Letter Cite This: Org. Lett. 2018, 20, 7926−7928
pubs.acs.org/OrgLett
Ophiorrhines A and B, Two Immunosuppressive Monoterpenoid Indole Alkaloids from Ophiorrhiza japonica Tao Feng,† Kai-Ting Duan,† Shi-Jun He,‡ Bin Wu,† Yong-Sheng Zheng,† Hong-Lian Ai,† Zheng-Hui Li,† Juan He,*,† Jian-Ping Zuo,*,‡ and Ji-Kai Liu*,† †
School of Pharmaceutical Sciences, South-Central University for Nationalities, Wuhan 430074, China State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
‡
Org. Lett. 2018.20:7926-7928. Downloaded from pubs.acs.org by UNIV OF WINNIPEG on 12/21/18. For personal use only.
S Supporting Information *
ABSTRACT: Two monoterpenoid indole alkaloids, ophiorrhines A (1) and B (2), were obtained from plant Ophiorrhiza japonica BI. Their structures were elucidated by extensive spectroscopic methods and single crystal X-ray diffraction. Compounds 1 and 2 possess a novel spirocyclic ring system. Its biosynthesis pathway is proposed. Compound 2 exhibits potent inhibitory activity against concanavalin A (Con A) induced T cell proliferation and lipopolysaccharide (LPS) induced B lymphocyte cell proliferation with IC50 values 13.3 and 7.5 μM, respectively. Compound 1 exhibits significant inhibition specifically against the LPS-induced proliferation of B lymphocyte cells with IC50 value 18.6 μM.
I
treat cough, arthralgia and myalgia, injuries, etc. However, there were few studies on chemical constituents such that only several steroids, triterpenoids, simple indole alkaloids, and phenolic acids were reported previously.13,16 In the current study, we describe two novel MIAs glycosides, ophiorrhines A (1) and B (2) (Figure 1), from O. japonica, including their isolation, structural elucidation, and proposed biosynthesis pathway, as well as immunosuppressive activity.
mmunosuppressants compose a very important class of drugs that are often used for organ transplantation and treatment of various autoimmune diseases like rheumatoid arthritis, multiple sclerosis, and psoriasis in clinic.1 At present there are a number of immunosuppressive drugs by inhibiting T cell proliferation, but the new, efficient, and safe immunosuppressive drugs by inhibiting B cell proliferation are still unavailable.1 Natural products with unique and diverse scaffolds are extraordinarily valuable sources of new therapeutic approaches and new drugs to treat some serious human diseases.2,3 A broad range of chemical diversity and biological specificity make natural products favorable leads for drug discovery and the identification and understanding of novel biochemical processes.4 Monoterpene indole alkaloids (MIAs), commonly found in the families of Apocynaceae, Rubiaceae, and Loganiaceae,5−7 are the largest and most diverse subgroup of alkaloids. MIAs comprise an important type of natural products including famous compounds such as camptothecins, ajmaline, vindoline, and quinine owing to their structural diversity and prominent pharmacological activities.8 It is understood that strictosidine serves as a central precursor in almost all MIAs, which itself originates from the condensation of tryptophan with secologanin.8−10 The rearrangements of the highly convertible 10-carbonterpene moiety derived from secologanin resulted in thousands of various MIA backbones.9 Ophiorrhiza plants in the family Rubiaceae are well-known to produce anticancer camptothecins, as well as β-carbolinetype MIAs.11−15 O. japonica has a wide distribution in Central and Southwest China. It is used as a folk herbal medicine to © 2018 American Chemical Society
Figure 1. Structures of ophiorrhines A (1) and B (2).
