Total Synthesis of an Isatis indigotica-Derived Alkaloid Using a Biomimetic Thio-Diels-Alder Reaction

Article abstract of DOI:10.1021/acs.orglett.8b01321

A biomimetic thio-Diels-Alder reaction between a dienylthiadiazole and 3-thioisatin leads to the Isatis indigotica-derived alkaloid (1), along with its diastereomer 2. This synthetic study, supported by molecular modeling, establishes the viability of the proposed biosynthesis by thio-Diels-Alder cycloaddition, a very rare reaction in nature. Moreover, the results described infer that the diastereomer 2 is an as-yet undiscovered natural product present in Isatis indigotica.

Full text of DOI:10.1021/acs.orglett.8b01321

Letter
pubs.acs.org/OrgLett
Cite This: Org. Lett. XXXX, XXX, XXX−XXX
Total Synthesis of an Isatis indigotica-Derived Alkaloid Using a
Biomimetic Thio-Diels−Alder Reaction
Emma K. Davison, Paul A. Hume, and Jonathan Sperry*
School of Chemical Sciences, University of Auckland, 23 Symonds Street, Auckland, New Zealand
S
* Supporting Information
ABSTRACT: A biomimetic thio-Diels−Alder reaction be-
tween a dienylthiadiazole and 3-thioisatin leads to the Isatis
indigotica-derived alkaloid (1), along with its diastereomer 2.
This synthetic study, supported by molecular modeling,
establishes the viability of the proposed biosynthesis by thio-
Diels−Alder cycloaddition, a very rare reaction in nature.
Moreover, the results described infer that the diastereomer 2 is
an as-yet undiscovered natural product present in Isatis
indigotica.
Scheme 1. Proposed Biosynthesis of 1 and the Putative
Diastereomer 2
he herbaceous plant Isatis indigotica is cultivated
throughout China for its medicinal properties. The
T
dried roots and leaves, named “Ban Lan Gen” and “Da Qing
Ye” respectively, have been used in Chinese traditional
medicine for hundreds of years to treat a variety of viral
diseases, infections, and fevers.1 Owing to the medicinal
properties of these plant preparations, several of the natural
products present in Isatis indigotica have been isolated and
tested for biological activity. The compounds isolated include
lignans,2 flavonoids3 and a remarkable array of alkaloids,4
including the structurally remarkable natural product 1.5 This
unnamed alkaloid, isolated as a scalemic mixture (2:1) of 1a
and 1b from the roots of Isatis indigotica in 2012, comprises a
unique heterocyclic framework containing spirodihydro-
thiopyran-oxindole and 1,2,4-thiadiazole moieties (Figure 1).6
Interestingly, the diastereomer of the natural product,
compound 2, was not isolated in the original report.
Interestingly, the putative biosynthetic intermediate 7 was
recently isolated9 from Isatis indigotica and is subsequently
shown to comprise four stereoisomers 7a−d in a 2:4:1:2 ratio,
individually named as insatindigothiadiazoles A−D. The ratio of
7a−d strongly supports their proposed biosynthetic assembly
from the union of 5a−b and 6a−b, and, hence, the 2:1 mixture
of epiprogoitrin (3a) and progoitrin (3b). The insatindigo-
thiadiazoles 7a−d are then thought to undergo a selective
monodehydration to generate diene 8. The 3-thioisatin 9,
Figure 1. Alkaloid 1 and its diastereomer 2.
A detailed biosynthesis of the alkaloid 1 was proposed in the
original report (Scheme 1).5 The putative biosynthetic
precursors, the glucosinolates epiprogoitrin (3a) and progoitrin
(3b), are present in I. indigotica in a 2:1 ratio.7 Myrosinase-
catalyzed hydrolysis of 3a and 3b is known to liberate the
thiohydroximate-O-sulfonate 4,7 which in turn is reported to
breakdown into nitrile 5 and imidothioate 6.8 Heterocyclization
between 5a−b and 6a−b would form the 1,2,4-thiadiazole 7.
