biphenyl substituted constituents from the rhizomes of curcuma

Mohammed Ali et al. IRJP 2011, 2 (12), 150-153

INTERNATIONAL RESEARCH JOURNAL OF PHARMACY, 2(12), 2011

INTERNATIONAL RESEARCH JOURNAL OF PHARMACY ISSN 2230 – 8407 Available online www.irjponline.com Research Article

BIPHENYL SUBSTITUTED CONSTITUENTS FROM THE RHIZOMES OF CURCUMA AMADA ROXB.

Akhlaq Mustafa1 and Mohammed Ali2*

1Drug Standardization Research Unit; Department of Chemistry, Faculty of Science, Jamia Hamdard, Hamdard Nagar, New Delhi-110062, India

2Department of Pharmacognosy and Phytochemistry, Faculty of Pharmacy, Jamia Hamdard, Hamdard Nagar, New Delhi-110062, India

Article Received on: 19/10/11 Revised on: 10/11/11 Approved for publication: 08/12/11 *Prof. Mohammed Ali, Department of Pharmacognosy and Phytochemistry, Jamia Hamdard, New Delhi- 110 062, India Email: [email protected] ABSTRACT Phytochemical investigation of the rhizomes of Curcuma amada Roxb. (Zingiberaceae) led to isolation of three new biphenyl substituted hydrocarbons identified as 1- (3’, 4’-dihydroxyphenyl)-8 - ( 3’’, 4’’ - dimethoxy - 5’’ - carboxylic acid phenyl)- octa-cis,cis-1, 4 - diene (amadanoic acid A) (2), 1- (3’, 4’ - dihydroxyphenyl) - 8 - ( 2’’, 6’’- dihydroxy -3’’, 4’’- dimethoxy - 5’’- carboxylic acid) –cis,cis- octa-1,4 - diene (amadanoic acid B) (3) and 1, 15 – dibenzyl – 3 - methyl pentadecane - 4β, 12β –diol (amadandiol) (5) along with the known compounds n-nonacosan-1-ol (1) and curcumin (4) . Their structures were elucidated by combination of chemical tests and spectral data analysis. Keywords: Curcuma amada Roxb., Zingiberaceae, rhizomes, Diphenyl hydrocarbons INTRODUCTION Curcuma amada Roxb. (Zingiberaceae), commonly known as 'Amada' or Amragandhi or mango-ginger, is a perennial herb found in Bengal, Tamilnadu and hills of the western Indian coast. It is planted in rotation of brinjal and lady’s finger and with turmeric in fields1. Its rhizomes are pale yellow inside with lighter colour outside, have distinct sweet smell of unripe mango when crushed. The rhizomes are used to manufacture pickles, culinary preparations, chutney, preserve, candy sauce and salad. Therapeutically, mango ginger is prescribed as an appetizer, aphrodisiac, febrifuge, laxative, refrigerant, aromatic, carminative, stomachic and to treat biliousness, itching, skin diseases, bronchitis, asthma, hiccough, inflammation due to injuries, rheumatism, contusions, sprains and nervous system disorders1-3. The fresh rhizome is composed of moisture (86%), ash (0.8), sugars (0.8%), fibers (1.4%), essential oil (0.1 %) and starch (6.9%) consisting of amylase (43%)4. Mango flavor is mainly attributed to car-3-ene and cis- ocimene present in the mango ginger rhizome5-7. The rhizomes also contained curcuminoids8, difurocurcumenonol, amadannlen9-12, d-curcumene, α-pinene, ocimene, linalool, linalyl acetate and safrole13,14, thiamine, vitamin-C, carotene, riboflavin, oleoresin, curcumin,

