Syntheses of benzophenone-xanthone hybrid polyketides and their antibacterial activities

Article abstract of DOI:10.1016/j.bmcl.2017.04.017

Muchimangins are benzophenone-xanthone hybrid polyketides produced by Securidaca longepedunculata. However, their biological activities have not been fully investigated, since they are minor constituents in this plant. To evaluate the possibility of muchimangins as antibacterial agent candidates, five muchimangin analogs were synthesized from 2,4,5-trimethoxydiphenyl methanol and the corresponding xanthones, by utilizing p-toluenesulfonic acid monohydrate for the Br?nsted acid-catalysis. The antibacterial assays against Gram-positive bacteria, Staphylococcus aureus and Bacillus subtilis, and Gram-negative bacteria, Klebsiella pneumoniae and Escherichia coli, revealed that the muchimangin analogs (±)-1,3,6,8-tetrahydroxy-4-(phenyl-(2′,4′,5′-trimethoxyphenyl)methyl)-xanthone (1), (±)-1,3,6-trihydroxy-4-(phenyl-(2′,4′,5′-trimethoxyphenyl)methyl)-xanthone (2), and (±)-1,3-dihydroxy-4-(phenyl-(2′,4′,5′-trimethoxyphenyl)methyl)-xanthone (3) showed significant activities against S. aureus, with MIC values of 10.0, 10.0, and 25.0?μM, respectively. Analogs (±)-1 and (±)-2 also exhibited antibacterial activities against B. subtilis, with MIC values of 50.0 and 12.5?μM, respectively. Furthermore, (+)-3 enhanced the antibacterial activity against S. aureus, with a MIC value of 10?μM.

Full text of DOI:10.1016/j.bmcl.2017.04.017

Accepted Manuscript
Syntheses of benzophenone-xanthone hybrid polyketides and their antibacterial
activities
Takeshi Kodama, Takuya Ito, Dya Fita Dibwe, So-Yeun Woo, Hiroyuki Morita
PII:
S0960-894X(17)30379-7
DOI:
http://dx.doi.org/10.1016/j.bmcl.2017.04.017
BMCL 24870
Reference:
To appear in:
Bioorganic & Medicinal Chemistry Letters
Received Date:
Revised Date:
Accepted Date:
16 February 2017
22 February 2017
5 April 2017
Please cite this article as: Kodama, T., Ito, T., Dibwe, D.F., Woo, S-Y., Morita, H., Syntheses of benzophenone-
xanthone hybrid polyketides and their antibacterial activities, Bioorganic & Medicinal Chemistry Letters (2017),
doi: http://dx.doi.org/10.1016/j.bmcl.2017.04.017
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers
we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and
review of the resulting proof before it is published in its final form. Please note that during the production process
errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Syntheses of benzophenone-xanthone hybrid polyketides and their
antibacterial activities
a
a, *
a
a
a, *
Takeshi Kodama , Takuya Ito , Dya Fita Dibwe , So-Yeun Woo , Hiroyuki Morita
a
Institute of Natural Medicine, University of Toyama, 2630-Sugitani, Toyama 930-0194, Japan
To whom correspondence should be addressed:
Hiroyuki Morita, Institute of Natural Medicine, University of Toyama, 2630-Sugitani, Toyama, 930-
0194, Japan; Tel: +81-76-434-7625, Fax: +81-76-434-5059; E-mail: hmorita@inm.u-toyama.ac.jp.
Takuya Ito, Institute of Natural Medicine, University of Toyama, 2630-Sugitani, Toyama, 930-0194,
Japan; Tel: +81-76-434-7627, Fax: +81-76-434-5059; E-mail: itot@inm.u-toyama.ac.jp.
1

