Biomimetic synthesis and anti-inflammatory effects of horsfiequinone A

Article abstract of DOI:10.1016/j.tetlet.2020.152756

Inspired by the oxy-tyrosinase type II copper enzyme, a biomimetic synthesis of the natural product horsfiequinone A (1) has been achieved using CuOTf/DBED/O₂ catalyzed oxidation as a key step. The synthetic route furnished 1 in 33% overall yield (64% brsm) from commercially available para-hydroxybenzaldehyde. Moreover, revisiting the biological activity of 1 resulted in the discovery of its in vitro inhibitory activity towards nitric oxide (NO) production in LPS-induced RAW264.7 cells with an IC₅₀ value of 4.42 ± 0.81 μM. The anti-inflammatory effect of 1 was further supported by an iNOS expression inhibition assay and molecular docking simulation.

Full text of DOI:10.1016/j.tetlet.2020.152756

Tetrahedron Letters 65 (2021) 152756
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Biomimetic synthesis and anti-inflammatory effects of horsfiequinone A
a
a
a
Mei Wang a, Yuan-Lie Liu b, Dashan Li a, , Wen-Wen Xiao , Yang Chen , Hong-Lin Zhang ,
⇑
a,
Rui Zhan b, , Li-Dong Shao
⇑
⇑
a Yunnan Key Laboratory of Southern Medicinal Resources, School of Chinese Materia Medica, Yunnan University of Chinese Medicine, Kunming 650500, China
b School of Chemistry and Chemical Engineering, Yunnan Normal University, Kunming 650050, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Inspired by the oxy-tyrosinase type II copper enzyme, a biomimetic synthesis of the natural product hors-
fiequinone A (1) has been achieved using CuOTf/DBED/O2 catalyzed oxidation as a key step. The synthetic
route furnished 1 in 33% overall yield (64% brsm) from commercially available para-hydroxybenzalde-
hyde. Moreover, revisiting the biological activity of 1 resulted in the discovery of its in vitro inhibitory
activity towards nitric oxide (NO) production in LPS-induced RAW264.7 cells with an IC50 value of
Received 21 November 2020
Revised 7 December 2020
Accepted 11 December 2020
Available online 5 January 2021
4.42 0.81
assay and molecular docking simulation.
lM. The anti-inflammatory effect of 1 was further supported by an iNOS expression inhibition
Keywords:
Diarylpropane
Horsfiequinone A
Biomimetic synthesis
Aerobic oxidation
Anti-inflammation
Ó 2021 Elsevier Ltd. All rights reserved.
Introduction
the corresponding hydroquinones [8–16]; however, the biomi-
metic transformations from phenols to hydroxyquinones have
been rarely reported [17,18]. Inspired by such pioneering works
that mimicked the oxidative function of the oxy-tyrosinase type
II copper enzyme [19–21], herein, we report a biomimetic syn-
thesis of 1 via direct oxidation by hydrogen peroxide (H2O2) and
stepwise aerobic oxidation catalyzed by Cu(I)/N,N0-di-tert-
butylethylenediamine (DBED). In addition, the in vitro anti-
inflammatory effect of 1 was also evaluated by serial bioassays
and molecular modelling.
Diarylpropanes (DAPs) and their asymmetric dimers contain-
ing para-benzoquinones are rarely distributed in naturally occur-
ring products, which have been isolated only from plant species
including Combretum griffithii [1], Horsfieldia tetratepala [2–4],
and Euonymus glabra Roxb [5]. Some of these DAPs showed inhi-
bitory activity against Mycobacterium tuberculosis[1] while
others, such as horsfiequinone B (2) (Fig. 1), exhibited selective
inhibition against the HL-60 cancer cell line with an IC50 value
of 4.28
lM [2]. In addition, horsfiequinone A (1) was proposed
to be the biogenetical precursor of other bioactive dimeric DAPs
[6]. However, as a major secondary metabolite in H. kingii, 1
was found to be inactive against various cancer cell lines [2],
and to date, no biological activity of 1 has been established. Since
H. kingii is an ethnopharmacological plant which has been used to
treat various inflammation related diseases [7], it appeared
important to revisit the anti-inflammatory activity of 1.
The 1,4-p-quinone moiety of 1 was previously derived from
hydroquinone, which was used as a pre-functional group trans-
formed to a quinone in the synthetic route [6]. As an alternative,
directly transforming phenols into hydroxyquinones is highly
desired due to its high atom economy. Extensive endeavors have
been made to prepare various hydroxyquinones via oxidation of
Results and discussion
Based on the proposed direct oxidation (Fig. 1), the synthesis
of 1 began with the preparation of compounds 3a and 3b
according to literature methods [6,7] from commercially avail-
able 4a and 4b via Wittig olefination/hydrolysis and Wittig ole-
fination/catalytic hydrogenation sequences in 84% and 70%
overall yield, respectively (Scheme 1). It is worth noting that
no protecting group was required for para-hydroxybenzalde-
hyde 4b in the Wittig reaction using tBuOK as a base instead
of LiHMDS. Next, we focused on the biomimetic oxidation of
DAPs 3a and 3b.
The treatment of 3a with hydrogen peroxide (2–5 eq.) in formic
acid (HCO2H) led to either a complex reaction mixture or complete
decomposition of the substrate (Table 1, entries 1 and 2) [22].
Replacing HCO2H with MeCN resulted in no reaction (Entry 3).
⇑
Corresponding authors.
E-mail addresses: lidashan@ynutcm.edu.cn (D. Li), sunnyzhanrui@126.com
(R. Zhan), shaolidong@ynutcm.edu.cn (L.-D. Shao).
https://doi.org/10.1016/j.tetlet.2020.152756
0040-4039/Ó 2021 Elsevier Ltd. All rights reserved.

