0031-9422@4)0@424-2 Phymdwmmry. Vol 37. No 3. pp. 757 -760. 1994 Copyright 0 1994 FJscvin .Gmce Ltd

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DIHYDROXY ACIDISSIMINOL AND ACIDISSIMINOL EPOXIDE, TWO TYRAMINE DERIVATIVES FROM LZMONZA ACZDZSSZMA

PARTHASARATHI GHOSH, MRINAL KAN-I-I GHOSH, SWAPNADIP THAKUR,* JASODHARA DAN (DATTA), TOSHIHIRO AKIHISA,~ TOSHITAKE TAMuRAt and YUMIKO KIMURA~

Department of Chemistry, The University of Burdwan, Burdwan-713 104, India; tCollege of Science and Technology, Nihon University, 1-8, Kanda Surugadai, Chiyoda-ku, Tokyo 101, Japan; SCollege of Pharmacy, Nihon University, 7-7-1, Narashinodai,

Funabashi, Chiba 274, Japan

(Received in reuisedjiirm 27 April 1994)

Key Word Index-Limonia acidissima; Rutaceae; fruit; tyramine derivative; dihydroxy acidissiminol; benzamide-N-{p-[(3,7-dimethyl-4,6,7-trihydroxy-2-octenyl)oxy] phenethyl}; acidissiminol epoxide.

Abstract-Dihydroxy acidissiminol, a new tyramine derivative, was isolated from the methanol extract of defatted fruits of Limcnia acidissimn. The structure of the compound was determined as benzamide-N-{p[(3,7-dimethyl-4,6,7- trihydroxy-2-octenyl)oxy] phenethyl} based on spectroscopic evidence. Two other tyramine derivatives isolated from the same source were identified as acidissiminol epoxide (benz.amide-N-{p-[(3,7dimethyl-6,7-epoxy-2-octenyl)oxy] phenethyl}) and N-benzoyl tyramine. Although acidissiminol epoxide is a known synthetic compound, this is the first identification of it as a natural product. The ‘H and ’ %J NMR spectra of all compounds were completely assigned.

INTRODUCTION

Limonia acidissima is well-known for its medicinal prop- erties [l] and is widely distributed in India. We have recently isolated four tyramine derivatives from the fruits of this species [2,3]. We now report the isolation and characterization of a new tyramine derivative, dihydroxy acidissiminol (I), along with the isolation and identifica- tion of acidissiminol epoxide (2) and N-benzoyl tyramine (3) from the fruits of the same source.

RE3JLTSANDDlSCUSSlON

The methanolic extract of defatted fruits was chro- matographed over silica gel, which on elution with methanol-chloroform ( 1: 99) furnished a crude semi- solid material. This semi-solid material on preparative TLC gave three compounds (l-3), viz., dihydroxy acidis- siminol (l), a semi-solid material, HR CI mass spectrum: m/z 428.2411 [M+H]+, highest-mass ion in HR EI mass spectrum: m/z 241.1095; acidissiminol epoxide (2), HR CI mass spectrum: m/z 410.2308 [M + HI+, highest- mass ion in HR EI mass spectrum: m/z 241.1087; N- benzoyl tyramine (3), HR CI mass spectrum: m/z 242.1156 [M + H] + and HR EI mass spectrum: m/z 241.1087 [M] +. Compounds 2 and 3 were identified as acidissiminol epoxide (benzamide-N-{p-[(3,7_dimethyl- 6,7-epoxy-2-octenyl)oxy]phenethyl}) and N-kmzoyltyra-

mine [benzamide-N-(p-hydroxy phenethyl)], respect-

*Author to whom correspondence should be addressed.

ively, by comparison of ‘H and 13C NMR, mass and IR spectral data with those of authentic compounds

