15-Hydroxygermacranolides as Sources of Structural Diversity: Synthesis of Sesquiterpene Lactones by Cyclization and Rearrangement Reactions. Experimental and DFT Study

Article abstract of DOI:10.1021/acs.joc.8b00407

A study on the electrophile-induced rearrangement of two 15-hydroxygermacranolides, salonitenolide and artemisiifolin, was carried out. These compounds underwent electrophilic intramolecular cyclizations or acid-mediated rearrangements to give sesquiterpene lactones with different skeletons such as eudesmanolides, guaianolides, amorphanolides, or other germacranolides. The cyclization that gives guaianolides can be considered a biomimetic route to this type of sesquiterpene lactones. The use of acetone as a solvent changes the reactivity of the two starting germacranolides to the acid catalysts, with a 4,15-diol acetonide being the main product obtained. The δ-amorphenolide obtained by intramolecular cyclization of this acetonide is a valuable intermediate for accessing the antimalarials artemisinin and its derivatives. Mechanistic proposals for the transformations are raised, and to provide support them, quantum chemical calculations [DFT B3LYP/6-31+G(d,p) level] were undertaken.

Full text of DOI:10.1021/acs.joc.8b00407

Subscriber access provided by Kaohsiung Medical University
1
5-Hydroxygermacranolides as Sources of Structural Diversity.
Synthesis of Sesquiterpene Lactones by Cyclization and
Rearrangement Reactions. Experimental and DFT Study
José María Álvarez-Calero, Enrique Ruiz, José Luis López-Pérez, Martin Jaraiz, José
E. Rubio, Zacarias D. Jorge, Margarita Suarez, and Guillermo Martínez Massanet
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.8b00407 • Publication Date (Web): 25 Apr 2018
Downloaded from http://pubs.acs.org on April 25, 2018
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a service to the research community to expedite the dissemination
of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in
full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully
peer reviewed, but should not be considered the official version of record. They are citable by the
Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore,
the “Just Accepted” Web site may not include all articles that will be published in the journal. After
a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web
site and published as an ASAP article. Note that technical editing may introduce minor changes
to the manuscript text and/or graphics which could affect content, and all legal disclaimers and
ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or
consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W.,
Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
1
5-Hydroxygermacranolides as Sources of Structural Diversity. Synthesis of
Sesquiterpene Lactones by Cyclization and Rearrangement Reactions. Experimental
and DFT Study
†
‡,§
, ,
∥ ⊥
#
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
José María ÁlvarezꢀCalero, Enrique Ruiz, José Luis LópezꢀPérez,* Martín Jaraíz, José
#
†
§
,†
E. Rubio, Zacarías D. Jorge, Margarita Suárez, and Guillermo M. Massanet*
†
Departamento de Química Orgánica, Facultad de Ciencias, Universidad de Cádiz, 11510
Puerto Real, Cádiz, Spain
‡
Departamento de Química, Instituto de Ciencias Básicas, Universidad Técnica de Manabí
(
UTM), Avenida Urbina y Che Guevara, 130103 Portoviejo, Manabí, Ecuador
§
Laboratorio de Síntesis Orgánica, Facultad de Química, Universidad de La Habana, 10400
La Habana, Cuba
∥Departamento de Farmacología, Facultad de Medicina, Universidad de Panamá, República
de Panamá
⊥Departamento de Ciencias Farmacéuticas, IBSALꢀCIETUS, Universidad de Salamanca,
Avda. Campo Charro s/n, 37007 Salamanca, Spain
#
Departamento de Electrónica, Universidad de Valladolid, Paseo Belén 15, 47011 Valladolid,
Spain
*g.martinez@uca.es
*lopez@usal.es
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 2 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
ABSTRACT: A study on the electrophileꢀinduced rearrangement of two 15ꢀ
hydroxygermacranolides, salonitenolide and artemisiifolin, was carried out. These compounds
underwent electrophilic intramolecular cyclizations or acidꢀmediated rearrangements to give
sesquiterpene lactones with different skeletons such as eudesmanolides, guaianolides,
amorphanolides or other germacranolides. The cyclization that gives guaianolides can be
considered as a biomimetic route to this type of sesquiterpene lactones. The use of acetone as
solvent changes the reactivity of the two starting germacranolides to the acid catalysts, with a
4
,15ꢀdiol acetonide being the main product obtained. The δꢀamorphenolide obtained by
intramolecular cyclization of this acetonide is a valuable intermediate for accessing the
antimalarials artemisinin and its derivatives. Mechanistic proposals for the transformations are
raised, and to provide support them, quantum chemical calculations [DFT B3LYP/6ꢀ
3
1+G(d,p) level] were undertaken.
INTRODUCTION
Sesquiterpene lactones are a large group of plant metabolites mainly isolated from numerous
genera of the family Asteraceae (Compositae) that possess a broad spectrum of biological
activities such as antiꢀinflammatory, antibacterial, antifungal, phytotoxic, cytotoxic, and
1
anticancer. In recent years, the anticancer properties shown by sesquiterpene lactones have
attracted a great deal of interest, and extensive research has been carried out to elucidate their
ACS Paragon Plus Environment

