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Thoreliolides A and B, two polyisoprenylated benzoylphloroglucinol derivatives with a new carbon skeleton from the fruits of Calophyllum thorelii
Tetrahedron Letters xxx (2016) xxx–xxx
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Thoreliolides A and B, two polyisoprenylated benzoylphloroglucinol derivatives with a new carbon skeleton from the fruits of Calophyllum thorelii Le-Thu T. Nguyen a, Ngan T. D. D. T. Lai a, Ly T. T. Nguyen a, Hieu T. Nguyen a, Duc M. Nguyen b, Lien-Hoa D. Nguyen a,⇑ a b
Natural Product and Medicinal Chemistry Lab, Faculty of Chemistry, VNUHCM–University of Science, 227 Nguyen Van Cu, District 5, Ho Chi Minh City, Viet Nam Faculty of Pharmacy, Ton Duc Thang University, 19 Nguyen Huu Tho, District 7, Ho Chi Minh City, Viet Nam
a r t i c l e
i n f o
Article history: Received 7 March 2016 Revised 25 April 2016 Accepted 4 May 2016 Available online xxxx Keywords: Calophyllum thorelii Guttiferae Polyisoprenylated acylphloroglucinols Thoreliolides A and B Cytotoxicity
a b s t r a c t Thoreliolides A and B, two new polyisoprenylated benzoylphloroglucinol derivatives with a novel carbon skeleton, together with four known compounds, were isolated from the fruits of Calophyllum thorelii and their structures elucidated using spectroscopic methods. Thoreliolide B showed moderate activity against three cancer cell lines, MCF-7, HeLa and NCI-H460, with IC50 values of 9.7, 16.6 and 24.2 lg/mL, respectively, using the sulforhodamine B assay. Ó 2016 Elsevier Ltd. All rights reserved.
Calophyllum thorelii Pierre (Guttiferae) is a large red hardwood tree, up to 30 in height.1 We have previously reported the isolation of a new xanthone, thorexanthone, from the fruits of the species collected in central Viet Nam.2 In addition, another new xanthone, calothorexanthone,3 two novel polyisoprenylated benzoylphloroglucinols, thoreliones A and B, and a new polyisoprenylated tetracyclic xanthone, oxy-thorelione A, were obtained from the bark.4 Continuing our chemosystematic investigation on this species, we herein describe the isolation and structure elucidation of two unusual polyprenylated benzoylphloroglucinol derivatives, thoreliolides A and B, together with four known compounds, from the fruits of the same plant. The cytotoxicity of the two new compounds using the sulforhodamine B (SRB) assay is also reported. Column chromatography of the petroleum ether extract of the fruits of C. thorelii led to the isolation of thoreliolides A (1) and B (2) (Fig. 1), along with four known compounds, thorelione A (3),4 guttiferone I,5 12b-hydroxy-des-D-garcigerrin A6 and 7 osajaxanthone. Thoreliolide A (1) was obtained as a yellow gum, [a]25 D +82.0 (c 0.45, EtOH), giving a positive reaction with ethanolic ferric chloride. The UV spectrum showed four maxima at 233, 268, 317 and ⇑ Corresponding author. Tel.: +84 8 38355270; fax: +84 8 38350096. E-mail addresses: (L.-H.D. Nguyen).
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365 nm whilst the IR spectrum exhibited absorption bands at 3415 (OAH), 1732 (C@O), 1626 (conjugated C@O), 1582 (aromatic C@C) and 1282 cm 1 (CAO). HRESIMS revealed the molecular formula to be C38H50O6 (m/z 625.3533 [M+Na]+), i.e., the molecule had fourteen degrees of unsaturation. The 1H and 13C NMR, and HMBC spectra of 1 (Table 1) contained resonances for a 1,2,4-trisubstituted benzene ring [dH 7.23 (1H, d, J = 1.9 Hz, H-12), 7.15 (1H, dd, J = 8.4, 1.9 Hz, H-16), 6.87 (1H, d, J = 8.4 Hz, H-15)], a conjugated carbonyl carbon (dC 194.8, C-10), two 3-methylbut-2-enyl groups [dH 5.27 (1H, m, H-35), 5.11 (1H, m, H-18), 2.94 and 2.90 (each 1H, m, H2-17), 2.28 and 1.82 (each 1H, m, H2-34), 1.76 (3H, s, H337), 1.73 (3H, s, H3-20), 1.72 (3H, s, H3-21), 1.67 (3H, s, H3-38)], a 2,2-dimethyl-3-isopropenylcyclopentyl ring [dH 4.79 and 4.60 (each 1H, s, H2-32), 2.38 (1H, m, H-26), 1.98 (1H, m, H-24), 2.02 (1H, m, H-27), 1.82 (1H, m, Ha-28), 1.71 (2H, m, H-27 and Hb-28), 1.63 (3H, s, H3-33), 1.08 (3H, s, H3-30), 0.73 (3H, s, H3-29); dC 145.9 (C-31), 109.8 (C-32), 51.1 (C-26), 43.1 (C-25), 38.9 (C-24), 38.0 (C-28), 30.3 (C-30), 29.5 (C-27), 23.2 (C-33), 16.5 (C-29)], two quaternary carbons (dC 46.6, C-8 and 39.0, C-5), a tertiary carbon (dC 42.0, C-6), a secondary carbon (dC 30.5, C-7), two tertiary methyls [dH 1.17 (3H, s, H3-22), 1.03 (3H, s, H3-23); dC 26.0 (C-22), 21.1 (C-23)], and an enolised-1,3-diketone moiety [dC 173.3 (C-1), 162.1 (C-3), 109.8 (C-2)]. The spectrum data of 1 were similar to those of thorelione A (3) previously isolated from
http://dx.doi.org/10.1016/j.tetlet.2016.05.021 0040-4039/Ó 2016 Elsevier Ltd. All rights reserved.
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L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx 21
20 28
21
21
20 18
OH 18
OH HO
13
HO
17 23
O
14
O 3
14
6 1
16
O
OH
34
25
O
38
O
12
14
27 24
5
O
O 9
3
7
10
27
25
29 33 34
33
17
7
32 31
30 37
HO
24
1
16
30 29
1
OH
22
5
1
2
27
9 8
16
24
4
3 10
22
8 10
22
23
O
9 5
23
17
O
13
4
20
28 35
O
OH
3
34
29 32
30
36
35
38
32 37 37
38
Figure 1. Structures of compounds 1–3.
