Molbank 2008, M581

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ISSN 1422-8599

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Short Note

Further Aporphine Alkaloids from Phoebe lanceolata

Deepak K. Semwal ¹*, Usha Rawat ¹ and G. J. P. Singh ²

¹  Department of Chemistry, H.N.B. Garhwal University Srinagar, Uttarakhand 246174, India

²  Department of Chemistry, Punjab University, Chandigarh, Punjab, India

*Author to whom correspondence should be addressed; E-mail: [email protected]; Tel. (+91) 9412947995

Received: 30 September 2008 / Accepted: 6 November 2008 / Published: 23 November 2008

Abstract: Stem bark of Phoebe lanceolata was extracted with ethanol and fractionated with ethyl acetate yielded soluble and insoluble fractions. Ethyl acetate insoluble fraction was subjected to column chromatography afforded two oxalyl-fused didehydroaporphine alkaloids, N-6/C-7 oxalyl-fused 2,9-dihydroxy-1,10-dimethoxy 6a,7-didehydroaporphine and N-6/C-7 oxalyl-fused 1,2,9,10-tetramethoxy 6a,7-didehydroaporphine along with well known β-sitosterol and β-sitosterol glucoside. The structures of isolated compounds were elucidated by chemical and spectral analysis.

Keywords: Phoebe lanceolata, Lauraceae, aporphine alkaloid, laurodionine

1. Introduction

Phoebe lanceolata belonging to family Lauraceae is an evergreen tree and well reputed in traditional medicine in India [1]. Ethanolic extract of the stem bark showed antidiabetic, antibacterial and antifungal activity (preliminary work done by us at different laboratory). We recently reported an aporphine alkaloid, nordelporphine [2] from this source and now outlined the isolation and characterization of two oxalyl-fused didehydroaporphine alkaloids.

2. Results and discussion

Compound 1 was isolated as black crystals, m.p. 280-283⁰C (uncorr.) deduced the molecular formula C₂₀H₁₅NO₆ from the molecular ion at m/z 365.7 in the LC-EIMS (positive mode). This compound was elucidated as N-6/C-7 oxalyl-fused 2,9-dihydroxy-1,10-dimethoxy 6a,7-didehydroaporphine by direct comparison (UV, IR and NMR) to published data for laurodionine [3] isolated from P. formosana. Compound 2 was isolated as black-brown crystals, m.p. 205⁰C deduced the molecular formula C₂₂H₁₉O₆N from the molecular ion at m/z 393.3 in the EIMS. The IR absorptions at υ 1734 cm^-1 was characteristic for carbonyl function. ¹H NMR spectrum revealed the presence of four methoxy (δ 3.92, 3.63, 3.76 and 3.85) and three aromatic protons (δ 7.81, 7.12 and 6.73).

Figure 1. Chemical structures of compound 1 and 2.

Figure 2. Important HMBC and NOESY correlations of compound 2.

¹³C NMR spectrum expressed the evidence for two carbonyl groups (δ 163.83 and 173.33). DEPT (135 ⁰) showed the presence of four CH₃, two CH₂, three CH and thirteen quaternary carbons. HMBC and NOESY correlations are shown in Fig. 2. These data were somewhat similar to that of 1 except the presence of two methoxy groups instead hydroxy groups. On methylation [4], compound 1 afforded black brown product identical to 2 indicated that the locations of methoxy groups in 2 were similar to those of hydroxy groups in 1. The HMBC correlation of OCH₃-1 (δ 3.92) to C-1 (δ 144.54), OCH₃-2 (δ 3.63) to C-2 (δ 159.28), OCH₃-9 (δ 3.76) to C-9 (δ 152.23) and OCH₃-10 (δ 3.85) to C-10 (δ 148.77) whereas the NOESY correlation of OCH₃-1 to OCH₃-2 and H-11 (δ 7.81); OCH₃-2 to OCH₃-1 and H-3 (δ 7.12); OCH₃-9 to OCH₃-10 and H-8 (δ 6.73) and OCH₃-10 to OCH₃-9 and H-11 were further confirmed the position of all methoxy groups. EIMS (positive mode) revealed a molecular ion at m/z 393, another ion at m/z 377 (loss of CH₄) and the most abundant ion [C₁₆H₅NO₄]^•+ at m/z 275 was due to loss of C₅H₁₀O₂. On the basis of these findings and the proposed structure described by Castedo et al., [5], the compound 2 was characterized as N-6/C-7 oxalyl-fused 1,2,9,10-tetramethoxy 6a,7-didehydroaporphine.

3. Experimental

3.1. General

Melting points were determined on Perfit melting point apparatus; UV spectra on Perkin-Elmer, Lambda- 25 spectrophotometer in MeOH; IR spectra on Perkin-Elmer, Spectrum RX I FT-IR spectrophotometer (KBr discs); NMR spectra on JEOL NMR spectrophotometer (300 MHz for ¹H and 125 MHz for ¹³C in DMSO, TMS as internal standard); LC-EIMS on Finnigan MAT spectrophotometer. Preparative TLC (0.5 mm thick layer) was carried out on silica gel (Merck 10-40 μ) spots were detected using UV at 254 and 365 nm and Dragendorff’s reagent.