myrcene,1,8-cineole, ar-curcumene, camphor, curzerenone10,15, β-farnesene, α-guaiene, guaia-6,9-diene and α-longipinene16, amadaldehyde17 and diterpene18. This paper describes isolation and structure elucidation of three new phytoconstituents along with n-nonacosan-1-ol and curcumin from the ethanolic extract of the rhizomes. MATERIAL AND METHODS Melting points of the compounds were determined on a Perfit melting point apparatus and are uncorrected. UV spectra were recorded on Beckman DU-6 spectrophotometer in methanol. IR spectra were measured on Jasco FT-IR-5 5000 spectrophotometer using KBr pellets. 1H NMR spectra were screened on Bruker spectrospin 400-MHz instrument using CDCl3 as solvent and tetramethyl silane (TMS) as an internal standard. 13C-NMR spectra were recorded on Bruker spectrospin 100-MHz with TMS as an internal standard. Mass spectra (MS) were scanned by effecting electron impact ( EI ) ionization at 70 eV on a JEOL- JMS-DX 303 instrument. Column chromatography was performed on silica gel (Qualigens, Mumbai,India) 60-12- mesh. TLC was run on silica gel G (Qualigens, Mumbai, India). Spots were visualized by exposure to iodine vapours, UV radiation and by spraying reagents. Plant material Rhizomes of C. amada were purchased from Aligarh market and

identified by Prof. M.P. Sharma, Department of Botany, Faculty of Science, Jamia Hamdard, Hamdard Nagar, New Delhi. A voucher specimen (No. PRL/JH/07/07) is deposited in the herbarium section of Drug Standardization Unit, CCRUM, Jamia Hamdard. Extraction and isolation of phytoconstituents Air-dried plant material (2 kg) was coarsely powdered, defatted with petroleum ether (60-80ºC) and then extracted with ethyl alcohol for 48 hrs in a Soxhlet apparatus. The extract on removal of the solvent yielded a dark reddish coloured viscous mass (118 g). The extract was dissolved in a minimum amount of methanol and adsorbed on silica gel to form slurry. The slurry was dried at 400 C and chromatographed over silica gel column prepared in petroleum ether (60-800 C). The column was eluted with petroleum ether, mixtures of petroleum ether and chloroform (9:1, 3:1, 1:1 and 1:3, v/v), pure chloroform and finally chloroform and finally mixtures of chloroform and methanol (99:1, 97:3, 95:5, 90:10, v/v). Various fractions were collected separately and checked by TLC for homogeneity. Similar fractions (having the same Rf values) were combined and crystallized. The isolated compounds were recrystallized to get pure compounds. The physiochemical and spectral data of the isolated compounds are reported below. n-Nonacosan-1-ol (1) Elution of the column with petroleum ether-chloroform (3:1) afforded colourless amorphous powder of 1, recrystallized from methanol : diethyl ether (1:1), 620 mg (0.0207 % yield); Rf : 0.41 (toluene); m.p. 88-89°C; IR umax: 3500, 2950, 2845, 1508, 1470, 1355, 1205, 1050, 795, 715 cm-1; 1H NMR: δ 3.57 (1H, d, J=6.56 Hz, H2-1a), 3.53 (1H, d, J=6.57 Hz, H2-1b), 1.53 (2H, m, H2-2), 1.42 (2H, brs, CH2), 1.19 (50H, brs, 25 × CH2), 1.06 (2H, brs, CH2), 0.83 (3H, t, J=6.23 Hz, Me-29); 13C NMR: δ 60.81 (C-1), 37.78 (CH2), 37.11 (CH2), 34.42 (CH2), 32.11 (CH2), 30.88 (CH2), 28.71 (CH2), 25.98 (19 × CH2), 21.59 (CH2), 12.98 (CH3); EIMS m/z (rel. int.): 424 [M]+ C29H60O (9.3), 307 (8.8), 283 (11.1), 262 (7.6), 256 (12.7), 227 (22), 213 (21.2), 199 (24.3), 185 (19.7), 127 (22.8), 113 (25.1), 99 (74.1), 85 (43.2), 71 (45), 69 (51.7), 57 (62.5). Amadanoic acid A (2) Elution of the column with chloroform furnished reddish coloured amorphous powder of 2, re-crystallized from MeOH, yield: 2.5 g, (0.13%) ; Rf: 0.69 (petroleum ether : CHCl3 7:3), m p : 185 -186º C, UV max (MeOH) : 236, 321 nm (log ε 5.8), IR umax (KBr): 3400, 3260 2950, 2860, 1695, 1645, 1580, 1450, 9732 cm-1 ; 1H NMR (CDCl3): δ 7.57 (I H, d, J = 3.0 Hz, H-2’), 7.51 (1H, d, J = 2.9 Hz, H-2’’), 7.06 (1H, d, J=2.9 Hz, H-6’’), 6.89 (1H, dd, J=8.0, 3.0 Hz,