Table 1 H (500 MHz) and
1
13
C (125 MHz) NMR data, and HMBC correlations for thoreliolides A (1) and B (2) in acetone-d6 (J in Hz in parentheses)
Position
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38
1
2
dH
HMBC correlations
dC
dH
HMBC correlations
dC
— — — — — 1.62 m Ha: 2.10 m Hb: 1.62 m — — — — 7.23 d (1.9) — — 6.87 d (8.4) 7.15 dd (8.4, 1.9) Ha: 2.94 m Hb: 2.90 m 5.11 m — 1.73 s 1.72 s 1.17 s 1.03 s 1.98 m
— — — — — C-7, C-23 C-5, C-24 C-6 — — — — C-10, C-13, C-14, C-16 — — C-11, C-13, C-14 C-10, C-12, C-14 C-4, C-5, C-9, C-18, C-19 C-4, C-5, C-9, C-18, C-19 C-17, C-20, C-21 — C-18, C-19, C-21 C-18, C-19, C-20 C-4, C-5, C-6, C-23 C-4, C-5, C-6, C-22 —
173.3 109.8 162.1 130.8 39.0 42.0 30.5
— C-25, C-31 C-24, C-25 C-24, C-26, C-28 C-6, C-24, C-25, C-27 C-24, C-26 C-24, C-25, C-25, C-30
43.1 51.1 29.5
— — — — — C-7 C-1, C-5, C-8, C-29 C-5 — — — — C-10, C-13, C-14, C-16 — — C-11, C-13, C-14, C-16 C-10, C-12, C-14 C-4, C-5, C-9, C-18, C-19 C-4, C-5, C-9, C-18, C-19 C-20, C-21 — C-18, C-19, C-21 C-18, C-19, C-20 C-4, C-5, C-6, C-23 C-4, C-5, C-6, C-22 C-6, C-25, C-26 C-6 C-27, C-28 — C-25, C-26, C-28
205.5 75.1 166.3 130.8 39.7 41.5 31.8
— 2.38 m Ha: 2.02 m Hb: 1.71 m Ha: 1.82 m Hb: 1.71 m 0.73 s
— — — — — 1.55 m Ha: 1.91 br d (13.5) Hb: 1.64 m — — — — 7.21 d (2.4) — — 6.76 d (8.2) 6.58 dd (8.2, 2.4) Ha: 3.08 m Hb: 2.92 m 5.17 m — 1.71 s 1.68 s 1.21 s 1.00 s 2.28 m 1.85 m 5.17 m — 1.63 s
38.0
1.72 s
C-25, C-26, C-27
26.1
16.5
C-24, C-25, C-25, C-29 — C-26, C-33 C-26, C-33 C-26, C-31, C-32 C-1, C-7, C-35, C-36 C-7, C-8 C-37, C-38 — C-35, C-36, C-38 C-35, C-36, C-37
30.3 145.9 109.8
C-1, C-8, C-9, C-30, C-31 C-1, C-8, C-30, C-34 — — C-2, C-30, C-31, C-33
46.2
1.08 s — Ha: 4.79 s Hb: 4.60 s 1.63 s Ha: 2.28 m Hb: 1.82 m 5.27 m — 1.76 s 1.67 s
Ha: 2.55 dd (13.6, 4.8) Hb: 1.59 t (13.6) 1.85 m — 1.17 s 1.45 s Ha: 2.28 m Hb: 1.73 m 5.17 m — 1.65 s 1.71 s
C-2, C-30, C-31, C-32 C-35, C-36 C-30 C-37, C-38 — C-35, C-36, C-38 C-35, C-36, C-37
46.6 143.9 194.8 128.0 117.6 145.3 151.5 115.3 124.3 26.0 124.9 131.3 18.0 26.0 26.0 21.1 38.9
23.2 29.4 124.8 133.4 18.1 26.2
the bark of the same plant.4 The notable differences were that the resonances for the isolated C-9 carbonyl carbon and the C-8 quaternary carbon in the phloroglucinol subunit of thorelione A (3) had been replaced by a carbon–carbon double bond carrying an oxygen atom [dC 143.9 (C-9) and 130.8 (C-4)] in compound 1.
51.0 142.7 190.1 129.1 116.5 146.0 150.9 115.2 121.8 26.5 124.4 131.8 18.1 25.9 26.2 21.0 29.2 123.5 133.8 18.0
45.3 52.0 17.0 23.7 28.7 124.3 133.4 18.0 25.9
In the HMBC spectrum of 1 (Fig. 2), the methylene protons of the first isoprenyl group (dH 2.94 and 2.90, H2-17) showed correlations to the fully-substituted mono-oxygenated olefinic carbons (dC 143.9 and 130.8) and a quaternary carbon (dC 39.0). As a result, the first was identified as C-9 which bore the oxygen atom, the
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L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx
21 21
20
20 18
OH 18
OH
23
H O
HO 14
O
12
22 6
H
3
16
24
34
HO
O
4
9
22
5
24
2
29
1
31
30 37
38
7
O
O 32
27
25
1
10
H
1
10
H
12
17 23
O 8
3
16
14
4 9 8
H
H O
HO
17
27
25
29 30 33
28 34
33
35
32 37
38
Figure 2. HMBC correlations for compounds 1 and 2.
Additional NOE interactions between Hb-7 (dH 1.62) and Ha-34 (dH 2.28 m) and H3-22 (dH 1.17) revealed that the isoprenyl attached to C-8 and the Cax-22 methyl group had the same orientation, i.e., the C-8 isoprenyl was equatorial. The structure of thoreliolide A was consequently established as 1. The second new compound, thoreliolide B (2), was obtained as a yellowish gum, [a]25 51.2 (c 0.75, EtOH), UV: (kmax) 234, 283 and D 314 nm, IR: (mmax) 3425 (OAH), 1750 (non-conjugated C@O), 1706 (conjugated C@O), 1604 (aromatic C@C), and 1286 cm 1 (CAO). HRESIMS showed a pseudo-molecular ion peak [M+H]+ at m/z 603.3647, suggesting that this compound and thoreliolide A (1) had the same molecular formula of C38H50O6. The 1H and 13C NMR spectra of 2 (Table 1) had signals due to a 3,4-dihydroxybenzoyl system, two 3-methylbut-2-enyl groups, two tertiary methyls [dH 1.21 (3H, s, H3-22), 1.00 (3H, s, H3-23); dC 26.2 (C-22), 21.0 (C-23)], and a C10 unit comprising a trisubstituted carbon–carbon double bond [dH 5.17 (1H, m, H-35); dC 133.4 (C-36), 124.3 (C-35)], two methylenes [dH 2.55 (1H, dd, J = 13.6, 4.8 Hz, Ha-29), 1.59 (1H, t, J = 13.6 Hz, Hb-29); dH 2.28 and 1.73 (each 1H, m, H2-34); dC 46.2 (C-29), 28.7 (C-34)], a methine [dH 1.85 (1H, m, H-30); dC 45.3 (C-30)], two allylic methyls [dH 1.71 (3H, s, H3-38), 1.65 (3H, s, H3-37); dC 25.9 (C-38), 18.0 (C-37)], two tertiary methyls [dH 1.45 (3H, s, H3-33) and 1.17 (3H, s, H3-32); dC 23.7 (C-33), and 17.0 (C-32)], and a quaternary carbon [dC 52.0 (C-31)]. In addition, the spectra showed the presence of a 4a,5,6,7-tetrahydrogen-3H-chromene-2,4-dione ring system consisting of a carbon–carbon double bond bearing an oxygen atom [dC 142.7 (C-9), 130.8 (C-4)], a 1,3-diketone moiety [dC 205.5 (C-1), 166.3 (C-3), 75.1 (C-2)], two quaternary carbons [dC 51.0 (C-8), 39.7 (C-5)], a tertiary carbon [dH 1.55 (1H, m, H-6); dC 41.5 (C-6)] and a secondary carbon [dH 1.91 (1H, br d, J = 13.5 Hz, Ha-7), 1.64 (1H, m, Hb-7); dC 31.8 (C-7)]. In the HMBC spectrum of 2 (Fig. 2), correlations of the methylene protons of the first isoprenyl side chain (H2-17), the second