Page 3 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
molecular mechanism of action as well as their potential use as chemopreventive and
2
chemotherapeutic agents. Some sesquiterpene lactones with anticancer potential are
3
parthenolide (germacranolide isolated from Tanacetum parthenium), thapsigargin
4
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(
guaianolide isolated from Thapsia garganica), artemisinin (endoperoxide, mainly known for
5
its antimalarial activity, isolated from Artemisia annua), helenalin (pseudoguaianolide
6
isolated from Psilostrophe cooperi), arglabin (guaianolide isolated from Artemisia
7
8
myriantha), or alantolactone (eudesmanolide isolated from Inula helenium).
Germacranolides are an abundant subset within sesquiterpene lactones which are considered
key biosynthetic precursors for other sesquiterpene skeletons such as elemanolides,
9
guaianolides and eudesmanolides. The major structural feature of this type of sesquiterpene
lactones is the trans,trans ring skeleton, trans,transꢀcyclodecaꢀ1(10),4ꢀdiene, which is well
known for its ability to undergo conformational changes that have been the subject to
1
0
theoretical studies. This conformational flexibility plays an important role in the reactivity
of these compounds, particularly in thermal, photochemical and acidic transannular
11
cyclizations, since the spatial arrangement of the different conformers determines the
9
b,12
outcome of the reactions.
Taking into account their biosynthetic interest, numerous germacranolides have been used as
13
models for synthesizing different skeletal types of sesquiterpenolides (Scheme 1a). These
reactions can be considered as a biomimetic synthesis of this type of secondary metabolites
and have been used by our group to propose a route for the biogenesis of sesquiterpenolides
1
3b
produced by plants of the Umbelliferae family. Most of the works on the mechanism of
cyclization are based on establishing the relationships between the conformation of the initial
germacranolide and the stereochemistry of the reaction products, but very few ones provide
reliable evidences of the mechanisms that lead to these products. In our efforts to develop new
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 4 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
synthetic applications of germacranolides via transannular cyclization and provide proofs
about the mechanisms, we herein present the biomimetic synthesis of several sesquiterpene
lactones starting from the 15ꢀhydroxygermacranolides salonitenolide and artemisiifolin
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(Scheme 1b). A computational study with DFT at B3LYP/6ꢀ31+G(d,p) level was conducted
to justify the formation of compounds described.
ACS Paragon Plus Environment

Page 5 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Scheme 1. Reactivity of Germacranolides
a) Reported reactivity of germacranolides
TsOH
CH Cl , rt, 1 h
2
2
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
H
H
O
O
O
O
O
O
O
O
H
TsOH
CH Cl , rt, 1 h
2
2
O
HO
H
HO
H
O
O
O
O
OH
OH
CHCl3
60 ºC, 5 h
H
OHC
O
OHC
O
O
O
b) This work
OH
O
O
O
OH
HO
HO
O
Salonitenolide
Artemisiifolin
R
H
O
OH
O
O
OR
O
O
H
H
H
O
OHC
OH
O
OHC
O
O
R = H, Ms
R = H, OH
O
O
O
R
OHC
O
O
O
H
H
R = OH, Cl
OH
O
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 6 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
RESULTS AND DISCUSSION
The presence of an oxygenated function at C15 confers a characteristic reactivity to the
germacranolides and makes them valuable starting materials for the hemisynthesis of different
1
3a,13c,13d
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
skeletons of sesquiterpene lactones.
With the purpose of increasing the potentiality of these compounds and their derivatives to
create structural diversity, we decided to explore their chemical behavior against different
acids and other electrophilic reagents. For this purpose, we selected as starting materials
14
15
salonitenolide (2) and artemisiifolin (3), two 15ꢀhydroxygermacranolides which can be
1
3a,13c
obtained in good yield from cnicin (1),
a germacranolide easily accessible from species
1
6
of Centaurea genus, (Figure 1). Both compounds have been prepared from cnicin (1)
although slight modifications of the reported methods have been introduced. Additionally
10b
artemisiifolin (3) has been isolated from Staehelina dubia (see Experimental Section).
Figure 1. Cnicin (1), salonitenolide (2), and artemisiifolin (3).
The transannular cyclizations were initiated with the treatment of salonitenolide (2) with silica
sulfuric acid (SSA) (Table 1, entry 1), a nontoxic heterogeneous solid acid catalyst that
1
7
presents great chemical versatility. The recent reported effectiveness of SSA to promote
1
8
threeꢀcomponent condensation reactions encouraged us for studying the reactivity of this
heterogeneous protic acid with germacranolides. Treatment of 2 with SSA in CHCl led to the
3
formation of stoebenolide (4) and its C4 epimer, 5, in only 15 min (Table 1, entry 1).
1
9
Stoebenolide (4) has been isolated from Centaurea stoebe, and synthesized by transannular
ACS Paragon Plus Environment

Page 7 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
1
3d
cyclization of 2 promoted by palladium(II). Both compounds were also obtained when 2
was treated with pꢀtoluenesulfonic acid (TsOH) in CHCl for 1 h (Table 1, entry 2),
3
demonstrating the great catalytic activity of the heterogeneous protic system.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Table 1. Reactivity of Salonitenolide (2)
electrophilic
reagent
a
entry
1
solvent
time
products(%)
SSA
CHCl3
15 min
(
1.0 equiv)
TsOH·H O
2
2
CHCl3
CH Cl
1 h
(
1.0 equiv)
b
MCPBA
3
1.5 h
2
2
(
1.0 equiv)
c
MsCl
2.0 equiv)
4
CH Cl
2 d
2
2
(
(
SSA
0.9 equiv)
5
6
Acetone
Acetone
3 d
6 d
TsOH·H O
1.0 equiv)
2
(
a
b
c
d
Isolated yield. K CO (3.7 equiv) used as base. Et N (6.0 equiv) used as base. The relative ratio
was established from H NMR (500 MHz) analysis of reaction crude.
2
3
3
1
Treatment of 2 with MCPBA in the presence of anhydrous K CO , to minimize mꢀ
2
3
chlorobenzoic acid generated during the reaction, gave the eudesmanolides 6 and its C4
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 8 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
epimer, 7, (Table 1, entry 3). 8αꢀHydroxyꢀ4ꢀepiꢀsonchucarpolide (6) is an antifungal
2
0
compound that has been isolated from different Centaurea species. The epimeric compound,
2
0c
21
7, has been found in Centaurea zuccariniana and Onopordum cynarocephalum, and has
not been previously synthesized. Eudesmanolide 7 induces apoptotic cell death in the A375
human melanoma cell line and a high DNA fragmentation correlated to a significant increase
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
1
of the caspaseꢀ3 enzyme activity. The same type of cyclization to give aldehydes has been
1
3a,22
described for other 1,10ꢀepoxyꢀ15ꢀhydroxygermacranolides.
23
Formation of 4 can be derived from the UU conformation of 2 through C1ꢀC10 double bond
protonation, cyclodecadiene cyclization, and hydride shift (Scheme 2). Aldehyde 5 should
come from slow epimerization at C4 via enol, promoted by the acidic medium. This fact was
confirmed by transformation of aldehyde 4 to the epimer 5 by its treatment with SSA in
CH Cl (see Experimental Section).
2
2
ACS Paragon Plus Environment

