Letter Cite This: Org. Lett. 2018, 20, 4040−4043
pubs.acs.org/OrgLett
Mangelonoids A and B, Two Pairs of Macrocyclic Diterpenoid Enantiomers from Croton mangelong Wei-Yi Zhang,† Jin-Xin Zhao,‡ Li Sheng,‡ Yao-Yue Fan,‡ Jing-Ya Li,‡ Kun Gao,*,† and Jian-Min Yue*,†,‡ †
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State Key Laboratory of Applied Organic Chemistry, College of Chemistry and Chemical Engineering, Lanzhou University, Lanzhou 730000, People’s Republic of China ‡ State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, People’s Republic of China S Supporting Information *
ABSTRACT: (+)- and (−)-Mangelonoids A and B, two pairs of novel macrocyclic diterpenoid enantiomers featuring an unprecedented bicyclo[9.3.1]pentadecane core and a rare bridgehead double bond, were isolated from Croton mangelong. Their structures were unambiguously established using spectroscopic data, X-ray crystallography, and electronic circular dichroism analysis. A plausible biosynthesis pathway for these compounds was proposed. (−)-Mangelonoid A exhibited NF-κB inhibition with an IC50 value of 7.27 ± 1.30 μM.
T
he genus Croton, a large member of Euphorbiaceae, comprises around 1300 species widespread in tropical and subtropical regions of both hemispheres.1 There are 21 species native to the southern provinces of China,2 among which some have been used in traditional Chinese medicine to treat dysmenorrhea, gastric ulcers, malaria, gastric cancer, and dysentery.3 Previous chemical investigations on the Croton plants contributed to an array of diterpenoids including tiglianes, cembranes, clerodanes, and kauranes, as well as other types of constituents.3a,4 These metabolites exhibited a wide variety of biological properties such as acetylcholinesterase inhibitory, antiplasmodial, cytotoxic, antiviral, and anti-inflammatory activities.3a,4 Croton mangelong is an arbor and is mainly distributed in the southern area of Yunnan Province.2 In our continuing efforts to explore structurally interesting and biologically significant secondary metabolites from the plants of Croton genus,3c,5 two pairs of macrocyclic diterpenoid enantiomers, (+)- and (−)-mangelonoids A and B, featuring an unprecedented bicyclo[9.3.1]pentadecane core and a rare naturally occurring bridgehead olefin (anti-Bredt system),6 were isolated from the EtOH extract of the twigs and leaves of C. mangelong and further enhanced the novelty and complexity of the macrocyclic diterpenoids. The structures and absolute configurations of these compounds were elucidated via MS, NMR, X-ray crystallographic data, and an electronic circular dichroism (ECD) method. All isolates were evaluated in a NFκB pathway luciferase assay for inhibitory effects, and (−)-1 exhibited good inhibition with an IC50 value of 7.27 ± 1.30 μM. Mangelonoid A (1) possessed a molecular formula of C20H32O2 with five double bond equivalents (DBEs) as established by the HRESIMS ion peak at m/z 631.4709 [2M + Na]+ (calcd 631.4702) and its 13C NMR data (Table 1). The 1 H NMR data (Table 1) revealed the presence of three olefinic protons (δH 4.55, 4.80, and 5.31), two singlet methyls (δH 1.54 © 2018 American Chemical Society
and 1.64), and two doublet methyls (δH 0.83 and 0.85). Analysis of the 13C NMR data with the aid of the DEPT and HSQC spectra resolved all 20 carbon resonances in the molecule, including three double bonds (δC 124.5, 125.0, 127.2, 133.8, 134.5, and 136.7), four methyls (δC 15.7, 15.7, 16.0, and 16.8), six methylenes, three methines (one oxygenated at δC 71.2), and one oxygenated quaternary carbon (δC 77.0). These functional groups accounted for three out of five DBEs, and the remaining DBEs required the existence of a bicyclic system in the molecule. Construction of the planar structure for 1 was accomplished by interpretation of 2D NMR data. Analysis of the 1H−1H COSY spectrum revealed five spin-coupling segments as H2-5 via H2-6 to H-7; H2-9 via H2-10 to H-11; H2-13 to H2-14; H316 via H-15 to H3-17; and H-3 via H-2 to H-20, as shown in Figure 1A (bold lines). These subunits were readily linked together with quaternary carbons by the HMBC spectrum (Figure 1A). The HMBC correlations of H-2/C-1 and C-20; Received: May 21, 2018 Published: June 11, 2018 4040
DOI: 10.1021/acs.orglett.8b01608 Org. Lett. 2018, 20, 4040−4043
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Organic Letters Table 1. 1H (400 MHz) and 13C (125 MHz) NMR Data of 1 and 2 in CDCl3 1 δH (mult, J, Hz) 1 2 3 4 5a 5b 6a 6b 7 8 9a 9b 10a 10b 11
