAN), Facultad de Farmacia,Universidad de Panam  aa, Panama, Republica de Panam  aa;  2 Institute of Natural Medicine, Toyama Medical andPharmaceutical University, Sugitani, Toyama, Japan Abstract A new lignan, 3,4,5 0 -trimethoxy-3 0 ,4 0 -methylenedioxy-7,9 0 :7 0 ,9 diepoxylignan ( 1 ) (6-[4-(3,4-dimethoxy-phenyl)-tetrahydro-furo[3,4- c ]furan-1-yl]-4-methoxy-benzo[1,3]dioxole) together with two known lignans, 7 0 - epi  -sesarte-min ( 2 ) and diayangambin ( 3 ), and a known flavonoid,5-hydroxy-7,4 0 -dimethoxyflavone ( 4 ), were isolated fromthe leaves of   Piper fimbriulatum  C. DC. Their structureswere assigned by a combination of one- and two-dimen-sional NMR techniques. 7 0 - epi  -Sesartemin ( 2 ) showed thehighest larvicidal activity against  Aedes aegypti   (LC 100 17.6 m g = ml) and weak antiplasmodial (IC 50  7.0 m g = ml)and antitrypanosomal (IC 50  39.0 m g = ml) activities. Noneofthecompoundswasactiveagainst Leishmaniamexicana . Keywords:  Aedes aegypti  , lignan,  Piper fimbriulatum , Plasmodium falciparum . Introduction Piper fimbriulatum  (Piperaceae) C. DC. is a slender shrubof 5m height commonly known as  canotillo  in CentralAmerica and is distributed in shaded sites in moist forestsand along streams and roadsides of Colombia, CostaRica, and Panama (Tebbs, 1989). In our ongoing searchfor larvicidal principles, the chloroform extract of theleaves of   Piper fimbriulatum  showed larvicidal activityagainst  Aedes aegypti   (LC 100  37.5 m g = ml). Bioassay-guided fractionation of the chloroform extract of theleaves of   P. fimbriulatum , using larvae of   Aedes aegypti  ,resulted in the isolation of one new compound 3,4,5 0 -trimethoxy-3 0 ,4 0 -methylenedioxy-7,9 0 :7 0 ,9 diepoxylignan( 1 ) (Fig. 1) in addition to two known lignans ( 2 ,  3 ) anda known flavonoid ( 4 ). Compounds  1 ,  2 , and  3  were alsotested for their antiparasitic activity against  Plasmodium falciparum ,  Leishmania mexicana , and  Trypanosomacruzi  . The structure elucidation of the new natural pro-duct  1  and the larvicidal and antiparasitic activities of compounds  1  –  3  are discussed herein. Accepted: March 1, 2005 Address correspondence to : Mahabir P. Gupta, Centro de Investigaciones Farmacogn  oosticas de la Flora Paname ~ nna, Facultad deFarmacia, Universidad de Panam  aa, Apartado 10767, Panama, Republica de Panam  aa. Tel.:  þ 507-269-7655; Fax:  þ 507-264-0789;E-mail: cytedqff@ancon.up.ac.pa Figure 1.  Structures of compounds 1–4.DOI: 10.1080/13880200590951865  # 2005 Taylor & Francis Ltd.Pharmaceutical Biology2005, Vol. 43, No. 4, pp. 378–381  Materials and Methods Melting points are uncorrected. Optical rotations weremeasured with a Perkin-Elmer 141 polarimeter. IR spec-tra were recorded on a Perkin Elmer 1310 spectrophoto-meter. NMR spectra were recorded on a Varian UNITY500 spectrometer in CDCl 3  at 500MHz and 125MHz for 13 C NMR. Mass spectra were obtained on a Jeol JMS-AX 505 