One-pot regio- and stereoselective synthesis of α′-methoxy- γ-pyrones: Biological evaluation as mitochondrial respiratory complex inhibitors

Article abstract of DOI:10.1021/jo201683u

The one-pot construction of functionalized α′-methoxy-γ- pyrones is detailed. Starting from α,α′-dimethoxy-γ- pyrone, molecular diversity is attained by a regio- and stereoselective desymmetrization using allyllithium followed by vinylogous aldol reaction

Full text of DOI:10.1021/jo201683u

Article
pubs.acs.org/joc
One-Pot Regio- and Stereoselective Synthesis of α′-Methoxy-γ-
pyrones: Biological Evaluation as Mitochondrial Respiratory Complex
Inhibitors
Helena Rosso,†,‡ Michael De Paolis,*,† Valerie C. Collin,§ Sriloy Dey,§ Sidney M. Hecht,§
́
̈
Cristina Prandi,‡ Vincent Richard,† and Jacques Maddaluno†
†Laboratoire des Fonctions Azotee
Saint-Aignan, France
‡Dipartimento di Chimica Generale e Chimica Organica, Universita di Torino, via P. Giuria 7, 10125, Torino, Italy
́ ́ ́ ́
s et Oxygenees Complexes de l’IRCOF, CNRS UMR 6014 & FR 3038, Universite de Rouen, Mont
§Center for BioEnergetics, The Biodesign Institute, and Department of Chemistry, Arizona State University, Tempe, Arizona 85287,
United States
S
* Supporting Information
ABSTRACT: The one-pot construction of functionalized α′-methoxy-γ-pyrones is detailed. Starting from α,α′-dimethoxy-γ-
pyrone, molecular diversity is attained by a regio- and stereoselective desymmetrization using allyllithium followed by vinylogous
aldol reaction. Mechanistic considerations including density functional theory calculations and insightful experiments have been
gathered to shed light on this complex multistep process. To illustrate the versatility of this methodology, some of the molecules
prepared were evaluated for their ability to inhibit NADH-oxidase and NADH-ubiquinone oxidoreductase. In the process, a
potent new inihibitor of NADH-oxidase activity (IC50 44 nM) was identified.
have recently introduced α,α′-dimethoxy-γ-pyrone 3 as a new
building block for the expedient synthesis of verticipyrone via
the direct preparation of α-allyl-α′-methoxy-γ-pyrone 4 from 3
(see Scheme 1).9 Product 4 results from a complex, multistep
process, beginning with the desymmetrization of electrophile 3
by the conjugate addition of allyllithium. Since product 4 is
relatively acidic, its formation is followed by its rapid
deprotonation with allyllithium leading to lithiated α-allyl-α′-
methoxy-γ-pyrone 5. Being electronically rich, this intermediate
behaves as a nucleophile and prevents further nucleophilic
attack of allyllithium to the γ-pyrone ring. Although the
protonation of 5 during the workup delivered 4 for the purpose
of the synthesis of verticipyrone, we anticipated that
modulation of the electrophiles used to intercept 5 would
allow the one-pot synthesis of biomolecules belonging to this
family of natural products. In terms of molecular diversity, this
strategy would constitute a unique tool in reaching, in one step,
valuable biomolecules from α,α′-dimethoxy-γ-pyrone 3.
INTRODUCTION
■
In recent years, several new natural products containing the α′-
methoxy-γ-pyrone scaffold have been isolated and shown to
exhibit interesting bioactivities.1 The modification of natural
products through diverted total synthesis is a powerful tool in
the discovery of analogues.2 In addition to having improved
bioactivities, these new molecules can also help in under-
standing the interactions involved between the natural products
and the biological targets. In this context, verticipyrone
constitutes a simple and interesting case of diverted total
synthesis (see Figure 1a). Verticipyrone ((E)-2-methoxy-3,5-
dimethyl-6-(3-methyl-2-undecenyl)-4H-pyran-4-one) was iso-
lated from the fungus Verticillium sp. FKI-1083 in a screen for
anthelmintic agents.3 The molecule was reported to inhibit
Ascaris suum NADH-fumarate reductase and bovine heart
NADH oxidase activity and NADH-ubiquinione oxidoreduc-
tase. Structurally related natural products such as cyercene A4
and salinipyrone,5 shown in Figure 1a, feature conjugated diene
side chains connected to α′-methoxy-γ-pyrone or γ-hydroxy-α′-
pyrone scaffolds. More recently, synthetic analogues of
verticipyrone have been prepared and tested on the biological
targets mentioned above,6,7 and products 1 and 2, which
exhibited greater inhibition than verticipyrone, were discovered
(see Figure 1b).
In continuation of our preliminary work, we report now a
stereocontrolled one-pot preparation of new molecules bearing
the α′-methoxy-γ-pyrone core and functionalized side chains
from 3 and commercially or readily available starting materials.
When it is possible, the one-pot reaction is an attractive
Received: August 24, 2011
Published: October 19, 2011
strategy for the rapid preparation of bioactive molecules.8 We
© 2011 American Chemical Society
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Figure 1. Verticipyrone and structurally related cyercene A and salinipyrone and selected bioactive synthetic derivatives 1 and 2.
Scheme 1. Desymmetrization of 3 by Allyllithium Leading to 4 via 5 and Interception of 5 with Carboxaldehydes
Scheme 2. Analogy of Regioselectivity Issues Arising from the Vinylogous Aldol Reaction of the α-Allyl-α′-methoxy-γ-pyrone
and an α,β-Unsaturated Ester
Scheme 3. One-Pot Preparation of α′-Methoxy-γ-pyrones 10−22 from 3
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Scheme 4. One-Pot Preparation of Esters 24−26 from 3
Scheme 5. Putative Pathway and Intermediates
Mechanistic insights into this process and biological evaluations
of some products are also disclosed.
Even hindered aliphatic aldehydes such as cyclohexylcarb-
oxaldehyde and pivalic aldehyde were reacted with 5 to produce
the secondary alcohols 20 and 21 in 35 and 26% yields,
respectively. Importantly, only ε-products were obtained with
carbonylated electrophiles.
RESULTS AND DISCUSSION
■
Although the use of metalated α,α′-dialkyl-γ-pyrone for
alkylation or aldolisation is well documented,10 the reactivity
of the lithium enolate of the α-allyl-α′-methoxy-γ-pyrone 5 and
the delocalized forms 5′ and 5″ had not been examined (see
Scheme 2a). The unknown preparation of 5 and the
polytopicity of isomers 5−5″ may explain this situation. Indeed,
a mixture of isomers 6−9 are expected to be produced after
treatment of 5−5″ with an electrophile. Apart from the O/C
chemoselectivity and the E/Z configuration of the double bond,
the nucleophilic attack may occur at three different positions, α-
, β-, or γ-, delivering several products of condensation.
Actually, these problems of regioselectivity are similar to
those encountered in the versatile and well-studied vinylogous
aldol reaction of an α,β-unsaturated ester (see Scheme 2b),
where the γ-functionalized α,β-unsaturated ester may be
obtained as a mixture with the α-isomer after quenching the
enolate with an aldehyde.11,12 The fact that the α′-methoxy-γ-
pyrone core is present in several natural products prompted our
interest in the reactivity of 5 with aldehydes.
Interestingly, γ-selectivity was observed when an alkylating
reagent such as allyl bromide was employed and resulted in the
formation of the unconjugated α-allyl-α′-methoxy-γ-pyrone 22
in 50% yield. The simplicity of the procedure deserves to be
highlighted here: α,α′-Dimethoxy-γ-pyrone 3 and tri(n-butyl)-
allylstannane were mixed together prior to the addition of n-
BuLi at low temperature. After 30 min, the electrophile was
added and the reaction was slowly warmed to room
temperature. The resulting lithium hydroxylates were quenched
with H3O+ to yield the products 10−22 or with another type of
electrophile such as benzoyl chloride, which afforded esters
24−26 in 53−38% yields, thus introducing a supplementary
point of molecular diversity (see Scheme 4).