Page 9 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Scheme 2. Proposed Mechanism for the Formation of Aldehydes 4 and 5*
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
*
NH4 and NH have been considered in calculations to emulate catalytic effects of the different acids used and
3
+
their conjugated bases generated during the reaction. Free energies relative to 2+NH +NH [B3LYP/6ꢀ
4
3
3
1+G(d,p) scrf=(smd, solvent=chloroform)//B3LYP/6ꢀ31+g(d,p)] in kcal/mol in blue, green and red. Selected
+
distances in TSs are shown in Å. To clarify the image, NH and NH4 of reagent, intermediate and products
3
included in the calculation have been deleted from the image.
To justify this finding a computational study was undertaken. A conjugate base moiety and
the solvent effect (chloroform) have been considered in the calculations by a single point
2
4
energy calculations using the SMD continuum model. The geometries and binding energies
for the carbocationꢀammonia complex analyzed by the B3LYP/6ꢀ31+G* method gave
25
satisfactory results. Compound 5, with the aldehyde group in equatorial disposition, turns
out to be 3.8 kcal/mol more stable than 4, which seems to disagree with the formation of a
larger ratio of 4.
This prompted us to study the formation path of both compounds, the results of which are
shown in Scheme 2. The process is initiated by the protonation of the C1ꢀC10 double bond of
2
3
the conformer with lower energy (UU). As a result, the carbocation at C10 triggers the
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 10 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
cyclization until the carbocation is stabilized on C4, producing the eudesmanolide cation
intermediate, 2a (See C.1.1 in Supporting Information). Subsequent 1,2ꢀhydride shift across
both faces of the C4ꢀC15 bond provides 4 and 5 after deprotonation catalyzed by the
conjugate base formed during the reaction. To overcome the last step, carbocation 2a must
rotate the C4ꢀC15 bond to achieve an orthogonal arrangement for each of the methylene
hydrogens that will lead in each case to compounds 4 and 5. The interconversion barrier of
carbocation 2a through the 1,2ꢀsuprafacial shift conducting to 5 is more than 2 kcal/mol
higher than its alternative, which is consistent with compound 4 being formed in higher ratio
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(
See C.1.2 and C.1.3 in Supporting Information).
In a similar manner, compound 6 comes from the transient and nonꢀisolable C1ꢀC10 epoxide
See Scheme S2 in Supporting Information). The reaction is initiated by the protonation of the
(
oxirane ring followed by an intramolecular cyclization that leads to the 2c intermediate
carbocation in a concerted manner through a 12.2 kcal/mol barrier (See C.2.1 in Supporting
Information). As we have described for the formation of 4 and 5, intermediate 2c evolves to 6
and 7 by a hydrogen shift over both sides of the C4ꢀC15 bond followed by deprotonation. The
barrier for the formation of 7 results to be 1.1 kcal/mol higher than the one leading to 6, which
would justify the higher formation ratio of 6 with respect to 7 (See C.2.2 and C.2.3 in
Supporting Information).
In order to explore the behavior of germacranolides which carry a C15 leaving group,
salonitenolide (2) was treated with mesyl chloride. In these conditions the cyclization occurs
spontaneously and goes in the opposite direction to that described above, to give the
26
unexpected natural guaianolide deacylsubexpinnatin (8) in 70% yield, and the mesylate 9 as
a minor product (Table 1, entry 4). The formation of the guaianolides comes from the
conformation UU of 2, through the cyclization shown in Scheme 3.
ACS Paragon Plus Environment