Abstract
Muchimangins are benzophenone-xanthone hybrid polyketides produced by Securidaca longepedunculata.
However, their biological activities have not been fully investigated, since they are minor constituents
in this plant. To evaluate the possibility of muchimangins as antibacterial agent candidates, five
muchimangin analogs were synthesized from 2,4,5-trimethoxydiphenyl methanol and the corresponding
xanthones, by utilizing p-toluenesulfonic acid monohydrate for the Brønsted acid-catalysis. The
antibacterial assays against Gram-positive bacteria, Staphylococcus aureus and Bacillus subtilis, and
Gram-negative bacteria, Klebsiella pneumoniae and Escherichia coli, revealed that the muchimangin
analogs (±)-1,3,6,8-tetrahydroxy-4-(phenyl-(2',4',5'-trimethoxyphenyl)methyl)-xanthone (1), (±)-1,3,6-
trihydroxy-4-(phenyl-(2',4',5'-trimethoxyphenyl)methyl)-xanthone (2), and (±)-1,3-dihydroxy-4-
(
phenyl-(2',4',5'-trimethoxyphenyl)methyl)-xanthone (3) showed significant activities against S. aureus,
with MIC values of 10.0, 10.0, and 25.0 M, respectively. Analogs (±)-1 and (±)-2 also exhibited
antibacterial activities against B. subtilis, with MIC values of 50.0 and 12.5 M, respectively.
Furthermore, (+)-3 enhanced the antibacterial activity against S. aureus, with a MIC value of 10 M.
Keywords: muchimangin, benzophenone-xanthone hybrid polyketide, Brønsted acid-catalysis,
antibacterial activity
2

Xanthone and benzophenone dimers are a widespread and structurally diverse family of
1,2
natural products frequently found in plants, fungi, and lichens. These natural product dimers possess
2,3
antimicrobial activity. Dicerandrols A-C are examples of the xanthone dimers produced by
Phomopsis longicolla, with antibacterial activities against the Gram-positive bacteria Staphylococcus
4
aureus and Bacillus subtilis. In contrast, microsphaerin A is an example of a benzophenone dimer
5
derived from Microsphaeropsis sp. with antibacterial activity against S. aureus. Interestingly, the
benzophenone dimer, phomalevone B isolated from Phoma sp., reportedly displayed a broad range of
6
antimicrobial activities against S. aureus, B. subtilis, Escherichia coli, and Candida albicans.
However, the benzophenone monomer lacks these antimicrobial activities.
Muchimangins are benzophenone-xanthone hybrid polyketides isolated from the roots of the
7
-11
medicinal plant, Securidaca longepedunculata Fresen (Polygalaceae)
(Fig. 1), used in traditional
Congolese medicine. These muchimangins are structurally divided into two types, the C-2 and C-4
diphenylmethyl xanthones. Among them, muchimangin B, a C-2 type diphenylmethyl xanthone, has
7
antiausteric activity against the human pancreatic cancer PANC-1 cell line. However, since
muchimangins are minor components in S. longepedunculata, their other biological activities have
remained unclear. In our ongoing research for the discovery of antibacterial agents, we synthesized
new five muchimangin analogs, 1,3,6,8-tetrahydroxy-4-(phenyl-(2',4',5'-trimethoxyphenyl)methyl)-
Fig. 1 Examples of the structures of muchimangins isolated from Securidaca longepedunculata.
3

xanthone (1), 1,3,6-trihydroxy-4-(phenyl-(2',4',5'-trimethoxyphenyl)methyl)-xanthone (2), 1,3-
dihydroxy-4-(phenyl-(2',4',5'-trimethoxyphenyl)methyl)-xanthone (3), 1,3-dihydroxy-6,8-dimethoxy-
4
-(phenyl-(2',4',5'-trimethoxyphenyl)methyl)-xanthone (4), and 1-hydroxy-3,6,8-trimethoxy-4-
(
phenyl-(2',4',5'-trimethoxyphenyl)-xanthone (5), and found that (±)-1 and (±)-2 display antibacterial
activities against S. aureus and B. subtilis, whereas (±)-3 shows selective antibacterial activity against
S. aureus. Herein, we report the syntheses of these analogs and their detailed antibacterial activities, as
well as the structure activity relationships of the analogs against S. aureus and B. subtilis.
Scheme 1. Syntheses of muchimangin analogs 1−5.
4