M. Wang, Yuan-Lie Liu, D. Li et al.
Tetrahedron Letters 65 (2021) 152756
ortho-quinone when 2,4-di-tert-butylphenol was used as a substrate
in previous work [23]. Additionally, both CuI and CuBr resulted in
complex reaction mixtures (Entries 14–15). Other ligands such as
triethylamine (TEA) and tetramethylethylenediamine (TMEDA)
were not suitable for this oxidation (Entries 16–17). To our delight,
the best result was obtained using CuOTf/DBED which gave 1 in
47% (92% brsm) yield (Entry 18). However, attempts to trans-
form 3a into the corresponding quinone 1 using the CuOTf/
DBED catalyst system were unsuccessful (Entry 19). The spec-
troscopic data of synthetic 1 were identical with those of natu-
ral horsfiequinone A.
Fig. 1. Horsfiequinone A (1), representative DAP dimer (2), and the proposed
biomimetic synthesis of 1.
Next, the in vitro anti-inflammatory effect of 1 was tested
against NO production in RAW264.7 macrophages. As shown
in Table 2, compound
(IC50 = 4.42 0.81 M) against NO production in LPS-induced
RAW264.7 cells without cytotoxicity (CC50 > 100 M), which
was more potent than the positive control -NMMA
(IC50 = 12.16 0.42 M). Encouraged by this result, we subse-
quently examined the role of compound 1 on the expression
of inducible NO synthase (iNOS) in LPS-induced RAW264.7 cells,
because iNOS accounts for most of the NO in inflammation [24].
As a result, 1 inhibited iNOS expression in a dose-dependent
manner in LPS-induced RAW264.7 cells suggesting the inhibi-
tory activity against NO production of 1 was likely caused by
its inhibitory effects on iNOS expression (Fig. 2).
1 exhibited a strong inhibition
Gratifyingly, trace amounts of 1 were detected using the mixed sol-
vents (MeCN/HCO2H = 10:1) (Entry 4). Subsequently, the yield of 1
was improved to 10% by increasing the ratio of HCO2H and con-
ducting the reaction at 40 °C (Entries 5–7). However, the product
was difficult to isolate from the complex reaction system, possibly
due to the highly reactive peroxy acid formed in situ. Consequently,
we explored the feasibility of stepwise oxidation of phenol 3b
using the [Cu(MeCN)4](PF6)/DBED catalyst system. Unfortunately,
compound 1 was obtained in less than 10% yield according to the
procedures reported by Lump and co-workers, whether the reac-
tion was conducted in a stepwise or one pot manner (Entries 8
and 9) [17,18]. Interestingly, the yield of 1 increased dramatically
to 40% (78% brsm) by changing the order of addition for substrate
3b and the [Cu(MeCN)4](PF6)/DBED catalyst system (Entry 10). In
addition, using the combination of MeOH/TfOH instead of H2SO4/
H2O and TMSCH2N2 afforded 1 in comparable yield (Entry 11).
The yield of 1 was improved (42%, 80% brsm) by increasing the
loading of MeOH/TfOH as well as using 100 wt% of 4 Å MS (Entry
12). The Cu source and various ligands were also tested. CuCl (Entry
13) gave trace amounts of 1, although it was able to generate the
l
l
L
l
Finally, a molecular docking simulation was performed to gain
further insights into the anti-inflammatory effect of compound 1
(Fig. 3). Two strong hydrogen bonds (2.07 and 2.46 Å) with resi-
dues ARG382 and VAL346, one
p-anion interaction with ASP376,
and one -alkyl interaction with PRO344 near the Heme-Fe [25]
p
were detected in the docking conformation with the lowest bind-
ing energy which may further support the NO production inhibi-
tory activity of 1.
Scheme 1. Biomimetic synthesis of 1 from DAPs 3a and 3b.
2