second was C-4 which carried the isoprenyl group, and the third was C-5. In addition, the two tertiary methyls gave cross-peaks to each other, to C-4, C-5 and a tertiary carbon (dC 42.0). Accordingly, C-5 carried the two methyls and the tertiary carbon was located at C-6. A methylene proton of the 2,2-dimethyl-3-isopropenylcyclopentyl ring (Ha-28) was correlated to C-6 and this C10 side chain was therefore attached to the C-6 carbon. The methylene protons at dH 2.10 and 1.62 showed correlations to C5, and they were thus assigned as H2-7 (dC 30.5). The methylene protons of the remaining isoprenyl group correlated to C-7, a quaternary carbon (dC 46.6) and the enolised carbon (dC 173.3). The former was hence C-8 which carried the isoprenyl group and the latter was C-1 which bore a hydroxyl group. The remaining olefinic carbon (dC 109.8) was C-2 and the conjugated carbonyl carbon (dC 162.1) had to be C-3 in order to form an enol 1,3-diketone moiety. To account for fourteen degrees of unsaturation in the molecule, the oxygen atom attached to C-9 had to be bonded to C-3 to form a conjugated lactone ring. The compound thus possessed a 4a,5,6,7-tetrahydro-4-hydroxy-2H-chromen-2-one ring system. Correlations observed in the 1H–1H COSY spectrum (Fig. 3) were consistent with the proposed structure. The relative stereochemistry of 1 was established based on 13C chemical shifts of the two tertiary methyls attached to C-5 and NOE interactions. In guttiferone B,8 the C-5 axial methyl (C-22) was shielded by the equatorial geranyl side chain attached to C-6 [dC 23.8 (C-23) and 16.5 (C-22)] due to a c-gauche interaction. Meanwhile in the cases of 7-epi-clusianone (4)9 and thorelione A (3),4 the side chain bonded to C-6 was axial, and the axial methyl located at C-5 was therefore less shielded (dC 27.0 and 27.5 ppm, respectively). Therefore, in thoreliolide A (1), the 13C shifts of the two tertiary methyls attached to C-5 [dC 21.0 (Ceq-23) and 26.0 (Cax-22)] showed that the side chain at C-6 was axial. This was confirmed by an NOE interaction (Fig. 3) observed between Ha-7 (dH 2.10) and H-24 (dH 1.98).
21 21
20
20
OH OH HO
13
18
O
14
O
17
14
4
8 1
O
34
OH
3
7
H
6
H
27
25
NOESY
32 30
38
33
31
30 37
COSY
24
O 29 H H
O
2
27 25
22
7
1
16
24
23
5
10
22
35
1
17 4 9 8
5
10
O
23
O
9
3
16
HO
18
13
34
28 35
33 32 37
38
Figure 3. COSY and key NOE correlations for compounds 1 and 2.
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L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx
the methyls and the remaining olefinic carbon was C-35. One proton of the remaining methylene group (dH 2.28; dC 28.7) correlated to C-35 and C-36, and the methylene was therefore C-34. The H2-34 protons showed cross-peaks to C-35, C-36 and a tertiary carbon (dC 45.3) whilst H2-29 correlated to C-34, the tertiary carbon and a quaternary carbon (dC 52.0). As a result, the tertiary carbon was C-30 and the quaternary carbon was C-31. The remaining two tertiary methyls correlated to each other [H3-32 (dH 1.17) to C33 (dC 23.7) and H3-33 (dH 1.45) to C-32 (dC 17.0)], to C-30, C-31 and a quaternary carbon (dC 75.1). They were thus bonded to C31 and the quaternary carbon was C-2. The structure of the subunit was confirmed by analysis of the 1H–1H COSY correlations (Fig. 3). The remaining carbon (dC 166.3) which showed no correlation had to be the lactone carbonyl carbon (C-3) in the 4a,5,6,7-tetrahydrogen-3H-chromene-2,4-dione ring system. Similar to the case of thoreliolide A (1), the 13C shifts of Cax-22 and Ceq-23 methyls (dC 26.2 and 21.0, respectively) indicated that the isoprenyl group at C-6 was axial and H-6 was hence equatorial. The geometry of the bicyclic ring system required substituents on C-2 and C-8 to be equatorial. The coupling constants of H2-29 [dH 2.55 (dd, J = 13.6 and 4.8 Hz, Ha-29), 1.59 (t, J = 13.6 Hz, Hb-29)] revealed H-30 to be axial, and the C-30 isoprenyl group was thus equatorial. This was confirmed by comparison of the 13C shifts of the two tertiary methyls (Cax-32 and Ceq-33) attached to C-5 with those of
Table 2 Cytotoxicity of 1 and 2 using the SRB assay11,a
MCF-7 HeLa NCI-H460 a b
1
2
Camptothecin
72.3 ± 1.0 n.d.b n.d.b
9.7 ± 0.3 16.6 ± 0.9 24.2 ± 3.6
N/A 0.013 ± 0.001 0.006 ± 0.001
Mean of three measurements. Not determined.
isoprenyl group (H2-24), and the two tertiary methyls (H3-22 and H3-23) revealed that the first isoprenyl side chain was attached to C-4 whilst the two methyls were bonded to C-5 and the second isoprenyl group was located at C-6. The two methylene protons at dH 1.91 and 1.64 correlated to C-5 and were therefore bonded to C7. The H2-7 protons gave cross-peaks to a quaternary carbon (dC 51.0), an isolated carbonyl carbon (dC 205.5) and a methylene (dC 46.2), which were thus assigned as C-8, C-1, and C-29, respectively. Correlations of the methylene protons of the C10 unit [dH 2.55 and 1.59 (H2-29)] with C-1, C-8 and C-9 established a bond between C-8 and C-9. Meanwhile, the two allylic methyls of the C10 unit (H3-37 and H3-38) correlated to each other and to two carbons of a trisubstituted carbon–carbon double bond (dC 133.4, s and 124.3, d). The fully substituted olefinic carbon (C-36) hence carried Enz B H OH OH O
O
5
O
HO
O
[O] Hydroxylation O
10
OH
O
5
3
4 9
+ H2 O
5
8 10
Retro-Claisen
8
1
4
O
Enz H
9
3
OH HO
6
OH
6
1
O
OH
H2 O Cyclisation
O
O
O
4
9
H
5
3 8
O
1
O
OPP
5
3
10
HO
O H
O
5 8
10
1
7
6
1
O
HO
4
9
3
6
8
7
1
O
4
9
6
8
HO
O
5
3
10
O
4
9
6
1
10
O
17
17
17
17
7
HO
PPO
C H2
Scheme 1. Plausible biosynthetic pathway for 1.