Page 11 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Scheme 3. Cyclization of Salonitenolide (2) to Give Guaianolides*
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
*
Free energies relative to 2Ms+Me NH [B3LYP/6ꢀ31+G(d,p) scrf=(smd, solvent=dichloromethane)//
3
B3LYP/6ꢀ31+g(d,p)] in kcal/mol in blue. Selected distances in TSs are shown in Å. Me N included in the
calculations has been eliminated for simplification of the picture. (For complete results of calculations see
Supporting Information: C.3.1 and C.3.2).
3
The catalytic role of amine present in the medium significantly lowers the transition state
barriers. IRC calculations indicate that the beginning of mesylate oxygenꢀC15 bond breaking
is favored by the approximation of the basic amine hydrogen to another mesylate oxygen
surmounting a first barrier of 13 kcal/mol. This enables the carbocation to swiftly move, after
overcoming a second small barrier of 1.4 kcal/mol, through 1,5ꢀcyclization until the
carbocation is located at C10. Simultaneously, the elimination of H14 takes place through its
trapping by the mesylate anion, thus completing the reaction (See C.3.1 and C.3.2 in
Supporting Information). When the dgdzvp2 basis set is used instead off 6ꢀ31+g(d,p), an
asynchronous concerted process is observed with only one saddle point between 2Ms and 8.
Simultaneous presence of guaianolides and 15ꢀhydroxygermacranolides in plants of the
1g,27
Compositae and Magnoliaceae families
might be explained through this type of
cyclization. Noteworthy is the isolation of picriside B (10) from Saussurea lappa, a plant on
2
8
which dehydrocostuslactone (11) is the main component (Figure 2). Therefore, these
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 12 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
transformations can be considered as an alternative way to guaianolide biosynthesis. González
29
et al. described a similar transformation of the germacranolide gallicin to give guaianolides.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Figure 2. Picriside B (10) and dehydrocostuslactone (11).
3
0
The influence of solvents over conformational equilibria is well known. For this reason, we
decided to study the electrophilic cyclizations of salonitenolide (2) using acetone as solvent.
The reaction with SSA or TsOH in this solvent (Table 1, entries 5 and 6), in addition to the
formation of acetonide 12, an evolution/inversion in the ratio 4/5 can be observed, which
would be in accordance with the results of the computational studies shown in Scheme 2,
since a longer time required for the consumption of starting material would imply a greater
evolution in the equilibrium of aldehyde 4 towards the thermodynamically more stable 5.
1
H NMR (500 MHz) spectrum in C D of 12 at rt shows the presence of three interconverting
6
6
conformers with ratio 74:22:3 (See Supporting Information). These conformational equilibria
were corroborated by NOESYꢀ1D since irradiating the proton signals of the major conformer,
the saturation of the frequency was transferred to the corresponding proton signals of the
other two conformers by the exchange process (see Supporting Information). This
3
1
phenomenon has also been observed by Faraldos et al., who have used it as a very useful
tool to assign the conformers of (+)ꢀgermacrene A in solution. The major conformer and the
intermediate one of 12 exhibit parallel UU orientation wherein the conformational movement
occurs at C2/C3, adopting chairꢀchair and boatꢀchair conformations, respectively. On the
other hand, the minor conformer presents crossed DU orientation adopting boatꢀchair
ACS Paragon Plus Environment

Page 13 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
conformation. Germacrene D, a sesquiterpene with germacraꢀ1(10),5ꢀdiene core also shows in
32
solution this conformational movement around C2/C3, and the corresponding conformers
3
3
have been previously predicted by computational calculations. Similar behavior have been
34
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
revealed for germacraꢀ1(10),5ꢀdienꢀ4ꢀols by molecular mechanic calculations, or reported
35
36
for other described sesquiterpenes and, even the natural diterpene obscuronatin. To better
understand the complexity of the conformational equilibrium observed for compound 12 in
NMR studies, a total conformational search with Molecular Mechanics force field MMFF
(Merck Pharmaceuticals) was carried out using the Mixed torsional/Lowꢀmode sampling
3
7
method. 1000 conformations were generated, and those presenting up to 10 kcal/mol above
the overall minimum were selected. Subsequently, an optimization at B3LYP/6ꢀ31+G(d,p)
level was performed with the lower energy conformers. Finally, the four lower energy
conformers were selected (See C.4 in Supporting Information) and optimized considering
benzene as solvent using the SMD continuum model to better reproduce experimental
conditions. In agreement with experimental NMR observations, very small energy differences
have been found between the three lower energy conformers (less than 0.5 kcal/mol) while the
fourth turns out to be more than 3 kcal/mol less stable. The three lower energy conformers
found in the conformational search correspond perfectly to those derived from NMR studies
including NOE experiments.
The unexpected formation of 12 from salonitenolide (2) implies relactonization from C6 to C8
via allylic rearrangement that must be promoted by acetone since, in the absence of this
solvent this transformation does not take place. The reaction starts (Scheme 4) with the
formation of the hemiacetal 12a through a barrier of 11.2 kcal/mol (See C.5.1 in Supporting
Information). Subsequently, in a concerted process, the formation of acetonide moiety of
intermediate 12b is produced via allylic rearrangement and nucleophilic opening of the
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 14 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
lactone ring, favored by protonation of the lactone carbonyl (See C.5.2 in Supporting
Information). Similar allylic rearrangement along with the formation of an acid has been
3
6
reported in diterpenes. Finally, relactonization affords acetonide 12.
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Scheme 4. Proposed Mechanism for the Formation of Acetonide 12*
+
*
NH4 and NH have been considered in calculations to emulate catalytic effects of the different acids used and
3
+
their conjugated bases during the reaction. Free energies relative to 2+Me CO+NH +NH [B3LYP/6ꢀ31+G(d,p)
2
4
3
scrf=(smd, solvent=acetone)//B3LYP/6ꢀ31+g(d,p)] in kcal/mol in blue. Selected distances in TSs are shown in
Å.
When acetonide 12 was subjected to acid treatment with TsOH in chloroform, cadalane 13
(Figure 3) was obtained in 69% yield. The configuration at C6 was inferred from NOE
experiments (See Supporting Information). The key NOE between H6 and H7 indicates that
both hydrogens are in cis disposition over the alpha side of the molecule, and therefore the
skeleton of the compound 13 corresponds to that of a δꢀamorphene.
14
2
10
1
3
4
9
8
6
7
15
5
O
12
H
11
1
3
13
O
Figure 3. Amorphene 13.
Amorphanes (relative configuration, H1α,H6α,H7α) are a sesquiterpene subgroup belonging
3
8
to the family of cadalanes, which have been isolated from different biological sources. The
ACS Paragon Plus Environment