The one-pot preparation of a variety of substituted α′-
methoxy-γ-pyrones constitutes a substantial improvement over
known strategies, which involve several steps for the
elaboration of the functionalized γ-pyrone core and the side
chain.13 Since the α,α′-dimethoxy-γ-pyrone 3 is readily available
in two steps on a multigram scale from dimethyl ketodicarbox-
ylate,9 this strategy opens a handy access to a large family of
compounds.
The putative mechanism of this transformation is depicted in
Scheme 5. The formation of allyllithium, from the reaction of n-
BuLi with tri(n-butyl)allylstannane,14 is the first step in this
process followed by the 1,4-addition to 3, affording the enolate
27. From 27, the primary expulsion of MeOLi can provide 4.
After deprotonation of 4 by the second equivalent of
allyllithium, 5 is expected to be formed. In 27, the molecular
orbitals alignement of the π-electrons of the enolate and the
orbital σ*C−O of the methoxy group favors the elimination of
MeOLi.
On the other hand, when the anion 28 generated by the
action of LiC-KOR base (t-BuOK/n-BuLi) upon diethoxybut-
2-ene was treated with 3 in the presence of CuCl, the trienic
keto ester 30 was isolated in 55% yield without the expected
In our previous study, we showed that the conjugate addition
of 2 equiv of in situ generated allyllithium to 3 is a simple and
efficient way to access lithiated α-allyl-α′-methoxy-γ-pyrone 5.
Using this procedure to prepare 5, its reactivity was evaluated
with aromatic aldehydes (see Scheme 3). The regio- and
stereoselectivity of the condensation were particularly good
because only ε-selectivity was observed and E-olefins 10−22
were obtained in yields ranging from 60 to 35%. 4-
Nitrobenzaldehyde gave the best result, delivering the ε-aldol
product 10 in 60% yield. An electron-rich benzaldehyde such as
4-methoxybenzaldehyde or 4-bromobenzaldehyde and hetero-
aromatic carboxaldehydes were also suitable electrophiles,
yielding the corresponding benzylic alcohols 11−18 (56−28%
yields).
Furthermore, the chemistry was found to tolerate an α,β-
unsaturated aldehyde to afford the allylic alcohol 19 (32%).
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Scheme 6. Reaction of Metal Alkoxydiene 28 to 3
product 31 (see Scheme 6).15,16 This trienic product could
arise from the ring-opening of the adduct 29 taking place
instead of the elimination of MeOLi. With an access to the
polyenic keto ester 30, the methodology of desymmetrization
demonstrated some versatility when the nature of the
nucleophile and the metal employed was modulated. Using
allylmagnesium bromide as a nucleophile illustrated further the
versatility of the methodology and the importance of the initial
1,4-addition. Hence, when compound 3 was treated with
allylmagnesium bromide in the presence of LiBr, the
polysubstituted γ-allyl-α′-methoxy-α-pyrone 33 was isolated in
55% yield (see Scheme 7). The formation of 33 may find its
the nucleophile to 3 is a mandatory step for the successful
formation of substituted α′-methoxy-γ-pyrones 10−22.
Density functional theory (DFT) calculations have been
carried out at the B3LYP/6-31G** level to investigate the
relative stability of the isomers 5−5″. In an effort to increase the
realism of the theoretical model, two discrete molecules of
THF were taken into account (see Figure 2). As expected,
enolate 5 appears to be more stable than 5′ (+10.6 kcal mol−1)
and 5″ (+18.9 kcal mol−1). If 5 cannot be ruled out as a possible
reacting intermediate, the ε-selectivity of the aldolisation
reaction may be explained by the higher reactivity of
intermediate 5′. Although it does not seem possible to go
through a Zimmerman−Traxler transition state involving 5 and
the aldehyde because of the rigid conformation of 5, such a
classical transition state does seem possible with 5′ and the
aldehyde, as illustrated in Scheme 8.
Scheme 7. Reaction of Allylmagnesium Bromide with 3
Scheme 8. Putative Zimmerman−Traxler Transition State
5′/ArCHO Explaining the ε-Selectivity
In order to deepen our understanding on the origin of the ε-
selectivity, the relative stability of the ε-(23) and γ-(23′)
isomers of the lithium hydroxylates arising from the aldol
reaction was investigated because an equilibrium between these
two isomers through retro-aldolisation/aldolisation processes
origin in the 1,2-addition of allylmagnesium bromide to 3. The
highly unstable diallylic tertiary alcohol 32 obtained may
undergo, upon workup, a Stork−Danheiser rearrangement to
produce 33.17 Consequently, it seems that the 1,4-addition of
Figure 2. DFT calculations of 5−5″ with two discrete molecules of THF.
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could be considered (see Scheme 9).18 This equilibrium was
likely to modify the distribution of isomers toward the
± 5.7 nM7 for the inhibition of NADH oxidase and NADH-
ubiquinone oxidoreductase activities, respectively.
In this study, the inhibition of complex I by seven additional
synthetic derivatives of verticipyrone was measured by
monitoring their effects on NADH-ubiquinone oxidoreductase
activity. NADH oxidase activity measures electron transport
between complex I and IV of the mitochondrial electron
transport chain.
Scheme 9. Relative Stability of 23 and Its Isomer 23′
Calculated by DFT
Electrons are initially transported from complex I (NADH:
ubiquinone oxidoreductase) to complex III (ubiquinol:
cytochrome c oxidoreductase) by coenzyme Q10 and then to
complex IV (cytochrome c oxidase) by the peripheral
membrane protein cytochrome c. Of the analogues tested in
the present study, structurally related compounds 10 and 11
showed the greatest affinity for complex I (Table 1).
thermodynamic product when the reaction reached room
temperature. This hypothesis implies that 23, the only isolated
product, is the most stable one. However, DFT computations,
ran at the same level as above on a model including two
discrete molecules of THF, led to the conclusion that 23′ is
more stable than 23 by 7.6 kcal mol−1 (see the Supporting
Information for details). Parallel to these computations, a
complementary reaction was run in which the temperature was
maintained at −40 °C after addition of the aldehyde. In these
conditions, the ε-isomer 23 was the sole product observed.19 In
light of these concording data, two conclusions can be drawn:
the equilibrium between 23 and 23′ seems unlikely and the
formation of 23 is under kinetic control.
Therefore, the one-pot process described here involves
probably at least five transformations: formation of allyllithium,
conjugate addition to the γ-pyrone, elimination of lithium
methoxide, deprotonation, and, finally, aldolisation or alkyla-
tion. The aldol reaction may take place via a Zimmermam−
Traxler transition state. With regard to the number of
transformations involved, the yields ranging from 60 to 26%
remain synthetically useful.
Table 1. Respiratory Chain Inhibition by Verticipyrone
Analogues
a
NADH oxidase
NADH-ubiquinone oxidoreductase
IC50 (μM)
compound
IC50 μM
Imax (%) half Imax (μM)
10
11
16
17
19
21
36
1.1 ± 0.08
0.044 ± 0.005
3.4 ± 0.3
>20
3.9 ± 1.1
2.8 ± 0.6
21.5 ± 1.7
56.3 ± 7.2
4.7 ± 0.4
59 ± 15
76.2 ± 3.7
91.4 ± 1.1
87.9 ± 1.1
89.9 ± 0.9
99.7 ± 1.7
90.4 ± 1.1
89.9 ± 6.3
2.7 ± 0.1
2.5 ± 0.5
17.0 ± 1.4
46.8 ± 5.7
4.7 ± 0.4
54 ± 16
0.6 ± 0.1
1.4 ± 0.2
3.7 ± 0.8
51.3 ± 7.4
44.8 ± 7.6
a
The maximum activity of NADH-ubiquinone oxidoreductase was
1.03 ± 0.06 μmol min−1 mg−1 ± SD and was determined from at least
five measurements.