HAD mass spectrometer at 70eV. Silica gel 60(Merck 70-230 mesh) (Darmstadt, Germany) and Sepha-dex LH ¼ 20 SIGMA (St. Louis, MO, USA) were usedfor column chromatography. Silica gel plates 60 F 254 O. 25mm thickness, Merck (Darmstadt, Germany) wereused for TLC. Plant material Piper fimbriulatum  C. DC. was collected in Altos deCampana National Park, Dichapetalaceae trail, on April11, 1997. Prof. Mireya D. Correa, Director of theHerbarium of the University of Panama (PMA), wherevoucher specimens (FLORPAN 2790) are deposited,established its taxonomic identification. Extraction and isolation Air-dried powdered leaves of   P. fimbriulatum  (619.0g)were extracted with CHCl 3  by percolation at room tem-perature and the extract concentrated to obtain a residue(63.0g; 10.0 % ). The chloroform extract was testedagainst larvae of   Aedes aegypti  . The chloroform extract(8.0g) was subjected to bioactivity-guided fractionationby repeated column chromatography using silica geland gradient of CHCl 3  MeOH (99:1) ! MeOH 100 % aseluent. Sixteen fractions (100ml each one) were obtainedand submitted to larvicidal assay. Two active fractions(PF-A; PF-B) were combined and submitted to columnchromatography with CH 2 Cl 2  EtOAc (95:5) ! CH 2 Cl 2  EtOAc (85:15). Four major compounds were elutedand purified by crystallization in MeOH from fractions5, 6, 7, and 8, respectively: 3,4,5 0 -trimethoxy-3 0 ,4 0 -methylenedioxy-7,9 0 :7 0 ,9 diepoxylignan ( 1 ) (116mg,0.018 % ), 7 0 - epi  -sesartemin ( 2 ) (158mg, 0.025 % ), diayan-gambin ( 3 ) (125mg, 0.020 % ), and 5-hydroxy-7,4 0 -dimethoxyflavone ( 4 ) (82mg, 0.013 % ). 3,4,5 0 -Trimethoxy-3 0 ,4 0 -methylenedioxy-7,9 0 :7  0 ,9diepoxylignan ( 1 ) Colorless needles from MeOH. mp (uncorr.): 134–136;[ a ] D þ 115.2 ( c  0.241, CHCl 3 ); UV (EtOH)  k max  (log  e )204 (5.39), 233 (4.54), 278 (4.16)nm; IR (KBr)  n  max 2920, 2850, 1630, 1520, 1460, 1270, 1140, 1090,730cm  1 ;  1 H NMR (CDCl 3 , 500MHz)  d  6.93 (1H,  d  , J  ¼ 1.9Hz, H-2), 6.89 (1H,  dd  ,  J  ¼ 1.9, 8.1, H-6), 6.84(1H,  d  ,  J  ¼ 8.1, H-5), 6.60 (1H,  s , H-6 0 ), 6.52 (1H,  s ,H-2 0 ), 5.98 (2H,  s , H-3 0 a), 4.83 (1H,  d  ,  J  ¼ 5.3, H-7 0 ),4.42 (1H,  d  ,  J  ¼ 7.3, H-7), 4.12 (1H,  d  ,  J  ¼ 9.6, H-9 a ),3.93 (3H,  s , H-5 0 a), 3.90 (3H,  s , H-3a), 3.88 (3H,  s , H-4a), 3.84 (1H,  d  ,  J  ¼ 9.4, H-9 0 b ), 3.83 (1H,  d  ,  J  ¼ 9.6,H-9 b ), 3.34 (1H,  m , H-9 0 a ), 3.32 (1H,  m , H-8 0 ), 2.91(1H,  m , H-8);  13 C NMR (CDCl 3 , 125MHz)  d  149.2 (C,C-4), 148.8 (C, C-3 0 ), 148.7 (C, C-3), 143.5 (C, C-5 0 ),134.1 (C, C-4 0 ), 133.5 (C, C-1), 132.9 (C, C-1 0 ), 118.5(CH, C-6), 110.9 (CH, C-5), 109.0 (CH, C-2), 104.8(CH, C-6 0 ), 101.4 (OCH 2 O, C-3 0 a), 99.9 (CH, C-2 0 ),87.6 (CH, C-7), 82.0 (CH, C-7 0 ), 70.9 (CH 2 , C-9), 69.6(CH 2 , C-9 0 ), 56.6 (OCH 3 , C-5 0 a), 55.9 (OCH 3 , C-4a),55.9 (OCH 3 , C-3a), 54.4 (CH, C-8), 50.1 (CH, C-8 0 ).HSQC, HMBC, and NOESY were used to confirm theseassignments. EIMS  m = z  400 (100), 219 (18), 191 (20), 181(10), 180 (10), 179 (15), 177 (15), 166 (10), 165 (80), 152(8), 151 (50). HREIMS  m = z  400.1550 (calcd forC 22 H 24 O 7 , 400.1522). 