O
O
Tautomerism O
4
9
5
3
4
9
6
8 2
1
HO
7
1
O
1. DMAPP
5
6
8 2
O
O 3
7
O
O
2
O
O
6 OPP
4
9
3
5 8
8 1
O
O
O
4
9
5
2. - H
C5
3
6 7
C5
10 1
O
O
7
C5
2
DMAPP Scheme 2. Plausible biosynthetic pathway for 2.
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L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx
propolone A.10 In propolone A, the equatorial isoprenyl side chain at C-30 shielded the axial methyl [dC 15.7 (Cax-32) and 23.7 (Ceq-33)] due to a c-gauche interaction. In thoreliolide B (2), the resonances of the C-31 methyls [dC 17.0 (Cax-32) and 23.7 (Ceq-33)] revealed that the isoprenyl group attached to C-30 was equatorial. NOE interactions (Fig. 3) observed in the NOESY spectrum agreed with the stereochemistry of the compound. Thoreliolide B therefore had structure 2. The absolute configurations of C-24, C-26, and compounds 1 and 2 have not been resolved. Previously we reported the cytotoxicity of thorelione A and guttiferone I isolated from the bark of C. thorelii4 towards three human cancer cell lines, MCF-7 (breast adenocarcinoma), HeLa (human cervical cancer), and NCI-H460 (human lung cancer) using the Sulforhodamine B (SRB) assay, performed according to the method of Skehan et al.11 The same test was applied to thoreliolides A (1) and B (2) and the results (Table 2) showed that compound 1 was inactive whilst compound 2 showed moderate activity against the three cell lines with IC50 values of 9.7 ± 0.3, 16.6 ± 0.9 and 24.2 ± 3.6 lg/mL, respectively. We previously suggested possible biosynthetic pathways for thoreliones A and B, and oxy-thorelione A, from 7-epi-clusianone (4) which was isolated from the bark of the same plant.4 Biogenetically, thoreliolide A (1) could also be derived from 7-epi-clusianone (4). Initially, hydroxylation of 4 produces 5 which undergoes ring opening of the b-diketones of the phloroglucinol subunit through a retro-Claisen reaction12 followed by dehydration and cyclisation to give 6. An internal nucleophilic attack to C-6 of the p bond in the C-6 isoprenyl side chain as a neighbouring group could proceed with retention of configuration,13 followed by subsequent nucleophilic attack on a DMAPP unit to form a carbocation which is cyclised and deprotonated to produce the 2,2-dimethyl-3isopropenylcyclopentyl ring (Scheme 1). For thoreliolide B (2), the isoprenyl group at C-8 of 6 possibly reacts with a DMAPP molecule followed by ring closure at C-2 to furnish the 3,3-dimethyl-4-isoprenylcyclohexanone ring (Scheme 2). In conclusion, the isolation of two naturally occurring polyisoprenylated benzoylphloroglucinol derivatives, thoreliolides A (1) and B (2), which possess a 4a,5,6,7-tetrahydro-4-hydroxy-2H-chromen-2-one or 4a,5,6,7-tetrahydrogen-3H-chromene-2,4-dione ring system is reported. We have proposed plausible biogenetic
5
pathways for the two new compounds from 7-epi-clusianone which was previously obtained from the bark of the same plant. Thoreliolide B showed moderate cytotoxic activity against three tumour cell lines, MCF-7, HeLa and NCI-H460. Acknowledgement This research is funded by Viet Nam National University–Ho Chi Minh City under the Grant Number C2014-18-11. Supplementary data Supplementary data (experimental procedures, 1D and 2D NMR spectra of compounds 1 and 2) associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.tetlet. 2016.05.021. References and notes 1. Pham, H. H. In The Flora of Viet Nam; Youth Publisher: Ho Chi Minh City, 1991; Vol. 1,. Band 1. 2. Nguyen, T. L. T.; Nguyen, M. D.; Nguyen, D. L. H. Vietnam J. Chem. 2010, 48, 360– 364. 3. Nguyen, T. L. T.; Nguyen, M. D.; Nguyen, D. L. H. Nat. Prod. Res. 2013, 27, 563– 567. 4. Nguyen, T. L. T.; Nguyen, T. H.; Barbicˇ, M.; Brunner, G.; Heilmann, J.; Pham, D. H.; Nguyen, M. D.; Nguyen, D. L. H. Tetrahedron Lett. 2012, 53, 4487–4493. 5. Nilar; Nguyen, D. L. H.; Venkatraman, G.; Sim, K. Y.; Harrison, L. J. Phytochemistry 2005, 66, 1718–1723. 6. Sordat-Diserens, I.; Marston, A.; Hamburger, M.; Hostettmann, K.; Rogers, C. Helv. Chim. Acta 1989, 72, 1001–1007. 7. Lopes, J. L. C.; Lopes, J. N. C.; Gilbert, B.; Bonini, E. S. Phytochemistry 1977, 16, 1101. 8. Gustafson, K. R.; Blunt, J. W.; Munro, M. H. G.; Fuller, R. W.; McKee, T. C.; Cardellina, J. H., II; McMahon, J. B.; Cragg, G. M.; Boyd, M. R. Tetrahedron 1992, 48, 10093–10102. 9. Piccinelli, A. L.; Cuesta-Rubio, O.; Chica, M. B.; Mahmood, N.; Pagano, B.; Pavone, M.; Barone, V.; Rastrelli, L. Tetrahedron 2005, 61, 8206–8211. 10. Rubio, C. O.; Cuellar, C. A.; Rojas, N.; Castro, V. H.; Rastrelli, L.; Aquino, R. J. Nat. Prod. 1999, 62, 1013–1015. 11. Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; McMahon, J.; Vistica, D.; Warren, J. T.; Bokesch, H.; Kenney, S.; Boyd, M. R. J. Natl. Cancer Inst. 1990, 82, 1107–1112. 12. Grogan, G.; Roberts, G. A.; Bougioukou, D.; Turner, N. J.; Flitsch, S. L. J. Biol. Chem. 2001, 276, 12565–12572. 13. Smith, M. B. In March’s Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 7th ed.; John Wiley & Sons: New Jersey, 2013; p 396.