M. Wang, Yuan-Lie Liu, D. Li et al.
Tetrahedron Letters 65 (2021) 152756
Table 1
Table 2
Biomimetic oxidation of DAPs 3a and 3b.
In vitro NO production inhibitory activity of 1.a
Entry
Substrate Reagents and conditions
Yield 1 (%)
(brsm in
Compound
IC50 (lM)
1
4.42 0.81
12.16 0.42
parenthesis)a
L-NMMA
1
2
3
4
5
6
7
3a
3a
3a
3a
3a
3a
3a
H2O2 (2 eq.), HCO2H, RT
H2O2 (5 eq.), HCO2H, RT
H2O2 (5 eq.), MeCN, RT – 40 °C
complex
decomposition
NRb
a
Inhibition of NO production.
L
-NMMA (NG-Monomethyl-
L
-arginine, monoac-
etate salt) was used as a positive control.
H2O2 (5 eq.), HCO2H/MeCN (10:1), RT trace
H2O2 (5 eq.), HCO2H/MeCN (2:1), RT
H2O2 (5 eq.), HCO2H/MeCN (1:1), RT
H2O2 (5 eq.), HCO2H/MeCN (1:1),
40 °C
trace
8 (12)
10 (15)
8d
3bc
3b
3b
3b
3b
3b
3b
3b
3b
3b
3b
3a
[Cu(MeCN)4](PF6) (0.05 eq.), DBED
(0.06 eq.), O2 (1 atm), CH2Cl2; H2SO4/
H2O (2:1); TMSCH2N2 (5 eq.)
[Cu(MeCN)4](PF6) (0.05 eq.), DBED
(0.06 eq.), O2 (1 atm), CH2Cl2; H2SO4/
H2O (2:1); TMSCH2N2 (5 eq.)
[Cu(MeCN)4](PF6) (0.10 eq.), DBED
(0.12 eq.), O2 (1 atm), CH2Cl2; H2SO4/
H2O (2:1); TMSCH2N2 (5 eq.)
[Cu(MeCN)4](PF6) (0.10 eq.), DBED
(0.12 eq.), O2 (1 atm), CH2Cl2; TfOH
(0.06 eq.), MeOH (1.05 eq.)
[Cu(MeCN)4](PF6) (0.10 eq.), DBED
(0.12 eq.), O2 (1 atm), 4 Å MS, CH2Cl2;
TfOH (0.3 eq.), MeOH (5 eq.)
CuCl (0.10 eq.), DBED (0.12 eq.), O2
(1 atm), 4 Å MS, CH2Cl2; TfOH
(0.3 eq.), MeOH (5 eq.)
CuI (0.10 eq.), DBED (0.12 eq.), O2
(1 atm), 4 Å MS, CH2Cl2; TfOH
(0.3 eq.), MeOH (5 eq.)
CuBr (0.10 eq.), DBED (0.12 eq.), O2
(1 atm), 4 Å MS, CH2Cl2; TfOH
(0.3 eq.), MeOH (5 eq.)
[Cu(MeCN)4](PF6) (0.10 eq.), TEA
(0.12 eq.), O2 (1 atm), 4 Å MS, CH2Cl2;
TfOH (0.3 eq.), MeOH (5 eq.)
[Cu(MeCN)4](PF6) (0.10 eq.), TMEDA
(0.12 eq.), O2 (1 atm), 4 Å MS, CH2Cl2;
TfOH (0.3 eq.), MeOH (5 eq.)
CuOTf (0.10 eq.), DBED (0.12 eq.), O2
(1 atm), 4 Å MS, CH2Cl2; TfOH
(0.3 eq.), MeOH (5 eq.)
5 (10)
8 (15)
40 (78)
36 (70)
42 (80)
trace
In conclusion,
a four-step biomimetic synthesis of hors-
fiequinone A (1) has been achieved using CuOTf/DBED catalyzed
aerobic oxidation as the key step. The concise synthetic route
may also be applicable to construct more complex DAP dimers