3.2. Plant material

Stem bark (6 kg) of P. lanceolata was collected from Kartikswami temple, Dist. Chamoli (Uttarakhand) and identified by Prof. R.D. Gaur, Department of Botany, H.N.B. Garhwal University Srinagar. A voucher specimen (GUH-17598) of the plant was deposited in the Departmental Herbarium.

3.3. Extraction and isolation

Coarsely powdered stem bark (2 kg) was extracted twice with 95% ethanol (5L) at 50⁰C (15 hours) on a heating mantle. After removal of the solvent under reduced pressure, the residue (230 g) was fractionated with EtOAc (repeated 3-4 times) yielded soluble and insoluble portions. The insoluble portion (120 g) was pre-adsorbed onto silica gel (50 g) and subjected to column chromatography over silica gel (500 g, Merck, 60-120 mesh). The elution was perform first with CHCl₃ and then with CHCl₃ containing increasing amount of MeOH. The fractions obtained were collected every 50 ml. The elution with CHCl₃–MeOH (47:3→9:1) afforded twelve fractions of 250 ml. These fractions were combined on the basis TLC analysis and subjected to preparative TLC in CHCl₃–MeOH (8:2) afforded compounds 1 (23 mg) and 2 (20 mg), purified by recrystallised with CHCl₃–MeOH (1:1).

3.5. N-6/C-7 oxalyl-fused 1,2,9,10-tetramethoxy 6a,7-didehydroaporphine (2)

Black-brown crystals (CHCl₃/MeOH); m.p. 205-207⁰C; [α]: +47⁰ (c 0.3, MeOH); M.F. C₂₂H₁₉NO₆, UV λ: 206, 268, 434 nm; IR υ: 2956, 1734, 1662, 1554, 1471 cm^-1. ¹H, ¹³C, HMBC and NOESY NMR data: see Table-1; LCMS: 393 [M]⁺, 377 [C₂₁H₁₅NO₆]^•+, 275 [C₁₆H₅NO₄]^•+, 148 [C₈H₆NO₂]⁺; Elemental analysis: (found C- 67.24%, H- 04.84%, N- 03.56 and O- 24.46%; calculated for C₂₂H₁₉NO₆ C- 67.17%, H- 04.87%, N- 03.56 and O- 24.40%).

Table 1. ^13C and ^1H NMR data of 2 in DMSO d^6
Position	δC ppm	δH ppm (J Hz)	DEPT	NOESY	HMBC
1	144.54	-	C
1a	126.20	-	C
1b	127.95	-	C
2	159.28	-	C
3	114.56	7.12 s	CH	3.63	127.95, 144.54
3a	133.82	-	C
4	25.87	2.85 t (3.8)	CH2	3.18	114.56, 127.95
5	36.88	3.18 t (3.8)	CH2	2.85	151.47, 163.83
6a	151.47	-	C
7	102.42	-	C
7a	124.76	-	C
8	104.06	6.73 s	CH	3.76	102.42, 115.83, 148.77
9	152.23	-	C
10	148.77	-	C
11	108.63	7.81 s	CH	3.85, 3.92	124.76, 126.20, 152.23
11a	115.83	-	C
12	163.83	-	C
13	173.33	-	C
OCH3-1	58.85	3.92 s	CH3	3.63, 7.81	144.54
OCH3-2	58.11	3.63 s	CH3	3.92, 7.12	159.28
OCH3-9	56.36	3.76 s	CH3	3.85, 6.73	152.23
OCH3-10	55.47	3.85 s	CH3	3.76, 7.81	148.77

3.6. Methylation of compound 1

Dimethyl sulfate (1.5 mg) and dry potassium carbonate (1.5 mg) were added to compound 1 (5 mg) in 5 ml of acetone. The mixture was stirred vigorously and refluxed for 30 minutes in round bottom flask on water bath. The concentrated filtrate afforded methylated product, M.P. 205-206⁰C as a black brown compound.

Acknowledgements

The work was supported by UGC New Delhi, India; Grant No. 33-282/2007(SR). Authors are grateful to Prof. M. S. M. Rawat for providing lab facilities and NIPER Chandigarh for scanning LCMS of isolated compounds.

References

1. Gaur, R.D. Flora of Garhwal North West Himalaya. Trans Media: Srinagar Garhwal. 1999, p. 59.

2. Semwal, D.K.; Rawat, U. Nordelporphine alkaloid from bark of Phoebe lanceolata. Orient. J. Chem. 2007, 23, 771-772.

3. Chen, C.C.; Huang, Y.L.; Lee, S.S.; Ou, J.C. A new oxalyl- fused aporphine alkaloid from Phoebe formosana. J. Nat. Prod. 1997, 60, 826-827.

4. Luo, Y.; Liu, Y.; Luo, D.; Li, B.; Zhang, G. Cytotoxic alkaloids from Boehmeria siamensis. Planta Med. 2003, 69, 842-845.

5. Castedo, L.; Sal, C.; Sad, J.M.; Suau, R. Synthesis of oxoaporphines. An unusual photocyclization-photoreduction of 2,3-diaryl-Δ²-pyrroline-4,5-diones. J. Org. Chem. 1982, 47, 513-517.

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