The Imax value for compound 11 (91.4 ± 1.1%) was slightly
greater than that observed for compound 10 (76.2 ± 1.1%),
whereas the half Imax values for the two compounds were quite
similar.
In the NADH oxidase assay, compound 11 was the most
potent of the species studied (IC50 0.044 ± 0.005 μM); its
potency was comparable to the best of the analogues reported
previously.7 Interestingly, compound 10 exhibited much less
inhibition (IC50 1.1 ± 0.08 μM) in the NADH oxidase assay.
The differences in results for these compounds in the two
assays underscores the roles that small differences in inhibitor
structure can have on the overall inhibitory effect on
mitochondrial electron transport chain function.
The conversion of some of the aldol products to dienes was
next carried out. Connected to the α′-methoxy-γ-pyrone core,
these conjugated products could be seen as analogues of
cyercene A or salinipyrone. Hence, the dehydration of electron-
rich 11 was conducted in CF3CO2H to deliver the (E/E)-diene
34 in 90% yield (see Scheme 10). Alternatively, the elimination
Scheme 10. Conversion of Alcohol 11 and Benzoates 24−26
to Dienes 34−37
The greater potency of compound 11 on NADH oxidase
activity may reflect additional binding interactions for this
compound with complex III or IV or possibly differences in the
qualitative nature of its interaction with complex I.
Formal insertion of a C(CH3)CH group within the
aliphatic side chain of 10 (to afford compound 19) slightly
decreased the affinity of the molecule for complex I and
resulted in a greater half Imax value, but the Imax value itself
was actually higher (99.7 ± 1.7%). Compound 19 also
exhibited strong inhibition of NADH oxidase. In an earlier
report, a compound with a shorter side chain was found to be
>100-fold less inhibitory to complex I and more than 300-fold
to the NADH oxidase activity.7 In contrast, in the present study
the effect of side chain length was much less pronounced,
possibly because of the presence of unsaturation within the side
chain, which must significantly reduce conformational flexi-
bility.
of the benzoate of 21, 22, and 23 took place in the presence of
t-BuOK at rt in lower yields (55−65%) and led stereoselectively
to (E/E)-dienes 35−37. The configurations of the double
1
bonds of products 34−37 have been determined by H NMR
analysis.20
The potential bioactivities of these compounds were of
interest. Indeed, verticipyrone is a known inhibitor of bovine
heart mitochondrial respiratory chain function having reported
IC50 values of 1.3 nM and 46 nM6 or 64.9 ± 1.7 nM and 230.9
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α,α′-dimethoxy-γ-pyrone 3 (100 mg, 0.54 mmol) and tributyl(allyl)-
stannane (360 mg, 337 μL, 1.08 mmol, 2 equiv) in THF (5 mL, 0.1
M) was cooled to −90 °C using a toluene/liquid nitrogen bath. After
n-BuLi (1.6 M in hexanes, 747 μL, 2.2 equiv) was added dropwise, the
solution went from colorless to shining red. After 20 min at this
temperature, the electrophile (0.65 mmol, 1.2 equiv) was added
dropwise. The resulting mixture was stirred between −90 and −78 °C
for 30 min, after which time the temperature was allowed to rise to rt.
Then a saturated NH4Cl solution (10 mL) was added, and the mixture
was extracted with CH2Cl2 (3 × 10 mL), washed with brine (2 × 10
mL), and dried with anhydrous MgSO4. After filtration and
evaporation of the solvent, the crude products were purified by
trituration or by flash chromatography.
Replacement of the benzene ring by a tert-butyl group
(compound 18) led to a dramatic loss of complex I inhibition
and to a smaller loss of NADH oxidase inhibitory activity. In
contrast, Leiris et al.7 found that removal of a phenyl ring
within the side chain had more minimal effects. The differences
may be due to overall chain length or to the presence of para
substituents in the present case.
The formal dehydration of compound 10 produces
compound 36, which exhibited much less potent inhibition of
NADH-ubiquinone oxidoreductase (IC50 51.3 ± 7.4 μM). The
hydroxyl group of 10 is thus essential for its affinity toward
complex I. Compound 34 also exhibited less inhibition of
electron transport through complex IV (IC50 3.7 ± 0.8 μM).
For compounds 16 and 17, the replacement of the benzene
ring by a thiophene and an N-methylpyrrole led to a dramatic
loss of affinity for complex I. The IC50 values determined were
21.5 ± 1.7 μM and 56.3 ± 7.2 μM, respectively. The Imax were
comparable for both compounds (87.9 ± 1.1 and 89.9 ± 0.9%,
respectively) and significantly higher than for 10. The half Imax
values (17.0 ± 1.4 and 46.8 ± 5.7 μM, respectively) reflected a
greater loss of affinity for the analogues having a methylated
pyrrole (17) than for the analogues having a thiophene ring
(16). In the NADH oxidase assay, compound 17 was essentially
inactive as an inhibitor (IC50 >20 μM), whereas compound 16
retained some inhibitory activity (IC50 3.4 ± 0.2 μM).
(E)-2-(4-Hydroxy-4-(4-nitrophenyl)but-1-enyl)-6-methoxy-3,5-di-
methyl-4H-pyran-4-one, 10. The residue was purified by trituration
with CH2Cl2/pentane to give 10 (118 mg, 63%) as a gray powder: mp
1
158−159 °C; H NMR (300 MHz) δ 8.20 (d, J = 8.7 Hz, 2H), 7.58
(d, J = 8.7 Hz, 2H), 6.70−6.12 (m, 2H), 5.01 (dd, J = 7.7, 4.6 Hz, 1H),
3.96 (s, 3H), 3.71 (bs, 1H), 3.14−2.36 (m, 2H), 1.93 (s, 3H), 1.82 (s,
3H); 13C NMR (75 MHz) δ 181.2, 161.9, 151.4, 147.3, 132.1 (2C),
126.6 (2C), 123.7 (2C), 122.9, 118.2, 99.5, 72.3, 55.4, 42.9, 9.6, 7.0; IR
(KBr disk) ν 3392, 2916, 1664, 1630, 1568, 1514, 1340, 1261, 1166,
1055, 854. HRMS (ESI) for C18H20NO6 Calcd: 346.1291 [M + H]+.
Found: 346.1299.
(E)-2-(4-Hydroxy-4-(4-methoxyphenyl)but-1-enyl)-6-methoxy-
3,5-dimethyl-4H-pyran-4-one, 11. The residue was purified by flash
column chromatography to give 11 (100 mg, 56%) as a white solid: Rf
= 0.4 (AcOEt/pentane 7/3, 1% Et3N) [UV, KMnO4]; mp 149−150
°C; 1H NMR (300 MHz) δ 7.21 (d, J = 8.7 Hz, 2H), 6.77 (d, J = 8.7
Hz, 2H), 6.37−6.29 (m, 2H), 4.71 (dd, J = 7.3, 5.3 Hz, 1H), 3.84 (s,
3H), 3.70 (s, 3H), 3.38 (bs, 1H), 2.80−2.30 (m, 2H), 1.84 (s, 3H),
1.72 (s, 3H); 13C NMR (75 MHz) δ 181.2, 161.8, 159.0, 151.7, 136.2,
133.5, 127.0 (2C), 122.0, 117.7, 113.7 (2C), 99.2, 73.0, 55.3 (2C),
42.8, 9.5, 7.0; IR (KBr disk) ν 3375, 1666, 1578, 1468, 1261, 1241,
1175. HRMS (ESI) for C19H23O5 Calcd: 331.1545 [M + H]+. Found:
331.1546.