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Thoreliolides A and B, two polyisoprenylated benzoylphloroglucinol derivatives with a new carbon skeleton from the fruits of Calophyllum thorelii Le-Thu T. Nguyen a, Ngan T. D. D. T. Lai a, Ly T. T. Nguyen a, Hieu T. Nguyen a, Duc M. Nguyen b, Lien-Hoa D. Nguyen a,⇑ a b
Natural Product and Medicinal Chemistry Lab, Faculty of Chemistry, VNUHCM–University of Science, 227 Nguyen Van Cu, District 5, Ho Chi Minh City, Viet Nam Faculty of Pharmacy, Ton Duc Thang University, 19 Nguyen Huu Tho, District 7, Ho Chi Minh City, Viet Nam
a r t i c l e
i n f o
Article history: Received 7 March 2016 Revised 25 April 2016 Accepted 4 May 2016 Available online xxxx Keywords: Calophyllum thorelii Guttiferae Polyisoprenylated acylphloroglucinols Thoreliolides A and B Cytotoxicity
a b s t r a c t Thoreliolides A and B, two new polyisoprenylated benzoylphloroglucinol derivatives with a novel carbon skeleton, together with four known compounds, were isolated from the fruits of Calophyllum thorelii and their structures elucidated using spectroscopic methods. Thoreliolide B showed moderate activity against three cancer cell lines, MCF-7, HeLa and NCI-H460, with IC50 values of 9.7, 16.6 and 24.2 lg/mL, respectively, using the sulforhodamine B assay. Ó 2016 Elsevier Ltd. All rights reserved.
Calophyllum thorelii Pierre (Guttiferae) is a large red hardwood tree, up to 30 in height.1 We have previously reported the isolation of a new xanthone, thorexanthone, from the fruits of the species collected in central Viet Nam.2 In addition, another new xanthone, calothorexanthone,3 two novel polyisoprenylated benzoylphloroglucinols, thoreliones A and B, and a new polyisoprenylated tetracyclic xanthone, oxy-thorelione A, were obtained from the bark.4 Continuing our chemosystematic investigation on this species, we herein describe the isolation and structure elucidation of two unusual polyprenylated benzoylphloroglucinol derivatives, thoreliolides A and B, together with four known compounds, from the fruits of the same plant. The cytotoxicity of the two new compounds using the sulforhodamine B (SRB) assay is also reported. Column chromatography of the petroleum ether extract of the fruits of C. thorelii led to the isolation of thoreliolides A (1) and B (2) (Fig. 1), along with four known compounds, thorelione A (3),4 guttiferone I,5 12b-hydroxy-des-D-garcigerrin A6 and 7 osajaxanthone. Thoreliolide A (1) was obtained as a yellow gum, [a]25 D +82.0 (c 0.45, EtOH), giving a positive reaction with ethanolic ferric chloride. The UV spectrum showed four maxima at 233, 268, 317 and ⇑ Corresponding author. Tel.: +84 8 38355270; fax: +84 8 38350096. E-mail addresses: (L.-H.D. Nguyen).
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365 nm whilst the IR spectrum exhibited absorption bands at 3415 (OAH), 1732 (C@O), 1626 (conjugated C@O), 1582 (aromatic C@C) and 1282 cm 1 (CAO). HRESIMS revealed the molecular formula to be C38H50O6 (m/z 625.3533 [M+Na]+), i.e., the molecule had fourteen degrees of unsaturation. The 1H and 13C NMR, and HMBC spectra of 1 (Table 1) contained resonances for a 1,2,4-trisubstituted benzene ring [dH 7.23 (1H, d, J = 1.9 Hz, H-12), 7.15 (1H, dd, J = 8.4, 1.9 Hz, H-16), 6.87 (1H, d, J = 8.4 Hz, H-15)], a conjugated carbonyl carbon (dC 194.8, C-10), two 3-methylbut-2-enyl groups [dH 5.27 (1H, m, H-35), 5.11 (1H, m, H-18), 2.94 and 2.90 (each 1H, m, H2-17), 2.28 and 1.82 (each 1H, m, H2-34), 1.76 (3H, s, H337), 1.73 (3H, s, H3-20), 1.72 (3H, s, H3-21), 1.67 (3H, s, H3-38)], a 2,2-dimethyl-3-isopropenylcyclopentyl ring [dH 4.79 and 4.60 (each 1H, s, H2-32), 2.38 (1H, m, H-26), 1.98 (1H, m, H-24), 2.02 (1H, m, H-27), 1.82 (1H, m, Ha-28), 1.71 (2H, m, H-27 and Hb-28), 1.63 (3H, s, H3-33), 1.08 (3H, s, H3-30), 0.73 (3H, s, H3-29); dC 145.9 (C-31), 109.8 (C-32), 51.1 (C-26), 43.1 (C-25), 38.9 (C-24), 38.0 (C-28), 30.3 (C-30), 29.5 (C-27), 23.2 (C-33), 16.5 (C-29)], two quaternary carbons (dC 46.6, C-8 and 39.0, C-5), a tertiary carbon (dC 42.0, C-6), a secondary carbon (dC 30.5, C-7), two tertiary methyls [dH 1.17 (3H, s, H3-22), 1.03 (3H, s, H3-23); dC 26.0 (C-22), 21.1 (C-23)], and an enolised-1,3-diketone moiety [dC 173.3 (C-1), 162.1 (C-3), 109.8 (C-2)]. The spectrum data of 1 were similar to those of thorelione A (3) previously isolated from
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L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx 21
20 28
21
21
20 18
OH 18
OH HO
13
HO
17 23
O
14
O 3
14
6 1
16
O
OH
34
25
O
38
O
12
14
27 24
5
O
O 9
3
7
10
27
25
29 33 34
33
17
7
32 31
30 37
HO
24
1
16
30 29
1
OH
22
5
1
2
27
9 8
16
24
4
3 10
22
8 10
22
23
O
9 5
23
17
O
13
4
20
28 35
O
OH
3
34
29 32
30
36
35
38
32 37 37
38
Figure 1. Structures of compounds 1–3.