Page 15 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
mechanism for the biosynthesis of amorphanes, and their other possible diasteromers
(bulgaranes, H1β,H6α,H7α; cadinanes, H1α,H6β,H7α; and muurolanes, H1β,H6β,H7α),
3
9
from farnesyl pyrophosphate (FPP) catalyzed by sesquiterpene cyclases is still unclear.
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Three possible mechanistic routes are proposed for the biosynthesis (Scheme 5). Path A
implies the (E,E)ꢀgermacrenꢀ6ꢀyl cation, derived from 1,10ꢀcyclization of FPP, which is
converted into germacrene D followed by transannular 1,6ꢀcyclization to afford the different
cadalane cations. Route B differs from the above in that the (E,Z)ꢀgermacrenꢀ6ꢀyl cation
intermediate has a structure suitable for undergoing direct transannular 1,6ꢀcyclization to
cadalenes. The third path is based on the formation of the bisabolꢀ6ꢀyl cation by 1,6ꢀ
cyclization followed by 6,7ꢀcyclization to give the cadalanꢀ11ꢀyl cation. Successive 1,3ꢀ
hydride shifts lead to the corresponding skeletons of cadalanes.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 16 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Scheme 5. Mechanistic Routes Proposed for the Biosynthesis of Cadalanes from FPP
Catalyzed by Sesquiterpene Cyclases
H
Path A
Path C
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1,10-Cyclization
1,6-Cyclization
PPO
Bisabol-6-yl
cation
(E,E)-Germacren-6-yl cation
FPP
-
H
6
,7-Cyclization
Path B
1,10-Cyclization
H
H
Germacrene D
Cadalan-11-yl
cation
1,3-H ~
H
H
(
E,Z)-Germacren-6-yl
cation
Cadalan-6-yl
cation
1,6-Cyclization
1,6-Cyclization
1,3-H ~
H
H
H
H
H
H
H
H
Amorphan-10-yl
cation
Bulgaran-10-yl
cation
Muurolan-10-yl
cation
Cadinan-10-yl
cation
Subsequently, mechanistic insights about the formation of amorphadiene and amorphenes
40
have been obtained using density functional calculations. The routes via bisabolyl and (E,Z)ꢀ
germacrenyl cations (paths B and C, Scheme 5) were examined, concluding that the first one
is the most accessible energetically, although both paths converge as a result of a low energy
1
,5ꢀhydride transfer involving a nonꢀclassical carbocation.
3
2
While transannular 1,6ꢀcyclizations of germacrene D to cadalanes have been investigated,
such cyclization studies for other germacrenes are scarce. (–)ꢀGermacrene D or (–)ꢀ
ACS Paragon Plus Environment

Page 17 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
isogermacrene D under acidic conditions produced different sesquiterpenes among which
32
cadalanes of the cadinane, muurolane and amorphane types were found. Treatment of
(1E,4S,5E,7R)ꢀ4,11ꢀdihydroxygermacraꢀ1(10),5ꢀdiene with protic acids only produced
4
1
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
cadinanes, whereas treatment of 15ꢀdeoxyꢀ8ꢀepiꢀsalonitenolide acetate with methanolic
42
potassium followed by acidification afforded a bulgarane skeleton. A mixture of δꢀ
amorphene and δꢀcadinene in 2:3 ratio was isolated by heating 14ꢀacetoxyꢀ15ꢀ
43
deoxyartemisiifolin in the presence of acid traces.
Theoretical calculations of the acidꢀcatalyzed cyclization of germacrene D have established
that relative ratios of cadinane and muurolane sesquiterpenoids found in essential oil
compositions as well as the experimental cyclization of germacrene D, reflect the energetic
44
differences of the sesquiterpenoids and their carbocation precursors.
Taking into account the above precedents, a computational study was undertaken to clarify
the mechanism of formation of δꢀamorphene 13 (Scheme 6). After a detailed study of the
different spatial arrangements of acetonide 12, the only productive conformation capable of
evolving towards δꢀamorphene 13 is the UU conformation in which the olefinic H6 is in alpha
orientation. This conformation is 2.9 kcal/mol less stable than the minimum energy
conformation (12M) due to the eclipsed disposition of H6 and H7. The acidic medium
triggers the cleavage of the acetonide moiety giving the allylic carbocation 13a (See C.6.1 in
Supporting Information). Transannular 1,6ꢀcyclization affords the amorphane skeleton that
evolves to the δꢀamorphene 13 by deprotonation across low energy barriers (See C.6.2 and
C.6.3 in Supporting Information).
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 18 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Scheme 6. Proposed Mechanism for the Formation of δ-Amorphene Lactone 13*
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
*
NH4 and NH have been considered in calculations to emulate catalytic effects of used TsOH and its
3
+
conjugated base generated during the reaction. Free energies relative to 12+NH4 [B3LYP/6ꢀ31+G(d,p)] in
kcal/mol in blue. Selected distances in TSs are shown in Å.
Artemisiifolin (3) (Figure 1), the C8ꢀlactonized isomer of salonitenolide (2), exists as a
mixture of four slowly interconverting conformers with different ratios of conformers
1
0b
(UD:UU:DU:DD) in CDCl (60:15:14:11) and Me COꢀd (29:30:27:14). Due to the fact
3
2
6
that this flexibility could play a fundamental role in reactivity, we decided to explore the
chemical behavior of artemisiifolin (3) in transannular cyclizations (Table 2), by subjecting
this compound to the reactions described above for salonitenolide (2) (Table 1).
ACS Paragon Plus Environment

