New Lignans from the Leaves and Stems of Kadsura philippinensis

Abstract

Three novel C19 homolignans, taiwankadsurins D (1), E (2) and F (4), and two new C18 lignans kadsuphilins N (3) and O (5) were isolated from the aerial parts of Taiwanese medicinal plant Kadsura philippinensis. The structures of compounds 1–5 were determined by spectroscopic analyses, especially 2D NMR techniques. The structure of compound 5 was further confirmed by X-ray crystallographic analysis. Compounds 1 and 2 have a 3,4-{1'-[(Z)-2''-methoxy-2''-oxoethylidene]}-pentano(2,3-dihydrobenzo[b]furano)-3-(2'''-methoxycarbonyl-2'''-hydroxy-2''',3'-epoxide) skeleton.

1. Introduction

Kadsura belongs to the family Schisandraceae and it is only distributed in eastern and southern Asia [1]. Species of Kadsura were used in Chinese folk medicine for the treatment of cold, rheumatoid arthritis and gastroenteritis and as an anodyne to relieve pain [2]. The major constituents of Kadsura plants were reported to be bioactive lignans, which possess antitumor, antiviral and anti-hepatitic activities [3,4,5,6,7,8]. K. philippinensis Elm. is an evergreen vine, mainly distributed at low altitude onremote islands of Taiwan such as Green Island [9]. Our previous phytochemical studies on the EtOAc extracts of K. philippinensis resulted in the isolation of two novel triterpene dilactones and many lignans [10,11,12,13,14,15,16,17]. In this paper, we report the isolation and structure elucidation of three new C19 homolignans, named taiwankadsurins D-F, and two new C18 lignans, designated kadsuphilins N and O.

2. Results and Discussion

The leaves and stems of K. philippinensis were extracted with mixture of CH₂Cl₂ and acetone, then suspended in H₂O and extracted with EtOAc. The EtOAc-soluble part was subjected to extensive chromatography including flash column, normal and reversed-phase HPLC, furnishing compounds 1–5 (Figure 1).

Figure 1. Chemical structures of compounds 1–6.

Figure 1. Chemical structures of compounds 1–6.

Taiwankadsurin D (1), (${[\alpha ]}_D^{25}$ +57°, CH₂Cl₂) had a molecular formula C₂₉H₃₂O₁₃, as derived from its HREIMS at m/z 611.1735 ([M+Na]⁺, calcd 611.1741) indicating 14 degrees of unsaturation. The UV absorption (273, 225 nm) and IR bands (1,731, 1,721 and 1,628 cm⁻¹) indicated a benzyl and α,β, unsaturated ester functionalities. The ¹H-NMR of 1 exhibited two methoxyl singlets (δ 3.93, 3.59), an acetyl singlet (δ 2.13), two methyl singlets (δ 1.31, 1.99), two methyl doublets (δ 1.36, J = 6.9 Hz; δ 2.05, J = 7.2 Hz), two oxymethylene protons (δ 5.00, 4.53, each d, J = 10.2 Hz) and two dioxymethylene protons (δ 5.97, 5.98, each s-like). According to ¹³C-NMR and DEPT spectra, compound 1 had total 29 signals including seven methyl, two methylene, six methine and fourteen quaternary carbons. Moreover, ¹H-NMR spectroscopic data of 1 showed characteristic signals of H-4 (δ 5.99), H-6 (δ 6.28) and H-9 (δ 6.55), and ¹³C-NMR data of C-1 (δ 97.5 s), C-2 (δ 171.0 s) and C-3 (δ 165.4 s) similar to those of taiwankadsurin A (6), suggesting that compound 1 is an analogue of the latter [10]. However, a benzoyl group in 6 was missing and replaced with an angeloyl group at C-6 in 1. Further HMBC correlations (Figure 2) of H-11/C-12, C-13, C-15 and H-20/C-14, C-15, C-16, confirmed that compound 1 possessed a dihydrobenzofuran system. The ethylidene-octane ring was also deduced from the HMBC correlations of H-9/C-7,C-10,C-11,C-15; Me-18/C-7,C-8,C-9; Me-17/C-6,C-7,C-8 and H-6/C-4,C-5, C-7, C-8. The acetyl and angeloyl groups attaching at C-9 and C-6 respectively, were resulted from the HMBC correlations of H-9 (δ 6.55) with the acetyl carbonyl, and H-6 (δ 6.28) with the angeloyl carbonyl. Furthermore, methoxyl groups (δ_Η 3.93, δ_Η 3.59) attaching at carbonyls C-2 (δ_C 171.0) and C-3 (δ_C 165.4) were deduced from their mutual HMBC correlations.

