Synthesis and biological profiling of half-calycanthaceous alkaloid analogues

Article abstract of DOI:10.1080/10286020.2020.1791100

During our continuous efforts to pursue antifungal agents, some calycanthaceous alkaloid analogs showed diverse and promising bioactivities. Therefore, 34 new calycanthaceous alkaloid derivatives were further prepared and screened for bioactivities. As a result of the evaluation against a great deal of plant pathogen fungi, bacteria and human pathogenic fungi, a majority of them displayed potent bioactivity. In particular, compound b6 displayed remarkably activity and might be novel potential leading compound for further development of antifungal agent. The relationship between structure and biological activity was also discussed.

Full text of DOI:10.1080/10286020.2020.1791100

Journal of Asian Natural Products Research
ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/ganp20
Synthesis and biological profiling of half-
calycanthaceous alkaloid analogues
Rui Zhu , Yang Wei , Ke Han , Xin-Wei Shi , Yong-Dong Gu , Hong-Jin Bai &
Shao-Jun Zheng
To cite this article: Rui Zhu , Yang Wei , Ke Han , Xin-Wei Shi , Yong-Dong Gu , Hong-Jin Bai
& Shao-Jun Zheng (2020): Synthesis and biological profiling of half-calycanthaceous alkaloid
analogues, Journal of Asian Natural Products Research, DOI: 10.1080/10286020.2020.1791100
To link to this article: https://doi.org/10.1080/10286020.2020.1791100
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JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
https://doi.org/10.1080/10286020.2020.1791100
Synthesis and biological profiling of half-calycanthaceous
alkaloid analogues
Rui Zhua, Yang Weia, Ke Hana, Xin-Wei Shib, Yong-Dong Gua, Hong-Jin Baic and
Shao-Jun Zhenga
aSchool of Environmental and Chemical Engineering, Jiangsu University of Science and Technology,
b
Zhenjiang 212003, China; Engineering Center of Qinling Mountains Natural Products, Shaanxi
c
Academy of Sciences, Institute of Botany of Shaanxi Province, Xi’an 710061, China; Key Laboratory
of Protection & Utilization of Biological Resources in Tarim Basin of Xinjiang Production and
Construction Corps/College of Life Sciences, Tarim University, Alar 843300, China
ABSTRACT
ARTICLE HISTORY
Received 8 March 2020
Accepted 27 June 2020
During our continuous efforts to pursue antifungal agents, some
calycanthaceous alkaloid analogs showed diverse and promising
bioactivities. Therefore, 34 new calycanthaceous alkaloid deriva-
tives were further prepared and screened for bioactivities. As a
result of the evaluation against a great deal of plant pathogen
fungi, bacteria and human pathogenic fungi, a majority of them
displayed potent bioactivity. In particular, compound b6 displayed
remarkably activity and might be novel potential leading com-
pound for further development of antifungal agent. The relation-
ship between structure and biological activity was also discussed.
KEYWORDS
Calycanthaceous alkaloids;
synthesis; plant pathogen
fungi; biological activity
1. Introduction
Plant diseases are caused mainly via fungal pathogens, threating to the global food
security. The majority of the currently antifungal agents possess serious drawbacks,
such as serious drug resistance, and severe drug interactions. Therefore, novel anti-
fungal agents with improved fungicidal potency are the key to control those
plant diseases.
As a well-established lead optimization method, structure modification is continu-
ally used to discover compounds with structural novelty, and increased potency. With
regard to pesticide design, structure optimization is also extensively recommended to
CONTACT Shao-Jun Zheng
sz281cam@just.edu.cn; Hong-Jin Bai
bhj67@163.com
Supplemental data for this article is available online at https://doi.org/10.1080/10286020.2020.1791100.
ß 2020 Informa UK Limited, trading as Taylor & Francis Group

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R. ZHU ET AL.
Figure 1. Structures of calycanthaceous alkaloids.
change the lead compounds. Calycanthaceous alkaloids [1–4] (Figure 1), bearing hexahy-
dropyrroloindole skeletons, are an important class of alkaloid which have been utilised as
traditional Chinese medicines for the treatment of funga [5], tumor, inflammatory, and
melanogenesis [6, 7]. Due to the wide spectrum of biological activities, a great deal of stud-
ies towards the synthesis and antimicrobial activity of calycanthaceous alkaloids have been
reported [8–17]. In our previous work, calycanthaceous alkaloids derivatives were designed
and synthesized. Among them, some derivatives showed diverse and promising bioactiv-
ities, such as, compound c4 showed remarkably high activity against Ralstonia solanacea-
rum, with a MIC value of 1.96mg mlꢀ1, compound b9 showed better against Verticillium
dahlia compared with chlorothalonil, with a MIC value of 7.81mg mlꢀ1, compound a1 dis-
played a significant activities against B. cereus, Escherichia sp and R. solanacearum, even
better than the positive control streptomycin and penicillin, with the same MIC value of
15.63mg mlꢀ1, and compounds c2 and c11 revealed potent activity against acetylcholin-
esterase, with MIC values of 0.01 and 0.1 ng mlꢀ1, respectively [18–24]. As a part of our
continuous work in pursuit of antifungal agents with novel structure and high efficiency,
we initiated a structural modification campaign focusing on introducing functional group
at the N-position to compound 3. Taking the synthetic accessibility into consideration,
three novel series of N-substituted calycanthaceous alkaloid derivatives were identified as
the target compounds for synthesis.
To the best of our knowledge, the biological activities of the synthetic derivatives
are reported here for the first time.
2. Results and discussion
2.1. Rational design and synthesis of calycanthaceous alkaloid analogues
According to our previous study, the introduction of functional group at the N-position
to compound 3 could affect the bioactivities. To explore novel antifungal agents with
novel structure and high efficiency, the calycanthaceous alkaloid analogs were prepared
from indole-3-acetonitrile via acylation at the N-position based on our reported meth-
1
ods and confirmed by H NMR, 13C NMR, and HRMS according to a previously
reported procedure in our group (Scheme 1) [19, 20].