An alkaloidal extract of O. japonica was subjected to column chromatography over silica gel and Sephadex LH-20 and purified by HPLC to produce two alkaloids 1 (23 mg) and 2 (7 mg). Ophiorrhine A (1) was isolated as colorless crystals (MeOH). Its molecular formula C28H32N2O10 was determined on the basis of the high-resolution ESI mass spectrum at m/z Received: November 1, 2018 Published: December 10, 2018 7926
DOI: 10.1021/acs.orglett.8b03489 Org. Lett. 2018, 20, 7926−7928
Letter
Organic Letters
proton H-1′ in the sugar moiety revealed a β-configuration of the glucose residue. D-Glucose was identified by single crystal X-ray diffraction; the letter also confirmed the novel carbon ring system ring, along with the absolute configuration of 1 (Flack parameter = 0.09(4). CCDC: 1875337, Figure 3).
579.1952 [M + Na] + (calcd for C28H32N2O10Na, 579.1949), corresponding to 14 degrees of unsaturation. The IR spectrum showed absorption bands for CO (1694 cm−1) and OH (3425 cm−1) groups, while the UV data at 280 nm indicated a conjugated system. The 13C NMR and DEPT spectra revealed 28 carbon resonances ascribable for two sp3 CH3, three sp3 CH2, 15 CH including five sp2 and ten sp3 ones, and eight quaternary carbons including one sp3 and seven sp2 ones (see Table S1 in the Supporting Information). In the 1H NMR spectrum, two aromatic doublets at δH 7.32 and 7.47 (d, J = 7.8 Hz) and two aromatic triplets at δH 7.01 and 7.04 (t, J = 7.8 Hz) suggested the existence of an ortho-disubstituted benzene ring, corresponding to an indole moiety. In addition, multiple signals at δH 3.16 to 3.88, as well as an anomeric proton at δH 4.59 (1H, d, J = 8.0 Hz, H-1′), indicated a sugar moiety, which was supported by the characteristic 13C NMR data (δC 98.5, 73.2, 76.5, 70.1, 76.9, 61.2). Analysis of these spectroscopic data with respect to those reported previously5−7 suggested that 1 is an MIA glucoside. Analysis of H−1H COSY spectrum revealed three spin systems as shown in Figure 2. Two spin systems were
Figure 3. ORTEP diagrams of 1 (left) and 2 (right).
Ophiorrhine B (2) was also isolated as colorless crystals (MeOH). The molecular formula C29H34N2O11 was established by the HRESIMS data at m/z 609.2041 [M + Na]+ (calcd for C29H34N2O11Na, 609.2055), 30 units more than that of 1. The spectroscopic data of 2 were closely related to those of 1. In the 1H and 13C NMR spectrum (see Table S1 in the Supporting Information), signals for one more OCH3 group were detected (δH 3.90, 3H, s; δC 53.7, CH3). These data suggested that 2 might be an OCH3 substituted product of 1. The HMBC correlations from δH 3.90 (3H, s, OCH3) and 1.99 (2H, d, J = 7.7 Hz, H-18) to δC 82.8 (s, C-6) suggested that the OCH3 should be placed at C-6, which allowed C-6 to be a quaternary carbon at δC 82.8. Detailed analysis of NMR data suggested that the other parts were the same as that of 1. The structure as well as the absolute configuration was, finally, determined by single crystal X-ray diffraction (Flack parameter = 0.15(4). CCDC: 1875336, Figure 3). Ophiorrhines A (1) and B (2) bear a novel bridged carbon ring system based on two new C−C bonds as depicted. Their biosynthetic process is understood to undergo an intramolecular [4 + 2] Diels−Alder cycloaddition via a proposed intermediate as shown in Scheme 1. To construct the
Figure 2. Key 2D NMR correlations of 1.