It was noted that the dioxygenated tertiary carbon C-1 connected to C-7 through an ether bridge to account for the last degree of unsaturation. The relative configuration of 1 was determined by the NOESY experiment and by comparing the NMR data of 1 with those of taiwankadsurin A (Figure 2). Assuming that H-9 was β-oriented due to quite similar NMR spectra of 1 and taiwankadsurin A [10], thus, cross peaks between H-4, H-9 and Me-5', and correlation between H-9 and H-8, rather than Me-18 suggested that H-8 and 6-O-angeloyl group should be positioned on the β−face of the molecule. On the other hand, correlation between Me-18(eq) and Me-17(eq) accounted for the α−disposition of the ether ring between C-1 and C-7. In addition, NOESY correlation between H-6 and the methoxyl protons at C-2 indicated that H-6 and the hydroxyl group attached at C-1 are α−oriented. On the basis of above findings, the relative configuration of 1 was assigned as 1R*, 6S*, 7S*, 8S*, 9R*, 16S*.

Figure 2. Selected HMBC (arrow) and NOESY (double headed arrow) correlations of 1.

Figure 2. Selected HMBC (arrow) and NOESY (double headed arrow) correlations of 1.

Taiwankadsurin E (2) is an isomer of 1 as inferred from the identical molecular weight in HRMS, similar UV and IR absorptions and NMR data. The ¹H-NMR spectrum (Table 1) of 2 had the same characteristic peaks with 1 except that H-6 was downfield shifted to δ_Η 6.91, while the methoxyl protons at C-2 was upfield shifted to δ_Η 3.61. Detail analysis of HMBC correlations of 2 revealed that the locations of angeloyl, acetyl and methoxyl groups were the same as 1. The configuration of 2 was established from NOESY experiment, in which most of the cross peaks were identical to those of 1. However, the correlation between H-6 and the methoxy at C-2 was missing in 2. Therefore, the structure of 2 was established, being an 1-epimer of 1.

Table 1. ¹H-NMR data (CDCl₃) of compounds 1–5 ^a,b.

Position	1 a	2 b	3 a	4 b	5 a
4	5.99, brs	6.06, d (2.4)	6.84, s	3.08, d (18.4 )	7.34, s
				3.17, d (18.4 )
6	6.28, d (2.7)	6.69, d (2.4)	5.76, s	5.42, s
8	2.23, m	2.23, m	1.97, m	2.00, m
9	6.55, d (2.7)	6.63, d (2.8)	5.48, s	4.82, brs	4.87, d (12.9)
11	6.45, s	6.44, s	6.47, s	6.28, s	6.71, s
17	1.31, s	1.34, s	1.37, s	0.97, s	1.31, s
18	1.04, d (6.9)	1.02, d (6.8)	1.30, d (6.9)	1.36, d (7.6)	0.98, s
19	5.97, s	5.93, s	5.94, s	5.83, s	5.90, d (1.5)
	5.98, s	5.94, s	6.03, s	5.98, s	5.91, d (1.5)
20	4.53, d (10.2)	4.59, d (10.0)		4.30, d (9.6)
	5.00, d (10.2)	4.98, d (10.0)		4.43, d (9.6)
OMe-1			3.46, s
OMe-2	3.93, s	3.57, s	3.84, s	3.66, s	3.77, s
OMe-3	3.59, s	3.58, s	3.92, s	4.07, s	4.11, s
OMe-14					3.81, s
OAc	2.13, s	2.13, s	1.49, s
1'
2'
3'	6.28, overlap	6.23, q (7.2)	1.92, m	7.32, m
4'	2.05, d (7.2)	2.06, d (7.2)	3.63, dd (5.0, 8.0)	7.35, m
			4.16, dd (5.0, 8.0)
5'	1.99, s	2.00, s	1.23, s	7.55, d (7.2)
6'			0.96,d (7.2)	7.35, m
7'				7.32, m
OH-9					4.28, d (12.9)

^a recorded at 300 MHz. ^b recorded at 400 MHz.