2.89, d (10.5) 4.55, d (10.5) 1.91 dd (12.6, 3.8) 2.10 ddd (12.5, 3.9, 3,7) 1.99, m 2.22, m 4.80, dd (10.3, 5.0) 1.83, 2.19, 1.94, 2.28, 5.31,
m m m m ddd (11.3, 5.7, 1.3)
12 13a 13b 14a
2.03, m 2.75, dd (15.5, 14.1) 1.62, dd (14.1, 5.0)
14b
1.81, m
15 16 17 18 19 20
1.48, 0.83, 0.85, 1.64, 1.54, 4.15,
m d (5.3) d (5.5) s s d (2.1)
2 δC 77.0 46.2 124.5 133.8 39.3 25.6 125.0 134.5 38.9 25.7 127.2 136.7 27.2 32.4
35.2 15.7 16.0 15.7 16.8 71.2
δH (mult, J, Hz) 2.90, ddd (10.2, 3.2, 1.5) 4.46, d (10.3) 1.95, 2.20, 1.99, 2.38, 4.75,
m m m m d (10.5)
2.00, m 2.18, m 2.03, m 2.29, m 4.88, ddd (11.9, 2.9, 1.5) 2.31, m 2.40, m 1.48, ddd (14.0, 13.7, 5.5) 1.89, ddd (14.0, 5.5, 1.5) 1.40, m 0.81, d (6.8) 0.84, d (6.9) 1.63, s 1.54, s 3.75, d (3.1)
δC 77.1 46.3 122.1 133.0 36.3 24.5 128.4 132.7 39.6
Figure 2. Key 1H−1H COSY, HMBC, and NOESY correlations of 2.
24.4 127.3 135.7 19.4 34.3
35.5 16.0 15.7 18.1 15.5 81.6
Figure 3. ORTEP drawings of compounds (−)-1 (left) and 2 (right).
Figure 1. Key 1H−1H COSY, HMBC, and NOESY correlations of 1.
H-20/C-12 and C-13; and H2-14/C-1, C-2, and C-13 delineated a typical cyclohexane fragment (A-ring). The cyclododecane moiety (B-ring) was delineated by the key HMBC cross-peaks of H3-18/C-3, C-4, and C-5; H3-19/C-7, C-8, and C-9; H2-13 and H-20/C-11 and C-12. Furthermore, the HMBC correlations from H-15, H3-16, and H3-17 to C-1 indicated that an isopropyl was attached to C-1. Thus, the gross structure of 1 was constructed as a macrocyclic diterpenoid with an unprecedented bicyclo[9.3.1]pentadecane scaffold. The partial relative configuration of 1 was determined by the detailed interpretation of the NOESY spectrum (Figure 1B).
Figure 4. Experimental and calculated ECD spectra of (+)-1 and (−)-1 (up) and (+)-2 and (−)-2 (down) in MeOH.
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DOI: 10.1021/acs.orglett.8b01608 Org. Lett. 2018, 20, 4040−4043
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Organic Letters
Magelonoid B (2) was obtained as colorless crystals and possessed a molecular formula of C20H32O2 based on the HRESIMS ion peak at m/z 631.4711 [2 M + Na]+ (calcd 631.4702) and the 13C NMR data (Table 1). The 1D and 2D NMR spectra of 2 highly resembled those of 1, and the gross structure of 2 was achieved by the 1H−1H COSY and HMBC analyses in the same manner as described for 1 (Figure 2A). By carefully comparing the 13C NMR data of 2 with those of 1, we found that the major differences occurred in two carbon resonances (C-13 and C-20) that directly connected to the Δ11 double bond. The upfield-shifted C-13 (ΔδC − 7.8) and downfield-shifted C-20 (ΔδC 10.4) in 2 when compared with those in 1 indicated that they could be geometrical isomers,10 and the Δ11 double bond in 2 may take E-geometry, which was further confirmed by the key NOESY cross-peak of H-20/H-11 (Figure 2B). Additionally, the NOESY correlations of H-2/H318 and H-7/H-9a indicated that the Δ3 and Δ7 double bonds in the structure of 2 also took E-geometry. The quality crystals of 2 were obtained and subjected to the X-ray diffraction analysis by using Cu Kα radiation (Figure 3), which confirmed the proposed planar structure and relative configuration of 2. Similar to 1, compound 2 was also a pair of enantiomers, which was verified by the P21/c space group of the crystals of 2 (Table S2). Subsequently, 2 was chirally separated via chiral HPLC to yield two optically pure enantiomers (+)-2 and (−)-2. Unfortunately, the quality crystals of optically pure enantiomers (+)-2 and (−)-2 for X-ray diffraction analysis were not acquired. The absolute configurations of (+)-2 and (−)-2 were determined using a quantum method by comparing the calculated ECD with experimental data. The calculated ECD spectra were mirror images and matched well with the experimental ECD spectra of (+)-2 and (−)-2, respectively (Figure 4). Thus, the absolute configurations of (+)-2 and (−)-2 were assigned as 1R, 2S, 20S and 1S, 2R, 20R, respectively. Mangelonoids A and B were novel macrocyclic diterpenoids and defined a new diterpene skeletal class, of which the main features are the bicyclo[9.3.1]pentadecane scaffold and a bridgehead double bond (anti-Bredt system). Natural products with a bridgehead double bond are still rare, with only some examples having been reported.6,11 Therefore, the fascinating architecture of mangelonoids A and B could attract the interests of synthetic chemists. The plausible biosynthetic precursors of mangelonoids A and B were considered to be cembranoids such as neocrotocembranal12 (Scheme 1). The protonated neocrotocembranal (i) enolized to intermediate ii and