Page 19 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Table 2. Reactivity of Artemisiifolin (3)
electrophilic
reagent
a
entry
1
solvent
CHCl3
time
2 h
products(%)
SSA
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(
1.1 equiv)
TsOH·H O
(1.0 equiv)
2
2
CHCl3
2 d
HO
O
OHC
5 (28%)
O
O
H
1
13 (30%)
O
b
MsCl
1.0 equiv)
c
3
CH Cl
24 h
24 h
2
2
(
d
MsCl
2.4 equiv)
c
4
CH Cl
2
2
(
(
SSA
1.2 equiv)
5
6
Acetone 2.5 d
TsOH·H O
1.0 equiv)
2
Acetone
3 d
(
a
b
c
Isolated yield. Et N (1.3 equiv) used as base. Additionally, the reaction crude was solved in
3
d
CDCl and kept for 3 d at rt. Et N (6.4 equiv) used as base.
3
3
When 3 was treated with SSA in chloroform, aldehyde 14 was the only compound detected
Table 2, entry 1). It seems evident that the reaction proceeds through the UU conformation in
(
a similar way to the cyclization of salonitenolide (2) (Table 1, entry 1). However, the reaction
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 20 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
of 3 with TsOH in the same halogenated solvent was slower, 2 days, from which the
45
amorphene lactone 13 and the natural heliangolide liabinolide (15) were obtained (Table 2,
entry 2).
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
The formation of heliangolide 15 (Scheme 7) involves the formation of the allylic cation 15a
(See C.7.1 in Supporting Information), and a 1,2ꢀhydride shift favored by the presence of the
tosylate anion in the medium, which takes the proton of the alcohol at C15 giving the
unsaturated β,γꢀaldehyde intermediate 15b with the aldehyde in equatorial disposition (See
C.7.2 in Supporting Information). This step, across a barrier of 13.3 kcal/mol, is
thermodynamically favorable by more than 15 kcal/mol. In addition, pꢀtoluenesulfonic acid
catalyzes a ketoꢀenol tautomerism to give the 15c intermediate (See C.7.3 in Supporting
Information). The acidꢀpromoted isomerization of butꢀ2ꢀenꢀ1,4ꢀdiol units into α,βꢀunsaturated
4
6
aldehydes has already been reported in the literature. Finally, catalyzed once again by pꢀ
toluenesulfonic acid, the enol 15c evolves to give an α,βꢀunsaturated aldehyde by protonation
at C6, thus giving heliangolide 15 (See C.7.4 in Supporting Information). As can be seen in
the diagram, the overall process is thermodynamically favorable by 25 kcal/mol, and only the
enolization from 15b to 15c shows a barrier that, although high, is surmountable and could be
the cause of the slowness of the process since it takes place in 2 days. The production of
heliangolide 15 as the only diasteromer would be supported by the fact that germacranolides,
bearing an aldehyde group at C15, under acidic conditions, change from trans to cis the
10a,13c
configuration of the C4ꢀC5 double bond.
ACS Paragon Plus Environment

Page 21 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Scheme 7. Proposed Mechanism for the Formation of Heliangolide 15*
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
*
In accordance with experimental conditions, TsOH has been considered in the calculation. Me NH has also
3
+
been considered. Free energies relative to 3+TsOH+Me NH
[B3LYP/6ꢀ31+G(d,p), scrf=(smd,
3
solvent=chloroform)//B3LYP/6ꢀ31+g(d,p)] in kcal/mol in blue. Selected distances in TSs are shown in Å.
Obtaining δꢀamorphene 13 by acid treatment of both artemisiifolin (3) and acetonide 12
suggests the formation of a common intermediate during both transformations. In the case of
artemisiifolin, the reaction mechanism (Scheme 8) begins with the loss of water, giving the
allylic carbocation 15a (also involved in the formation of heliangolide 15), that, after a
conformational change over the partial double bond C5ꢀC6, evolves to the common key
intermediate 13a. Transannular 1,6ꢀcyclization and deprotonation give the final product 13.
This mechanism is supported by the computational results described in Schemes 6 and 7.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 22 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Scheme 8. Proposed Mechanism for the Formation of δ-Amorphene 13
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
The differences in reactivity and reaction times observed in the treatments of artemisiifolin (3)
with SSA and TsOH in CHCl (Table 2, entries 1 and 2), demonstrate the importance of the
3
heterogeneous nature of the catalyst.
Cyclization via epoxidation of artemisiifolin (3) has been previously described by Bruno et
1
3a
al.
Of the two epoxides obtained by these authors, only the compound with UU
conformation experienced cyclization in acid medium to give guaianolides (Figure 4). This
can be explained taking into account that the spatial arrangement of the C1ꢀC10 double bond
10f,12b,47
and the epoxide is the “crossed” one which is the appropriate for the cyclization.
Figure 4. (4R,5S)ꢀEpoxyartemisiifolin.
In order to explore if the reaction with sulfonyl chlorides to give guaianolides is a common
behaviour of 15ꢀhydroxygermacranolides, artemisiifolin (3) was treated with 1.0 equiv of
1
mesyl chloride (Table 2, entry 3). The H NMR spectrum of the reaction crude in CDCl3
showed the presence of different mesylated compounds. With the aim of leaving these labile
ACS Paragon Plus Environment