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
3
Scheme 1. Synthetic route to the title compounds a1–a12 and b1–b22 [19].
2.2. Antimicrobial activity
Chlorothalonil, fluconazole, carbendazim, gentamicin, penicillin, streptomycin, or
amphotericin B were used as the positive control. The fungicidal activities of all the
target compounds were evaluated against V. dahliae, Fusarium oxysperium sp. vasin-
fectum, Cytospora juglandis, Aspergillus flavus, Penicillium citrinum, Fusarium oxyspo-
rum, Colletotrichum orbiculare, Aspergillus niger, B. cinerea Pers, Curvularia lunaia,
Escherichia sp, Pseudomonas aeruginosa, R. solanacearum, B. cereus, Staphylococcus
aureus, Candida krusei, Crytococcus neoformans and Candida tropicalis, and the
results are provided in Table 1.
The antibacterial results are displayed in Table 1. The biological testing showed
that a majority of the compounds showed potent antifungal activities. Compounds
b1, b6, b7, b8, b9, b9, and b13 revealed excellent activity against V. dahlia, with MIC
values of 62.5, 15.6, 62.5, 31.3, 31.3, 62.5, and 31.3 mg mlꢀ1, respectively. Among
them, Compound b6 showed more effective activity against V. dahlia than that of
chlorothalonil (MIC ¼ 31.3 mg mlꢀ1), with a MIC value of 15.6 mg mlꢀ1. All of the
synthesied compounds displayed activity against C. juglandis, A. flavus, F. oxysporum,
and A. niger, but only compound a3 is similar to that of chlorothalonil against C.
juglandis, all with the same MIC value of 62.5 mg mlꢀ1. Compound b1 manifested
comparable control efficacy against P. citrinum to that of amphotericin (MIC ¼
31.3 mg mlꢀ1), with a MIC value of 31.3 mg mlꢀ1. Compound b1 displayed moderate
activity against Escherichia sp, with a MIC value of 31.3 mg mlꢀ1. The activity of com-
pound b1 is more potent than that of streptomycin against Escherichia sp, with a
MIC value of 31.3 mg mlꢀ1. Compounds a10 and b9 indicated more activity against
Escherichia sp than that of Streptomycin, with a MIC value of 62.5 mg mlꢀ1
.
Compounds a2, b1, and b2, displayed moderate activity against B. cereus, with MIC
values of 15.6, 31.3 and 62.5 mg mlꢀ1, respectively. Compounds b1 and b2 displayed
moderate activity against S. aureus, with MIC values of 31.3 and 62.5 mg mlꢀ1
,
respectively.
Although it is difficult to extract apparent structure-activity relationships from the
bioassay results, some conclusions can still be obtained. Firstly, compounds (a2, a3,
b1, b2, and b6-b9) with alkyl carbonyl substituted groups at N-position indicated

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R. ZHU ET AL.