7-epi-Sesartemin ( 2 ) UV (EtOH)  k max  (log  e ) 208 (4.94), 238 (4.00), 275 (3.46); 1 H NMR (CDCl 3 , 500MHz)  d  6.59 (1H,  d  ,  J  ¼ Hz,H-6 0 ), 6.52 (1H,  d  ,  J  ¼ 0.85Hz, H-2 0 ), 6.51 (2H,  s , H-2,H-6), 5.98 (2H,  s , H-3 0 a), 4.83 (1H,  d  ,  J  ¼ 5.12Hz,H-7 0 ), 4.41 (1H,  d  ,  J  ¼ 7.26, H-7), 4.14 (1H,  d  , J  ¼ 9.61Hz, H-9b), 3.93 (3H,  s , H-5 0 a), 3.91–3.85 (2H, m , H-9a, H-9 0 a), 3.89 (6H,  s , H-3a, H-5a), 3.84 (3H,  s ,H-4a);  13 C NMR (CDCl 3 , 500MHz)  d  153.4 (C, C-3,C-5), 148.80 (C, C-3 0 ), 143.5 (C, C-5 0 ), 137.4 (C, C-4),136.70 (C, C-1), 134.1 (C, C-4 0 ), 132.9 (C, C-1 0 ), 104.8(CH, C-6 0 ), 102.8 (CH, C-2), 101.4 (CH 2 , C-3 0 a), 99.8(CH, C-2 0 ), 87.8 (CH, C-7 0 ), 81.9 (CH, C-7), 70.9 (CH 2 ,C-9), 69.7 (CH 2 , C-9 0 ), 60.8 (CH 3 , C-4a), 56.6 (CH 3 ,C-5 0 a), 56.1 (CH 3 , C-3a, C-5a), 54.5 (CH, C-8), 50.0(CH, C-8 0 ); EIMS  m = z  430 [M] þ (100), 400 (15), 249(12), 224 (12), 207 (13), 195 (23), 181 (30), 165 (27). Diayangambin ( 3 ) UV (EtOH)  k max  (log  e ) 208 (5.38), 240 (4.29), 274 (3.63); 1 H NMR (CDCl 3 , 500MHz)  d  6.61 (4H,  s , H-2, H-6,H-2 0 , H-6 0 ), 4.92 (2H,  d  ,  J  ¼ 4.9Hz, H-7, H-7 0 ), 3.89(12H,  s , CH 3 O-4a, CH 3 O-4 0 a), 3.85 (6H,  s , CH 3 O-3a,CH 3 O-5a, CH 3 O-3 0 a, CH 3 O-5 0 a), 3.74 (2H,  dd  ,  J  ¼ 1.3,9.6Hz, H-9b, H-9 0 b), 3.59 (2H,  dd  , H-9a, H-9 0 a), 3.21(2H,  m , H-8, H-8 0 );  13 C NMR (CDCl 3 , 500MHz)  d 153.2 (C, C-3, C-5, C-3 0 , C-5 0 ), 136.9 (C, C-4, C4 0 ),134.5 (C, C-1, C-1 0 ), 103.0 (C, C-2, C-6, C-2 0 , C-6 0 ),84.0 (CH, C-7, C-7 0 ), 68.8 (CH 2 , C-9, C-9 0 ), 60.8(CH 3 O, C-4a, C-4 0 a), 56.0 (CH 3 O, C-3a, C-5a, C-3 0 a,C-5 0 a), 49.4 (CH, C-8, C-8 0 ). EIMS  m = z  446 [M] þ (100), 250 (25), 197 (38), 181 (62), 169 (25). 5-Hydroxy-7,4 0 -dimethoxyflavone ( 4 ) 1 H NMR (CDCl 3 , 500MHz)  d  10.50 (1H,  s , OH-5a), 7.82(2H,  d  ,  J  ¼ 9.1, H-3 0 , H-5 0 ), 7.00 (2H,  d  ,  J  ¼ 9.1, H-2 0 ,Larvicidal lignan from  Piper fimbriulatum  379  H-6 0 ), 6.55 (1H,  s , H-3), 6.45 (1H,  d  ,  J  ¼ 2.1, H-8), 6.34(1H,  d  ,  J  ¼ 2.1, H-6), 3.88 (3H,  s , CH 3 O-7a), 3.87 (3H,  s ,CH 3 O-4a);  13 C NMR (, 500MHz)  d  182.5 CDCI 3 (C, C-4), 165.4 (C, C-7), 164.0 (C, C-5), 162.6 (C, C-2), 162.2(C, C-4 0 ), 157.7 (C, C-4a, C-8a), 128.0 (CH, C-2 0 , C-6 0 ),123.5 (C, C-1 0 ), 114.5 (CH, C-3 0 , C-5 0 ), 104.33 (CH, C-3), 98.1 (CH, C-6), 92.6 (CH, C-8), 55.8 (C, CH 3 O-7a),55.6 (C, CH 3 O-4 0 a); EIMS  m = z  298 [M] þ (100). Larvicidal activity The larvicidal activity was determined on larvae of   Aedesaegypti   in a 96-well plate according to the methodof Ceplenau (1993), modified by Solı´s et al. (1996).Tetrametrin was used as a standard larvicidal agent. Antimalarial activity The antimalarial activity was