Received: April 26, 2018
© XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.8b01321
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
proposed to be derived from glucobrassicin (10),4 then
undergoes thio-Diels−Alder reaction with diene 8 followed
by double bond isomerization to give 1a and 1b. The ratio of
the naturally occurring enantiomers 1a and 1b (2:1) strongly
suggests that the 2‴ stereocenter is derived from the 2:1
mixture of glucosinolates 3a and 3b, but some uncertainty
surrounds the diastereoselectivity of the thio-Diels−Alder
cycloaddition. It was proposed in the original report that the
stereocenter in diene 8 is too distant from the spirocenter to
exert any diastereocontrol in this cycloaddition, and as a result,
the diastereomer 2 must also exist in Isatis indigotica
(presumably as a 2:1 ratio of 2a:2b).
An ongoing interest in thiadiazole natural products10 and
biomimetic synthesis11 led us to instigate a synthetic program
to test the biosynthetic hypothesis outlined in Scheme 1,
focusing on the thio-Diels−Alder reaction. To the best of our
knowledge, examples of thio-Diels−Alder reactions in nature
are limited to the assembly of vinyldithiins from thioacrolein in
Allium sativum.12,13
Figure 2. DFT computed Gibbs free energies and transition state
structures for thio-Diels−Alder reactions of 9 and 13.
Our preliminary investigations focused on using a model
diene in the biomimetic thio-Diels−Alder cycloaddition as a
means to examine the viability of this proposed reaction
(Scheme 2).14,15 Commercially available 3-bromo-5-chloro-
of these products were investigated using density functional
theory (DFT) calculations. The DFT calculations employed
the M06-2X exchange-correlation functional and polarized
triple-ζ 6-311+G (d,p) basis set. Solvent effects were taken into
account by employing a polarized continuum model. Further
details regarding computational methodology can be found in
the Supporting Information (SI).
Scheme 2. Model Thio-Diels−Alder Reaction
The DFT calculations support regioselective formation of 15
via intermediates ( )-16a and ( )-16b due to a free energy
difference of ∼6−7 kcal/mol between transition states TS1a/
TS1b and TS2a/TS2b. The difference in energy was attributed
to favorable orbital overlap in TS1a/TS1b, including significant
secondary orbital interactions between the π-systems of the
reactants. In contrast, these interactions are not present in
transition states TS2a/TS2b. The frontier orbital energies of 9
(HOMO = −8.0 eV, LUMO = −2.8 eV) and 13 (HOMO =
−8.1 eV, LUMO = −1.7 eV) indicate that the reaction
proceeds via a normal Diels−Alder cycloaddition, as opposed
to an inverse electron-demand pathway.19 However, it should
be noted that the difference between the LUMOdienophile
−
HOMOdiene and LUMOdiene−HOMOdienophile gaps is ∼1 eV, so
both interactions are likely to play a role in the reaction.20 This
assessment is supported by the fact that the transition state
frontier molecular orbitals do not resemble a simple linear
combination of the HOMO of 13 and the LUMO of 9.
1,2,4-thiadiazole (11) underwent C5-selective Suzuki cou-
pling16 with the dienyl boronate ester 1217 to give the
somewhat unstable diene 13 as the model substrate for the
thio-Diels−Alder cycloaddition.18 The dienophile 9 was also
very unstable and was prepared in situ. Thus, 2-oxindole (14)
was treated with N-chlorosulfenylsuccinimide (succNSCl)15
and triethylamine to form an intermediate sulfenamide, at
which point the diene 13 was added. The dienophile 9 was
generated upon addition of pyridine, and the reaction mixture
was stirred for 15 h at room temperature. A single product was
formed, identified as the spirodihydrothiopyran-oxindole
( )-15 by X-ray crystallography. This model study was
considered a success; the cycloaddition had proceeded with
the desired regioselectivity and the initial cycloadduct (i.e., 16)
underwent isomerization in situ, as would be required for the
eventual synthesis of the natural product.