Table 1. ¹H-NMR data (CDCl₃) of compounds 1–5 ^a,b.

Position	1 a	2 b	3 a	4 b	5 a
4	5.99, brs	6.06, d (2.4)	6.84, s	3.08, d (18.4 )	7.34, s
				3.17, d (18.4 )
6	6.28, d (2.7)	6.69, d (2.4)	5.76, s	5.42, s
8	2.23, m	2.23, m	1.97, m	2.00, m
9	6.55, d (2.7)	6.63, d (2.8)	5.48, s	4.82, brs	4.87, d (12.9)
11	6.45, s	6.44, s	6.47, s	6.28, s	6.71, s
17	1.31, s	1.34, s	1.37, s	0.97, s	1.31, s
18	1.04, d (6.9)	1.02, d (6.8)	1.30, d (6.9)	1.36, d (7.6)	0.98, s
19	5.97, s	5.93, s	5.94, s	5.83, s	5.90, d (1.5)
	5.98, s	5.94, s	6.03, s	5.98, s	5.91, d (1.5)
20	4.53, d (10.2)	4.59, d (10.0)		4.30, d (9.6)
	5.00, d (10.2)	4.98, d (10.0)		4.43, d (9.6)
OMe-1			3.46, s
OMe-2	3.93, s	3.57, s	3.84, s	3.66, s	3.77, s
OMe-3	3.59, s	3.58, s	3.92, s	4.07, s	4.11, s
OMe-14					3.81, s
OAc	2.13, s	2.13, s	1.49, s
1'
2'
3'	6.28, overlap	6.23, q (7.2)	1.92, m	7.32, m
4'	2.05, d (7.2)	2.06, d (7.2)	3.63, dd (5.0, 8.0)	7.35, m
			4.16, dd (5.0, 8.0)
5'	1.99, s	2.00, s	1.23, s	7.55, d (7.2)
6'			0.96,d (7.2)	7.35, m
7'				7.32, m
OH-9					4.28, d (12.9)

^a recorded at 300 MHz. ^b recorded at 400 MHz.

