Research Journal of Chemical Vol. 2(9), 9-13, Sept. (2012) International Science Congress Association New Sesquiterpene Hydroquinones from Marine Brown Alga Joshi Bipin C., Department of Chemistry and Biochemistry, California Received 11th Abstract One new sesquiterpene hydroquinone, together with four known sesquiterpene hydroquinones and one hydrocarbon have been isolated from the bioassay- guided fractionation of the CH (zonarioides ). The structural elucidation is based on the analysis of spectroscopic data. This is the first report of isolation of 6 from this alga. Keywords: Dictyopteris undulata, Dictyotaceae Introduction Brown algae belonging to the family Dictyotaceae have given rise to a great number of secondary metabolites possessing antibiotic, antifungal, antiviral, cytotoxic and antitumor activities1-5 . Marine brown algae of the genus been reported to contain sesquiterpenoids and C hydrocarbons6-18. In our search of antimycobacterial marine natural products we have found that the dichloromethane extract of undulata possesses antimycobacterial activity. We report herein the isolation and structural elucidation of one new sesquiterpene hydroquinone 1 together with four known compounds Material and MethodsPlant Material: The marine brown alga collected at the Catalina Island, California (USA) in August 1997. The alga was frozen shortly after collection and kept frozen until the workup. Extraction and Isolation: The dry marine brown alga undulata was exhaustively extracted with CH temperature and the solvent was removed in vacuo at 30 yield a dark green residue (6.22 g). The crude residue showed antimycobacterial activity against M. marinum concentration of 0.2g/ml. The active crude residue (6.20g) was vaccum fla sh chromatographed over silica gel (400 g), eluting with gradient of increasing polarity of ethyl acetate in TMP to 10% MeOH in EtOAc yielding twelve fractions. Fraction 3 (TMP: 10%EtOAc, 510 mg) and fraction 11 (EtOAc: 5%MeOH, 431 mg) showed activity ag ainst marinum. Fraction 3 was purified by chromatography on a column of silica gel with stepwise increases of EtOAc content (1-20%) in hexane. The fraction eluted with 4%, 6%, 10%, 12%, 14% EtOAc in hexane gave compounds Chemical Sciences _________________________________ ______ International Science Congress Association New Sesquiterpene Hydroquinones from Marine Brown Alga Dictyopteris undulata Bipin C., Kazaoka M. and Trischman Jacqueline A. Department of Chemistry and Biochemistry, California State University San Marcos, San Marcos, 92096, CA, USA Available online at: www.isca.in May 2012, revised 21st May 2012, accepted 23rd May 2012 hydroquinone, together with four known sesquiterpene hydroquinones and one hydrocarbon have guided fractionation of the CH Cl extract of the marine brown alga Dictyopteris undulata ). The structural elucidation is based on the analysis of spectroscopic data. This is the first report of isolation of Dictyopteris undulata, Dictyotaceae , sesquiterpene hydroquinone. Brown algae belonging to the family Dictyotaceae have given rise to a great number of secondary metabolites possessing antibiotic, antifungal, antiviral, cytotoxic and antitumor . Marine brown algae of the genus Dictyopteris have to contain sesquiterpenoids and C 11 In our search of antimycobacterial marine natural products we have found that the