determined with a chloro-quine-resistant (Indochina clone W2) strain of   Plasmo-dium falciparum  according to the novel DNA-basedmicrofluorimetric method of Corbett et al. (2004). Chlo-roquine was used as a standard antimalarial agent. Antileishmanial activity The antileishmanial activity was determined on WHOreference strain of   Leishmania mexicana  MOHM = B2 = 82 = BELZ promastigotes according to the protocol of Cornelly et al. (2003). Amphotericin B was used as astandard antileishmanial agent. Antitrypanosomal activity The intracellular assay was carried out according toBuckner et al.(1996) on the recombinant Tulahuen cloneC4 of   Trypanosoma cruzi   (trypomastigotes), whichexpresses  b -galactosidase ( b Gal) as a reporter enzyme,provided by F. Buckner (University of Washington,Seattle, WA, USA). Nifurtimox was tested as a standardantitrypanosomal agent. Results and Discussion The isolation of the three lignans ( 1  –  3 ) and one flavonoid( 4 ) was carried out according to the procedures describedin ‘‘Material and Methods.’’ Three known compounds7 0 - epi  -sesartemin ( 2 ) (MacRae & Towers, 1985), diayan-gambin ( 3 ) (Russell & Fenemore, 1973), and 5-hydroxy-7,4 0 -dimethoxyflavone ( 4 ) (Yang et al., 1995) wereidentified by comparison of their spectral data with thosepublished in the literature.The HREIMS spectrum showed a [M] þ peak at m = z  400.1550 corresponding to the molecular formulaC 22 H 24 O 7 . The  1 H NMR,  13 C NMR and HMQC dataof   1  showed five aromatic protons [ d H = d C  6.93(H-2) = 109.0 (C-2), 6.89 (H-6) = 118.5 (C-6), 6.84 = 110.9(C-5), 6.60 (H-6 0 ) = 104.8 (C-6 0 ), 6.52 (H-2 0 ) = 99.9 (C-2 0 )];one methylenedioxy group [ d H = d C 5.98 (H-3 0 a) = 101.4(C-3 0 a)]; four methine protons [ d H = d C  4.83 (H-7 0 ) = 82.0(C-7 0 ); 4.42 (H-7) = 87.6 (C-7); 3.32 (H-8 0 ) = 50.1 (CH,C-8 0 ), 2.91(H-8) = 54.4 (C-8)]; three methoxy groups[ d H = d C  3.93 (H-5 0 a) = 56.6 (C-5 0 a), 3.90 (H-3a) = 55.9(OCH 3 , C-3a), 3.88 (H-4a) = 55.9 (C-4a)]; two methylenegroups [ d H = d C  4.12 (H-9 a ), 3.83 (H-9 b ) = 70.9 (C-9), 3.84(H-9 0 b ), 3.34 (H-9 0 a ) = 69.6 (C-9 0 )]; and quaternary car-bons [ d C  149.2 (C-4), 148.8 (C-3 0 ), 148.7 (C-3), 143.5(C-5 0 ), 134.1 (C-4 0 ), 133.5 (C-1), 132.9 (C-1 0 )]. The keyHMBC correlations of H-7 b  with C-1, C-8, C-9, H-7 0 a with C-1 0 , C-2 0 , C-6 0 , C-8 0 , C-9 0 , H-9 b  with C-8, and H-9 0 a  with C-8 0 strongly supported the presence of a bis-tetrahydrofuran ring. Additional HMBC correlationsare shown in Figure 2. The spatial relationships in themolecule were then deduced from the NOESY spectrumof   1 . In particular, the cross-peaks implicating H-8 = H-8 0 ,H-8 = H-9 a  , H-8 0 = H-9 0 a  , H8 0 = H-7 0 a  indicated that theseprotons were all on the same face of the molecule andhad  a  orientation, whereas those observed with H-7 b = H-9 b , H-7 b = H-9 0 b  established that these other substi-tuents were on the other