Kadsuphilin N (3), (${[\alpha ]}_D^{26}$ −2.4, CH₂Cl₂), had a molecular formula of C₃₀H₃₆O₁₂ as deduced from a pseudo-molecular ion [M+Na]⁺ at m/z 611.2107 in the HRESIMS. The UV absorption bands at 212, 259 and 292 nm suggested that 1 possessed a biphenyl chromophore. The IR absorption indicated the presence of hydroxyl (3,479 cm⁻¹) and carbonyl (1,738 cm⁻¹) groups. The ¹³C-NMR spectroscopic data and DEPT analysis revealed that compound 3 contains 30 carbons, including ten quaternary sp² carbons (δ_C 121.2, 121.7, 130.8, 132.8, 137.5, 139.0, 141.8, 148.6, 151.5 and 152.3), two ester carbons (δ_C 168.7 and 172.4), two quaternary sp³ oxygen-bearing carbons (δ_C 73.8 and 76.6), two oxygen-bearing methylene carbons (δ_C 72.4 and 101.5), two sp² methine carbons (δ_C 102.5 and 111.0), two sp³ methine carbons (δ_C 42.4 and 44.1), two oxygen-bearing sp³ methine carbons (δ_C 83.9 and 86.6), and eight methyl groups (δ_C 12.8, 17.8, 20.3, 21.4, 28.4, 56.2, 60.5 and 60.7). The HMBC correlations of H-11/C-9, C-10, C-12, C-13, C-15; H-9/C-10, C-15; H-4/C-2, C-3, C-5, C-6, C-16; H-6/C-5, C-16; Me-17/C-6, C-7, C-8; Me-18/C-7, C-8, C-9 implied that compound 3 indeed possessed a schizandrin type dibenzocyclooctadiene system [16]. Moreover, HMBC correlations of H₂-19/C-12, C-13; OMe-1/C-1; OMe-2/C-2; OMe-3/C-3 and H-9/ acetyl carbonyl assigned the methylenedioxy group and three methoxyl groups attached to the aromatic ring and an acetyl group at C-9. In addition, the ester linkage could be proved by correlations of H-6/C-1'; Me-5'/C-1', C-2', C-3'; Me-6'/C-2', C-3', C-4' and H-4'/C-14. From the above interpretation, the structure of 3 could be established as 9-acetylgomisin D. The configuration was determined by CD spectrum and NOESY experiment. The strong positive Cotton effect at 229 nm and the negative Cotton effect at 245 nm assigned the S-configuration of the biphenyl system [18]. The NOESY correlations of H-9/H-8, H-11, H-8/Me-17 and H-6/H-4, Me-17(eq) revealed that the cyclooctadiene ring had a twist-boat-chair form and H-8, H-9 and Me-17 were β-oriented while H-6 and Me-18 were α-configuration (Figure 3). The correlations of H-3'/Me-5' and the NMR data were in good agreement with the configuration of ester linkage that was also present in gomisin D [19].

Figure 3. Selected HMBC (arrow) and NOESY (double headed arrow) correlations of 3.

Figure 3. Selected HMBC (arrow) and NOESY (double headed arrow) correlations of 3.

Taiwankadsurin F (4) was isolated as a pale yellow amorphous solid. The molecular formula C₂₉H₂₈O₁₀ was deduced from a pseudo-molecular ion at m/z 559.1580 [M+Na]⁺ in HRESIMS. The UV spectrum showed absorptions at λ_max 255, 220 nm and IR bands at ν_max 3,510, 1,735, 1,725 cm⁻¹ suggested that compound 4 contained phenyl, benzoyl, α,β-unsaturated ketone and hydroxyl functionalities. The ¹H and ¹³C-NMR spectroscopic data (Table 1 , Table 2) revealed that 4 possessed a substituted cyclohex-2-enone moiety and a spirodihydrobenzofuran ring in a homolignan skeleton similar to kadsuphilol G [14]. The difference could be the angeloyloxy side chain, which was substituted with a benzoyloxy group. This scaffold was supported from HMBC correlations of H-19/ C-12, C-13; H-11/ C-9, C-12, C-13, C-15; Me-18/C-7, C-8, C-9; Me-17/C-6, C-7, C-8; H-4/C-2, C-3, C-5, C-16 and H-20/C-1, C-14, C-15. Moreover, the key HMBC correlations of H-6/ benzoyl carbonyl (δ_C 165.4) and H-9/ C-5 assigned the benzoyl group at C-6. It was found that an ether linkage appeared between C-5 and C-9 due to calculation of double bond equivalence. The relative configuration of 4 was determined by comparing the coupling constants of 4 with those of kadsuphilol G and NOESY experiments. Thus a twist-boat-chair configuration was elucidated on the basis of CD observation, in which a positive Cotton effect was found at 216 nm and a negative one at 249 nm. The NOESY correlations of H-11/H-8 /H-9 and H-8/ Me-17 suggested that H-8, H-9 Me-17 were all in β-face while Me-18 was α-oriented (Figure 4). Because compound 4 had a TBC-S configuration, the oxygen bridge could be assigned as α-disposed. On the basis of the above interpretation, the structure of compound 4 was established and the name taiwankadsurin F was given.

Table 2. ¹³C-NMR data (CDCl₃) of compounds 1–5 ^a,b.