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
5
more activities and a broad antifungal spectrum than the compounds (a5-a12 and
b11-b14) with aryl carbonyl substituted groups, even better than the positive control
carbendazim, chlorothalonil, and streptomycin. Secondly, compounds (a2, b1, and
b2) with shorter alkyl carbonyl substituted groups at N-position showed a greater
activities than the compounds (a3, and b6-b9) with longer alkyl carbonyl substituted
groups. Thirdly, almost all synthesised compounds showed better activity against a
broad range of plant pathogen fungi than bacteria and human pathogenic fungi.
Fourth, compounds of series b displayed distinctive activity against V. dahlia. Fifth,
the biological activity shows that the electron donor substituents are beneficial to the
improvement of biological activity.
In conclusion, a total of 34 novel tetrahydropyrroloindole-based calycanthaceous
alkaloid analogs were prepared via acylation at the N-position, and the activity against
a great deal of plant pathogen fungi, bacteria and human pathogenic fungi were eval-
uated. The biological testing showed that a majority of the compounds indicated
potent antifungal activities, in which compound b6 was the optimal one and identi-
fied as the most potential candidate for further study. Further structure optimization
of calycanthaceous alkaloids and the QSAR studies are well underway in our group.
3. Experimental
3.1. General experimental procedures
All chemicals were purchased from market and purified based on standard routes
1
before use. The H NMR (500 MHz), and 13C NMR (125 MHz) spectra were obtained
on an AM ꢀ 500 FT ꢀ NMR spectrometer (Bruker Corporation, Karlsruhe,
Switzerland) with CDCl3 as the solvent and TMS as the internal standard. Melting
points were taken on an Electrothermal digital apparatus (Beijing, China) and are
uncorrected. MS were recorded under ESI conditions using a LCQ Fleet instrument
(Thermo Fisher, Waltham, MA, USA). Analytical thin-layer chromatography (TLC)
was performed with silica gel plates using silica gel 60 GF254 (Qingdao Haiyang
Chemical Co., Ltd., Qingdao, China). The title compounds were synthesized under a
nitrogen atmosphere. Yields were not optimized.
3.2. Synthesis
3.2.1. Synthesis of intermediates 1-3
Intermediates 1-3 were synthesized according to our previously reported
method [18–24].
3.2.2. Synthesis of compounds a1–a12 and b1–b22
Intermediate 3 was added in pyridine (15 ml) in a round flask. Then, the correspond-
ing desired reagent was added portionwise at 0 ꢁC. After refluxing for 2 h, the mixture
was warmed to room temperature. Then, the resulting mixture was reacted for 1 h.
When TLC monitoring showed that the intermediate 3 had disappeared, the resulting
mixture was quenched. The solvent was evaporated under reduced pressure and then
washed with water, extracted with ethyl acetate and the extracts were dried over

6
R. ZHU ET AL.
Na2SO4. At last, the solvent was concentrated and purified to afford the compounds
a1–a12 and b1–b22 in yields from 78% to 91%.
3.2.2.1. Synthesis of intermediate a1. The acetyl chloride was added portionwise to a
stirred solution of intermediate 3 in pyridine (15 ml) in a round flask at 0 ꢁC. After
refluxing for 1.5 h, the mixture was warmed to room temperature. Then, the resulting
mixture was reacted for 1 h. When TLC monitoring showed that the intermediate 3
had disappeared, the resulting mixture was quenched. The solvent was evaporated
under reduced pressure and then washed with water, extracted with ethyl acetate and
the extracts were dried over Na2SO4. At last, the solvent was concentrated and puri-
fied to afford compounds a1.
1-(3a,8-bis(2-methylbenzyl)-3,3a,8,8a-tetrahydropyrrolo[2,3-b]indol-1(2H)-yl)e-
than-1-one(a1) Yellow oil, 1H- NMR (500 MHz, CDCl3) d 7.19 ꢀ 6.88 (m, 9H),
6.77 ꢀ 6.56 (m, 2H), 6.19 (d, J ¼ 7.9 Hz, 1H), 5.54 (s, 1H), 4.87 ꢀ 4.15 (m, 2H),
3.60 ꢀ 3.51 (m, 1H), 3.31 ꢀ 3.14 (m, 2H), 2.95 (d, J ¼ 13.7 Hz, 1H), 2.35 (s, 2H),
2.30 ꢀ 2.15 (m, 3H), 2.03 ꢀ 1.72 (m, 6H). 13C- NMR (125 MHz, CDCl3) d 170.02(C),
150.34(C), 137.41(C), 137.03(C), 135.72(C), 135.54(C), 131.30(C), 130.52(CH),
130.4(CH), 130.1(CH), 128.6 (CH), 126.7 (CH), 126.4 (CH), 125.9 (CH), 125.7 (CH),
125.7 (CH), 123.2 (CH), 117.1 (CH), 106.3 (CH), 83.3 (CH), 57.0 (C), 47.6 (CH2),
44.0 (CH2), 39.9 (CH2), 36.8 (CH2), 22.8 (CH3), 19.9 (CH3), 19.4 (CH3). HRESIMS:
m/z 411.2434 [M þ H]þ (calcd for C28H31N2O, 411.2436).
The data of the other compounds and the NMR spectral details can be found in
the Supplementary data.
3.3. Biological activity
The bioactivities of calycanthaceous alkaloid derivatives were tested based on the
reported procedure [25]. The concentrations were 250, 125, 62.5, 31.3, 15.6, 7.8, 3.9
and 2 mg mlꢀ1, respectively. The antibacterial test cells were incubated at 37 ꢁC for
24 h. All tests were manipulated in triplicate and repeated if the results differed.
Disclosure statement
The authors declare no conflict of interest.
Funding
This work was financially supported by the National Natural Science Foundation of China
(21502073), the Natural Science Foundation of Jiangsu Province (Grants No BK 20150465 and
BK20180978), and the Key Research and Development Program (Modern Agriculture) of
Zhenjiang City (NY2018002), Xinjiang Production Construction Corps Key Laboratory of
Protection and Utilization of Biological Resources in Tarim Basin (BRYB1703), and Science
and Technology Research Project of Shaanxi Province Academy of Sciences (2018nk-01), and
Zhoushan Public Welfare Science and Technology Project (2019C31066).

JOURNAL OF ASIAN NATURAL PRODUCTS RESEARCH
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