side of the molecule and had  b orientation (Figure 3). The linkage between C-8 and C-8 0 was confirmed to be  cis , as is typical of naturally occur-ring bis-tetrahydrofuran lignans. From the NOESY dataand the optical rotation, the compound  1  was concludedto be an unsymmetrically substituted bis-tetrahydrofuranlignan, possessing an axial aromatic ring at C-7 and anequatorial aromatic ring at C-7 0 (Russell & Fenemore,1973). On the basis of the above spectroscopic data, thestructureof  1 wasassignedas3,4,5 0 -trimethoxy-3 0 ,4 0 -methy-lenedioxy-7,9 0 :7 0 ,9 diepoxylignan (6-[4-(3,4-dimethoxy-phenyl)-tetrahydro-furo[3,4- c ]furan-1-yl]-4-methoxy-benzo[1,3]dioxole) ( 1 ), a new natural product.All spectral and optical rotation data obtained for  2 , 3 , and  4  are in close agreement with those reported for7 0 - epi  -sesartemin (MacRae & Towers, 1985), diayangam-bin (Russell & Fenemore, 1973), and 5-hydroxy-7,4 0 -dimethoxyflavone (Yang et al., 1995). Compound  2 was isolated from the bark of   Virola elongata  (MacRae& Towers, 1985), compound  3  was first reported in Macropiper excelsum  (Russell & Fenemore, 1973), andcompound  4  was isolated from  Biota orientalis  (Yanget al., 1995). De Leo´n et al. (2002) reported  in vitro  and in vivo  immunomodulatory and anti-inflammatoryactivities of diayangambin ( 3 ).Table 1 shows the larvicidal activity and  in vitro  anti-parasitic activity against  Plasmodium falciparum ,  Leish-mania donovani  , and  Trypanosoma cruzi   of compounds 1  –  3 . Compound  2  was the most active of the three testedcompounds against  P. falciparum  (W2: chloroquine-resistant strain) and  T. cruzi   with IC 50  values of 7.0380 P.N. Solı´s et al.  and 39 m g = ml, respectively, and  Aedes aegypti   larvae(LC 100  17.6 m g = ml).The larvicidal activity against  Aedes aegypti   has beenpreviously reported in  Piper acutisleginum  (Olsen et al.,1993), while  Piper rusbyi   showed antileishmanial activityagainst  L. amazonensis ,  L. braziliensis , and  L. donovani  (Fournet et al., 1994), and antitrypanosomal activityagainst  T. cruzi   (Fournet et al., 1994). Acknowledgments We thank the International Foundation for Science forfunding a project (grant no. F = 1081-2) (PNS) and theInternational Matsumae Foundation of Japan for a post-doctoral fellowship grant to P.N.S. This investigationalso received financial support from the UNI-CEF = UNDP = World Bank = WHO Special Programmefor Research and Training in Tropical Diseases (TDR)(Project ID A2076) the Organization of American States,and Fundaci  oon Natura. We are grateful to Dr. EduardoOrtega-Barrı´a (INDICASAT, Panama) for the antipara-sitic screening of compounds. References Buckner FS, Verlinde CLMJ, La Flamme AC, Van VoorhisWC (1996): Efficient technique for screening drugs foractivity against  Trypanosoma cruzi   using parasitesexpressing  b -galactosidase.  