Position	1 a	2 b	3 a	4 b	5 a
1	97.5, C	97.6, C	151.5,C	193.0, C	196.3, C
2	171.0, C	170.1, C	141.8,C	132.5, C	140.6, C
3	165.4, C	165.5, C	152.3, C	157.4, C	159.0, C
4	117.2, CH	118.3, CH	111.0, CH	40.7, CH2	123.4, CH
5	150.5, C	149.8, C	130.8, C	77.6, C	143.7, C
6	72.5, CH	72.5, CH	86.6 , CH	77.3, CH	200.7, C
7	79.2, C	78.4, C	73.8, C	72.5, C	80.7, C
8	45.4, CH	45.5, CH	44.1, CH	43.7, CH	60.4, C
9	70.3, CH	70.2, CH	83.9, CH	77.3, CH	75.7, CH
10	127.9, C	127.8, C	132.8,C	127.9, C	144.5, C
11	98.7, CH	99.9, CH	102.5, CH	95.9, CH	100.4, CH
12	150.4, C	149.8, C	148.6, C	151.3, C	151.2, C
13	129.1, C	128.9, C	137.5, C	129.5, C	136.4, C
14	144.9, C	142.6, C	139.0, C	140.9, C	139.5, C
15	118.0, C	120.5, C	121.2, C	121.3, C	125.7, C
16	57.0, C	58.6, C	121.7, C	56.9, C	69.5, C
17	28.2, CH3	28.5, CH3	28.4, CH3	23.1, CH3	19.9, CH3
18	8.9, CH3	8.5, CH3	17.8, CH3	15.3, CH3	15.1, CH3
19	101.8, CH2	101.5, CH2	101.5, CH2	101.3, CH2	101.7, CH2
20	80.4, CH2	78.5, CH2		78.2, CH2
OMe-1			60.5, CH3
OMe-2	53.6, CH3	54.0, CH3	60.7, CH3	60.7, CH3	60.2, CH3
OMe-3	51.8, CH3	51.7, CH3	56.2, CH3	58.9, CH3	58.3, CH3
OMe-14					59.4, CH3
OAc	168.9, C	168.9, C	168.7, C
	21.0, CH3	20.3, CH3	20.3, CH3
1'	166.0, C	166.1, C	172.4, C	165.4, C
2'	126.3, C	126.5, C	76.6, C	129.5, C
3'	142.3, CH	141.7, CH	42.4, CH	128.3, CH
4'	16.0, CH3	15.9, CH3	72.4, CH2	129.7, CH
5'	20.4, CH3	21.0, CH3	21.4, CH3	133.9, CH
6'			12.8, CH3	129.7, CH
7'				128.3, CH

^a recorded at 75 MHz. ^b recorded at 100 MHz.

Table 2. ¹³C-NMR data (CDCl₃) of compounds 1–5 ^a,b.

