Unveiling the 6,8-Rearrangement in 8-Phenyl-5,7-dihydroxyflavylium and 8-Methyl-5,7-dihydroxyflavylium through Host–Guest Complexation

Article abstract of DOI:10.1002/ejoc.201701009

8-Phenyl-5,7-dihydroxyflavylium and 8-methyl-5,7-dihydroxyflavylium were synthesized to observe the 6,8-rearrangement. 8-Phenyl and 8-methyl residues were introduced by Suzuki–Miyaura reaction of 8-iodochrysin, then reduced by LiAlH₄ to give the corresponding 3-deoxyanthocyanidins. At pH 1.0 the stable form is the 8-substituted flavylium cation in both compounds. At higher pH values the quinoidal bases are the stable species and some evidence for the 6,8-rearrangement was obtained, but the respective spectral variations are very small. This was overcome by using a modified cyclodextrin (captisol), which favors the formation of the trans-chalcone at the expense of the quinoidal bases. The trans-chalcone was isolated and dissolved in CD₃OD/DCl (pD < 1) to give a mixture of the two flavylium isomers. This was confirmed by ESI-MS analysis (recorded in positive ion mode) of the two isomers after separation by HPLC, which gave the same peak ([M]⁺ m/z 253). The 6-isomer slowly reverts to the most stable 8-isomer. The 6,8-rearrangement was also observed after irradiation of the trans-chalcone (in the presence of captisol) at pH 5. Acidification of this photostationary state gave a mixture of both flavylium cations. The UV/Vis absorption of the flavylium cation (6-isomer) was blueshifted in comparison to the 8-isomer.