With the model study successful, we turned our attention to
the natural product itself. Our strategy was modeled on the
proposed biosynthesis outlined in Scheme 1, and as such, we
targeted the selective dehydration of the insatindigothiadiazole
natural products (7) as a means to access the key diene 8
(Scheme 3). Silylation of known hydroxyamide ( )-1821 gave
( )-19 which was converted to the thioamide ( )-20 using
Lawesson’s reagent (LR). Oxidative dimerization of ( )-20
using iodosobenzene diacetate22 followed by desilylation gave
an inseperable mixture of insatindigothiadiazoles A−D (7a−d,
1:1:1:1), the NMR spectroscopic data of which was in excellent
agreement with that in the isolation report.9 Next, the selective
dehydration was attempted; upon treatment of diol mixture
7a−d with 1 equiv of methanesulfonyl chloride and 2 equiv of
base, a single product was formed, which was identified as the
mesylate ( )-21. As only 1 equiv of mesyl chloride was used in
this process, we posit that the diols 7a−d undergo selective
mesylation−elimination to give the desired product ( )-8,
which itself is mesylated by the liberated methanesulfonic acid23
In principle, the cycloaddition between 9 and 13 could lead
to four compounds that correspond to the endo- and exo-
adducts of each “ortho” (16a/b) and “meta” regiosiomer (17a/
b), respectively (Figure 2). In order to gain further insight into
this key reaction, the thio-Diels−Alder reactions leading to each
B
DOI: 10.1021/acs.orglett.8b01321
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
and hence explain why it was not detected in the original
report. It is possible that the thio-Diels−Alder cycloaddition of
8 and 9 is under enzymatic control, thus favoring the formation
of 1 over 2. The reaction could also follow a stepwise, conjugate
addition pathway (perhaps facilitated by a metal ion) that
favors the diastereomer 1.
Scheme 3. Synthesis of Diene ( )-8 via
Insatindigothiadiazoles (7a−d)
To conclude, the biomimetic synthesis of a structurally
unprecedented alkaloid isolated from Isatis indigotica is
described. A thio-Diels−Alder reaction between insatindigo-
thiadiazole-derived diene ( )-8 and the 3-thioisatin 9 gave the
natural product ( )-1, along with its diastereomer ( )-2, in a
1:1 ratio. The regiochemical outcome and lack of diaster-
eocontrol in this cycloaddition is supported by molecular
modeling, and as such, it is probable that diastereomer 2 is also
present in Isatis indigotica, as a scalemic mixture of 2a and 2b
(2:1). Moreover, the realization of the original biosynthesis
proposal represents a very rare example of a thio-Diels−Alder
reaction occurring in nature.
(from the elimination) to give ( )-21. This side reaction was of
little consequence, as the mesylate ( )-21 was readily
converted to the desired alcohol ( )-8 upon stirring in
aqueous tetrahydrofuran. Distinctive HMBC correlations from
both protons of the methylene group to the thiadiazole C3
confirmed the diene component had been successfully installed
at the desired C5 position.24
ASSOCIATED CONTENT
■
S
* Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.or-
glett.8b01321.
With the diene ( )-8 in hand, attention was focused toward
the key biomimetic thio-Diels−Alder reaction (Scheme 4). In
Full experimental procedures and NMR spectra of all
novel compounds; full details of the computational
methodology employed (PDF)
Scheme 4. Biomimetic Thio-Diels−Alder Reaction
Accession Codes
CCDC 1825337 contains 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 data_
request@ccdc.cam.ac.uk, or by contacting The Cambridge
Crystallographic Data Centre, 12 Union Road, Cambridge CB2
1EZ, UK; fax: +44 1223 336033.
AUTHOR INFORMATION
■
Corresponding Author
*E-mail: j.sperry@auckland.ac.nz.
ORCID
Jonathan Sperry: 0000-0001-7288-3939
Notes
line with the model study, the diene ( )-8 and the dienophile 9
were stirred together at room temperature. After ∼3 days, a 1:1
mixture of the natural product ( )-1 and its diastereomer
( )-2 was formed; the absence of diastereocontrol in this
biomimetic thio-Diels−Alder reaction corroborates the postu-
late5 that the diastereomer 2 is also likely to be present in Isatis
indigotica. This assessment is further supported by DFT
modeling of the thio-Diels−Alder reaction between diene 8
and dienophile 9 (SI). In line with the model study (Scheme
2), the regioselectivity of the cycloaddition between 8 and 9 is
controlled by stabilizing π−π interactions in the transition state.