CONCLUSIONS
■
The one-pot preparation of molecules containing the α′-
methoxy-γ-pyrone core has been achieved. This procedure is
remarkably simple, short, and open to molecular diversity. The
whole transformation is based upon the unprecedented
reactivity of lithiated α-allyl-α′-methoxy-γ-pyrone 5; the latter
reacts regio- and stereoselectively with electrophiles such as
carboxaldehydes or allyl bromide. Furthermore, the conjugated
dienes α′-methoxy-γ-pyrones have been prepared without
transition metals from the aldol products. Interestingly, several
of the compounds, including 10, 11, and 19, are strong
inhibitors of NADH oxidase and NADH-ubiquinone oxidor-
eductase activities.
(E)-2-(4-(4-Bromophenyl)-4-hydroxybut-1-enyl)-6-methoxy-3,5-
dimethyl-4H-pyran-4-one, 12. The residue was purified by tritu-
ration with CH2Cl2/pentane to give 12 as a white powder (82 mg,
1
40%): mp 137−138 °C; H NMR (300 MHz) δ 7.37 (d, J = 8.4 Hz,
2H), 7.18 (d, J = 8.4 Hz, 2H), 6.35−6.28 (m, 2H), 4.75 (t, J = 6.2 Hz,
1H), 3.86 (s, 3H), 3.73 (bs, 1H), 2.63−2.52 (m, 2H), 1.84 (s, 3H),
1.73 (s, 3H); 13C NMR (75 MHz) δ 181.3, 161.8, 151.6, 143.1, 132.9,
131.4 (2C), 127.6 (2C), 122.3, 121.2, 117.8, 99.3, 72.6, 55.3, 42.8, 9.5,
7.0; IR (KBr disk) ν 3352, 1663, 1596, 1573, 1466, 1263, 1163. HRMS
(ESI) for C18H20BrO4 Calcd: 379.0545 [M + H]+. Found: 379.0559.
(E)-2-(4-Hydroxy-4-phenylbut-1-enyl)-6-methoxy-3,5-dimethyl-
4H-pyran-4-one, 13. The residue was purified by flash column
chromatography to give 13 (78 mg, 48%) as a pale yellow solid: Rf =
0.6 (AcOEt/pentane 7/3, 1% Et3N) [UV, KMnO4]; mp 128.5−129.0
EXPERIMENTAL SECTION
■
THF was distilled from sodium/benzophenone ketyl, and argon was
passed through a pad of anhydrous CaSO4/crystals of silica gel prior to
use. α,α′-Dimethoxy-γ-pyrone,9 tri(n-butyl)allylstannane,21 3,4-ethyl-
enedioxythiophene-2-carboxaldehyde,22 and (E)-2-methyl-3-(4-
nitrophenyl)acrylaldehyde23 were synthesized according to literature
procedures. All reactions requiring anhydrous conditions were
performed in oven (120 °C) and/or heat gun dried glassware under
an atmosphere of desiccated argon. Syringes and needles were dried
(90−120 °C) and cooled to rt prior use in desiccators containing P2O5
or anhydrous CaSO4/crystals of SiO2 as desiccants. 1H NMR (300 and
200 MHz) and 13C NMR (75 and 50.2 MHz) spectra were recorded at
293 K in CDCl3 unless otherwise mentioned and using (CH3)4Si and
residual CHCl3 as internal references. Chemical shifts (δ) are
expressed in parts per million (ppm) and the coupling constants in
hertz (Hz). Thin layer chromatography was performed on aluminum-
backed plates precoated with silica gel (Merck, Silica Gel 60 F254).
Compounds were visualized by exposure to UV light (254 nm) and by
dipping the plates in a solution of potassium permanganate (KMnO4)
in water. Unless otherwise stated, chromatographic separations were
carried out under pressure on Merck silica gel 60 Å (70−230 mesh)
pretreated, in some cases, with Et3N (1%). The neutral alumina
employed was Broeckmann grade III.
1
°C; H NMR (200 MHz) δ 7.39−7.06 (m, 5H), 6.40−6.18 (m, 2H),
4.75 (t, J = 6.3 Hz, 1H), 3.81 (s superimposed to a bs, 4H), 2.71−2.47
(m, 2H), 1.82 (s, 3H), 1.71 (s, 3H); 13C NMR (50.2 MHz) δ 181.1,
161.4, 151.3, 143.9, 133.3 (2C), 127.8, 127.4, 125.3, 121.8 (2C), 117.3,
99.1, 73.0, 54.9, 42.6, 9.1, 6.6; IR (KBr disk) ν 3316, 1667, 1633, 1570,
1467, 1417, 1259, 1168. HRMS (ESI) for C18H21O4 Calcd: 301.1440
[M + H]+. Found: 301.1427.
(E)-2-(4-Hydroxy-4-(pyridin-4-yl)but-1-enyl)-6-methoxy-3,5-di-
methyl-4H-pyran-4-one, 14. The residue was purified by an acid/
base workup (2 M HCl or citric acid/10% w/w NaOH) followed by
ammonia-impregnated preparative TLC to give 14 (76 mg, 47%) as a
light brown oil: Rf = 0.2 (CH2Cl2/MeOH/NH3, 9:1) [UV, KMnO4];
1H NMR (300 MHz) δ 8.60 (s br, 2H), 7.34 (d br, J = 3.5 Hz, 2H),
6.47 (d, J = 16.0 Hz, 1H), 6.38 (m, 1H), 4.90 (m, 1H), 3.95 (s, 3H),
2.70 (m, 2H), 1.97 (s, 3H), 1.85 (s, 3H); 13C NMR (50.2 MHz) δ
180.9, 161.6, 153.7, 151.4, 149.1, 132.5, 122.2, 120.8, 117.5, 99.0, 71.2,
55.0, 42.2, 9.2, 6.7; IR (film) ν 3332, 3054, 1665, 1602, 1416, 1265,
1166. HRMS (ESI) for C17H20NO4 Calcd: 302.1392 [M + H]+.
Found: 302.1387.
General Procedure I for the Syntheses of Compounds 10−
22. In a Schlenck vessel under an argon atmosphere, a solution of
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(E)-2-(4-(Furan-2-yl)-4-hydroxybut-1-enyl)-6-methoxy-3,5-di-
methyl-4H-pyran-4-one, 15. The residue was purified by flash
column chromatography to give 15 (50 mg, 32%) as a yellow-orange
solid: Rf = 0.4 (AcOEt/pentane 7/3, 1% Et3N) [UV, KMnO4]; mp
(E)-2-(4-Hydroxy-5,5-dimethylhex-1-enyl)-6-methoxy-3,5-di-
methyl-4H-pyran-4-one, 21. The residue was purified by flash
column chromatography to give 21 (39 mg, 26%) as a white solid:
Rf = 0.5 (AcOEt/pentane 7/3, 1% Et3N) [UV, KMnO4]; mp 137.1−
1
1
116.5−118.6 °C; H NMR (200 MHz) δ 7.34 (dd, J = 1.8, 0.9 Hz,
138.0 °C; H NMR (200 MHz) δ 6.52−6.36 (m, 2H), 3.98 (s, 3H),
1H), 6.45−6.36 (m, 2H), 6.31−6.25 (m, 2H), 4.85 (t, J = 6.4 Hz, 1H),
3.92 (s, 3H), 3.66 (bs, 1H), 2.82−2.76 (m, 2H), 1.91 (s, 3H), 1.79 (s,
3H); 13C NMR (50.2 MHz) δ 180.9, 161.6, 155.9, 151.4, 141.7, 132.4,
122.1, 117.7, 110.0, 106.0, 99.1, 66.6, 55.2, 39.2, 9.3, 6.7; IR (KBr disk)
ν 2926, 1667, 1572, 1470, 1422, 1341, 1263, 1170. HRMS (ESI) for
C16H18O5 Calcd: 291.1232 [M + H]+. Found: 291.1225.