Position	1 a	2 b	3 a	4 b	5 a
1	97.5, C	97.6, C	151.5,C	193.0, C	196.3, C
2	171.0, C	170.1, C	141.8,C	132.5, C	140.6, C
3	165.4, C	165.5, C	152.3, C	157.4, C	159.0, C
4	117.2, CH	118.3, CH	111.0, CH	40.7, CH2	123.4, CH
5	150.5, C	149.8, C	130.8, C	77.6, C	143.7, C
6	72.5, CH	72.5, CH	86.6 , CH	77.3, CH	200.7, C
7	79.2, C	78.4, C	73.8, C	72.5, C	80.7, C
8	45.4, CH	45.5, CH	44.1, CH	43.7, CH	60.4, C
9	70.3, CH	70.2, CH	83.9, CH	77.3, CH	75.7, CH
10	127.9, C	127.8, C	132.8,C	127.9, C	144.5, C
11	98.7, CH	99.9, CH	102.5, CH	95.9, CH	100.4, CH
12	150.4, C	149.8, C	148.6, C	151.3, C	151.2, C
13	129.1, C	128.9, C	137.5, C	129.5, C	136.4, C
14	144.9, C	142.6, C	139.0, C	140.9, C	139.5, C
15	118.0, C	120.5, C	121.2, C	121.3, C	125.7, C
16	57.0, C	58.6, C	121.7, C	56.9, C	69.5, C
17	28.2, CH3	28.5, CH3	28.4, CH3	23.1, CH3	19.9, CH3
18	8.9, CH3	8.5, CH3	17.8, CH3	15.3, CH3	15.1, CH3
19	101.8, CH2	101.5, CH2	101.5, CH2	101.3, CH2	101.7, CH2
20	80.4, CH2	78.5, CH2		78.2, CH2
OMe-1			60.5, CH3
OMe-2	53.6, CH3	54.0, CH3	60.7, CH3	60.7, CH3	60.2, CH3
OMe-3	51.8, CH3	51.7, CH3	56.2, CH3	58.9, CH3	58.3, CH3
OMe-14					59.4, CH3
OAc	168.9, C	168.9, C	168.7, C
	21.0, CH3	20.3, CH3	20.3, CH3
1'	166.0, C	166.1, C	172.4, C	165.4, C
2'	126.3, C	126.5, C	76.6, C	129.5, C
3'	142.3, CH	141.7, CH	42.4, CH	128.3, CH
4'	16.0, CH3	15.9, CH3	72.4, CH2	129.7, CH
5'	20.4, CH3	21.0, CH3	21.4, CH3	133.9, CH
6'			12.8, CH3	129.7, CH
7'				128.3, CH

^a recorded at 75 MHz. ^b recorded at 100 MHz.

Figure 4. Selected HMBC (arrow) and NOESY (double headed arrow) correlations of 4.

Figure 4. Selected HMBC (arrow) and NOESY (double headed arrow) correlations of 4.

Kadsuphilin O (5) was obtained as pale yellow crystals, with molecular formula C₂₂H₂₂O₉ as determined by HRESIMS (12 degrees of unsaturation). IR absorption bands at 3,420, 1,716 and 1,620 cm⁻¹ indicated the presence of hydroxyl, carbonyl and aromatic moieties. The ¹H-NMR data (Table 1) and HMQC spectrum showed characteristic signals for two aromatic (δ_H 6.71, 7.34), one methoxyene-dioxy (δ_H 5.90, 5.91 as an AB quartet), one oxygen-bearing methine (δ_H 4.87), two tert-methyl (δ_H 0.98, 1.31) and three methoxyl (δ_H 3.77, 3.81 and 4.11) protons. A methine doublet at δ_H 4.28 (J = 12.9 Hz) revealed the presence of a hydroxy due to no correlation was found in HMQC. ¹³C-NMR data and DEPT spectra revealed that compound 5 contained five pairs of double bonds (δ_C 100.4, 123.4, 125.7, 136.4, 139.5, 140.6, 143.7, 144.5, 151.2, 159.0), two ketone carbonyl carbons (δ_C 196.3 and 200.7) and three quaternary carbons (δ_C 60.4, 69.5, 80.7), one oxygenated methine carbon (δ_C 75.7), a methylenedioxy carbon (δ_C 101.7), two methyl carbons (δ_C 15.1 and 19.9) and three methoxyl carbons (δ_C 58.3, 59.4, 60.2). Thus compound 5 possessed five ring systems after deduction of seven double bonds. In the HMBC spectrum, correlations of H-11/C-9, C-12, C-13, C-14, C-15; H-19/C-12, C-13; H-4/C-2, C-3, C-5, C-6, C-16; Me-17/C-6, C-7, C-8; Me-18/C-7, C-8; H-9/C-7, C-15 and C-9-OH/C-9 suggested a dibenzocyclo-octadiene framework with a ketone substituted at the C-6 position. Furthermore, the linkage between C-8 and C-16 was deduced by the correlations of H-9 and Me-18 with C-16, and the remaining ketone group could be assigned to the C-1 position. This finding was further confirmed by comparing the NMR data with those of heteroclitin G [20]. The relative configuration of 5 was determined by NOESY correlation and CD. The CD spectrum of 5 was similar to that of kadsutherin C [21]. The negative Cotton effect at 240 nm and the positive Cotton effect at 218 nm accounted for S-configuration for the biphenyl skeleton. Assuming that the H-9 of 5 was in a β-orientation similar to heteroclitin G, the NOESY correlations of HO-9/Me-18 and Me-18/Me-17 indicated that they were on the α-face and OH-7 was β-oriented. Therefore, the configuration of the bipentacyclic ring was established. The structure of 5 was finally confirmed by a single-crystal X-ray diffraction analysis, from which a perspective drawing of 5 is provided in Figure 5.