dichloromethane extract of Dictyopteris activity. We report herein the isolation and structural elucidation of one new sesquiterpene together with four known compounds 2-6. The marine brown alga D. undulata was collected at the Catalina Island, California (USA) in August 1997. The alga was frozen shortly after collection and kept The dry marine brown alga D. was exhaustively extracted with CH Cl at room temperature and the solvent was removed in vacuo at 30 C to yield a dark green residue (6.22 g). The crude residue showed M. marinum at a g/ml. The active crude residue (6.20g) sh chromatographed over silica gel (400 g), eluting with gradient of increasing polarity of ethyl acetate in TMP to 10% MeOH in EtOAc yielding twelve fractions. Fraction 3 (TMP: 10%EtOAc, 510 mg) and fraction 11 (EtOAc: ainst Mycobacterium Fraction 3 was purified by chromatography on a column of silica gel with stepwise increases of EtOAc content The fraction eluted with 4%, 6%, 10%, 12%, 14% EtOAc in hexane gave compounds 4, 6, 2, 3, and 1,respectively. F raction 11 was subjected to slica gel column chromatography and compound 5 acetate in iso-octane (2,2,4- Trimethylpentane or TMP) was then crystallized in EtOAc. General experimental procedures was performed with Si gel (230 - Aldrich. IR spectra were recorded as KBr disks on a Nicolet magna 550FTIR spectrophotometer. were recorded at ambient temperature on a BrukerARX NMR spectromete r operating at 300 recorded as values in parts per million (ppm), and were indirectly referenced to tetramethylsilane (TMS) via the solvent signal (e.g.7.26 for H, 77.0 for 13 C in CDCl EI-MS was obtained on Hew lett Packard series II 5890 GC TLC was carried out with glass precoated silica gel GF Spots were visualized under UV light and by spraying with 50% aqueous sulphuric acid followed by heating. Results and DiscussionThe CHClcrude extract of ictyopteris vitro antimycobacterial activity against marinum and was subjected to activity using vacuum flash chromatography followed by silica gel chromatography and Prep- TLC to yield Compound 1 was obtained as as colored gum and showed absorption bands for hydroxy (3400 cm 1050 cm-1 ) functional groups and an exocyclic double bond (1640 cm-1 ) in the IR spectrum. The low resonance MS of gave a molecular ion peak at 328 [M] formula was established as C2232 O CDCl showed three methyl singlets at (CH-14), and 0.91 (CH 15), one methyl singlet attributed to a methoxy group at 3.51 (CH O), two exocyclic methylene ______ _______ ISSN 2231-606X Res.J.Chem. Sci. 9 New Sesquiterpene Hydroquinones from Marine Brown Alga State University San Marcos, San Marcos, 92096, CA, USA hydroquinone, together with four known sesquiterpene hydroquinones and one hydrocarbon have extract of the marine brown alga Dictyopteris undulata ). The structural elucidation is based on the analysis of spectroscopic data. This is the first report of isolation of raction 11 was subjected to slica gel column was eluted with 50% ethyl Trimethylpentane or TMP) , which General experimental procedures : Column chromatography - 300 mesh), purchased from Aldrich. IR spectra were recorded as KBr disks on a Nicolet magna 550FTIR spectrophotometer. H and 13C NMR spectra were recorded at ambient temperature on a BrukerARX -300 FT- r operating at 300 MHz. Chemical shifts were values in parts per million (ppm), and were