Page 23 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
species to evolve, the reaction crude was kept in the deuterated solvent for 3 d at rt. After
chromatographic separation, guaianolide 16 and cadalane 17 were obtained in low yields.
Compound 16 is an example of C8ꢀlactonized guaianolide that has not been reported before.
1
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
The absolute configuration of C5 was perfectly established by H NMR wherein the spatial
disposition anti between the protons H5 and H6 with J = 8.7 Hz was observed. NOE
experiments by irradiating H5, H6, H7 and H8 corroborated the absolute configuration of
guaianolide 16.
The initial step in the formation of compound 16 from the UU conformation of 15ꢀ
mesylartemisiifolin, through 1,5ꢀcyclization until the carbocation is located at C10 (See C.8.1
in Supporting Information), takes place in the same way as the formation of guaianolide 8
described above in Scheme 3. However, the deprotonation leading to the final product takes
place at C1 in this case (See C.8.2 in Supporting Information), rather than in C14 as occurs in
the formation of compound 8. This deprotonation is a consequence of the approximation of
the mesylate anion generated during the reaction by the alpha face of the sesquiterpenoid
instead of approximation by beta face which yields 8. These finding clearly indicate that the
global reaction is not a concerted process, as corroborated by the computational studies
carried out for guaianolide 8 (See C.3 in Supporting Information).
In order to justify the regioselective deprotonation that leads to guaianolides 8 and 16,
additional computational studies were undertaken (Scheme 9). The deprotonation at C14 that
gives 8 is clearly favored respect to the elimination of H1 that would yield a hypothetical
guaianolide with an endocyclic double bond. In fact, the barrier for the former is more than 7
kcal/mol lower (See C.3.2 and C.3.3 in Supporting Information and Scheme 9), while both
compounds exhibit similar thermodynamic stability (See C.3.2.2 and C.3.3.3 in Supporting
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 24 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Information). For the formation of guaianolide 16 and its hypothetical isomer with an
exocyclic double bond derived from deprotonation at C14, both deprotonation barriers are
very low (about 1 kcal/mol) (See C.8.2 and C.8.3 in Supporting Information); however,
compound 16 is 6.6 kcal/mol more stable than the other isomer and its reverse barrier is high
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(19.1 kcal/mol) (See C.8.2.3 and C.8.3.3 in Supporting Information and Scheme 9).
Scheme 9. Potential Energy Profiles for Rationalizing the Observed Regioselectivity for
Compounds 8 and 16*
* Free energies relative to 8a or 16a [B3LYP/6ꢀ31+G(d,p), scrf=(smd, solvent=dichloromethane)//B3LYP/6ꢀ
31+g(d,p)] in kcal/mol.
1
The H NMR spectrum of cadalane 17 is very similar to that of amorphane 13 with the main
differences being the deshielding of 0.11 ppm of H5 and the shape of the proton signal at C15
(See Figures S1B and S1C in Supporting Information). This fact, together with the similarity
of NOEs indicate that 17 is also a δꢀamorphene. The diastereotopic nature of the protons at
1
3
C15, the great shielding of this carbon in C NMR (ꢀδ = –17.4 ppm) and the different state
of aggregation of this compounds (white solid) made us think of a possible chlorinated δꢀ
ACS Paragon Plus Environment

Page 25 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
amorphene. This possibility was corroborated by the appearance in HRMS of the molecular
+
ions [M+H] 265.0995 and 267.0970 with ratio 3:1.
1
When the amount of mesyl chloride was increased to 2.4 equiv (Table 2, entry 4), H NMR
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
spectrum of reaction showed mainly a major product and the chlorinated amorphane 17 in 4:1
ratio (See Figure S1A in Supporting Information). When this same sample was kept in the
NMR tube for 3 d, only the δꢀamorphene 17 was observed, practically pure (See Figure S1B
in Supporting Information). This result evidences that this chlorinated cadalane derives from
the initial major product whose structure is assigned to a possible 15ꢀchloroꢀ6ꢀ
mesyloxygermacranolide by the presence of two vinyl protons at 5.00 ppm (bd, J = 10.3 Hz,
H1) and 4.88 (d, J = 10.9 Hz, H5), one deshielded proton at 5.12 ppm (t, J = 10.5 Hz, H6),
and one chloromethylene group at 4.18 and 3.94 ppm as doublets with J = 11.6 Hz.
The high tendency of the mesyloxy group at C15 to undergo nucleophilic substitution S 2 by
N
chloride instead of transannular cyclization to give guaianolides, as in the case of
salonitenolide (2) (Table 1, entry 4), could be explained considering that artemisiifolin (3) in
solution exists as a mixture of four slowly interconverting conformers, UD being the major
conformer that cannot undergo transannular cyclization. Transformations of primary allylic
alcohols to chlorinated derivatives using the MsCl/Et N system have been reported in the
3
48
literature.
A proposal for the formation of chlorinated δꢀamorphene 17 is depicted in Scheme 10. The
mechanism is the same to that proposed for the formation of hydroxylated δꢀamorphene 13
from artemisiifolin (3) (Scheme 8) but starting from 15ꢀchloroꢀ6ꢀmesyloxygermacranolide.
The involvement of the same carbocation intermediate is again key for transannular 1,6ꢀ
cyclization to afford the amorphane skeleton.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 26 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
Scheme 10. Proposed Mechanism for the Formation of Chlorinated δ-Amorphene 17
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
Finally, as with salonitenolide (2), the assays of artemisiifolin (3) against the above catalysts
(SSA or TsOH) were carried out in acetone, giving in all cases the acetonide 12 as the only
product in 17% and 66% yields, respectively (Table 2, entries 5 and 6). In principle, starting
from butꢀ2ꢀenꢀ1,4ꢀdiol unit, the formation of a sevenꢀmembered acetonide is expected, but
nevertheless a 1,3ꢀdioxolane is produced. This same reactivity has also been reported for butꢀ
4
6c
2ꢀenꢀ1,4ꢀdiols by using BF ·Et O as Lewis acid in acetone, however the authors achieved
3
2
the synthesis of the sevenꢀmembered acetonide operating at –78 ºC. Recently, transformations
of butꢀ2ꢀenꢀ1,4ꢀdiols into 1,3ꢀdioxolanes have been reported by using aldehydes as
4
9
condensing agent in the presence of a gold(I) catalyst.
A plausible mechanism for the formation of acetonide 12 from artemisiifolin (3) in acetone is
depicted in Scheme 11. The transformation begins with the nucleophilic attack of the
hydroxyl group at C15 to the protonated carbonyl of acetone to afford a hemiketal
intermediate which evolves to acetonide 12 by intramolecular ringꢀclosing reaction, allylic
transposition and the loss of water, in tandem. Generation of carbocation at C6, derived from
the loss of water, could also be considered.
ACS Paragon Plus Environment