bearing the 1,4-p-quinone moiety. Importantly, revisiting the
anti-inflammatory activity of horsfiequinone A (1) provided evi-
dence for the folk use of H. kingii in treating various inflammation
related diseases.
9d,f
10e,f
11e,f
12e,f
13e,f
14e,f,g
15e,f,g
16e,f,g
17e,f,g
18e,f
19
Declaration of Competing Interest
The authors declare that they have no known competing finan-
cial interests or personal relationships that could have appeared
to influence the work reported in this paper.
complex
complex
trace
Acknowledgements
This research was partly supported by the National Natural
Science Foundation of China (grant numbers 81960631,
31860097, and 81803396), the Yunnan Fundamental Research Pro-
ject (202001AS070038), the Top Young Talent of Ten Thousand
Talents Program of Yunnan Province (R. Z and L.-D. S), the Start-
up fund of Yunnan University of Chinese Medicine (grant number
2019YZG03), and the Discipline Funding of School of Chinese
Materia Medica, Yunnan University of Chinese Medicine (grant
number 2020TD15).
NR
47 (92)
NR
CuOTf (0.10 eq.), DBED (0.12 eq.), O2
(1 atm), 4 Å MS, CH2Cl2; TfOH
(0.3 eq.), MeOH (5 eq.)
aIsolated yield; brsm: based on recovered starting material.
bNo reaction.
Appendix A. Supplementary data
cWhen 3b was used as the substrate, 1 was obtained after acidifying the dimeric
General procedures for the synthesis of 1, 1H and
C NMR
13
product 6 and the subsequent methylation of 1a (the route is shown in Scheme 1
and detailed in the ESI).
dCu(I)/DBED was added to substrate 3b.
eSubstrate 3b was added to Cu(I)/DBED.
fOne pot.
spectrum of synthetic compounds, and detailed procedures for bio-
logical experiments and molecular docking are available in the ESI.
Supplementary data to this article can be found online at
https://doi.org/10.1016/j.tetlet.2020.152756.
gRecovered 3b.
Fig. 2. Effects of 1 on LPS-induced iNOS expression in RAW264.7 cells. Values represent the mean
***/###P < 0.001).
SD of three independent experiments. (# vs control, * vs LPS,
3

M. Wang, Yuan-Lie Liu, D. Li et al.
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4