Table 1 H (500 MHz) and
1
13
C (125 MHz) NMR data, and HMBC correlations for thoreliolides A (1) and B (2) in acetone-d6 (J in Hz in parentheses)
Position
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38
1
2
dH
HMBC correlations
dC
dH
HMBC correlations
dC
— — — — — 1.62 m Ha: 2.10 m Hb: 1.62 m — — — — 7.23 d (1.9) — — 6.87 d (8.4) 7.15 dd (8.4, 1.9) Ha: 2.94 m Hb: 2.90 m 5.11 m — 1.73 s 1.72 s 1.17 s 1.03 s 1.98 m
— — — — — C-7, C-23 C-5, C-24 C-6 — — — — C-10, C-13, C-14, C-16 — — C-11, C-13, C-14 C-10, C-12, C-14 C-4, C-5, C-9, C-18, C-19 C-4, C-5, C-9, C-18, C-19 C-17, C-20, C-21 — C-18, C-19, C-21 C-18, C-19, C-20 C-4, C-5, C-6, C-23 C-4, C-5, C-6, C-22 —
173.3 109.8 162.1 130.8 39.0 42.0 30.5
— C-25, C-31 C-24, C-25 C-24, C-26, C-28 C-6, C-24, C-25, C-27 C-24, C-26 C-24, C-25, C-25, C-30
43.1 51.1 29.5
— — — — — C-7 C-1, C-5, C-8, C-29 C-5 — — — — C-10, C-13, C-14, C-16 — — C-11, C-13, C-14, C-16 C-10, C-12, C-14 C-4, C-5, C-9, C-18, C-19 C-4, C-5, C-9, C-18, C-19 C-20, C-21 — C-18, C-19, C-21 C-18, C-19, C-20 C-4, C-5, C-6, C-23 C-4, C-5, C-6, C-22 C-6, C-25, C-26 C-6 C-27, C-28 — C-25, C-26, C-28
205.5 75.1 166.3 130.8 39.7 41.5 31.8
— 2.38 m Ha: 2.02 m Hb: 1.71 m Ha: 1.82 m Hb: 1.71 m 0.73 s
— — — — — 1.55 m Ha: 1.91 br d (13.5) Hb: 1.64 m — — — — 7.21 d (2.4) — — 6.76 d (8.2) 6.58 dd (8.2, 2.4) Ha: 3.08 m Hb: 2.92 m 5.17 m — 1.71 s 1.68 s 1.21 s 1.00 s 2.28 m 1.85 m 5.17 m — 1.63 s
38.0
1.72 s
C-25, C-26, C-27
26.1
16.5
C-24, C-25, C-25, C-29 — C-26, C-33 C-26, C-33 C-26, C-31, C-32 C-1, C-7, C-35, C-36 C-7, C-8 C-37, C-38 — C-35, C-36, C-38 C-35, C-36, C-37
30.3 145.9 109.8
C-1, C-8, C-9, C-30, C-31 C-1, C-8, C-30, C-34 — — C-2, C-30, C-31, C-33
46.2
1.08 s — Ha: 4.79 s Hb: 4.60 s 1.63 s Ha: 2.28 m Hb: 1.82 m 5.27 m — 1.76 s 1.67 s
Ha: 2.55 dd (13.6, 4.8) Hb: 1.59 t (13.6) 1.85 m — 1.17 s 1.45 s Ha: 2.28 m Hb: 1.73 m 5.17 m — 1.65 s 1.71 s
C-2, C-30, C-31, C-32 C-35, C-36 C-30 C-37, C-38 — C-35, C-36, C-38 C-35, C-36, C-37
46.6 143.9 194.8 128.0 117.6 145.3 151.5 115.3 124.3 26.0 124.9 131.3 18.0 26.0 26.0 21.1 38.9
23.2 29.4 124.8 133.4 18.1 26.2
the bark of the same plant.4 The notable differences were that the resonances for the isolated C-9 carbonyl carbon and the C-8 quaternary carbon in the phloroglucinol subunit of thorelione A (3) had been replaced by a carbon–carbon double bond carrying an oxygen atom [dC 143.9 (C-9) and 130.8 (C-4)] in compound 1.
51.0 142.7 190.1 129.1 116.5 146.0 150.9 115.2 121.8 26.5 124.4 131.8 18.1 25.9 26.2 21.0 29.2 123.5 133.8 18.0
45.3 52.0 17.0 23.7 28.7 124.3 133.4 18.0 25.9
In the HMBC spectrum of 1 (Fig. 2), the methylene protons of the first isoprenyl group (dH 2.94 and 2.90, H2-17) showed correlations to the fully-substituted mono-oxygenated olefinic carbons (dC 143.9 and 130.8) and a quaternary carbon (dC 39.0). As a result, the first was identified as C-9 which bore the oxygen atom, the
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21 21
20
20 18
OH 18
OH
23
H O
HO 14
O
12
22 6
H
3
16
24
34
HO
O
4
9
22
5
24
2
29
1
31
30 37
38
7
O
O 32
27
25
1
10
H
1
10
H
12
17 23
O 8
3
16
14
4 9 8
H
H O
HO
17
27
25
29 30 33
28 34
33
35
32 37
38
Figure 2. HMBC correlations for compounds 1 and 2.
Additional NOE interactions between Hb-7 (dH 1.62) and Ha-34 (dH 2.28 m) and H3-22 (dH 1.17) revealed that the isoprenyl attached to C-8 and the Cax-22 methyl group had the same orientation, i.e., the C-8 isoprenyl was equatorial. The structure of thoreliolide A was consequently established as 1. The second new compound, thoreliolide B (2), was obtained as a yellowish gum, [a]25 51.2 (c 0.75, EtOH), UV: (kmax) 234, 283 and D 314 nm, IR: (mmax) 3425 (OAH), 1750 (non-conjugated C@O), 1706 (conjugated C@O), 1604 (aromatic C@C), and 1286 cm 1 (CAO). HRESIMS showed a pseudo-molecular ion peak [M+H]+ at m/z 603.3647, suggesting that this compound and thoreliolide A (1) had the same molecular formula of C38H50O6. The 1H and 13C NMR spectra of 2 (Table 1) had signals due to a 3,4-dihydroxybenzoyl system, two 3-methylbut-2-enyl groups, two tertiary methyls [dH 1.21 (3H, s, H3-22), 1.00 (3H, s, H3-23); dC 26.2 (C-22), 21.0 (C-23)], and a C10 unit comprising a trisubstituted carbon–carbon double bond [dH 5.17 (1H, m, H-35); dC 133.4 (C-36), 124.3 (C-35)], two methylenes [dH 2.55 (1H, dd, J = 13.6, 4.8 Hz, Ha-29), 1.59 (1H, t, J = 13.6 Hz, Hb-29); dH 2.28 and 1.73 (each 1H, m, H2-34); dC 46.2 (C-29), 28.7 (C-34)], a methine [dH 1.85 (1H, m, H-30); dC 45.3 (C-30)], two allylic methyls [dH 1.71 (3H, s, H3-38), 1.65 (3H, s, H3-37); dC 25.9 (C-38), 18.0 (C-37)], two tertiary methyls [dH 1.45 (3H, s, H3-33) and 1.17 (3H, s, H3-32); dC 23.7 (C-33), and 17.0 (C-32)], and a quaternary carbon [dC 52.0 (C-31)]. In addition, the spectra showed the presence of a 4a,5,6,7-tetrahydrogen-3H-chromene-2,4-dione ring system consisting of a carbon–carbon double bond bearing an oxygen atom [dC 142.7 (C-9), 130.8 (C-4)], a 1,3-diketone moiety [dC 205.5 (C-1), 166.3 (C-3), 75.1 (C-2)], two quaternary carbons [dC 51.0 (C-8), 39.7 (C-5)], a tertiary carbon [dH 1.55 (1H, m, H-6); dC 41.5 (C-6)] and a secondary carbon [dH 1.91 (1H, br d, J = 13.5 Hz, Ha-7), 1.64 (1H, m, Hb-7); dC 31.8 (C-7)]. In the HMBC spectrum of 2 (Fig. 2), correlations of the methylene protons of the first isoprenyl side chain (H2-17), the second