The IR spectrum of 1 indicated the presence of hy- droxyl, amide, amide carbonyl, aromatic and trisubstitu- ted olefinic functions. Its “CNMR spectrum exhibited three methyl (612.4, 23.7, 26.2), four methylene (634.8, 35.7, 41.3, 64.5), eight methine (corresponding to 12 carbons) [two oxy (677.4, 78.7) and six olefinic and aromatic(6114.9, 121.1, 126.8, 128.6, 129.8, 131.4; corres- ponding to 10 carbons)], and six quatemary [one oxy (672.6), four olefinic and aromatic (6 131.0, 134.6, 142.0, ‘157.3), and one carbonyl (6167.6)] carbon signals as determined with the aid of a 13C DEPT experiment. The number and pattern of the ’ 3C signals were comparable with those of 2. The diagnostic fragmentations in both the CI and EI mass spectra of 1 were also comparable with those of 2, whereas 1 displayed a [M + H]+ 18 mu (corresponding to water) higher than that of 2 in the CI mass spectrum. This suggests that 1 possesses the same structural features as 2, with the exception of the C-6 and C-7 functionality in the geranyl moiety, viz., the 6,7-epoxy group in 2 is substituted by the 6,7-diol functionality in 1. The new tyramine derivative, dihydroxy acidissiminol(1) was, therefore, considered to have the structure benzamide-N-{p[(3,7-dimethyl4,6,7-trihydroxy-2-octenyl) oxy]phenethyl}.

The ‘H and 13CNMR data of l-3 are shown in Table 1. The signal assignments were performed with the aid of “CDEPT, ‘H-*H COSY, ‘H-‘3CCOSY, NOESY, HMQC and HMBC experiments. The “C chemical shift value 672.6 for the hydroxy-bearing C-7 of

757

758 P. GHOSH et al.

H H H H

H H H H

Me OH OH

1 R =-CH2--_cH--_ I I

I 2 3 -&-CHz-CH~C--MC

I

s

6 I

GH Me

Me

I 2R =-CH2-_cH--_C- -CH2-

Scheme 1.

Table I. ‘H NMR (400 MHz, CDCI,) and 13C NMR (100 MHz, CDCI,) data of dihydroxy acidissiminol (I), acidissim- inol epoxide (2) and N-benzoyl tyramine (3)

Proton Carbon - - _._~ -..- - .-.-

Position 1 2 3 1 2 3

1’

2’,6 3’,5’

4

CO-NH

N-CH,

Ar-CH,

1”

2”6” 3Y5” 4”’

Ar-OH

1

2 3

3-Me 4

5

6

7

7-Me

8

4-OH 6,7-OH

- 7.69 (ZH, d, 7.2) 7.41 (2H, f, 6.4)

7.49 (1 H, t, 7.0)

6.10 (lH, m)*

3.70 (2H, q, 6.6)t 2.88 (2H, t, 7.0) -

6.87 (2H, d, 8.4)

7.16 (2H, d, 8.4) -

-

4.58 (ZH, d, 6.2)

5.80 (1 H, I, 6.0) -

1.76 (3H, s)

4.35 (I H, dd, 3.6, 9.2) 1.7 (IH, m)

2.3 (lH, m)

3.64 (IH, m) -

1.18 (3H, s)$

1.22 (3H, s)$

1.55 (s),

1.55 (S).

7.69 (2H, d, 8. I)

7.40 (2H, t, 7.7)

7.48 (I H, t, 7.0)

6.21 (lH, m)*

3368 (2H. q. 6.W 2.86 (2H. t, 7.0)

6.86 (ZH, d, 8.1)

7.13 (ZH, d, 8.4) -.

4.60 (2H, d, 6.6)

5.85 (1 H, I, 6.2) _._

1.74 (3H, s)

4.37 (1 H, t, 7.3)

1.77 (IH, m)

2.48 (1 H. tt, 6.0, 7.3)

3.90 (IH, m) -

1.20 (3H, s) 1.26 (3H, s)

1.56 (s)*

_- 7.69 (2H, d, 7.3)

7.41 (2H, L, 7.3)

7.49 (IH, r, 7.3)

6.10 (lH, m)+ 3.69 (2H, q. 6.6)t 2.87 (2H, t. 7.3)

6.80 (2H, d, 8.8)

7.11 (2H.d.8.1) -

4.92 (1 H, hr. s)*

_

_

_ -

134.6 134.8

126.8 126.8

128.6 128.5

131.4 131.3 167.6 167.5

41.3 41.3

34.8 34.8

157.3 157.3

114.9 115.2

129.8 129.8

131.0 131.2

64.5 64.5

121.1 120.1

142.0 141.0

12.4 13.0 77.4 79.0

35.7

78.7

72.6

23.7

26.2

39.4

78.1

83.3

25.8

22.4 -.