3.37 (dd, J = 10.4, 2.1 Hz, 1H), 2.49 (ddd, J = 14.5, 5.7, 2.1 Hz, 1H),
2.23 (m, 2H), 1.96 (s, 3H), 1.83 (s, 3H), 0.95 (s, 9H); 13C NMR (50.2
MHz) δ 180.9, 161.6, 151.6, 135.4, 121.3, 117.3, 99.0, 78.5, 55.1, 35.4,
34.9, 25.5, 9.3, 6.8; IR (KBr disk) ν 3435, 2961, 1596, 1575, 1471,
1344, 1264, 1163. HRMS (ESI) for C16H25O4 Calcd: 281.1753 [M +
H]+. Found: 281.1752.
(E)-2-(4-Hydroxy-4-(thiophen-2-yl)but-1-enyl)-6-methoxy-3,5-di-
methyl-4H-pyran-4-one, 16. The residue was purified by flash
column chromatography to give 16 (71 mg, 43%) as a pale yellow
solid: Rf = 0.55 (AcOEt/pentane 7/3, 1% Et3N) [UV, KMnO4]; mp
3-(Hexa-1,5-dien-3-yl)-5-methoxy-2,6-dimethylcyclohexa-2,5-di-
enone, 22. The residue was purified by flash column chromatography
to give 22 (65 mg, 51%) as a brown oil: Rf = 0.4 (neutral alumina,
Et2O/pentane 7/3) [UV, KMnO4]; 1H NMR (300 MHz) δ 5.85 (ddd,
J = 15.3, 10.3, 7.1 Hz, 1H), 5.68 (ddt, J = 17.1, 10.1, 7.0 Hz, 1H),
5.16−4.95 (m, 4H), 3.92 (s, 3H), 3.60 (dt, J = 15.3, 7.0 Hz, 1H),
2.58−2.36 (m, 2H), 1.93 (s, 3H), 1.81 (s, 3H); 13C NMR (75 MHz),
δ 180.9, 162.0, 157.5, 135.9, 134.6, 118.9, 117.3, 116.8, 99.3, 55.3, 44.6,
36.3, 9.6, 6.9; IR (film) ν 2928, 1746, 1667, 1601, 1463, 1324, 1252;
MS (IE) 234 (M+, 37), 193 (35), 165 (100), 105 (41), 83 (46), 77
(36). HRMS (IE, 70 eV) for C14H18O3 Calcd: 234.1256 [M]+. Found:
234.1262.
1
127.5−128.8 °C; H NMR (200 MHz) δ 7.15 (dd, J = 4.8, 1.4 Hz,
1H), 7.05−6.77 (m, 2H), 6.92−6.82 (m, 2H), 5.04 (t, J = 6.3 Hz, 1H),
3.86 (s, 3H) superimposed to 4.1−3.8 (bs, 1H), 2.85−2.63 (m, 2H),
1.85 (s, 3H), 1.73 (s, 3H); 13C NMR (50.2 MHz) δ 181.0, 161.7,
151.5, 147.9, 132.6, 126.4, 124.3, 123.4, 122.2, 117.7, 99.1, 69.1, 55.1,
42.7, 9.3, 6.8; IR (KBr disk) ν 3313, 1666, 1632, 1594, 1571, 1468,
1422, 1170, 1038. HRMS (ESI) for C16H19O4S Calcd: 307.1004 [M +
H]+. Found: 307.1004.
(E)-2-(4-Hydroxy-4-(1-methyl-1H-pyrrol-2-yl)but-1-enyl)-6-me-
thoxy-3,5-dimethyl-4H-pyran-4-one, 17. The residue was purified
by flash column chromatography to give 17 (46 mg, 28%) as a pale
yellow solid: Rf = 0.55 (AcOEt/pentane 7/3, 1% Et3N) [UV,
KMnO4]; mp 104.3−106.1 °C; 1H NMR (200 MHz) δ 6.63−6.56 (m,
1H), 6.54−6.39 (m, 2H), 6.11 (dd, J = 3.6, 1.8 Hz, 1H), 6.08−6.00
(m, 1H), 4.80 (t, J = 6.7 Hz, 1H), 3.96 (s, 3H), 3.69 (s, 3H), 2.84 (t, J
= 6.3 Hz, 2H), 2.71 (bs, 1H), 1.96 (s, 3H), 1.82 (s, 3H); 13C NMR
(50.2 MHz) δ 180.9, 161.6, 151.4, 133.6, 133.0, 123.1, 122.0, 117.7,
106.4, 106.0, 99.1, 65.6, 55.2, 40.0, 34.0, 9.3, 6.8; IR (KBr disk) ν 3311,
1664, 1380, 1340, 1261, 1169, 1039. HRMS (ESI) for C17H22NO4
Calcd: 304.1549 [M + H]+. Found: 304.1553.
General Procedure II for the Syntheses of Esters 24−26.
Following the general procedure I with the exception of the workup,
which is as follows: The solution was cooled to 0 °C and treated with
benzoyl chloride (153 mg, 126 μL, 1.08 mmol, 2 equiv). The mixture
was stirred for 4 h, during which time the temperature was allowed to
rise to rt. Then a saturated NaHCO3 solution (10 mL) was added, and
the mixture was extracted with CH2Cl2 (3 × 10 mL), washed with
brine (2 × 10 mL), and dried with anhydrous MgSO4. After filtration
and removal of the solvent under reduced pressure, the crudes were
purified by flash column chromatography.
(E)-4-(6-Methoxy-3,5-dimethyl-4-oxo-4H-pyran-2-yl)-1-(4-
nitrophenyl)but-3-enyl Benzoate, 24. The title product was obtained
as a yellow solid (54 mg, 54%): Rf = 0.7 (AcOEt/pentane 7/3, 1%
Et3N) [UV, KMnO4]; mp 72.5−73.2 °C; 1H NMR (200 MHz) δ 8.19
(d, J = 8.8 Hz, 2H), 8.03 (dd, J = 8.4, 1.3 Hz, 2H), 7.71−7.50 (m, 3H),
7.50−7.34 (m, 2H), 6.55−6.26 (m, 2H), 6.19 (dd, J = 7.6, 5.5 Hz,
1H), 3.84 (s, 3H), 3.09−2.79 (m, 2H), 1.89 (s, 3H), 1.78 (s, 3H); 13C
NMR (50.2 MHz) δ 180.6, 165.2, 161.5, 150.7, 147.5, 146.7, 133.5,
129.6, 129.4, 129.1, 128.4, 126.9, 123.7, 123.4, 118.4, 99.3, 74.0, 55.0,
39.6, 9.2, 6.7; IR (film) ν 1722, 1666, 1608, 1347, 1267, 1108. HRMS
(ESI) for C25H24NO7 Calcd: 450.1553 [M + H]+. Found: 450.1564.