The synthetic route for the muchimangin analogs is outlined in Scheme 1. Commercially
available 1,2,4-trihydroxybenzene (6) and dimethyl sulfate were coupled in the present of sodium
hydride to give 1,2,4-trimethoxybenzene. Subsequently, the acylated 1,2,4-trimethoxybenzene was
treated with benzoyl chloride and aluminium chloride in dichloromethane, to produce 2,4,5-
12
trimethoxybenzophenone (7) in 96% yield. Compound 7 was reduced by sodium borohydride in
acetonitrile, to furnish part of the muchimangin analog skeleton, 2,4,5-trimethoxydiphenyl methanol
13
(
8), in 77% yield.
14
14
15
The 1,3,6,8-tetrahydroxy (9)-, 1,3,6-trihydroxy (10)-, 1,3-dihydroxy (11), and 1,3-
16
dihydroxy-6,8-dimethoxy (12) -xanthones were synthesized from the corresponding salicylic acid and
phenol in phosphorus pentoxide-methanesulfonic acid (Eaton reagent, v/v = 1:10) at 80 C, according
17
to the published method. Compounds 8 (1.0 equiv.) and 9 (1.0 equiv.) were coupled by Brønsted
acid-catalyzed nucleophilic substitution, using p-toluenesulfonic acid monohydrate (TsOH·H O, 0.4
2
18
equiv.) in acetonitrile, to obtain (±)-1 as racemic mixture in 66% yield. The structure of (±)-1 was
elucidated on the basis of 1D and 2D NMR, and HR-ESI-MS analyses. The HMBC correlations from
1
-OH ( 12.10, s) to C-1 ( 161.2), C-2 ( 100.0), and C-8b ( 102.4), and from H-7' ( 6.27, s) to
H
C
C
C
H
C-3 ( 162.73), C-4 ( 108.1), and C-4a ( 154.7) confirmed the connectivity between C-7' in the
C
C
C
diphenylmethyl unit and C-4 in the xanthone unit. The muchimangin analogs (±)-2, (±)-3, and (±)-4,
with C-7' of the diphenylmethyl unit linked to C-4 of the xanthone skeleton, were also prepared from
the corresponding xanthones 10, 11, and 12 and the diphenyl methanol 8, respectively, using the same
procedure as that for the synthesis of 1. The yields of (±)-2, (±)-3, and (±)-4 were 38%, 35%, and 40%,
respectively. In addition, the muchimangin analog (±)-5 was synthesized from 1-hydroxy-3,6,8-
trimethylxanthone (13) and the diphenyl methanol 8 in 42% yield, using the same procedure as that for
(
±)-1, after 9 was methylated with dimethyl sulfate in tetrahydrofuran to give 13. Despite the use of
TsOH·H O, which is reported to be an effective Brønsted acid catalyst for the formation of carbon-
2
18-20
carbon and carbon-heteroatom bonds,
none of the C-2 diphenyl methyl substituted muchimangin
analogs was produced in these preparations. Presumably, the steric repulsion between the C-1 and C-3
5

substitutions of the xanthones prevented the nucleophilic substitution of the diphenyl methyl unit to
the C-2 of the xanthone scaffold.
The in vitro antibacterial activities of the synthesized muchimangin analogs (±)-1−5, as well
as the synthetic intermediates 6−13, were assessed against two Gram-positive bacteria, S. aureus
NBRC 100910 and B. subtilis NBRC 13719, and two Gram-negative bacteria, Klebsiella pneumoniae
2
1-24
NBRC 14940 and E. coli NBRC 102203 (Table 1).
As in the case of the reported benzophenone
monomer, the benzophenone monomer 7 did not display any antibacterial activities against the tested
strains. Furthermore, all compounds were inactive against the tested Gram-negative bacteria. In
contrast, (±)-1 showed antibacterial activity against the Gram-positive bacterium, S. aureus, with a
MIC value of 10.0 M. However, the antibacterial activity of the corresponding xanthone 9 against S.
aureus was 5 times weaker than that of (±)-1. The muchimangin analogs (±)-2 and (±)-3 also displayed
antibacterial activities against S. aureus, with MIC values of 10.0 M and 25.0 M, respectively.
Compounds 10 and 11, the corresponding xanthone skeletons of (±)-2 and (±)-3, respectively, showed
weak antibacterial activities against S. aureus, in contrast to those of (±)-2 and (±)-3. Furthermore, the
muchimangin analogs (±)-4 and (±)-5 lacked antibacterial activity against S. aureus. In the case of the
antibacterial activity against B. subtilis, the muchimangin analog (±)-2 showed the highest activity
with a MIC value of 12.5 M, among the tested compounds. In addition, the muchimangin analog (±)-
1
displayed weak antibacterial activity. However, the muchimangin analogs (±)-3, (±)-4, and (±)-5
lacked antibacterial activity against B. subtilis. Furthermore, 9−13, corresponding to the xanthone
skeletons of (±)-3, (±)-4, and (±)-5, also did not show antibacterial activity against this strain.
Thus, the antibacterial assays revealed that the benzophenone-xanthone hybrid structure
enhanced the antibacterial activities. Furthermore, the structure activity relationship analysis of the
tested muchimangin analogs suggested that the presence of the hydroxy group at C-6 would be
important for the growth inhibitory activities against S. aureus and B. subtilis. To further clarify the
enantiomeric efficacy of the antibacterial activities of 1, 2, and 3, the potent and active muchimangin
Table 1. Antibacterial activities of muchimangin analogs 1−5 and synthetic intermediates 6−13.
6