However, the free energy difference between the transition
states leading to 1 and 2 was calculated to be <0.6 kcal/mol,
within the accuracy limit of the computational method
employed. Thus, it appears that the formation of both
diastereomers 1 and 2 is equally likely at ambient temperature,
which is in line with our experimental results and the proposal
in the original report. However, it is important to note that, in
nature, there are other factors influencing the reaction that may
preclude the production of diastereomer 2 in Isatis indigotica
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We thank the University of Auckland for the award of a
doctoral scholarship (E.K.D.) and the Royal Society of New
Zealand for a Rutherford Discovery Fellowship (J.S.)
REFERENCES
■
(1) (a) Zheng, H. Z.; Dong, Z. H.; Yu, Q. Modern Study of Traditional
Chinese Medicine; Xueyuan Press: Beijing, 1997; Vol. 1, pp 328−334.
(b) An, Y. Q.; Jia, X. B.; Yuan, H. J.; Chen, Y.; Jin, X. Y. Chin. Tradit.
Herb. Drugs 2008, 39, 616. (c) Peng, S. P.; Gu, Z. L. Chin. Wild Plant
Resour. 2005, 24, 4.
(2) (a) Li, B.; Chen, W. S.; Zhao, Y.; Zhang, H. M.; Dong, J. X.; Qiao,
C. Z. Chin. Tradit. Herb. Drugs 2005, 36, 326. (b) He, L. W.; Li, X.;
Chen, J. W.; Sun, D. D.; Ju, W. Z.; Wang, K. C. Acta Pharm. Sin. 2006,
41, 1193. (c) Zuo, L.; Li, J. B.; Xu, J.; Yang, J. Z.; Zhang, D. M.; Tong,
Y. L. Chin. J. Chin. Mater. Med. 2007, 32, 688.
C
DOI: 10.1021/acs.orglett.8b01321
Org. Lett. XXXX, XXX, XXX−XXX

Organic Letters
Letter
(3) (a) He, Y.; Lu, J.; Lin, R.-C. Chin. Tradit. Herb. Drugs 2003, 34,
777. (b) Liu, J. F.; Zhang, X. M.; Xue, D. Q.; Jiang, Z. Y.; Gu, Q.;
Chen, J. J. Chin. J. Chin. Mater. Med. 2006, 31, 1961.
Trans. 1 1992, 1261. (d) Kirby, G. W.; Lochead, A. W.; Williamson, S.
J. Chem. Soc., Perkin Trans. 1 1996, 977. (e) Breu, J.; Hocht, P.; Rohr,
̈
U.; Schatz, J.; Sauer, J. Eur. J. Org. Chem. 1998, 1998, 2861. (f) Vedejs,
E.; Galante, R. J.; Goekjian, P. G. J. Am. Chem. Soc. 1998, 120, 3613.
(g) Prabhakaran, J.; Lhermitte, H.; Das, J.; Sasi-Kumar, T. K.;
Grierson, D. S. Synlett 2000, 658. (h) Heras, M.; Gulea, M.; Masson, S.
Chem. Commun. 2001, 611. (i) Pfund, E.; Lequeux, T.; Vazeux, M.;
Masson, S. Tetrahedron Lett. 2002, 43, 2033. (j) Heras, M.; Gulea, M.;
Masson, S.; Philouze, C. Eur. J. Org. Chem. 2004, 2004, 160.
(4) (a) Wu, X.; Liu, Y.; Sheng, W.; Sun, J.; Qin, G. Planta Med. 1997,
63, 55. (b) Wei, X. Y.; Leung, C. Y.; Wong, C. K. C.; Shen, X. L.;
Wong, R. N. S.; Cai, Z. W.; Mak, N. K. J. Nat. Prod. 2005, 68, 427.
(c) Liu, J.-F.; Jiang, Z.-Y.; Wang, R.-R.; Zheng, Y.-T.; Chen, J.-J.;
Zhang, X.-M.; Ma, Y.-B. Org. Lett. 2007, 9, 4127. (d) Wu, Y.; Zhang,
Z.-X.; Hu, H.; Li, D.; Qiu, G.; Hu, X.; He, X. Fitoterapia 2011, 82, 288.