Page 27 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Scheme 11. Proposed Mechanism for the Formation of Acetonide 12 from Artemisiifolin
(3) in Acetone
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
The structural relationship of amorphane 13 with some of the intermediates of the synthesis of
50
artemisinin and its derivatives is evident. Since artemisiifolin (3) can be obtained at
multigram scale from Staehelina dubia, and, in turn, the latter leads to 13 in two steps in good
yield, we have found a valuable intermediate to access these antimalarial compounds.
CONCLUSIONS
The presence of a hydroxyl group at C15 in germacranolides gives to these metabolites a
particular reactivity in the cyclization reactions that allows obtaining different
sesquiterpenolides. Under acid catalysis, this hydroxyl contributes to give 15ꢀ
oxoeudesmanolides. Instead, when this hydroxyl is transformed in a good leaving group the
cyclization goes in the opposite direction to produce guaianolides. This last reaction can be
considered as a biomimetic synthesis of guaianolides.
The nature of the solvent has great influence on the outcome of the reaction. Thus, when
acetone is used as solvent, the main reactions observed instead of the cyclization, were the
rearrangement concomitant with C8ꢀrelactonization of salonitenolide (2), or the allylic
transposition of artemisiifolin (3) to give the acetonide 12.
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 28 of 56
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
It is worth noting that cyclization of 12 gives a lactone with the skeleton of δꢀamorphene in
good yield. The stereoꢀstructural relationship of the amorphene 13 with some of the
intermediates of the synthesis of artemisinin and its derivatives, and its ready availability from
artemisiifolin (3), make it a valuable intermediate to access this antimalarial compounds.
All the mechanistic proposals in this paper have been evaluated by computational studies.
EXPERIMENTAL SECTION
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
General Information. Reagents and solvents were purchased from commercial suppliers and
were used as received. All experiments were carried out under air atmosphere, unless
otherwise indicated. Reactions were monitored through TLC on commercial silica gel plates
precoated with silica gel F254. Visualization of the developed plate was performed by
fluorescence quenching and pꢀanisaldehyde stain. Column chromatography was performed on
silica gel (60, particle size 40–63 ꢁm) as the stationary phase and the solvents employed were
of analytical grade, and reaction crudes were added by wetꢀloading method due to sensitivity
of products against silica gel acidity. Deactivated silica gel was prepared by addition of
deionized water (15% w/w) and homogenization. NMR spectra were recorded on 400 or 500
1
MHz spectrometers using standard pulse sequences. H NMR chemical shifts (δ) are
expressed in ppm referenced internally to residual solvent signal (CHCl , δ = 7.26 ppm;
3
1
3
acetone, δ = 2.05 ppm; benzene, δ = 7.16 ppm). C NMR chemical shifts (δ) are expressed in
ppm relative to the central resonance of deuterated solvent (CDCl , δ = 77.0 ppm; acetoneꢀd ,
3
6
δ = 29.9 ppm; C D , δ = 128.4 ppm). Coupling constant (J) are given in Hz. Signal splitting
6
6
were designated as singlet (s), doublet (d), triplet (t), quartet (q), doublet of doublets (dd),
doublet of triplets (dt), doublet of doublet of doublets (ddd), multiplet (m), broad singlet (bs).
Two dimensional NMR spectroscopy (gCOSY, gHSQC, gHMBC, NOESYꢀ2D) and NOESYꢀ
ACS Paragon Plus Environment

Page 29 of 56
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
1
13
1
D were used to assist the assignment of signal in the H and C NMR spectra. Melting
points were measured on a melting point apparatus and are uncorrected. Optical rotations
were obtained at 20 ºC in CHCl and calibrated with pure solvent as a blank. Fourier
3
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
transform infrared spectroscopy (FTIR) spectra were recorded using NaCl plates, and data are
–1
reported in cm . Mass spectra were recorded on a UPLCꢀQTOF mass spectrometer. Silica
5
1
sulfuric acid (SSA) was prepared according to the literature.
Isolation of (6R,7R,8S,1’’R)-8-[(1’’,2’’-dihydroxyethyl)acryloyl]-15-hydroxygermacra-
1
(10),4,11(13)-trien-6,12-olide (cnicin, 1). Centaurea calcitrapa L. was collected in June
2
013 at Torre del Puerco, Chiclana de la Frontera (Cádiz), Spain. The dry aerial part (890 g)
was submitted to 40 h maceration in CH Cl (3.5 L). The solid residues were removed by
2
2
filtration and the solvent was evaporated under reduced pressure to afford a greenish viscous
extract (16 g). The above extract was supported on deactivated silica gel, and was purified by
deactivated silica column chromatography (EtOAc:Hexane = 1:2, 1:1, 2:1 to 1:0) to afford,
after crystallization from EtOAc, cnicin (1) as white amorphous solid (1.7 g, 0.2% yield from
1
13
52
dry plant). H and C NMR spectra are in accordance with the literature.
Preparation of (6R,7R,8S)-8,15-dihydroxygermacra-1(10),4,11(13)-trien-6,12-olide
(salonitenolide, 2). Due to instability problems shown by salonitenolide (2) in the cyclization
reactions, its obtention from cnicin (1) was modified with respect to those previously reported
methods. In these cases salonitenolide (2) was used immediately in the cyclization reactions
and yields were calculated from cnicin (1). General procedure: An aqueous 0.25 M Na CO3
2
solution (1.2 equiv) was dropped to a solution of cnicin (1) (0.1 mmol) in 2:3 dioxane/water
(
2 mL). The mixture was stirred for 15 min at rt and was poured into a vigorously stirred
mixture of EtOAc (15 mL), saturated aqueous NaHCO (10 mL) and brine (10 mL). The
3
biphasic system was stirred for 10 min, the organic phase was separated and the aqueous
ACS Paragon Plus Environment