ascribable for the indole aromatic ring and the sugar moiety, respectively. The third one, involving protons for seven carbons of C-6, C-18, C-19, C-20, C-21, C-15, and C-14, revealed an abnormal carbon skeleton with respect to those known glucosidic MIAs from plants of the same genus.12 In the HMBC spectrum, a singlet at δH 2.95 (3H, s) for N4-methyl showed key correlations to δC 178.3 (s, C-5) and 68.3 (s, C-3), which suggested a lactam moiety at C-5 and an sp3 quaternary carbon at C-3. Then the HMBC correlations from δH 4.15 (1H, t, J = 2.6 Hz, H-6) to C-5 and δC 111.1 (s, C-7), as well as the1H−1H COSY correlation between H-6 with H-18 (δH 1.85, 2H, ddd, J = 6.8, 2.5, 2.1 Hz, H-18), suggested that a new C−C bond was established between C-6 and C-18. In addition, a key HMBC correlation from δH 1.91 (1H, m, H19) to C-3 was observed, indicating another new C−C bond between C-3 and C-19. The above new carbon bonds allowed the establishment of a bridged ring E and a five-membered ring F. The other 1D and 2D NMR data suggested that the other parts of 1 were the same to the known glycosidic MIAs including a seco-iridoidal ring D with a sugar moiety, as well as a methyl ester functional group.12 Due to the bridged ring system, the relative configuration of the alkaloidal part in 1 was easily identified by the ROESY experiment. As shown in Figure 1, the ROESY correlation of H-15/H-20 suggested that they were in the same face (assigned as α orientation). Based on this, the observed ROESY correlation between H-19 and H-21 suggested that H21 was β oriented. The J value (8.0 Hz) of the anomeric
Scheme 1. Proposed Biosynthetic Pathway for 1 and 2
intermediate, two possible pathways were given (Scheme 1). The first is suggested to start from the oxidation of a structurally related precursor 5-oxodolichantoside, which was isolated as the major constituent from the same resource. The secondary pathway is suggested to construct a 3-keto intermediate by a condensation of tryptamine and secologanin, then formed an imine ion. The latter, under an alkaline 7927
DOI: 10.1021/acs.orglett.8b03489 Org. Lett. 2018, 20, 7926−7928
Letter
Organic Letters condition, produced the same intermediate as proposed in the first pathway. Compounds 1 and 2 showed no cytotoxicity against five human cancer cell lines (HL-60, A-549, SMMC-7721, SW480, MCF-7). They were evaluated for their in vitro inhibition activities on concanavalin A (Con A) induced T cell proliferation and lipopolysaccharide (LPS) induced B cell proliferation.17,18 Compound 2 was found to exhibit potent inhibitions against concanavalin A (Con A) induced T cell proliferation and lipopolysaccharide (LPS) induced B lymphocyte cell proliferation with IC50 values 13.3 and 7.5 μM, respectively. Compound 1 exhibited significant inhibition specifically against the LPS-induced proliferation of B lymphocyte cells with IC50 value 18.6 μM. (Table 1).
Ji-Kai Liu: 0000-0001-6279-7893 Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This work was financially supported by the National Natural Science Foundation of China (31870513, 81872762, 31560010, 81773590, 81561148013), the National Key Research and Development Program of China (2017YFC1704007), the Key Projects of Technological Innovation of Hubei Province (No. 2016ACA138), and the Fundamental Research Funds for the Central University, South-Central University for Nationalities (CZT18014, CZT18013, CZP18005, CZQ17010, CZQ17008). The authors thank the Analytical & Measuring Center, School of Pharmaceutical Sciences, South-Central University for Nationalities for the spectral measurement.