- -

134.6

126.8

128.6

131.4

167.5

41.3

34.8

154.3 115.6

130.0

132.0

_

_

-

-.

-

The chemical shifts (s) are given in ppm with TMS (‘H NMR) or CDCI, (“C NMR) as internal standard. Figures in

parentheses denote J values (Hz).

*Signal disappeared by D,O exchange. tcoupling pattern changed into triplet (J = 6.6 Hz). SAssignments may be interchanged.

Tyramine derivatives from Limonia acidissima 159

Table 2. ‘“C-‘H long range correlations observed in the HMBC of l-3

‘H(6) Correlated carbons

Assignment 1 2 3 1 2 3

2’,6 1.69 7.69 7.69 3’s 7.41 7.40 7.41 4 1.49 1.48 7.49 N-CH, 3.69 3.68 3.69

Ar-CH, 2.88 2.86 287

2”,6” 6.87 6.86 6.80 Y.5” 7.16 7.13 7.11

1 2 3-Me 4

5

7-Me 8

4.58 5.80 1.76 4.35 1.67 2.33

1.18 1.22

4.60 -

5.85 1.74 -

4.31 1.77 -

2.48

4’,CO-NH 4’CO-NH 1’ 1’ 2’,6 2’,6 CO-NH, CO-NH, Ar-CH, Ar-CH, 4” 4” N-CH,, N-CH,, 3”,5” 3”,5” 4” 4’

I”# 1”,4”

Ar-CH,, Ar-CH *, ,,

1, I, 1 ,

2”,6” 2”,6” 1”,2,3 1”,2,3 4 4

2.3.4 23.4 2 2

3t4.6 3,6,7

1.20 - 6,7,8 6.7.8 1.26 6,7,7-Me 6,7,7-Me

4’,CO-NH 1’ 2’,6 CO-NH Ar-CH, 4” N-CH, 3”,5” 4’

1”,4”

Ar-CH,, 1 ,#

-

-

1 was consistent with that of the hydroxy-bearing tertiary carbon of 2.3~dimethyl-2-butanol (C-l, 626.3; C-2, a72.2) [4], thus supporting the proposed structure for 1. Table 2 showed the cross-peaks observed in the HMBC spectra of 1-3 which are correlated with the long range couplings between the given carbons and protons. This and the other 2DNMR data were consistent with the proposed structure of 1.

Although 2 has recently been synthesized from 6 by alkaline hydrolysis [3], this is the first identification of 2 as a natural product.

EXPERIMENTAL

General. Mps: uncorr. Prep. silica gel TLC plates were developed 3 x with hexane-EtOAc (1: 1). Spectral ana- lyses were performed using identical instruments as de- scribed previously [2,3]. Compounds 2 and 3 were used as ref. samples [3].

Plant material. Limonia acidissima L. was collected locally and voucher specimens (386 and 388) are kept at the herbarium of the Department of Botany, The Univer- sity of Burdwan.

Extraction and isolation. Air-dried and crushed defatted fruits (petrol W-80”) (2 kg) were extracted for 48 hr in a Soxhlet using MeOH and the solvent removed under red. pres. The residue was subjected to CC over silica gel (BDH 60-120). Elution was carried out with pure solvents and their mixts in order of increasing polarity. The MeOH-CHCI, (1:99) eluate (21 mg) on prep. TLC yielded 3 bands from which 1 (R, 0.08; 3.3 mg), 2 (Rf 0.36; 1.1 mg), 3 (R, 0.55; 2.8 mg) were obtained.