(E)-4-(6-Methoxy-3,5-dimethyl-4-oxo-4H-pyran-2-yl)-1-phenyl-
but-3-enyl Benzoate, 25. The title product was obtained as a yellow
oil (51 mg, 47%): Rf = 0.7 (AcOEt/pentane 7/3, 1% Et3N) [UV,
(E)-2-(4-(2,3-Dihydrothieno[3,4-b][1,4]dioxin-5-yl)-4-hydroxybut-
1-enyl)-6-methoxy-3,5-dimethyl-4H-pyran-4-one, 18. The residue
was purified by flash column chromatography to give 18 (67 mg, 28%)
as a white solid: Rf = 0.3 (AcOEt/pentane 7/3, 1% Et3N) [UV,
1
KMnO4]; mp 79.2−80.0 °C; H NMR (200 MHz) δ 6.49 (m, 2H),
6.26 (s, 1H), 5.10 (t, J = 6.2 Hz, 1H), 4.19 (s, 3H), 3.97 (s, 3H),
2.85−2.74 (m superimposed to 2.71 bs, 3H), 1.98 (s, 3H), 1.84 (s,
3H); 13C NMR (50.2 MHz) δ 181.0, 161.6, 151.5, 141.1, 137.2, 132.7,
122.0, 120.2, 117.6, 99.0, 97.3, 66.0, 64.5, 55.1, 41.0, 9.3, 6.8; IR (KBr
disk) ν 3312, 1664, 1560, 1437, 1382, 1261, 1166, 1065. HRMS (ESI)
for C18H21O6S Calcd: 365.1059 [M + H]+. Found: 365.1075.
2-((1E,5E)-4-Hydroxy-5-methyl-6-(4-nitrophenyl)hexa-1,5-dien-
yl)-6-methoxy-3,5-dimethyl-4H-pyran-4-one, 19. The residue was
purified by flash column chromatography to give 19 (64 mg, 32%) as a
pale yellow solid: Rf = 0.4 (AcOEt/pentane 7/3, 1% Et3N) [UV,
1
KMnO4]; H NMR (200 MHz) δ 8.05 (dd, J = 8.1, 1.2 Hz, 2H),
7.60−7.20 (m, 8H), 6.54−6.22 (m, 2H), 6.13 (dd, J = 7.4, 5.9 Hz,
1H), 3.80 (s, 3H), 3.08−2.76 (m, 2H), 1.90 (s, 3H), 1.80 (s, 3H); 13C
NMR (50.2 MHz) δ 180.7, 165.4, 161.5, 150.9, 139.3, 133.0, 130.8,
129.8, 129.3, 128.4, 128.3, 128.1, 126.1, 122.7, 118.0, 99.1, 75.0, 54.9,
39.8, 9.2, 6.7; IR (film) ν 3061, 2955, 1719, 1634, 1464, 1269, 1110.
HRMS (ESI) for C25H25O5 Calcd: 405.1702 [M + H]+. Found:
405.1695.
1
KMnO4]; mp 147−148 °C; H NMR (300 MHz) δ 8.14 (d, J = 8.6
Hz, 2H), 7.36 (d, J = 8.6 Hz, 2H), 6.64 (s, 1H), 6.50−6.42 (m, 2H),
4.36 (dd, J = 7.2, 4.5 Hz, 1H), 3.92 (s, 3H), 3.57 (bs, 1H), 2.78−2.47
(m, 2H), 1.94 (s, 6H), 1.80 (s, 3H); 13C NMR (75 MHz) δ 180.2,
160.9, 150.7, 145.0, 143.4, 143.3, 132.4 (2C), 128.5 (2C), 122.5 (2C),
121.1, 116.8, 98.3, 74.9, 54.3, 38.2, 13.6, 8.6, 6.0; IR (KBr disk) ν 3352,
1664, 1595, 1514, 1341, 1260, 1167. HRMS (ESI) for C21H23NO6
Calcd: 386.1604 [M + H]+. Found: 386.1602.
(E)-1-(Furan-2-yl)-4-(6-methoxy-3,5-dimethyl-4-oxo-4H-pyran-2-
yl)but-3-enyl Benzoate, 26. The title product was obtained as a pale
yellow oil (81 mg, 38%): Rf = 0.6 (AcOEt/pentane 3/2, 1% Et3N)
1
(E)-2-(4-Cyclohexyl-4-hydroxybut-1-enyl)-6-methoxy-3,5-dimeth-
yl-4H-pyran-4-one, 20. The residue was purified by flash column
chromatography to give 20 (55 mg, 33%) as a pale yellow solid: Rf =
0.6 (AcOEt/pentane 7/3, 1% Et3N) [UV, KMnO4]; mp 123.4−125.2
[UV, KMnO4]; H NMR (200 MHz) δ 8.07−7.95 (m, 2H), 7.59−
7.24 (m, 4H), 6.52−6.25 (m, 4H), 6.20 (t, J = 7.0 Hz, 1H), 3.83 (s,
3H), 3.03 (dt, J = 7.0, 2.7 Hz, 2H), 1.90 (s, 3H), 1.79 (s, 3H); 13C
NMR (50.2 MHz) δ 180.7, 165.4, 161.5, 151.3, 150.9, 142.6, 133.1,
130.2, 129.5, 129.4, 128.2, 122.9, 118.1, 110.2, 108.9, 99.1, 67.8, 54.9,
36.0, 9.2, 6.7; IR (film) ν 2955, 1719, 1667, 1609, 1263, 1168. HRMS
(ESI) for C23H23O6 Calcd: 395.1495 [M + H]+. Found: 395.1479.
(4Z,6E)-Methyl-6-ethoxy-5-methoxy-2,4-dimethyl-3-oxonona-
4,6,8-trienoate, 30. In a dry Schlenck tube, a solution of freshly
sublimated t-BuOK (245 mg, 2.3 mmol, 2.5 equiv) in THF (7 mL)
was cooled to −78 °C under N2 atmosphere. (E)-1,1-Diethoxybut-2-
ene (130 mg, 0.9 mmol, 1 equiv) in THF (2 mL) and n-BuLi (1.4 mL,
1
°C; H NMR (200 MHz) δ 6.65−6.22 (m, 2H), 3.94 (s, 3H), 3.47
(ddd, J = 9.0, 5.6, 3.5 Hz, 1H), 2.78 (bs, 1H), 2.57−2.19 (m, 2H), 1.91
(s, 3H), 1.78 (s, 3H) superimposed to 1.84−1.52 (m, 6H), 1.49−0.87
(m, 5H); 13C NMR (50.2 MHz) δ 181.0, 161.6, 151.6, 134.6, 121.4,
117.2, 98.9, 74.8, 55.1, 43.5, 37.0, 28.9, 28.0, 26.2, 26.0, 25.9, 9.3, 6.8;
IR (KBr disk) ν 3315, 2925, 1665, 1601, 1461, 1337, 1261, 1166.
HRMS (ESI) for C18H27O4 Calcd: 307.1909 [M + H]+. Found:
307.1907.
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2.3 mmol, 1.6 M in hexane, 2.5 equiv) were successively added, and
the mixture was stirred for 2 h, during which the temperature rose to
−40 °C. Simultaneously, in a 25 mL flask, a mixture of 3 (284 mg, 1
mmol, 1.1 equiv) and CuCl (99 mg, 1 mmol, 1.1 equiv) was stirred in
THF (8 mL) under N2 atmosphere at rt for 2 h. The Schlenck tube
was refrigerated to −78 °C, and the mixture contained in the flask was
transferred into the Schlenck tube. The resulting mixture was stirred at
−78 °C for 1 h. Saturated NH4Cl solution (10 mL) was added, and
the mixture was extracted with AcOEt (3 × 10 mL), washed with
water (10 mL), brine (2 × 10 mL), and dried over anhydrous K2CO3.