(e) Chen, M.; Gan, L.; Lin, S.; Wang, X.; Li, L.; Li, Y.; Zhu, C.; Wang,
Y.; Jiang, B.; Jiang, J.; Yang, Y.; Shi, J. J. Nat. Prod. 2012, 75, 1167.
(f) Yang, L.; Wang, G.; Wang, M.; Jiang, H.; Chen, L.; Zhao, F.; Qiu,
F. Fitoterapia 2014, 95, 175. (g) Liu, Y.-F.; Chen, M.-H.; Guo, Q.-L.;
Lin, S.; Xu, C.-B.; Jiang, Y.-P.; Li, Y.-H.; Jiang, J.-D.; Shi, J.-G. J. Asian
Nat. Prod. Res. 2015, 17, 689. (h) Liu, Y.-F.; Chen, M.-H.; Lin, S.; Li,
Y.-H.; Zhang, D.; Jiang, J.-D.; Shi, J.-G. J. Asian Nat. Prod. Res. 2016,
18, 1. (i) Meng, L.; Guo, Q.; Liu, Y.; Chen, M.; Li, Y.; Jiang, J.; Shi, J.
Acta Pharm. Sin. B 2017, 7, 334. (j) Meng, L.-J.; Guo, Q.-L.; Xu, C.-B.;
Zhu, C.-G.; Liu, Y.-F.; Chen, M.-H.; Lin, S.; Li, Y.-H.; Jiang, J.-D.; Shi,
J.-G. J. Asian Nat. Prod. Res. 2017, 19, 529. (k) Liu, S.-F.; Zhang, Y.-Y.;
Zhou, L.; Lin, B.; Huang, X.-X.; Wang, X.-B.; Song, S.-J. Phytochemistry
2018, 149, 132.
(5) Chen, M.; Lin, S.; Li, L.; Zhu, C.; Wang, X.; Wang, Y.; Jiang, B.;
Wang, S.; Li, Y.; Jiang, J.; Shi, J. Org. Lett. 2012, 14, 5668.
(6) For 1,2,4-thiadiazole natural products, see: (a) Heitz, S.; Durgeat,
M.; Guyot, M.; Brassy, C.; Bachet, B. Tetrahedron Lett. 1980, 21, 1457.
(b) Pham, C.-D.; Weber, H.; Hartmann, R.; Wray, V.; Lin, W.; Lai, D.;
Proksch, P. Org. Lett. 2013, 15, 2230. (c) Yang, Z.; Huang, N.; Xu, B.;
Huang, W.; Xie, T.; Cheng, F.; Zou, K. Molecules 2016, 21, 232.
(7) Xie, Z.; Shi, Y.; Wang, Z.; Wang, R.; Li, Y. J. Agric. Food Chem.
2011, 59, 12467.
́
(k) Denance, M.; Legay, R.; Gaumont, A.-C.; Gulea, M. Tetrahedron
Lett. 2008, 49, 4329. (l) Watanabe, Y.; Sakakibara, T. Tetrahedron
2009, 65, 599. (m) Murai, T.; Morikawa, K.; Maruyama, T. Chem. -
Eur. J. 2013, 19, 13112.
(19) Zhao, Y.; Truhlar, D. G. Theor. Chem. Acc. 2008, 120, 215.
(20) Sustmann, R. Pure Appl. Chem. 1974, 40, 569.
(21) Knapp, S.; Gibson, F. S. J. Org. Chem. 1992, 57, 4802.
(22) Yan, M.; Chen, Z.-C.; Zheng, Q.-G. J. Chem. Res. 2003, 2003,
618.
(23) For mesylations using methanesulfonic acid, see: (a) Ashby, J.;
Griffiths, D.; Paton, D. J. Heterocycl. Chem. 1978, 15, 1009.
(b) Lapitskaya, M. A.; Demin, P. M.; Zatonsky, G. V.; Pivnitsky, K.
K. Russ. Chem. Bull. 2004, 53, 2617.
(24) In the 13C NMR spectrum of 1,2,4-thiadiazoles; the signal for
C3 appears 10−15 ppm further upfield than C5. Cameron, T. S.;
Decken, A.; Fang, M.; Parsons, S.; Passmore, J.; Wood, D. J. Chem.