Figure 5. Selected HMBC correlations and X-ray perspective drawing of 5.

Figure 5. Selected HMBC correlations and X-ray perspective drawing of 5.

3. Experimental

3.1. General

Melting points were measured on a Büchi melting point B-540 apparatus and are uncorrected. Optical rotations were recorded on a JASCO DIP-1000 polarimeter. IR and UV spectra were measured on HORIBA FT-720 and U-3210 spectrophotometers, respectively. The ¹H- and ¹³C-NMR, COSY, HMQC, HMBC, and NOESY spectra were recorded respectively on a Bruker FT-300 spectrometer (300 MHz for ¹H and 75 MHz for ¹³C) or on a Bruker AVANCE 400 (400 MHz for ¹H and 100 MHz for ¹³C) using TMS as an internal standard. The chemical shifts were given in δ values (ppm) and coupling constants in Hz. Low-resolution FABMS were recorded on a VG Quattro 5022 mass spectrometer, and HREIMS were measured on a JEOL JMS-SX 102 spectrometer. Silica gel 60 (Merck) was used for column chromatography (CC), and precoated silica gel plates (Merck, Kieselgel 60 F-254, 1 mm) were used for preparative TLC.

3.2. Plant Material

The leaves and stems of K. philippinensis were collected at Green Island, Taiwan, in November, 2002. A voucher sample (specimen code: TP 93-2) was deposited at the School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan.

3.3. Extraction and Isolation

K. philippinensis was extracted with mixture of CH₂Cl₂ and acetone and partitioned between EtOAc and H₂O (1:1). The EtOAc-soluble part was subjected to Si gel column chromatography (n-hexane/EtOAc, 1:0 to 0:1), and after monitoring by ¹H-NMR, the middle fraction (fr. 21) was further eluted on LH-20 (MeOH) to give five subfractions (fr.21-1~5). Fr.21-5 was chromatographed on a flash column (Si gel, n-hexane/EtOAc, 15:1-0:1) and further separated by normal phase HPLC (n-hexane/CH₂Cl₂/MeOH, 35:65:1) to furnish taiwankadsurins D (1, 13 mg) and E (2, 2 mg). Kadsuphilin N (3, 14 mg) was isolated from fr.21-2, which was chromatographed on a flash column (n-hexane/ acetone/EtOAc, 15:1:1 to 1:1:1) and further purified with normal phase HPLC (n-hexane/CH₂Cl₂/ MeOH, 30:70:1) and reverse phase HPLC (MeOH/H₂O, 65:35) alternatively. Fraction fr.21-4 was separated on a Si gel column (n-hexane/EtOAc, 25:1 to 0:1) and a reverse phase HPLC (MeOH/H₂O, 65:35) column to yield taiwankadsurin F (4, 4 mg) and kadsuphilin O (5, 7 mg).

3.4. Spectroscopic Data

Taiwankadsurin D (1). ${[\alpha ]}_D^{26}$ +57 ° (c 0.5, CH₂Cl₂); UV λ_max (MeOH) 225, 273 nm; CD (MeOH, c = 0.2) nm (ε) 222 (−1.30), 254 (+1.27); IR (neat) ν_max 3,450, 2,938, 1,731, 1,721, 1,628 cm⁻¹; ¹H-NMR and ¹³C-NMR (CDCl₃, 300/75 MHz) see Table 1 , Table 2, respectively; HRESIMS m/z 611.1735 (calcd for C₂₉H₃₂O₁₃Na, 611.1741).