indirectly referenced to tetramethylsilane (TMS) via the solvent C in CDCl ). Low resonance lett Packard series II 5890 GC -MS. TLC was carried out with glass precoated silica gel GF 254 plates. Spots were visualized under UV light and by spraying with 50% aqueous sulphuric acid followed by heating. ictyopteris undulate showed in vitro antimycobacterial activity against mycobacterium and was subjected to activity -guided fractionation using vacuum flash chromatography followed by silica gel TLC to yield compounds1-6 was obtained as as colored gum and showed absorption bands for hydroxy (3400 cm -1) and ether (1254 and ) functional groups and an exocyclic double bond ) in the IR spectrum. The low resonance MS of 1 328 [M] , and the molecular O . The H NMR spectrum in showed three methyl singlets at 0.83 (CH -13), 0.85 15), one methyl singlet attributed to a O), two exocyclic methylene Research Journal of Chemical Sciences ____ _ Vol. 2(9), 9-13, Sept. (2012) International Science Congress Association proton singlets at 4.70 and 4.83 (H- 12), and an hydroxy proton singlet at 4.60 (OH- 5’), two doublets and one singlet for aromatic protons at 6.63, 6.52 and 6.61 respectively, as well as signals with complex cou pling patterns attributed to methylene and methine protons. The 13 C NMR spectrum showed 22 carbon signals including six aromatic carbons. The DEPT spectrum indicated the presence of four methyls (one methoxy), seven methylenes (one exocyclic double bond), methines, and six quaternary carbons. The signal at the 13 C NMR was characteristic of an exocyclic double bond. These data suggested that compound 1 was a sesquiterpene hydroquinone. On the basis of spectral data structure proposed fo r this compound, which was further confirmed by the spectral data of closely related compound spectra of 1 were similar to those of 2 except that the hydroxyl signal of 2 was replaced by methoxy signal at The remaining H and 13C- NMR absorptions are consistent with the structural feature of zonarol 2 . A small amount was extracted with various solvents to confirm presence of methoxy zonarol 1 in alga rather than artifact of extraction, fractionation techniques. ConclusionOn th e basis of spectroscopic data compound a new sesquiterpene hydroquinone and five known compounds were identified as zonarol 2 , isozonarol Yahazunol 5 , and a hydrocarbon neophytadiene comparing their physical and spectral data with those reported in the literature. Neophytadiene has been isolated for the first time from this alga. It is common that the same taxonomic species distributed in different regions contain different chemical constituents. The zonarol 2 and Yahazunol further confirmed by the spectral data of their diacetates 5a, respectively. Compound 1: IR(cm-1): 3400 (w), 2980(s), 2860(m), 2830(m),1640 (w), 1595(w), 1495(s), 1455(m), (w), 1254 (w), 1050(m). LR EI-MS : 55, 69, 81, 95, 109, 123, 135, 161, 178, 191, 207, 181, 315, 328[M]. C2232. Compound 2: FAB MS : 43(29), 48(33), 53(22), 55(60), 67(27), 69(45), 77(22), 79(26), 81(34), 83(21), 91(30), 95(38), 123(100), 109(27), 107(23), 137(32), 161(16), 163(13), 178(13), 191(53), 192(8), 314( 33, M), C21 H IR(cm-1): 3595 (w), 3355(w), 2925(s), 2860(m), 1640 1595(w), 1495(s), 1455(m),1380(m), 1315 (w), 1285 (w), 1140(m),1070 (w). Neophytadiene: LR EI-MS : 67 (100%), 69 (17%), 77 (17%), 79 (37%), 81 (50%), 