Antimicrob Agents Che-mother 40 : 2592–2597.Cepleanu F. Validation and applications of three bench-topbioassays for screening of crude extracts and sub-sequent activity-guided isolation. Ph.D. Thesis. Univer-sity of Lausanne, Lausanne, Switzerland, 1993, pp.102–119.Corbett Y, Herrera L, Gonz  aalez J, Cubilla L, Capson TL,Coley PD, Kursar TA, Romero LI, Ortega-Barria E(2004): A novel DNA-based microfluorimetric methodto evaluate antimalarial drug activity.  Am J Trop Med Hyg 70 : 119–124.Cornelly W, Espinoza OA, Montenegro H, Cubilla L, Cap-son TA, Ortega-Barrı´a E, Romero LI (2003). Hydroso-luble formazan XTT: Its application to natural productdrug discovery for  Leishmania J Microbiol Meth 55 :813–816.De Le  oon E, Olmedo DA, Solı´s PN, Gupta MP, TerencioMC (2002): Diayangambin exerts immunosuppressiveand anti-inflammatory effects  in vitro  and  in vivo . Planta Med 68 : 1128–1131.Fournet A, Barrios AA, Mu ~ nnoz Z (1994): Leishmanicidaland trypanocidal activities of Bolivian medicinal plants. J Ethnopharmacol 41 : 19–37.MacRae WD, Towers GHN (1985): Non-alkaloidalconstituents of   Virola elongata  bark.  Phytochemistry 24 : 561–566.Olsen CE, Tyagi OD, Boll PM, Hussaini FA, Parmar VS,Sharma NK, Taneja P, Jain SC (1993): An aristolactamfrom  Piper acutisleginum  and revision of the struc-tures of piperolactam B and D.  Phytochemistry 33 :518–520.Russell GB, Fenemore PY (1973): New lignans fromleaves of   Macropiper excelsum. Phytochemistry 12 :1799–1803.Solı´s PN, Olmedo DA, Avila JE, Correa MD, Gupta MP(1996): Un nuevo ensayo larvicida para detectar plantascon actividad anti- Aedes aegypti  . XV Congreso Cientı´-fico Nacional. Universidad de Panam  aa, Repu´blica dePanam  aa, October 21–25. Poster No. 18.Tebbs MC (1989): Revision of   Piper  (Piperaceae) in the NewWorld 1. Review of characters and taxonomy of   Piper section of   Macrostachys .  Bull Br Mus Nat Hist Bot19 : 117–158.Yang HO, Suh DY, Han BH (1995): Isolation and character-ization of platelet-activating factor receptor bindingantagonists from  Biota orientalis .  Planta Med 61 : 37–40. Table 1.  Larvicidal and antiparasitic activities of isolated compounds.Compound  A. aegypti   LC 100 m g = ml  L. mexicana  IC 50 m g = ml  T. cruzi   IC 50 m g = ml  P. falciparum  IC 50 m g = mlCompound  1  25.0  > 40  > 50 27.07 0 - epi  -Sesartemin ( 2 ) 17.6  > 40 39.0 7.0Diayangambin ( 3 )  > 500  > 40  > 50 15.0Tetrametrin 0.62 NT NT NTAmphotericin B NT 0.08 NT NTNifurtimox NT NT 25.0 NTChloroquine NT NT NT 0.08IC 50 , 50 % inhibitory concentration; LC 100 , lethal concentration; NT, not tested. Larvicidal lignan from  Piper fimbriulatum  381
Search
Similar documents
View more...
Tags
Related Search
We Need Your Support
Thank you for visiting our website and your interest in our free products and services. We are nonprofit website to share and download documents. To the running of this website, we need your help to support us.

Thanks to everyone for your continued support.

No, Thanks