H-6’), 6.80 (1H, d, J=8.0, H-5’), 6.15 (1H, d, J=4.6 Hz, H-1), 6.03 (1H, m,w1/2=7.2 Hz, H-2), 5.82 (1H, m, w1/2=7.5 Hz, H-4), 5.35 (1H, m,w1/2=8.1 Hz, H-5), 3.92 (3 H, brs, OMe), 3.88 (3 H, brs, OMe), 2.88 (2 H, brs, H2-3) 2.13 (2 H, m, H2-8), 1.80 (2 H, m, H2-6), 1.25 (2 H, m, H2-7) . ; 13C NMR (CDCl3): δ 141.27 (C-1’), δ 126.55 (C-2’), 155.12 (C-3’), 153.63 (C-4’), 123.48 (C-5’), 121.28 (C-6’), 125.81 (C-1), 119.17 (C-2),41.52 (C-3), 118.57 (C-4), 116.90 (C-5) 35.19 (C-6), 29.86 (C-7), 39.88 (C-8), 148.52 (C-1’’), 120.78 (C-2’’), 159.28 (C-3’’), 156.52 (C-4’’), 147.32 (C-5’’), 121.27 (C-6’’), 182.84 (C-7’’), 55.93 (OMe), 55.49 (OMe). EIMS m/z (rel. int): 398 [M]+ (C23H26O6) (7.3) 381 (9.6), 367 (46.7), 350 (63.9), 335 (7.2), 326 (8.7), 285 (12.8), 272 (21.6), 223 (14.2), 192 (31.7), 189 (43.1), 181 (16.3), 178 (100), 175 (19.3), 161 (16.1), 149 (18.1), 145 (51.1), 137 (49.3), 135 (15.9), 132 (23.8), 118 (22.6), 103 (9.3), 89 (18.3), 77 (13.1). Amadanoic acid B (3) Further elution of the column with chloroform furnished brown coloured amorphous powder of 3, recrystalised from MeOH, yield: 440 mg (0.022%) Rf : 0.89 (CHCl3- petroleum ether, 4 : 1), mp: 170-171º C, UV λmax (MeOH) : 233,325 nm (log ε 6.9), IR umax (KBr) : 3460, 3300, 2960, 2855, 1698, 1634, 1510, 1457, 1360, 1225, 795 cm-1; 1H NMR (CDCl3) : δ 7.35, (1H, d, J=2.35 Hz, H-2’), δ 7.05, (1H, d, J=8.5 Hz, H-5’), 6.68 (1H, dd, J=8.5, 2.35 Hz, H-6’), 6.15 (1H, d, J=8.5 Hz, H-1), 5.91 (1 H, m, w1/2=7.6 Hz, H -2), 5.86 (1 H, m, w1/2= 7.9 Hz, H -4), 5.36 (1 H, m, w1/2=8.3 Hz, H -5), 3.95 (3H, brs, OMe), 3.87 (3H, brs, OMe), 2.81 (2H, m, H-3), 2.21 (2H, m, H2 - 8), 1.82 (2H, m, H2-6), 1.25 (2 H, brs, H2-7). ; 13C NMR (CDCl3) : δ 143.69 (C-1’), 127.11(C-2’), 157.02 (C-3’), 155.63 (C-4’), 124.06 (C-5’), 122.15 (C-6’), 125.13 (C-1), 118.66 (C-2), 42.38 (C-3), 116.36 (C-4), 115.28 (C-5), 34.09 (C-6), 29.70 (C-7), 38.07(C-8), 145.13 (C-1’’),157.05 (C-2’’), 159.45 (C-3’’), 155.93 (C-4’’), 147.36 (C-5’’), 154.15 (C-6’’), 183.16 (C-7’’), 55.16 (OMe), 55.03 (OMe). EIMS m/z (rel. int.) : 430 [M]+ (C23H26O8) (3.1), 366 (20.6), 348 (23.6), 337 (14.3), 319 (10.9), 264 (13.3), 260 (39.8), 241 (11.2), 237 (10.1), 227 (11.4), 217 (20.2), 213 (13.8), 211 (13.5), 210 (15.7), 200 (17.5), 197 (13.8), 196 (14.8), 189 (38.0), 182 (21.9), 180 (11.8), 175 (100), 169 (250), 166 (14.3), 165 (13.3), 158 (27.7), 149 (44.0), 138 (19.2), 135 (74.9), 132 (31.9), 121 (51.3), 118 (71.6), 115 (53.8), 109 (21.3), 96 (60.5), 90 (63.6), 84 (71.8), 71 (88.6), 57 (91.2). Curcumin (4) Elution of the column with chloroform-methanol (99:1) afforded yellow crystalline mass of 4, recrystallized from methanol, 630 mg (0.021 % yield); Rf: 0.86 (chloroform: methanol:: 9:1); m.p. 180-1830C; UV lmax: 240 nm (log e 3.2); IR umax (KBr): 3450, 3400, 2945, 2860, 2360, 1690, 1650, 1570, 1310, 980 cm-1; 1H NMR : d 7.58 (1H, d, J=15.7 Hz, H-8), 7.53 (1H, d, J=15.7 Hz, H-12), 7.06 (4H, brs, H-2, H-5, H-15, H-18), 7.04 (1H, brs, D2O exchangeable, OH), 6.91 (2H, d, J=8.5 Hz, H-6, H-19), 6.50 (1H, d, J=15.7 Hz, H-7), 6.45 (1H, d, J=15.7 Hz, H-13), 5.82 (1H, brs, D2O exchangeable, OH), 3.90 (8H, brs, OMe-3; OMe-17, H2-11); 13C NMR : d 126.44 (C-1), 120.67 (C-2), 140.18 (C-3), 148.48 (C-4), 115.11 (C-5), 109.82 (C-6), 115.11 (C-7), 122.38 (C-8), 182.75 (C-9), 55.38 (C-10), 182.75 (C-11), 122.38 (C-12), 115.11 (C-13), 122.38 (C-14), 120.67 (C-15), 140.18 (C-16), 147.26 (C-17), 120.67 (C-18), 100.62 (C-19), 55.38 (2 × OMe); EIMS m/z (rel. int.): 368 [M]+ (C21H20O6) (41.6), 350 (43.5), 271 (15.8), 230 (11.3), 216 (11.5), 191 (27.4), 188 (37.4), 177 (18.2), 176 (100), 149 (26.3), 146 (7.2), 144 (32.1), 134 (12.5), 129 (11.2), 123 (17.1), 118 (10.2), 108 (9.2). Amadandiol (5) Fractions eluted with chloroform-methanol (49 : 1) furnished greenish yellow amorphous powder of 5, recrystallized from MeOH, yield : 80 mg (0.004%), Rf : 0.52 (petroleum ether-CHCl3 3:1), m p: 155 -156º C, UV λmax (MeOH) : 231, 319 nm (log ε 4.8), IR umax