Compound
MIC (M)
Compound
MIC (M)
S. aureus
S. aureus
B. subtilis
50.0
B. subtilis
>100
>100
>100
>100
>100
>100
>100
>100
1.25
(±)-1
(+)-1
(−)-1
(±)-2
(+)-2
(−)-2
(±)-3
(+)-3
(−)-3
(±)-4
(±)-5
10.0
10.0
12.5
10.0
10.0
10.0
25.0
10.0
50.0
>100
>100
6
7
>100
>100
>100
50.0
100
50.0
100
8
12.5
9
10.0
10
11
12
13
12.5
100
>100
>100
>100
>100
>100
>100
>100
1.25
1.25
a
Ampicillin
Kanamycin
a
1.25
a
Positive control.
analogs (±)-1, (±)-2, and (±)-3 were separated on a chiral stationary phase column [CHIRALPAK-AD-
H, 10 mm × 250 mm (Daicel); detection: UV 340 nm; flow rate: 3.0 mL/min; column temperature:
40 °C], using hexane/ethanol/acetic acid as an eluent to furnish each enantiomer, and their
antibacterial activities were evaluated against S. aureus and B. subtilis. The enantiomers (+)-1 and (−)-
2
4
1
were eluted at retention times of 29.1 min and 20.7 min and showed optical rotations of [α]D +25
(
c 0.02 CHCl ) and −28 (c 0.02 CHCl ), respectively. The muchimangin analogs (±)-2 and (±)-3 were
3
3
also separated using the same procedure, to furnish the corresponding enantiomers (+)-2, (−)-2, (+)-3,
and (−)-3. The antibacterial assays revealed that both enantiomers (+)-1 and (−)-1 display antibacterial
activities against S. aureus at levels comparable to that of (±)-1. In the case of the antibacterial assay
against B. subtilis, (+)-1 exhibited the same MIC value as that of (±)-1, while the activity was
significantly decreased with (−)-1. In contrast, the enantiomers (+)-2 and (−)-2 retained the same
efficacy as that of (±)-2 against both strains. However, the antibacterial activity of (+)-3 against S.
aureus was 2.5 and 5 times higher than those of (±)-3 and (−)-3, respectively, although the
enantiomers lacked antibacterial activity against B. subtilis, as in the case of (±)-3. Thus, (+)-3
possessed the highest activity against S. aureus, followed by (±)-1 and (±)-2, suggesting the
importance of the stereo-differentiation for the further development of antibacterial muchimangin
7

analogs.
In conclusion, we demonstrated the antibacterial activities of the muchimangin analogs, as
well as the syntheses of the C-4 benzophenone-substituted xanthones, using the corresponding
salicylic acids and phloroglucinols. Further evaluations of the structure activity relationships of the
muchimangin analogs might contribute to the development of structurally interesting anti-S. aureus
and anti-B. subtilis agents.
Acknowledgement
This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of
Education, Culture, Sports, Science and Technology, Japan (T.I. and H.M.).
A. Supplementary material
Supplementary data associated with this article can be found, in the online version, at XXX.
References and Notes
1
2
3
4
.
.
.
.
Masters, K. S.; Bräse, S. Chem. Rev. 2012, 112, 3717.
Wu, S. B.; Long, C.; Kennelly, E. J. Nat. Prod. Rep. 2014, 31, 1158.
Wezeman, T.; Bräse, S.; Masters, K. S. Nat. Prod. Rep. 2015, 32, 6.
Chaesung, L.; Kim, J.; Choi, J. N.; Ponnusamy, K.; Jeon, Y.; Kim, S. U.; Kim, J. G.; Lee, C. H.
J. Microbiol. Biotechnol. 2010, 20, 494.
5.
Yoganathan, K.; Cao, S.; Crasta, S. C.; Aitipamula, S.; Whitton, S. R.; Ng, S.; Buss, A. D.;
Butler, M. S. Tetrahedron 2008, 64, 10181.
6
7
8
.
.
.
Shim, S. H.; Baltrusaitis, J.; Gloer, J. B.; Wicklow, D. T. J. Nat. Prod. 2011, 74, 395.
Dibwe, D. F.; Awale, S.; Kadota, S.; Tezuka, Y. Tetrahedron Lett. 2012, 53, 6186.
Dibwe, D. F.; Awale, S.; Kadota, S.; Morita, H.; Tezuka, Y. Bioorg. Med. Chem. 2013, 21,
7663.
8