further transformed to iii, which was a mixture of the geometrical isomers at the Δ11 double bond. The C-2 and C-12 in iii were then connected to form the bicyclo[9.3.1]pentadecane core via an intramolecular Prins reaction.13 Intermediate iii was a flat ring without chiral center, so it was free to fold in either direction of the molecular planar in the Prins reaction, which resulted in a pair of enantiomers (iv and v). The 12-membered ring forced the C-2−C-3 bond to occupy the axial position in the intermediates iv and v. The deprotonation of iv and v finally gave rise to (+)-1, (+)-2, (−)-1, and (−)-2, respectively. Nuclear factor-κB (NF-κB) is an important transcription factor that plays central roles in inflammation, oxidative stress, and carcinogenesis by regulating the expression of genes critically involved in the process of inflammation, immunity, aging, cell survival and apoptosis, and metabolic diseases.14 All
Scheme 1. Hypothetical Biogenetic Pathways for (+)-1, (−)-1, (+)-2, and (−)-2
The NOESY cross-peaks of H-2/H3-18; H-7/H-9a; and H-11/ H-13a indicated that the Δ3 and Δ7 double bonds in 1 took Egeometry, and the Δ11 double bond took Z-geometry, respectively. However, the relative configurations of the three chiral carbons (C-1, C-2, and C-20) could not be determined with confidence on the basis of the NOESY data. Notably, the specific optical rotation value of 1 was close to zero, which alerted us that 1 was likely racemic. Chiral resolution was then performed via chiral HPLC to yield two optically pure enantiomers, (+)-1 and (−)-1, respectively. To elucidate the relative and absolute configurations of (+)-1 and (−)-1, we managed to acquire crystals of both optically pure enantiomers for X-ray diffraction analysis. Fortunately, quality crystals of (−)-1 were obtained, and the X-ray crystallography study was performed by using the anomalous dispersion of Cu Kα radiation successfully. The X-ray crystallography (Figure 3) not only confirmed the proposed planar structure but also determined the absolute configuration of (−)-1 with excellent absolute structure parameter [0.03(10)].7 Further to this, the theoretical ECD spectra for (+)-1 and (−)-1 were calculated by using the time-dependent density functional theory (TDDFT) at the B3LYP/dgdzvp level with PCM in MeOH,8,9 and the calculated ECD spectra matched well with the experimental ECD spectra of (+)-1 and (−)-1, respectively (Figure 4). Thus, the absolute configurations of (+)-1 and (−)-1 were assigned as 1R, 2S, 20S and 1S, 2R, 20R, respectively. 4042
DOI: 10.1021/acs.orglett.8b01608 Org. Lett. 2018, 20, 4040−4043
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isolates were evaluated in NF-κB pathway luciferase assay for the inhibitory effects.15 Under the concentration of 20 μg/mL (65.79 μM), compounds (+)-1, (−)-1, (+)-2, and (−)-2 showed inhibition rates of 33.99, 84.24, 40.79, and 48.78%, respectively. Compound (−)-1 further exhibited an IC50 value of 7.27 ± 1.30 μM in the subsequent test. These compounds were also evaluated in vitro for their inhibition against A549 and HL-60 tumor cell lines as well as inhibitory effects on protein tyrosine phosphatase 1B (PTP1B), but none of them were active.
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ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.orglett.8b01608. Experimental procedures; bioassay; X-ray crystallographic data for (−)-1 and (±)-2; 1D and 2D NMR, CD, IR, and HRESIMS of (+)-1, (−)-1, (+)-2, and (−)-2 (PDF) Accession Codes
CCDC 1833955−1833956 contain the supplementary crystallographic data for this paper. These data can be obtained free of charge via www.ccdc.cam.ac.uk/data_request/cif, or by emailing
[email protected], or by contacting The Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: +44 1223 336033.
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AUTHOR INFORMATION
Corresponding Authors
*Tel.: +86-0931-8912592. E-mail:
[email protected] (K. Gao). *Tel.: +86-21-50806718. E-mail:
[email protected] (J. M. Yue). ORCID
Kun Gao: 0000-0002-3856-3672 Jian-Min Yue: 0000-0002-4053-4870 Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This work was supported by the National Natural Science Foundation of China (nos. 21532007 and 21778027). We thank Prof. Y.-K. Xu of Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences (CAS), for the identification of the plant material.
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REFERENCES
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DOI: 10.1021/acs.orglett.8b01608 Org. Lett. 2018, 20, 4040−4043