(KBr) : 3500, 3370, 2950, 2860, 1648, 1555, 1538, 1470, 1212, 1108, 1062, 795 cm-1. ; 1H NMR (CDCl3) : δ 7.51 (1H, m, H – 2’), 7.35 (1H, m, H-2’’), 7.13 (1H, m, H-6’), 7.11 (1H, m, H-6’’), 7.02 (2H, m, H-3’, H-5’), 6.97 (2H, m, H-3’’, H-5’’), 6.67 (1H, m, H-4”), 6.64 (1H, m, H-4’), 3.89 (1H, brs, w1/2=18.5 Hz, H-12), 3.75 (1H, brs, w½ =17.3 Hz, H-4), 2.61 (2H, m, H2-1), 2.31 (2H, m, H2-15), 2.03 (2H, m, CH2-2), 1.86 (1H, m, H-3), 1.66 (2H, m, CH2), 1.59 (2H, brs, CH2), 1.55 (2H, m, CH2), 1.42 (2H, brs, CH2), 1.33 (2H, brs, CH2), 1.28 (2H, brs, CH2) 1.25 (2H, brs, CH2), 1.20 (2H, brs, CH2), 0.87 (3H, d, J=6.5 Hz, Me-16). ; 13C NMR (CDCl3) : δ 34.25 (C-1), 22.26 (C-2), 37.27(C-3), 71.83 (C-4), 26.13 (C-5), 27.21 (C-6), 29.50 (C-7), 29.34 (C-8), 29.68 (C-9), 22.67 (C-10), 31.91 (C-11), 68.13 (C-12), 31.43 (C-13), 24.91 (C-14), 38.36 (C-15), 140.98 (C-1’), 129.06 (C-2’), 119.34 (C-3’), 113.51 (C-4’), 123.57 (C-5’), 127.90 (C-6’), 139.72 (C-1”), 129.09 (C-2”), 119.32 (C-3”), 124.21 ( C-4”), 124.46 (C-5”), 126.31 (C-6’’), 14.09 (Me-16). EIMS m/z (rel. int) : 410 [M]+ (C28H42O2) (2.1), 291 (15.3), 277 (11.3), 261 (26.5), 216 (14.1), 189 (13.2), 183 (10.8), 173 (12.4), 163 (11.2), 160 (17.2), 149 (27.8), 146 (19.9), 133 (80.8), 119 (63.8), 105 (39.2), 95 (38.3), 80 (42.7), 68 (83.2), 56 (91.4), 43 (100). RESULTS AND DISCUSSION Compounds 1 and 4 are known phytoconstituents characterized as n-nonacosan-1-ol and curcumin, respectively. Compound 2, designated as amadanoic acid A, was obtained as a red coloured powder from chloroform eluants. It gave chemical tests of phenol and yielded effervescences with sodium bicarbonate solution. Its IR spectrum exhibited characteristic absorption bands for hydroxyl groups (3400 cm-1), carboxylic function (3260,1695 cm-1) and unsaturation (1645 cm-1). On the basis of mass and 13C NMR spectra, the molecular weight of 2 was established at m/z 398 corresponding to biphenyl substituted octadiene-type molecule, C23H26O6. The mass spectrum displayed prominent ion peaks at m/z 381 [M-OH]+, 367 [M-OMe]+, 350 [M-OMe-OH]+, and 335 [350-Me]+. The ion peaks at m/z 217 [(HO)2-C6H3-(C5H6)-(CH2)3]+,189 [(HO)2-C6H3 (C5H6)-CH2]+, 175 [(OH)2-C6H3-CH=CH-CH2-CH=CH], 149 [(HO)2-C6H3-CH=CH-CH2], 135 [(HO)2-C6H3-CH=CH2]+ 132 [149-OH]+ and 118 [135-OH]+ suggested that dihydroxy substituted phenyl ring was present at one of the terminal carbon chain conjugated with an olefinic linkage and another vinylic linkage at the fourth carbon of octadiene chain. The ion peaks generated at m/z 181 [C6H2–(OMe)2-COOH]+ and 223 [(CH2)3 C6H2 (OMe)2-COOH]+ supported the location of dimethoxy benzoic acid at another terminal of the carbon chain. The 1H NMR spectrum of 2 showed three one-proton m-coupled doublets at δ 7.57 (J=3.0 Hz), 7.51 (J=2.9 Hz), and 7.06 (J=2.9 Hz), assigned to aromatic H-2’, H-2’’and H -6’’, respectively. A one-proton ortho-coupled doublet at δ 6.80 (J=8.0 Hz) was ascribed to H – 5’. A one-proton ortho-, meta-coupled double doublet at δ 6.89 (J=8.0, 3.0 Hz) was associated with H-6’. Four another deshielded signals as a doublet at δ 6.15 (J=4.61 Hz) and as multiplets at δ 6.03 (w1/2 = 7.2 Hz), 5.82 (w1/2 = 7.5 Hz) and 5.35 (w1/2 = 8.1 Hz) were attributed to cis-oriented H-1, H-2, H-4 and H-5 vinylic protons, respectively. Two three-proton broad signals at δ 3.92 and 3.88 were due to methoxy group attached to C-3’ and C-4’ benzene carbons. The remaining methylene protons appeared between δ 2.88 -1.25. The 13C NMR spectrum of 2 exhibited aromatic and vinylic carbons between δ 159.28 -116.90, carboxylic carbon at δ 182.84 (C- 7’’), methoxy carbons at δ 55.93 and 55.49 and ethylene carbons from δ 41.52 to 29.86. The DEPT spectrum of 2 exhibited the presence of two methyl, four methylene, nine methine and eight quaternary carbons. The 1H–1H COSY spectrum of 2 showed correlations of H-1 with H-2, H2–3, H-2’ and H–6’; H-5’ with H-6’; and H-2’’ with H-6’’. The HMBC of 2 showed interactions of C-3’ with H-2’ and H-5’. ; C- 1’ with H- 2’, H- 6’ and H- 1 ; and C- 7’’ with H- 6’’. On the basis of these evidences, the structure of 2 has been formulated as 1 – (3’, 4’–