second was C-4 which carried the isoprenyl group, and the third was C-5. In addition, the two tertiary methyls gave cross-peaks to each other, to C-4, C-5 and a tertiary carbon (dC 42.0). Accordingly, C-5 carried the two methyls and the tertiary carbon was located at C-6. A methylene proton of the 2,2-dimethyl-3-isopropenylcyclopentyl ring (Ha-28) was correlated to C-6 and this C10 side chain was therefore attached to the C-6 carbon. The methylene protons at dH 2.10 and 1.62 showed correlations to C5, and they were thus assigned as H2-7 (dC 30.5). The methylene protons of the remaining isoprenyl group correlated to C-7, a quaternary carbon (dC 46.6) and the enolised carbon (dC 173.3). The former was hence C-8 which carried the isoprenyl group and the latter was C-1 which bore a hydroxyl group. The remaining olefinic carbon (dC 109.8) was C-2 and the conjugated carbonyl carbon (dC 162.1) had to be C-3 in order to form an enol 1,3-diketone moiety. To account for fourteen degrees of unsaturation in the molecule, the oxygen atom attached to C-9 had to be bonded to C-3 to form a conjugated lactone ring. The compound thus possessed a 4a,5,6,7-tetrahydro-4-hydroxy-2H-chromen-2-one ring system. Correlations observed in the 1H–1H COSY spectrum (Fig. 3) were consistent with the proposed structure. The relative stereochemistry of 1 was established based on 13C chemical shifts of the two tertiary methyls attached to C-5 and NOE interactions. In guttiferone B,8 the C-5 axial methyl (C-22) was shielded by the equatorial geranyl side chain attached to C-6 [dC 23.8 (C-23) and 16.5 (C-22)] due to a c-gauche interaction. Meanwhile in the cases of 7-epi-clusianone (4)9 and thorelione A (3),4 the side chain bonded to C-6 was axial, and the axial methyl located at C-5 was therefore less shielded (dC 27.0 and 27.5 ppm, respectively). Therefore, in thoreliolide A (1), the 13C shifts of the two tertiary methyls attached to C-5 [dC 21.0 (Ceq-23) and 26.0 (Cax-22)] showed that the side chain at C-6 was axial. This was confirmed by an NOE interaction (Fig. 3) observed between Ha-7 (dH 2.10) and H-24 (dH 1.98).
21 21
20
20
OH OH HO
13
18
O
14
O
17
14
4
8 1
O
34
OH
3
7
H
6
H
27
25
NOESY
32 30
38
33
31
30 37
COSY
24
O 29 H H
O
2
27 25
22
7
1
16
24
23
5
10
22
35
1
17 4 9 8
5
10
O
23
O
9
3
16
HO
18
13
34
28 35
33 32 37
38
Figure 3. COSY and key NOE correlations for compounds 1 and 2.
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L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx
the methyls and the remaining olefinic carbon was C-35. One proton of the remaining methylene group (dH 2.28; dC 28.7) correlated to C-35 and C-36, and the methylene was therefore C-34. The H2-34 protons showed cross-peaks to C-35, C-36 and a tertiary carbon (dC 45.3) whilst H2-29 correlated to C-34, the tertiary carbon and a quaternary carbon (dC 52.0). As a result, the tertiary carbon was C-30 and the quaternary carbon was C-31. The remaining two tertiary methyls correlated to each other [H3-32 (dH 1.17) to C33 (dC 23.7) and H3-33 (dH 1.45) to C-32 (dC 17.0)], to C-30, C-31 and a quaternary carbon (dC 75.1). They were thus bonded to C31 and the quaternary carbon was C-2. The structure of the subunit was confirmed by analysis of the 1H–1H COSY correlations (Fig. 3). The remaining carbon (dC 166.3) which showed no correlation had to be the lactone carbonyl carbon (C-3) in the 4a,5,6,7-tetrahydrogen-3H-chromene-2,4-dione ring system. Similar to the case of thoreliolide A (1), the 13C shifts of Cax-22 and Ceq-23 methyls (dC 26.2 and 21.0, respectively) indicated that the isoprenyl group at C-6 was axial and H-6 was hence equatorial. The geometry of the bicyclic ring system required substituents on C-2 and C-8 to be equatorial. The coupling constants of H2-29 [dH 2.55 (dd, J = 13.6 and 4.8 Hz, Ha-29), 1.59 (t, J = 13.6 Hz, Hb-29)] revealed H-30 to be axial, and the C-30 isoprenyl group was thus equatorial. This was confirmed by comparison of the 13C shifts of the two tertiary methyls (Cax-32 and Ceq-33) attached to C-5 with those of
Table 2 Cytotoxicity of 1 and 2 using the SRB assay11,a
MCF-7 HeLa NCI-H460 a b
1
2
Camptothecin
72.3 ± 1.0 n.d.b n.d.b
9.7 ± 0.3 16.6 ± 0.9 24.2 ± 3.6
N/A 0.013 ± 0.001 0.006 ± 0.001
Mean of three measurements. Not determined.
isoprenyl group (H2-24), and the two tertiary methyls (H3-22 and H3-23) revealed that the first isoprenyl side chain was attached to C-4 whilst the two methyls were bonded to C-5 and the second isoprenyl group was located at C-6. The two methylene protons at dH 1.91 and 1.64 correlated to C-5 and were therefore bonded to C7. The H2-7 protons gave cross-peaks to a quaternary carbon (dC 51.0), an isolated carbonyl carbon (dC 205.5) and a methylene (dC 46.2), which were thus assigned as C-8, C-1, and C-29, respectively. Correlations of the methylene protons of the C10 unit [dH 2.55 and 1.59 (H2-29)] with C-1, C-8 and C-9 established a bond between C-8 and C-9. Meanwhile, the two allylic methyls of the C10 unit (H3-37 and H3-38) correlated to each other and to two carbons of a trisubstituted carbon–carbon double bond (dC 133.4, s and 124.3, d). The fully substituted olefinic carbon (C-36) hence carried Enz B H OH OH O
O
5
O
HO
O
[O] Hydroxylation O
10
OH
O
5
3
4 9
+ H2 O
5
8 10
Retro-Claisen
8
1
4
O
Enz H
9
3
OH HO
6
OH
6
1
O
OH
H2 O Cyclisation
O
O
O
4
9
H
5
3 8
O
1
O
OPP
5
3
10
HO
O H
O
5 8
10
1
7
6
1
O
HO
4
9
3
6
8
7
1
O
4
9
6
8
HO
O
5
3
10
O
4
9
6
1
10
O
17
17
17
17
7
HO
PPO
C H2
Scheme 1. Plausible biosynthetic pathway for 1.