Table 1. Immunosuppressive Tests of 1 and 2 ConA-induced T-cell proliferation
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LPS-induced B-cell proliferation
compd
CC50 (μM)
IC50 (μM)
SIa
IC50 (μM)
SIa
1 2 CsA
143.00 90.92 >2.50
>200 13.34 0.03
83.33
18.60 7.51 0.32
7.63 12.11 >7.81
(1) Dangroo, N. A.; Singh, J.; Dar, A. A.; Gupta, N.; Chinthakindi, P. K.; Kaul, A.; Khuroo, M. A.; Sangwan, P. L. Eur. J. Med. Chem. 2016, 120, 160−169. (2) Cragg, G. M.; Grothaus, P. G.; Newman, D. J. J. Nat. Prod. 2014, 77, 703−723. (3) Butler, M. S. J. Nat. Prod. 2004, 67, 2141−2153. (4) Koehn, F. E.; Carter, G. T. Nat. Rev. Drug Discovery 2005, 4, 206. (5) Ishikura, M.; Abe, T.; Choshi, T.; Hibino, S. Nat. Prod. Rep. 2013, 30, 694−752. (6) Saxton, J. E. Nat. Prod. Rep. 1996, 13, 327−363. (7) Leonard, J. Nat. Prod. Rep. 1999, 16, 319−338. (8) De Luca, V.; Salim, V.; Atsumi, S. M.; Yu, F. Science 2012, 336, 1658. (9) Stöckigt, J.; Panjikar, S. Nat. Prod. Rep. 2007, 24, 1382−1400. (10) Smith, G. N. Chem. Commun. 1968, 912−914. (11) Kitajima, M.; Yoshida, S.; Yamagata, K.; Nakamura, M.; Takayama, H.; Saito, K.; Seki, H.; Aimi, N. Tetrahedron 2002, 58, 9169−9178. (12) Kitajima, M.; Ohara, S.; Kogure, N.; Santiarworn, D.; Takayama, H. Tetrahedron 2013, 69, 9451−9456. (13) Kitajima, M. J. Nat. Med. 2007, 61, 14−23. (14) Aimi, N.; Tsuyuki, T.; Murakami, H.; Sakai, S.; Haginiwa, J. Tetrahedron Lett. 1985, 26, 5299−302. (15) Aimi, N.; Hoshino, H.; Nishimura, M.; Sakai, S.; Haginiwa, J. Tetrahedron Lett. 1990, 31, 5169−72. (16) Fujita, E.; Sumi, A. Yakugaku Zasshi 1967, 87, 1153−1155. (17) Zhang, S. B.; Huang, Y.; He, S. J.; Chen, H. P.; Wu, B.; Li, S. Y.; Zhao, Z. Z.; Li, Z. H.; Wang, X.; Zuo, J. P.; Feng, T.; Liu, J. K. J. Org. Chem. 2018, 83, 10158−10165. (18) Fan, Y. Y.; Gan, L. S.; Liu, H. C.; Li, H.; Xu, C. H.; Zuo, J. P.; Ding, J.; Yue, J. M. Org. Lett. 2017, 19, 4580−4583.
a
SI (selectivity index) is determined as the ratio of the concentration of the compound that reduced cell viability to 50% (CC50) to the concentration of the compound needed to inhibit the proliferation by 50% relative to the control value (IC50).
In conclusion, two glycosidic MIAs, ophiorrhines A and B with a new carbon skeleton, were obtained from plant Ophiorrhiza japonica. To the best of our knowledge, the proposed intramolecular [4 + 2] Diels−Alder cycloaddition involving the double bond of C-18/C-19 in MIAs is reported for the first time. The discovery of this type of bioactive MIAs provides new possibilities to explore new immunosuppressants by inhibiting B cell proliferation from natural sources.
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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.8b03489. General experimental procedures, plant materials, extraction and isolation, bioactivity assay, and spectroscopic data for 1 and 2 (PDF) Accession Codes
CCDC 1875336−1875337 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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REFERENCES
AUTHOR INFORMATION
Corresponding Authors
*E-mail:
[email protected] (J.H.). *E-mail:
[email protected] (J.P.Z.). *E-mail:
[email protected] (J.K.L.). ORCID
Tao Feng: 0000-0002-1977-9857 Zheng-Hui Li: 0000-0003-1284-0288 7928
DOI: 10.1021/acs.orglett.8b03489 Org. Lett. 2018, 20, 7926−7928