9
1
.
Dibwe, D. F.; Awale, S.; Kadota, S.; Morita, H.; Tezuka, Y. Tetrahedron Lett. 2014, 55, 1916.
Dibwe, D. F.; Awale, S.; Kadota, S.; Morita, H.; Tezuka, Y. J. Nat. Prod. 2014, 77, 1241.
Dibwe, D. F.; Awale, S.; Kadota, S.; Morita, H.; Tezuka, Y. Nat. Prod. Commun. 2014, 9, 655.
Kamino, S.; Ichikawa, H.; Wada, S.; Horio, Y.; Usami, Y.; Yamaguchi, T.; Koda, T.; Harada,
A.; Shimanuki, K.; Arimoto, M.; Doi, M.; Fujita, Y. Bioorg. Med. Chem. Lett. 2008, 18, 4380.
Waterlot, C.; Couturier, D.; Backer, M. D.; Rigo, B. Can. J. Chem. 2000, 78, 1242.
Liu, Y.; Zou, L.; Ma, L.; Chen, W. H.; Wang, B.; Xu, Z. L. Bioorg. Med. Chem. 2006, 14, 5683.
Zhou, T.; Shi, Q.; Chen, C. H.; Huang, L.; Ho, P.; Morris-Natschke, S. L.; Lee, K. H. Eur. J.
Med. Chem. 2012, 47, 86.
0.
11.
1
2.
1
1
1
3.
4.
5.
1
1
1
1
6.
7.
8.
9.
Brahmachari, G.; Gorai, D.; Mondal, S.; Gangopadhyay, A.; Chatterjee, D. J. Chem. Res.
Synopses 2003, 362.
Kiang Lim, C.; Tho, L. Y.; Hoe Lim, C.; Mooi Lim, Y.; Adnan Ali Shah, S.; Faizal Weber, J. F.
Lett. Drug. Des. Discov. 2001, 8, 523.
Sanz, R.; Martínez, A.; Miguel, D.; Álvarez-Gutiérrez, J. M.; Rodríguez, F. Adv. Synth. Catal.
2
006, 348, 1841.
Urgaonkar, S.; La Pierre, H. S.; Meir, I.; Lund, H.; RayChaudhuri, D.; Shaw, J. T. Org. Lett.
005, 7, 5609.
2
2
2
0.
1.
Iovel, I.; Mertins, K.; Kischel, J.; Zapf, A.; Beller, M. Angew. Chem. Int. Ed. 2005, 44, 3913.
Antimicrobial Assay: The microdilution method described by Eloff was used to determine the
minimum inhibitory concentration (MIC) values for the synthetic compounds against bacteria,
with some modifications. The respective MIC values of the samples were determined using
two Gram-positive bacteria, S. aureus NBRC 100910 and B. subtilis NBRC 13719, and two
Gram-negative bacteria, Klebsiella pneumoniae NBRC 14940 and E. coli NBRC 102203.
Briefly, 2 µL of pure compounds (1 mM) in duplicate, dissolved in dimethyl sulfoxide (DMSO,
concentration less than 5%), were added to 198 µL of bacterial culture in YP medium (1%
polypeptone, 0.2% yeast extract, and 0.1% MgSO ·7H O) and serially diluted two-fold in 96-
4
2
9

6
well microtiter plates. The densities of the bacterial cultures were approximately 1.5 × 10
cfu/mL, obtained by dilution (1:1,000) from the overnight bacterial cultures in YP medium.
The plates were incubated overnight at 37 °C, and the MICs were determined as the lowest
concentration inhibiting microbial growth. The plates were further incubated at 37 °C
overnight. Ampicillin and kanamycin were used as reference reagents for the bacterial strains.
Eloff, J. N. Planta Medica 1998, 64, 711.
2
2
2
2.
3.
4.
Nisa, K.; Ito, T.; Subehan; Matsui, T.; Kodama, T.; Morita, H. Phytochem. Lett. 2016, 15, 42.
Awouafack, M. D.; Ito, T.; Tane, P.; Kodama, T.; Tanaka, M.; Asakawa, Y.; Morita, H.
Phytochem. Lett. 2016, 15, 220.
1
0

1
1