82 (16%), 95 (49%), 109 (15%), 123 (20%), 278(1% M). C2038. _ _____________________________________________ _ International Science Congress Association 12), and an hydroxy proton 5’), two doublets and one singlet for 6.52 and 6.61 respectively, as well pling patterns attributed to C NMR spectrum showed 22 carbon signals including six aromatic carbons. The DEPT spectrum indicated the presence of four methyls (one methoxy), seven methylenes (one exocyclic double bond), five methines, and six quaternary carbons. The signal at 107.6 in C NMR was characteristic of an exocyclic double bond. was a sesquiterpene hydroquinone. On the basis of spectral data structure 1 was r this compound, which was further confirmed by the spectral data of closely related compound . The NMR except that the hydroxyl was replaced by methoxy signal at 3.51 and 50.9. NMR absorptions are consistent with . A small amount was various solvents to confirm presence of methoxy in alga rather than artifact of extraction, fractionation e basis of spectroscopic data compound 1 was identified as and five known compounds , isozonarol , Zonarone 4, , and a hydrocarbon neophytadiene 19 by comparing their physical and spectral data with those reported in the literature. Neophytadiene has been isolated for the first It is common that the same taxonomic species distributed in different regions contain different and Yahazunol 5 were further confirmed by the spectral data of their diacetates 2a and 3400 (w), 2980(s), 2860(m), 1455(m), 1380(m), 1315 : 55, 69, 81, 95, 109, 123, 135, 161, 178, 191, 207, 43(29), 48(33), 53(22), 55(60), 67(27), 69(45), 77(22), 79(26), 81(34), 83(21), 91(30), 95(38), 123(100), 109(27), 107(23), 137(32), 161(16), 163(13), H 30. 3595 (w), 3355(w), 2925(s), 2860(m), 1640 (w), 1595(w), 1495(s), 1455(m),1380(m), 1315 (w), 1285 (w), : 67 (100%), 69 (17%), 77 (17%), 79 (37%), 81 (50%), 82 (16%), 95 (49%), 109 (15%), Acetylation of compound 2 A solution of 20 mg of ml pyridine was treated with 2 ml acetic anhydride and the reaction mixture left at room temperature for 24 hr. Dilution with water and extraction with ethylacetate yielded a diacetate 2a(15.2 mg) Acetylation of compound 5 : A solution of 5.0 mg of ml pyridine was treated with 2 ml acetic anhydride and the reaction mixture left at room temperature for 22 hr. Dilution with water and extraction with ethylacetate yielded a diacetate 5a(4.8 mg). Acknowledgement This research is a result of fianancial support from NIH under MBRS Grant. In addition, B.C.J. would like to acknowledge the support of the NIH for providing a Postdoctoral fellowship. References1. Faulkner D.J., Marine natural products 19, 1–48 (2002)2.Amico V., Oriente G. , Piattelli M Fattorusso E., Magno S . and on the dolabellane skeleton from Tetrahedron,36, 1409–1414 (1980) 3.Sun H.H. , McEnroe F Acetox ycrenulide, a new bicyclic cyclopropane diterpenoid from the brown seaweed Org. Chem., 48, 1903–1906 (1983) 4.Ishitsuka M.O., Kusumi T . and xenicane and norxenicane lactones from the brown alga Dictyota dichotoma, J. Org. Chem. 5.Siamopoulou P. , Bimplakis A Cos P., Berghe D.V. and Roussis V. brown algae Dictyota dichotoma Phytochemistry,65, 2025– 2030 6.Fenical W. and Sims J. J. Fungitoxic Hydroquinones from the Brown Seaweed Dictyopteris zonarioides, J. Org. Chem. (1973)7.Fenical W. and Mcconnel O. isochromazonarol, new