Commun. 1999, 1801. Also refer to the Supporting Information of ref
6b.
(8) Polat, U. J. Biol. Environ. Sci. 2010, 4, 39.
(9) Chen, M.-H.; Lin, S.; Wang, Y.-N.; Zhu, C.-G.; Li, Y.-H.; Jiang, J.-
D.; Shi, J.-G. Chin. Chem. Lett. 2016, 27, 643.
(10) Davison, E. K.; Sperry, J. Org. Chem. Front. 2016, 3, 38.
(11) (a) Wang, C.; Sperry, J. Tetrahedron 2014, 70, 3430. (b) Boyd,
E. M.; Sperry, J. Org. Lett. 2015, 17, 1344. (c) Calvert, M. B.; Sperry, J.
Chem. Commun. 2015, 51, 6202.
(12) (a) Block, E.; Iyer, R.; Grisoni, S.; Saha, C.; Belman, S.; Lossing,
F. P. J. Am. Chem. Soc. 1988, 110, 7813. (b) Block, E.; Ahmad, S.;
Catalfamo, J. L.; Jain, M. K.; Apitz-Castro, R. J. Am. Chem. Soc. 1986,
108, 7045. (c) Nohara, T.; Kiyota, Y.; Sakamoto, T.; Manabe, H.; Ono,
M.; Ikeda, T.; Fujiwara, Y.; Nakano, D.; Kinjo, J. Chem. Pharm. Bull.
2012, 60, 747.
(13) For reviews on Diels−Alder reactions in natural product
biosynthesis, see: (a) Stocking, E. M.; Williams, R. M. Angew. Chem.,
Int. Ed. 2003, 42, 3078. (b) Oikawa, H.; Tokiwano, T. Nat. Prod. Rep.
2004, 21, 321. (c) Minami, A.; Oikawa, H. J. Antibiot. 2016, 69, 500.
(14) For the synthesis of spirothiopyran-oxindoles, see: (a) Wang, S.;
Jiang, Y.; Wu, S.; Dong, G.; Miao, Z.; Zhang, W.; Sheng, C. Org. Lett.
2016, 18, 1028. (b) Ji, C.; Wang, S.; Chen, S.; He, S.; Jiang, Y.; Miao,
Z.; Li, J.; Sheng, C. Bioorg. Med. Chem. 2017, 25, 5268. (c) Wang, S.;
Chen, S.; Guo, Z.; He, S.; Zhang, F.; Liu, X.; Chen, W.; Zhang, S.;
Sheng, C. Org. Biomol. Chem. 2018, 16, 625. (d) Wang, S.; Guo, Z.;
Chen, S.; Jiang, Y.; Zhang, F.; Liu, X.; Chen, W.; Sheng, C. Chem. -
Eur. J. 2018, 24, 62.
(15) For the synthesis of a spirodihydrothiopyran-oxindole by thio-
Diels−Alder reaction with a symmetrical diene, see: Shermolovich, Y.
G.; Emets, S. V.; Tolmachev, A. A. Chem. Heterocycl. Compd. 2003, 39,
1076.
(16) Wehn, P. M.; Harrington, P. E.; Eksterowicz, J. E. Org. Lett.
2009, 11, 5666.
(17) Coombs, J. R.; Zhang, L.; Morken, J. P. Org. Lett. 2015, 17,
1708.
(18) For the use of thiocarbonyls as dienophiles in the Diels−Alder
reaction, see: (a) Vedejs, E.; Perry, D. A.; Houck, K. N.; Rondan, N. G.
J. Am. Chem. Soc. 1983, 105, 6999. (b) Katada, T.; Eguchi, S.; Sasaki,
T. J. Org. Chem. 1986, 51, 314. (c) Adam, D.; Freer, A. A.; Isaacs, N.
W.; Kirby, G. W.; Littlejohn, A.; Rahman, M. S. J. Chem. Soc., Perkin
D
DOI: 10.1021/acs.orglett.8b01321
Org. Lett. XXXX, XXX, XXX−XXX