After filtration and evaporation of the solvent, the crude product was
purified by flash chromatography (petroleum ether/AcOEt 80:20 and
1% of Et3N) to give 30 (140 mg, 55%) as a pale yellow oil: Rf = 0.6
2-Methoxy-3,5-dimethyl-6-((1E,3E)-4-phenylbuta-1,3-dienyl)-4H-
pyran-4-one, 35. The title product was obtained from 25 as a white
solid (51 mg, 60%): Rf = 0.6 (AcOEt/pentane 3/2, 1% Et3N) [UV,
KMnO4]; mp 245.6−247.5 °C; 1H NMR (200 MHz) δ 7.49−7.29 (m,
5H), 7.11−6.90 (m, 2H), 6.81 (d, J = 15.0 Hz, 1H), 6.61 (d, J = 14.3
Hz, 1H), 4.09 (s, 3H), 2.07 (s, 3H), 1.88 (s, 3H); 13C NMR (50.2
MHz) δ 180.6, 161.5, 151.9, 136.9, 136.3, 133.4, 128.6, 128.4, 127.4,
126.6, 120.9, 118.8, 99.5, 55.2, 9.5, 6.8; IR (KBr disk) ν 1653, 1595,
1464, 1413, 1262. HRMS (ESI) for C18H19O3 Calcd: 283.1334 [M +
H]+. Found: 283.1314.
2-Methoxy-3,5-dimethyl-6-((1E,3E)-4-(4-nitrophenyl)buta-1,3-di-
enyl)-4H-pyran-4-one, 36. The title product was obtained from 26 as
a yellow solid (54 mg, 55%): Rf = 0.7 (AcOEt/pentane 3/2, 1% Et3N)
[UV, KMnO4]; mp 209.1−210.0 °C; 1H NMR (200 MHz) δ 8.22 (d, J
= 8.8 Hz, 2H), 7.58 (d, J = 8.8 Hz, 2H), 7.21−6.90 (m, 2H), 6.84 (d, J
= 14.2 Hz, 1H), 6.74 (d, J = 14.2 Hz, 1H), 4.09 (s, 3H), 2.09 (s, 3H),
1.89 (s, 3H); 13C NMR (50.2 MHz) δ 180.4, 161.5, 151.2, 147.0,
142.6, 133.8, 132.1, 131.7, 127.0, 124.0, 123.8, 120.2, 99.9, 55.3, 9.6,
6.9; IR (KBr disk) ν 1653, 1595, 1335, 1259. HRMS (ESI) for
C18H18NO5 Calcd: 328.1185 [M + H]+. Found: 328.1175.
1
[UV, KMnO4)]; H NMR (300 MHz) δ 6.18 (dt, J = 16.7, 10.4 Hz,
1H), 5.70 (d, J = 10.4 Hz, 1H), 5.15 (d, J = 16.7 Hz, 1H), 4.97 (d, J =
10.4 Hz, 1H), 4.13 (m, 1H), 3.85 (q, J = 7.0 Hz, 2H), 3.69 (s, 3H),
3.59 (s, 3H), 1.70 (s, 3H), 1.35 (m, 6H); 13C NMR (75 MHz) δ
197.3, 172.6, 156.2, 149.4, 131.5, 117.0, 115.5, 108.6, 63.6, 56.0, 52.7,
51.9, 14.6, 14.1, 13.5; IR (film) ν 2943, 1740, 1666, 1455, 1374, 1202.
HRMS (ESI) for C15H23O5 Calcd: 283.1545 [M + H]+. Found:
283.1538.
2-((1E,3E)-4-(Furan-2-yl)buta-1,3-dienyl)-6-methoxy-3,5-dimeth-
yl-4H-pyran-4-one, 37. The title product was obtained from 27 as a
pale yellow solid (53 mg, 65%) without distinguishable other isomers
4-Allyl-6-methoxy-3,5-dimethyl-2H-pyran-2-one, 33. In a 25 mL
Schlenck tube, a mixture of 3 (284 mg, 1 mmol) and flame-dried LiBr
(435 mg, 5 mmol, 5 equiv) was stirred in THF (8 mL) under an argon
atmosphere at rt for 1 h. The suspension was cooled to −78 °C, a
solution of allylmagnesium bromide (1.5 mL, 1.5 mmol, 1.5 equiv, 1 M
in Et2O) was added dropwise, and the reaction mixture was stirred at
−78 °C for 2 h. Saturated NH4Cl solution (10 mL) was added, and
the mixture was extracted with CH2Cl2 (3 × 10 mL), washed
sequentially with water (10 mL) and brine (2 × 10 mL), and dried
over anhydrous Na2SO4. After filtration and evaporation of the solvent,
the crude product was purified by flash chromatography (pentane/
AcOEt 4/1, 1% Et3N) to give 33 as a white solid (107 mg, 55%): mp
1
of the double bonds by H NMR analysis: Rf = 0.6 (AcOEt/pentane
1
3/2, 1% Et3N) [UV, KMnO4]; mp 135.8−136.0 °C; H NMR (200
MHz) δ 7.40 (s, 1H), 7.00−6.75 (m, 2H), 6.55 (d, J = 15.0 Hz, 1H),
6.53 (d, J = 15.0 Hz, 1H), 6.46−6.34 (m, 2H), 4.03 (s, 3H), 2.02 (s,
3H), 1.85 (s, 3H); 13C NMR (50.2 MHz) δ 180.6, 161.4, 152.3, 151.9,
143.0, 133.0, 125.8, 123.7, 120.7, 118.7, 111.9, 110.5, 99.4, 55.2, 9.5,
6.8; IR (KBr disk) ν 1653, 1602, 1415, 1339, 1284, 1168, 991. HRMS
(ESI) for C16H17O4 Calcd: 273.1127 [M + H]+. Found: 273.1132.
Submitochondrial Particle (SMP) Preparation. Mitochondria
were prepared as described.24,25 One bovine heart was ground and
blended in sucrose buffer (0.25 M sucrose, 10 mM Tris-HCl, pH 7.8,
containing 0.2 mM EDTA) at 4 °C. Cellular debris was removed by
centrifugation at 1200g for 20 min. The supernatant was filtered
through two layers of cheesecloth. The mitochondria were harvested
by centrifugation at 26000g for 15 min and then homogenized in the
same buffer. The mitochondria were harvested by centrifugation at
12000g for 30 min and then stored at −80 °C in a sucrose buffer.
Submitochondrial particles (SMPs), were prepared as described.25,26
The mitochondria were sonicated with a Sonic Dismembrator (Fisher
Scientific) in 0.25 M sucrose containing 5 mM MgCl2, 1 mM ATP, 10
mM MnCl2, and 1 mM sodium succinate in 10 mM Tris-HCl, pH 7.8,
at 4 °C. Cell debris was pelleted by centrifugation at 20000g for 7 min
at 4 °C. The SMPs were harvested by centrifugation at 152000g for 30
min at 4 °C and stored at −80 °C in 0.25 M sucrose cotaining 5 mM
MgCl2, 2 mM ATP, 2 mM glutathione, and 1 mM sodium succinate in
10 mM Tris-HCl, pH 7.5. Protein concentration was determined by
BCA titration (Pierce) using albumin for the standard curve.
1
75−76 °C; Rf = 0.8 (pentane/AcOEt 4/1) [UV, KMnO4]; H NMR
(200 MHz) δ 5.86−5.66 (m, 1H), 5.03 (m, 1H), 4.88 (m, 1H), 3.89
(s, 3H), 3.43−3.33 (m, 2H), 2.10 (s, 3H), 2.06 (s, 3H); 13C NMR
(50.2 MHz) δ 157.8, 157.3, 150.3, 133.5, 115.6, 109.6, 108.2, 54.0,
33.5, 10.8, 10.6; IR (film) ν 2928, 1693, 1643, 1469, 1134, 915; MS
(EI, 70 eV) m/z [M − 1]+ 193 (100), 178, 160, 150, 132, 122, 107.