dihydroxyphenyl)- 8-(3’’, 4’’-dimethoxy-5’’- carboxylic acid phenyl) – cis, cis-octa –1, 4–diene, This is an unreported phenolic phytoconstituents from Curcuma and another species.

HO

HO

OMe

OMe

COOH

CH CH CH21'

6'5'

4'

3'

2

2'

1CH

3 4 5CH

6 7 8

2 2 2

1"

2"3"

4"

5"6"

R

R

CH CHCH

2. R=H

3. R=OH

CH CH1'

6'

3'

2

2'

1 3 4 5CH CH

2 2

CH OH3

16

(CH )2 7

CH

OH

(CH )2 2 2

1"

2"3"

6"

CH15

12

5

Compound 3, named amadanoic acid B, was obtained as brown amorphous powder from chloroform eluants. It yielded the effervescences with sodium bicarbonate solution and greenish-blue colour with ferric chloride solution. Its IR spectrum showed absorption bands for phenolic groups (3460 cm-1), carboxylic function (3300, 1698, cm-1) and unsaturation (1634 cm-1). Its molecular weight has been established at m//z 430 on the basis of mass and 13C NMR spectra, which corresponded to a biphenolic molecule, C23H26O8. The prominent ion peaks arising at m/z 189 [(HO)2-C6H3-(C5H6)-CH2]+, 175 [(HO)2-C6H3-(C3H4)-CH=CH]+, 149 [(HO)2-C6H3-CH=CH-CH2]+ and 135 [(HO)2-C6H3-CH=CH]+ indicated the existence of a dihydroxy substituted benzene ring at one of the terminal of octadiene. The ion peaks at m/z 241 [CH2-CH2-(OH)2-C6 -(OMe)2-COOH]+, 227 [241-CH2]+, 213 [(HO)2-C6-(OMe)2-(COOH)]+, 210 [241-OMe]+,197 [241-CO2]+, 196 [227-OMe]+ and 182 [213-OMe]+ supported dihydroxy dimethoxy substituted benzoic acid moiety at another terminal of the n-octadiene chain. The 1H NMR spectrum of 3 exhibited two one - proton doublets at δ 7.35 (J=2.35 Hz) and 7.05 (J=8.5 Hz) assigned to meta-coupled H-2’ and ortho-coupled H-5’ protons, respectively. A one-proton double doublet at δ 6.68 (J=8.5, 2.35 Hz) was associated with ortho-meta-coupled H-6’. Four one-proton signals as a doublet at δ 6.15 (J = 8.5 Hz) and as multiplets at δ 5.91(w1/2 = 7.6 Hz), 5.86 (w1/2 = 7.9 Hz) and 5.36 (w1/2 = 8.3 Hz) were attributed to cis-oriented H-1, H-2, H-4 and H-5 vinylic protons, respectively. Two three-proton broad signals at δ 3.95 and 3.87 attested the presence of two methoxy groups in the molecule. The remaining methylene proton resonated between δ 2.81-1.25. The 13C NMR spectrum of 3 showed deshielded 12 carbon signals for aromatic rings (δ 159.45 -121.13), one carboxylic carbon (δ 183.16), two methoxy carbons (δ 55.16, 55.03), four vinylic carbons (δ 125.15,118.66, 116.36 and 115.28) and four methylene carbons (δ 45.38, 34.29, 24.70 and 38.07). The DEPT spectrum of 3 displayed the presence of two methyl, four methylene, seven methine and ten quaternary carbons. The 1H–1H COSY spectrum of 3 exhibited correlations of H-1 with H -2’, H – 6’, H -2 and H2 – 3 ; and H–5’ with H-6’. The HMBC spectrum of 3 showed interactions of C–4’ with H–2’, H–5’ and H-6’; C-1’ with H–2’, H–6’ and H–1; and C-1’’ with H2-8. On the basis of spectral data analysis, the structure of‘ 3 has been formulated as 1- (3’, 4’–dihydroxyphenyl)–8-(2’’, 6’’– dihydroxy –3’’, 4’’- dimethoxy – 5’’- carboxylic acid) – cis, cis- octa –1, 4–diene. This is a new phenolic acid. Compound 5, named amadandiol, was obtained as a greenish yellow amorphous powder from chloroform – methanol (49:1) eluants. Its