chromanols seaweed Dictyopteris undulate Experientia, 31, 1004-1005 1975) 8. Cimino G, Stefano SD, Fenical W, Minale L, Sims JJ. Experientia, 31, 1250-1251 ( 1975) 9.Ochi M., Kotsuki H. , Muraka K structure of Yahazu nol, a New Seqiterpene Hydroquinone from the Brown Seaweed undulata, Bull. Chem. Soc. Jap, _ ________ ISSN 2231-606X Res. J. Chem. Sci. 10 A solution of 20 mg of in 1.0 ml pyridine was treated with 2 ml acetic anhydride and the reaction mixture left at room temperature for 24 hr. Dilution with water and extraction with ethylacetate yielded a diacetate A solution of 5.0 mg of in 1.0 ml pyridine was treated with 2 ml acetic anhydride and the reaction mixture left at room temperature for 22 hr. Dilution with water and extraction with ethylacetate yielded a diacetate This research is a result of fianancial support from NIH under MBRS Grant. In addition, B.C.J. would like to acknowledge the support of the NIH for providing a Postdoctoral fellowship. Faulkner D.J., Marine natural products , Nat. Prod. Rep., , Piattelli M ., Tringali C., . and Mayol L., Diterpenes based on the dolabellane skeleton from Dictyota dichotoma, (1980) , McEnroe F .J. and Fenical W., ycrenulide, a new bicyclic cyclopropane -containing diterpenoid from the brown seaweed Dictyota crenulata,J. (1983) . and Kakisawa H., Antitumor xenicane and norxenicane lactones from the brown alga J. Org. Chem. , 53, 5010–5013 (1988) , Bimplakis A ., Iliopoulou D., Vagias C., Roussis V. , Diterpenes from the Dictyota dichotoma and Dictyota linearis, 2030 (2004), Zonarol and Isozonarol, Fungitoxic Hydroquinones from the Brown Seaweed J. Org. Chem. , 38, 2383-2386 Mcconnel O. , Chromazonarol and isochromazonarol, new chromanols from the brown Dictyopteris undulate (zonarioides), 1975) Cimino G, Stefano SD, Fenical W, Minale L, Sims JJ. 1975) , Muraka K . and Tokoroyama T. The nol, a New Seqiterpene -substituted Hydroquinone from the Brown Seaweed Dictyopteris Bull. Chem. Soc. Jap, 52(2), 629-630 (1979) Research Journal of Chemical Sciences ____ _ Vol. 2(9), 9-13, Sept. (2012) International Science Congress Association 10.Fenical W., Sims J.J., Wing R.M. , Radlick P a sesquiterpene from the brown seaweed zonarioide,Phytochemistry, 11, 1161- 1164 11.Kajiwara T., Hatanaka A., Tanaka Y. , Kawai T M., Tsuneya T. and Fujimura T., Volatile constituents from marine brown algae of Japanese Dictyopteris Phytochemistry, 28, 636-639 (1989)12.Suzuki M., Kowata N. and Kurosawa E. Chemical Society of Japan, 54, 2366- 2368 13.Suzuki M., Kowata N., Kurosawa E. , Kobayashi H Tanaka I., Chemistry Lett.,228, 2187- 2190 14.Segawa M., Yamano K. and Shrahama H. type sesquiterpene from the brown alga divaricata, Phytochemistry, 29, 973- 974 1 H NMR and 13C NMR data for Compound No.  C 1 39.10, t 2 19.39, t 3 42.50, t 4 33.61,s 5 55.63, d 6 24.36, t 7 38.18, t ------ 8 148.71, s 9 56.02, d 10 40.13, s 11 24.78, t 12 107.60, t 13 14.49, q 14 21.72, q 15 1’ 2’ 33.61, q 130.10, s 149.10, s 3’ 115.79,d 4’ 5’ 112. 