Taiwankadsurin E (2). ${[\alpha ]}_D^{26}$ −11° (c 0.2, CH₂Cl₂); UV λ_max (MeOH) 233, 276 nm; CD (MeOH, c = 0.2) nm (ε) 228 (−0.78), 247 (+0.27); IR (neat) ν_max 3,457, 1,728, 1,717 cm⁻¹; ¹H-NMR and ¹³C-NMR (CDCl₃, 400/100 MHz) see Table 1 , Table 2, respectively; HRESIMS m/z 611.1737 (calcd for C₂₉H₃₂O₁₃Na, 611.1741).

Kadsuphilin N (3). ${[\alpha ]}_D^{25}$ −2.4° (c 1.3, CH₂Cl₂); UV λ_max (MeOH) 212, 259, 292 nm; CD (MeOH, c = 0.16) nm (ε) 229 (+33.56), 245 (−2.97), 293 (−5.54); IR (neat) ν_max 3,479, 1,738, 1,624, 1,594 cm⁻¹; ¹H-NMR and ¹³C-NMR (CDCl₃, 300/75 MHz) see Table 1 , Table 2, respectively; HRESIMS m/z 611.2107 (calcd for C₃₀H₃₆O₁₂Na, 611.2104).

TaiwankadsurinF (4). ${[\alpha ]}_D^{25}$ −13.2° (c 0.6, CH₂Cl₂); UV λ_max (MeOH) 220, 255 nm; CD (MeOH, c = 0.3) nm (ε) 216 (+17.34), 249 (−8.77), 290 (−1.53); IR (neat) ν_max 3,510, 1,735, 1,725, 1,660, 1,580 cm⁻¹; ¹H-NMR and ¹³C-NMR (CDCl₃, 400/100 MHz), see Table 1 , Table 2, respectively; HRESIMS m/z 559.1573 (calcd for C₂₉H₂₈O₁₀Na, 559.1580).

Kadsuphilin O (5). ${[\alpha ]}_D^{25}$ −8.0° (c 0.6, CH₂Cl₂); MP 167 °C; UV λ_max (MeOH) 215, 246, 283 nm; CD (MeOH, c = 0.22) nm (ε) 218 (+7.66), 240 (−28.11), 282 (−6.75); IR (neat) ν_max 3,420, 1,716, 1,620 cm⁻¹; ¹H-NMR (CDCl₃, 300 MHz) and ¹³C-NMR (CDCl₃, 75 MHz), see Table 1 and Table 2, respectively; HRESIMS m/z 453.1158 (calcd for C₂₂H₂₂O₉Na, 453.1161). Crystal data: C₂₂H₂₂O₉, M = 430.40, trigonal system, space group P2₁, a = 10.706(2), b = 8.218(2), c = 10.9345(9) Å, V = 960.1(3) ų, Z = 2, d = 1.489 Mg/cm³. A crystal of dimensions 0.60 × 0.60 × 0.20 mm was used for measurements on a RIGAKU AFC7S diffractometer with a graphite monochromator (ω-2θscans, 2θ_max= 52.0°), Mo Kα radiation. The total number of independent reflections measured was 2,134, of which 2026 were observed (|F|² ≥ 2σ|F|²). The crystal structure was solved by the direct method SHELX-86 [22] and expanded using difference Fourier techniques, refined by the program SHEXTL-97 [23] and full-matrix least-squares calculations. Final indices: R_f = 0.030, R_w = 0.0784, w = 1/[σ² (F_o²) + (0.070P)² + 0.1457P], where P = (F_o² + 2 F_c²)/3). Copies of the deposited crystal data (CCDC 829589) can be obtained, free of charge, from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: +44 (0) 1223 336033 or E-Mail: [email protected].

4. Conclusions

Phytochemical investigation of the aerial part of Taiwanese Kadsura philippinensis has resulted in isolation of five new lignans 1–5, including three novel C19 homolignans, designated taiwankadsurins D, E and F. Their structures have been established by spectroscopic analyses, especially 2D NMR techniques. In addition, the structure of compound 5 was further confirmed by X-ray crystallographic analysis.