IR spectrum exhibited absorption bands at 3500, 3370 cm-1 for hydroxyl groups and at 1648, 1538 cm-1 for aromatic ring. Its molecular ion peak was determined at m/z 410 on the basis of mass and 13C NMR spectra corresponding to diphenyl substituted methyl pentadecane type molecule, C28H21O2. The prominent ion peaks arising at m/z 291 [C6H5-(CH2)2-CH (CH3)CH-(OH)-(CH2)7-CH-(OH)]+, 261[C6H5-CH2-CH2-CH-(CH3)-CH-(OH)(CH2)7]+ , 163 [C6H5-CH2-CH2-CH-(CH3)-CHOH)]+, 133[C6H5-CH2-CH2-CH- (CH3)]+ and 105 [C6H5-CH2-CH2]+ suggested the presence of a phenyl ring at one of the terminal carbon, methyl group at C-3 and hydroxy groups at C-4 and C- 12. The ion peaks formed at m/z 277 [CH(OH)-(CH2)7-CH-(OH) - (CH2)3-C6H5]+, and 119 [(CH2)3-C6H5]+ supported the existence of another phenyl ring at the second terminal of the alkane chain and hydroxyl groups at C-4 and C- 12. The 1H NMR spectrum of 5 showed six deshielded one-proton multiplets at δ 7.51, 7.35, 7.13, 7.11, 6.67 and 6.64 assigned to H – 2’, H-2’’, H – 6’, H-6’’, H-4’’ and H-4’, respectively. Two multiplets at δ 7.02 and 6.97, integrated two-protons each, were ascribed to aromatic H-3’, H-5’and H-3’’ and H-5’’, respectively. Two one-proton broad multiplets at δ 3.89 and 3.75 were attributed to H-12 and H-4 carbinol protons, respectively, and their corresponding half width of 18.5 Hz and 17.3 Hz indicated α-orientation of the carbinol protons. The remaining methylene protons appeared between δ 2.61-1.20. A three- proton doublet at δ 0.87(J = 6.5 Hz) attested the presence of C-16 secondary methyl group. The 13C NMR spectrum of 5 showed twelve deshielded aromatic carbons between δ 140.98-113.51, two carbinol carbons (δ68.13, 71.83), one methyl carbon (δ14.09) and methylene carbons between δ 38.36-22.67. The multiplicity of each carbon was determined by DEPT studies. The 1H – 1H COSY spectrum of 5 exhibited correlations of H2-1 with H2 -2, H – 2’ and H – 6’ ; H3 -16 with H-3 and H -4 ; H -12 with methylene protons ;and H- 2’’ with H – 3’’, H -4’’, H-6’’ and H2 -15. The HMBC spectrum of 5 showed interactions of C–1’ with H–2’, H–6’ and H2 -1 ; C-16 with H–3, H2 -2 and H–4; C-12 with methylene protons ; and C-1’’ with H-2’’ and H-6’’. These data led to establish the structure of 5 as 1, 15- biphenyl- 3–methyl pentadecane- 4β,12β –diol. This is a new biphenyl substituted alkane - diol isolated from a natural or synthetic source for the first time. CONCLUSION Phytochemical investigation of C. amada rhizomes led to the isolation of three new aromatic compounds. This species is different from the other Curcuma species in phytoconstituents which may be responsible for its medicinal properties. The varied phytoconstituents may confirm the therapeutic values for its combating abilities and for its uses in Ayurveda and other traditional medicines. ACKNOWLEDGMENT The authors are thankful to the Head SAIF, Central Drug Research Institute, Lucknow, India for scanning mass spectra of the compounds. REFERENCES 1. Anonymous, The Wealth of India, Raw Materials, National Institute of Science

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biphenyl substituted constituents from the rhizomes of curcuma