87, d 148.90, s 6’ 116.51, d OH -------- OH -------- OCH 3 50.90, s _ _____________________________________________ _ International Science Congress Association , Radlick P .C., Zonarene, a sesquiterpene from the brown seaweed Dictyopteris 1164 (1972) , Kawai T ., Ishhara Volatile constituents from marine brown algae of Japanese Dictyopteris , Kurosawa E. , Bulletin of the 2368 (1981) , Kobayashi H . and 2190 (1990) Shrahama H. , A germacrane- sesquiterpene from the brown alga Dictyopteris 974 (1990)15.Koenig G.M. and Wright A. D. 183 (1995)16.Boland W. and Mueller D.G. , 310 (1987)17.Yamada K., Ojika M. and Tan H. 1634 (1980)18.Moore R.E., Pettus J.A.J . and 39, 2201-2207 (1974)19.Pasadas G.A., Alcaide I.V. , Rios J Vicario I.M. and Camacho quantification of the hydrocarbons subcutaneous fresh fat of Iberian pig by off combination of high performance liquid chromatography and gas chromatography, J. Chromatogr.A (2006)Table-1 H NMR and 13C NMR data for Compound 1 and 2 (300 MHz in CDCl 1 2  H  C 1.19 m, 1.76 m39.14, t 1.54 m 42.09, t 1.23 m, 1.41 m 42.09, t ----------- 33.62, s 1.04 m 55.62, d 1.56 m 24.36, t 2.72 d (3) 38.18, t 2.39 dd (15, 3) ------- -------- 148.71, s 2.18 m 56.01, d --------- 40.14, s 2.72 d (3) 23.59, t 4.70 s, 4.83 s 107.61, t 0.83 s 14.50, q 0.85 s 21.72, q 0.91 s 33.62, q 129.76, s 147.50, s 6.63 d (6) 116.51, d 6.52 d (3) --------- 112.88, d 147.50, s 6.61 s 115.80, d 4.60 brs 4.36 s --------- -------- 3.51 s ------- _ ________ ISSN 2231-606X Res. J. Chem. Sci. 11 D. , Mag. Res.Chem., 33, 178- , Tetrahedron Lett., 28, 307- Tan H. Chemistry Lett., , 1633- . and Mistysyn J., J. Org. Chem., , Rios J .J., Constante E.G., L.M.Characterization and quantification of the hydrocarbons fraction of the subcutaneous fresh fat of Iberian pig by off -line combination of high performance liquid chromatography J. Chromatogr.A 1123(1)82-91 (300 MHz in CDCl )  H 1.18 m, 1.75 m 1.22 m, 1.40 m 1.22 m, 1.40 m --------- 1.18 m 1.36 m 2.05 d (9) 2.41 (12) -------- 1.75 m --------- 2.72 d (6) 4.70 s, 4.83 s 0.83 s 0.85 s 0.91 s ------- ------- 6.63 d (3) 6.53 d (3) 6.61 s 4.52 s ------- Research Journal of Chemical Sciences ___________________________________________________________ ISSN 2231-606XVol. 2(9), 9-13, Sept. (2012) Res. J. Chem. Sci. International Science Congress Association 12 Table-2 H NMR and 13C NMR data for Compound 2a and 5a ( 300 MHz in CDCl3) 2a 5a No. H H 1 39.89, t 1.12 d (6),1.04 m 40.1, t 1.85 m 2 19.36, t 1.43 m 18.5, t 1.51 m 3 42.07, t 1.22 m,1.18 m 41.7, t 1.15, m 4 38.01, s ------- 33.3, s ------- 5 55.48, d 1.19 m 56.0, d 1.11 m 6 24.27, t 1.37 m 20.3, t 1.64 m 7 39.29, t 2.37 d (9) 44.1, t 1.89 m 8 148.01, s ------- 73.8, s ------- 9 55.60, d 2.07 m 62.3, d 1.56 m 10 39.89 -------- 39.2,s -------- 11 23.83, t 2.75 d (6) 24.6, t 2.62 dd (17.1, 2.6) 12 108.53, t 4.52 s, 4.80 s 24.3, q 1.26 s 13 14.52, q 0.80 s 15.3, q 0.90 s 14 20.97, q 0.85 s 21.5, q 0.81 s 15 33.60, q 0.91 s 33.4, q 0.88 s 1’ 135.66, s ------- 137.8, s ------- 2’ 146.42, s ------- 145.6, s ------- 3’ 122.44, d 6.90 d (3) 123.4, d 6.99 d (8.7) 4’ 119.37, d 6.99 d (3) 123.0, d 6.90 dd (8.7, 2.6) 5’ 147.76, s 148.1, s 6’ 122.67, d 6.93 s 119.4, d 7.13 d (2.6) Acetate -CH3 21.17 2.30 s 21.2, q 2.35 s 21.73 2.35 s 21.1, q 2.30 s Acetate C=O 169.31 ------- 169.4, s ------- 169.31 ------- 169.6, s ------- Research Journal of Chemical Sciences ___________________________________________________________ ISSN 2231-606XVol. 2(9), 9-13, Sept. (2012) Res. J. Chem. Sci. International Science Congress Association 13 13 H 10111214151'2'3'5'6' OR 4' 13 H 10111214151'2'3'5'6' OH HO4' = CH , R = H 3 2 1 = R =H2a 1 = R = COCH 13 H 10111214151'2'3'5'6'4' O 13 H 10111214151'2'3'5'6' OR RO4' OH R=H5a R= COCH 6