HRMS (EI): for C11H13O3 Calcd: 193.0865 [M − 1]+. Found:
193.0860.
2-((1E,3E)-4-(Furan-2-yl)buta-1,3-dienyl)-6-methoxy-3,5-dimeth-
yl-4H-pyran-4-one, 34. In 25 mL, single-neck round-bottom flask,
the (E)-2-(4-hydroxy-4-(4-methoxyphenyl)but-1-enyl)-6-methoxy-3,5-
dimethyl-4H-pyran-4-one 11 (100 mg, 0.3 mmol) was stirred in the
presence of TFA (2 mL) at 0 °C. The temperature was then allowed
to rise to rt. After 30 min, an aqueous solution of NaOH (10% w/v)
was added, and the mixture was extracted with CH2Cl2 (3 × 10 mL),
washed with brine (10 mL), and dried with anhydrous MgSO4. After
filtration and evaporation of the solvent, the product 34 was isolated
without further purification as a yellow solid (90 mg, 95%) without
distinguishable other isomers of the double bonds by 1H NMR
analysis: Rf = 0.7 (AcOEt/pentane 7/3) [UV, KMnO4]; mp 156−152
NADH: Ubiquinone Oxidoreductase Activity Measured
Using SMPs. The inhibition of NADH-ubiquinone oxidoreductase
activity was determined under the same experimental conditions
reported.7,27 Twenty-five micrograms of SMPs were incubated at 39
°C for 5 min with the test compound in 1 mL of 0.25 M sucrose
containing 1 mM MgCl2, 2 μM antimycin A, 2 mM KCN, and 50 mM
phosphate buffer, pH 7.4. The reaction was initiated by the addition of
50 μM NADH and 50 μM coenzyme Q1. The enzymatic activity, at 39
°C, was monitored at 340 nm.
NADH Oxidase Activity Assay. Bovine heart mitochondria were
obtained by a large-scale procedure.24 Inverted submitochondrial
particles (SMPs) were prepared by the method of Matsuno-Yagi and
Hatefi28 and stored in a buffer containing 0.25 M sucrose in 10 mM
Tris-HCl, pH 7.4, at −80 °C. The inhibitory effects of test compounds
on bovine heart mitochondrial complex I, II, and IV were evaluated.
The compounds were dissolved in dimethylsulfoxide (DMSO) and
then used to make serial dilutions. Maximal DMSO concentrations
never exceeded 2% and had no influence on the control enzymatic
activity. Bovine heart SMPs were diluted to 0.5 mg/mL. After the pre-
equilibration of 30 μg/mL of SMP with inhibitor for 5 min at 30 °C in
1
°C; H NMR (200 MHz) δ 7.40 (d, J = 8.7 Hz, 2H), 7.01 (dd, J =
15.0, 9.6 Hz, 1H), 6.89 (d, J = 8.7 Hz, 2H), 6.81 (d, J = 9.7 Hz, 1H),
6.75 (d, J = 15.3 Hz, 1H), 6.54 (d, J = 15.1 Hz, 1H), 4.07 (s, 3H), 3.83
(s, 3H), 2.06 (s, 3H), 1.88 (s, 3H); 13C NMR (50.2 MHz) δ 180.9,
161.7, 160.1, 152.3, 136.8, 134.1, 129.3, 128.2, 125.6, 119.9, 118.5,
114.3, 99.6, 55.4, 55.4, 9.7, 7.0; IR (KBr disk) ν 1654, 1608, 1510,
1407, 1255, 1179, 1033, 979. HRMS (ESI) for C19H21O4 Calcd:
313.1440 [M + H]+. Found: 313.1456.
General Procedure III for the Syntheses of Dienes 35−37. In
a 25 mL single necked round-bottom flask, a solution of ester (0.3
mmol) in THF (3 mL) was stirred in the presence of t-BuOK (0.4
mmol, 45 mg, 1.2 equiv) at rt, and the reaction progress monitored by
TLC. After 2 h a small amount of water was added. The mixture was
extracted with CH2Cl2 (3 × 10 mL), washed with brine (10 mL), and
dried with anhydrous MgSO4. After filtration and evaporation of the
solvent, the crude was purified by flash chromatography.
9436
dx.doi.org/10.1021/jo201683u|J. Org. Chem. 2011, 76, 9429−9437

The Journal of Organic Chemistry
Article
2.5 mL of 50 mM Hepes, pH 7.5, and 5 mM MgCl2, the oxidation of
50 μM of NADH at 30 °C was monitored spectrophotometrically with
a Beckman Coulter DU-530 (340 nm, ε 6.22 mM−1 cm−1). The initial
rates were calculated from the linear portion of the traces.
(13) For a recent review on the strategies for the contruction of
α′methoxy-γ-pyrones, see: Sharma, P.; Powell, K. J.; Burnley, J.;
Awaad, A. S.; Moses, J. E. Synthesis 2011, 2865−2892.
(14) Seyferth, D.; Weiner, M. A. J. Org. Chem. 1959, 24, 1395−1396.
(15) Venturello, P. J. Chem. Soc., Chem. Commun. 1992, 1032−1033.
(16) Without CuCl, unreacted 3 was recovered.
ASSOCIATED CONTENT
■
(17) Stork, G.; Danheiser, R. L. J. Org. Chem. 1973, 38, 1775−1776.
(18) We thank one referee for drawing our attention on this point.
(19) No aldol reaction took place when the reaction was maintained
at −78 °C during one hour after addition of the aldehyde. When the
same procedure was applied at −40 °C, only the ε-isomer 23 was
obtained.
S
* Supporting Information
1H and 13C NMR spectra of new compounds and absolute
energies and atom coordinates for DFT calculations of 5−5″
and 23−23′. This material is available free of charge via the
Internet at http://pubs.acs.org.
(20) Spectroscopic analysis confirmed the E/E configuration with a
3
3
coupling constant of JH1−H2 = JH3−H4 = 15.0 Hz.
(21) Migita, T.; Nagai, K.; Kosugi, M. Bull. Chem. Soc. Jpn. 1983, 56,
2480.
AUTHOR INFORMATION
■
Corresponding Author
(22) Cava, M. P.; Lakshmikantham, M. V.; Lyon, M. A.; Hucke, A.;
Mohanakrishnan, A. K. Tetrahedron 1999, 55, 11745.
(23) Hirata, Y.; Nakata, H.; Yamada, K.; Okuhara, K.; Naito, T.
Tetrahedron 1961, 14, 252−274.
(24) Smith, A. L. Methods Enzymol. 1967, 10, 81−86.
(25) Pallotti, F.; Lenaz, G. Methods Cell Biol. 2007, 80, 3−44.
(26) Linnane, A. W.; Titchener, E. B. Biochim. Biophys. Acta 1960,
39, 469−478.
(27) Estornell, E.; Fato, R.; Pallotti, F.; Lenaz, G. FEBS Lett. 1993,
332, 127−131.
(28) Matsuno-Yagi, A.; Hatefi, Y. J. Biol. Chem. 1985, 260, 14424−
14427.
*E-mail: michael.depaolis@univ-rouen.fr.
ACKNOWLEDGMENTS
■
This work was financially supported by the Reg
́
ion Haute-
Normandie, the Universite
́
de Rouen, and the Centre National
de la Recherche Scientifique (CNRS). H.R. is grateful to the
Ministere de la Recherche and Regione Piemonte Cipe 2007
̀
(Biobits project) and the UFI/UIF (Vinci program) for
fellowships. The authors thank Dr. Catherine Fressigne
́
́
(Universite de Rouen, IRCOF) for DFT calculations and the
CRIHAN (project 2004-004) for computing facility.
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