O
O
Tautomerism O
4
9
5
3
4
9
6
8 2
1
HO
7
1
O
1. DMAPP
5
6
8 2
O
O 3
7
O
O
2
O
O
6 OPP
4
9
3
5 8
8 1
O
O
O
4
9
5
2. - H
C5
3
6 7
C5
10 1
O
O
7
C5
2
DMAPP Scheme 2. Plausible biosynthetic pathway for 2.
Please cite this article in press as: Nguyen, L.-T. T.; et al. Tetrahedron Lett. (2016), http://dx.doi.org/10.1016/j.tetlet.2016.05.021
L.-T. T. Nguyen et al. / Tetrahedron Letters xxx (2016) xxx–xxx
propolone A.10 In propolone A, the equatorial isoprenyl side chain at C-30 shielded the axial methyl [dC 15.7 (Cax-32) and 23.7 (Ceq-33)] due to a c-gauche interaction. In thoreliolide B (2), the resonances of the C-31 methyls [dC 17.0 (Cax-32) and 23.7 (Ceq-33)] revealed that the isoprenyl group attached to C-30 was equatorial. NOE interactions (Fig. 3) observed in the NOESY spectrum agreed with the stereochemistry of the compound. Thoreliolide B therefore had structure 2. The absolute configurations of C-24, C-26, and compounds 1 and 2 have not been resolved. Previously we reported the cytotoxicity of thorelione A and guttiferone I isolated from the bark of C. thorelii4 towards three human cancer cell lines, MCF-7 (breast adenocarcinoma), HeLa (human cervical cancer), and NCI-H460 (human lung cancer) using the Sulforhodamine B (SRB) assay, performed according to the method of Skehan et al.11 The same test was applied to thoreliolides A (1) and B (2) and the results (Table 2) showed that compound 1 was inactive whilst compound 2 showed moderate activity against the three cell lines with IC50 values of 9.7 ± 0.3, 16.6 ± 0.9 and 24.2 ± 3.6 lg/mL, respectively. We previously suggested possible biosynthetic pathways for thoreliones A and B, and oxy-thorelione A, from 7-epi-clusianone (4) which was isolated from the bark of the same plant.4 Biogenetically, thoreliolide A (1) could also be derived from 7-epi-clusianone (4). Initially, hydroxylation of 4 produces 5 which undergoes ring opening of the b-diketones of the phloroglucinol subunit through a retro-Claisen reaction12 followed by dehydration and cyclisation to give 6. An internal nucleophilic attack to C-6 of the p bond in the C-6 isoprenyl side chain as a neighbouring group could proceed with retention of configuration,13 followed by subsequent nucleophilic attack on a DMAPP unit to form a carbocation which is cyclised and deprotonated to produce the 2,2-dimethyl-3isopropenylcyclopentyl ring (Scheme 1). For thoreliolide B (2), the isoprenyl group at C-8 of 6 possibly reacts with a DMAPP molecule followed by ring closure at C-2 to furnish the 3,3-dimethyl-4-isoprenylcyclohexanone ring (Scheme 2). In conclusion, the isolation of two naturally occurring polyisoprenylated benzoylphloroglucinol derivatives, thoreliolides A (1) and B (2), which possess a 4a,5,6,7-tetrahydro-4-hydroxy-2H-chromen-2-one or 4a,5,6,7-tetrahydrogen-3H-chromene-2,4-dione ring system is reported. We have proposed plausible biogenetic
5
pathways for the two new compounds from 7-epi-clusianone which was previously obtained from the bark of the same plant. Thoreliolide B showed moderate cytotoxic activity against three tumour cell lines, MCF-7, HeLa and NCI-H460. Acknowledgement This research is funded by Viet Nam National University–Ho Chi Minh City under the Grant Number C2014-18-11. Supplementary data Supplementary data (experimental procedures, 1D and 2D NMR spectra of compounds 1 and 2) associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.tetlet. 2016.05.021. References and notes 1. Pham, H. H. In The Flora of Viet Nam; Youth Publisher: Ho Chi Minh City, 1991; Vol. 1,. Band 1. 2. Nguyen, T. L. T.; Nguyen, M. D.; Nguyen, D. L. H. Vietnam J. Chem. 2010, 48, 360– 364. 3. Nguyen, T. L. T.; Nguyen, M. D.; Nguyen, D. L. H. Nat. Prod. Res. 2013, 27, 563– 567. 4. Nguyen, T. L. T.; Nguyen, T. H.; Barbicˇ, M.; Brunner, G.; Heilmann, J.; Pham, D. H.; Nguyen, M. D.; Nguyen, D. L. H. Tetrahedron Lett. 2012, 53, 4487–4493. 5. Nilar; Nguyen, D. L. H.; Venkatraman, G.; Sim, K. Y.; Harrison, L. J. Phytochemistry 2005, 66, 1718–1723. 6. Sordat-Diserens, I.; Marston, A.; Hamburger, M.; Hostettmann, K.; Rogers, C. Helv. Chim. Acta 1989, 72, 1001–1007. 7. Lopes, J. L. C.; Lopes, J. N. C.; Gilbert, B.; Bonini, E. S. Phytochemistry 1977, 16, 1101. 8. Gustafson, K. R.; Blunt, J. W.; Munro, M. H. G.; Fuller, R. W.; McKee, T. C.; Cardellina, J. H., II; McMahon, J. B.; Cragg, G. M.; Boyd, M. R. Tetrahedron 1992, 48, 10093–10102. 9. Piccinelli, A. L.; Cuesta-Rubio, O.; Chica, M. B.; Mahmood, N.; Pagano, B.; Pavone, M.; Barone, V.; Rastrelli, L. Tetrahedron 2005, 61, 8206–8211. 10. Rubio, C. O.; Cuellar, C. A.; Rojas, N.; Castro, V. H.; Rastrelli, L.; Aquino, R. J. Nat. Prod. 1999, 62, 1013–1015. 11. Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; McMahon, J.; Vistica, D.; Warren, J. T.; Bokesch, H.; Kenney, S.; Boyd, M. R. J. Natl. Cancer Inst. 1990, 82, 1107–1112. 12. Grogan, G.; Roberts, G. A.; Bougioukou, D.; Turner, N. J.; Flitsch, S. L. J. Biol. Chem. 2001, 276, 12565–12572. 13. Smith, M. B. In March’s Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 7th ed.; John Wiley & Sons: New Jersey, 2013; p 396.
Please cite this article in press as: Nguyen, L.-T. T.; et al. Tetrahedron Lett. (2016), http://dx.doi.org/10.1016/j.tetlet.2016.05.021