Dihydroxy acidissiminol(1). Amorphous semi-solid. IR v;i cm- ‘: 3420 (-OH stretch), 3300 (-NH stretch), 1638 (amide carbonyl), 1577, 1542, 1510 (aromatic C-C stretch), 823 ( > C =C-H). HRCIMS: m/z ( > m/z 60, rel. int.) CM +H]+ 428.2411 (4, CzsHs,O,N,), 410.2291 (18, CzsH,,O,N,), 392.2222 (11, C,,H,,OsNt). 322.1801 (5, Cz1H2~0zN,), 241.1143 (100, C,,H160zN,), 201.1092 (19, C,,,Hr,OJ, 183.1044 (9, C,,H,,O,), 169.1200 (21, C,,,Hr,Oz), 151.1100 (6, C,cH,,Or), 134.0653 (4, CsHsO,N,), 120.0566 (38, CsHsOr), 105.0321 (21, C,H501), 85.0600 (33, CsHsO,), 81.0697 (33, C,H,), 69.0668 (63, CsH9). HREIMS: m/z (rel. int.) 241.1095 (9, highest-mass ion, C,,H,,OzN,), 169.1251 (10, CteH,,Oz), 151.1147 (2, CIOH,sOI), 134.0635 (4, CsHsOrN,), 120.0560 (100, CsHsO,), 105.0341 (57, C,H,O,), 77.0385 (23, C6H5), 55.0565 (30, C,H,), 43.0556 (20, GH,).

Acidissiminol epoxide (2). Mp 146-149” (recrystallized from MeOH). IR V”,“:cm-‘: 3420 (-OH), 3334 (-NH), 1640 (-CONH), 1578, 1534, 1510 (aromatic C-C), 1110 (C-O-C), 826,808) (>C=C-H): HRCIMS: m/z (>m/z 100, rel. int.) [M+H]’ 410.2308 (30, CzsHszO~N,), 392.2175(18,C2SH3,,0sN,),322.1787(15,C2rH2~OzN,), 242.1153 (98, C,,H,,O,N,), 169.1217 (89, C,,H,,Oz), 151.1084 (18, ClOH,,O,), 134.0630 (5, CsHsOrN,), 120.0551 (100, CsHsO,), 105.0334 (78, C,HsOr). HREIMS: m/z (rel. int.) 241.1087 (23, highest-mass ion, C,,H,,OzN,), 169.1245 (72, C,,,Hr,Oz), 151.1113 (4, C,,H,,O,), 134.0656 (4, C,H,O,N,), 120.0575 (100, CsHsO,), 105.0325 (83, C,H,O,), 83.0496 (43, CsH,O,), 71.0492 (80, C4H,0,), 55.0540 (97, C,H,), 43.0542 (37, C,H,).

760 P. GH~H et al.

N-Benzoyl tyramine (3). Mp 157-159” (recrystallized 2. Ghosh, P., Bandyopadhyay, A. K., Thakur, S., Akihisa, from MeOH), (lit. [3], mp 159-160”). Spectral data of N- T., Simizu, N., Tamura, T. and Matsumoto, T. (1989) .I. benzoyl tyramine have been described previously [3]. Nat. Prod. 52, 1323.

Acknowledgements-Thanks are due to UGC, New Delhi, India, for the grant of a Teacher-Fellowship to Mrinal Kanti Ghosh and Jasodhara Dan.

REFERENCES

1. Chopra, R. N., Nayar, S. L. and Chopra, I. C. (1956) in Glossary of Indian Medicinal Plants, p. 154. C.S.I.R., New Delhi.

3. Ghosh, P., Sil, P., Das, S., Thakur, S., Kokke, W. C. M. C., Akihisa, T., Simizu, N., Tamura, T. and Matsum- oto, T. (1991) J. Nat. Prod. 54, 1389.

4. Roberts, J. D., Weighert, F. J., Kroschwitz, J. I. and Reich, H. J. (1970) J. Am. Chem. Sot. 92, 1338.