Full text of DOI:10.1002/ejoc.201701009

A Journal of
Accepted Article
Title: Unveiling the 6,8-rearrangement in 8-phenyl-5,7-
dihydroxyflavylium and 8-methyl-5,7-dihydroxyflavylium through
host-guest complexation
Authors: Kumi Yoshida, Nuno Basílio, João Carlos Lima, Luís Cruz,
Victor de Freitas, Fernando Pina, Hiroki Ando, Yuki Kimura,
and Kin-Ichi Oyama
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To be cited as: Eur. J. Org. Chem. 10.1002/ejoc.201701009
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10.1002/ejoc.201701009
European Journal of Organic Chemistry
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Unveiling the 6,8-rearrangement in 8-phenyl-5,7-
dihydroxyflavylium and 8-methyl-5,7-dihydroxyflavylium through
host-guest complexation
Nuno Basílio,[a] João Carlos Lima,[a] Luís Cruz,[b] Victor de Freitas,[b] Fernando Pina,*[a] Hiroki
Ando,[c] Yuki Kimura,[c] Kin-Ichi Oyama,[d] Kumi Yoshida,*[e]
Abstract: 8-phenyl-5,7-dihydroxyflavylium and 8-methyl-5,7-dihydroxyflavylium were synthesized to observe the 6,8-rearrangement. 8-phenyl
and 8-methyl residue were introduced by Suzuki-Miyaura reaction of 8-iodochrysin, then, reduced by LiAlH4 to give the corresponding 3-
deoxyanthocyanidins. At pH=1.0 the stable form is the 8-substituted flavylium cation in both compounds. At higher pH values the quinoidal
bases are the stable species and some evidence for the 6,8-rearrangement was achieved, but the respective spectral variations are very small.
This was overcome by using a modified cyclodextrin (captisol), which favors the formation of the trans-chalcone at expenses of the quinoidal
bases. The trans-chalcone was isolated and dissolved in CD3OD/DCl (pD<1) to give a mixture of the two flavylium isomers. This was confirmed
by ESI-MS spectrum of the two isomers upon separation by HPLC, recorded in positive ion mode, gives the same peak ([M]+ m/z 253). The 6-
isomer slowly reverts to the most stable 8-isomer. The 6,8-rearrangement was also observed after irradiation of the trans-chalcone (in the
presence of captisol) at pH=5. Acidification of this photostationary state, gives a mixture of both flavylium cations. The UV-vis absorption of
flavylium cation (6-isomer) is blue shifted in comparison to 8-isomer.
A convenient way to study the multistate of chemical reactions
represented in Scheme 1 is to carry out a direct pH jump defined
Introduction
as the addition of base to equilibrated solutions of the flavylium
Flavylium derivatives that comprise anthocyanins, anthocyanidins
cation and follow the relaxation process towards the new
and 3-deoxyanthocyanidins, as well as many other compounds of
equilibrium. Similarly, reverse pH jumps obtained by the addition
synthetic origin, follow the same multistate of species reversibly
of acid to equilibrated solutions at higher pH values can be used.
interconverted by external stimuli, such as pH and light, see
Considering a direct pH jump, the flavylium cation, AH+, can
Scheme 1 for apigeninidin.[1,2] Flavylium cation is only one of the
transfer a proton to water forming the quinoidal base A (by far the
components of the multistate, the species thermodynamically
faster reaction of the multistate) and in a competitive reaction
favorable at sufficiently acidic solutions (Scheme 1).
hydrate to give the hemiketal, B. The species B, may open the
ring (tautomerism) to give cis-chalcone, Cc, that can isomerize to
lead trans-chalcone, Ct. In spite of the complexity of the multi-
equilibrium, the system can be simplified considering a single acid
base reaction involving the species AH+ and its conjugate base
CB defined as the sum of the other species A, B, Cc and Ct,
eq.(1)[2-5]
The mole fraction distribution of the conjugated base, CB, is
dramatically dependent on the substitution pattern of the 2-
phenylbenzopyrylium core. In other words, could be almost
constituted by A as in 4-methyl-7-hydroxyflavylium, a mixture of
Ct and A as in apigeninidin, or B as major species together with
a mixture of the minor species, A, Cc and Ct as in the case of
anthocyanins.[2]
The flavylium multistate is reversible. A reverse pH jump towards
sufficiently acidic medium gives back the flavylium cation as the
sole species. [6]
[a]
[b]
Dr. Nuno Basílio,(a) Dr. João Carlos Lima,(a) Prof. Dr. Fernando Pina
LAQV, REQUIMTE, Departamento de Química, Faculdade de
Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516
Caparica (Portugal)
E-mail: fp@fct.unl.pt
Dr. Luís Cruz, Prof. Dr. Victor de Freitas
REQUIMTE/LAQV, Departamento de Química e Bioquímica,
Faculdade de Ciências, Universidade do Porto, Rua do Campo
Alegre, 687, 4169-007 Porto, (Portugal)
Hiroki Ando, Dr. Yuki Kimura
Graduate School of Information Sciences, Nagoya University,
Chikusa, Nagoya 464-8601 (Japan)
[c]
[d]
[e]
Dr. Kin-Ichi Oyama
Chemical Instrumentation Facility, Research Center for Materials
Science, Nagoya University, Chikusa, Nagoya 464-8602 (Japan)
Prof. Dr. Kumi Yoshida
Graduate School of Informatics, Nagoya University, Chikusa,
Nagoya 464-8601 (Japan)
The interpretation of the flavylium multistate chemistry is easily
achieved by means of an energy level diagram obtained from the
equilibrium constants of the several processes, Scheme 2.[7,8]
After a direct pH jump an equilibrium between flavylium cation
(AH+) and quinoidal base (A) is achieved in microseconds, much
E-mail: yoshidak@is.nagoya-u.ac.jp.
Supporting information for this article is given via a link at the end of
the document.
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Scheme 1. Multistate of apigeninidin in acidic to moderately acidic aqueous solutions. Light absorption by the chalcones and hemiketal, leads to the cis-trans
isomerization and ring opening respectively. 6,8-rearrangement; R≠H.
AH+ + H2O
CB + H3O+ K’a=Ka + Kh + KhKt + KhKtKi
(1)
With [CB]=[A]+[B]+[Cc]+[Ct]
irradiation of Ct (blue arrows) that photo-isomerize to Cc. The
species Cc can go back to Ct or forward to form the colored
species AH+/A. The amount of the colored species is a balance
between the amplitude of the isomerization barrier and pH. The
pH dependence is due to the acid catalysis of the ring-opening
closure of Cc and B, and the hydration-dehydration involving
AH+ and B. After formation of the colored species they revert
back to the equilibrium following steps 2 (red arrows)
faster than the mixing time of the solutions (black arrow).
These two species (AH+ and A) behave as a single one during
all subsequent kinetic steps. In a sequence of further kinetic
process the system reaches the equilibrium (red color arrows).
This last process is dependent on the existence or not of a cis-
trans isomerization barrier and is controlled by the slowest step
of the multistate (hydration or isomerization depending on the
pH). The mole fraction distribution of the species A, B, Cc and
Ct are function of their relative energy levels and change
dramatically with the nature and position of the substituents of
the flavylium core. A photochromic system is obtained upon
One interesting feature of flavylium compounds possessing a
hydroxyl substituent in 5-position is the possibility of observing
a 6,8-rearrangement, Scheme 2.[9]
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Scheme 2. Thermodynamic and kinetic processes in flavylium multistates. After a direct pH jump the flavylium cation and quinoidal base equilibrate during the
mixing time of the base, step 1. A second more or less complex process leads to the equilibrium, step 2. Irradiation of trans-chalcone produces the isomer cis that
could give back the trans or the colored species AH+/A by a reverse process of step 2. At this point the system is out of the equilibrium (photochemical signal),
which is reached by steps 1 and 2
Whenever there is a hydroxyl in 5-position of the flavylium cation,
the ring closure of cis-chalcone to give hemiketal and flavylium
cation has two possibilities (Scheme 1). In the case of symmetric
flavylium derivatives i.e. the substitution in 6-position is the same
of 8-position the two possible isomers are undistinguishable.
However, when the substituents in 6- and 8-position are different
two isomers can be obtained unless one of them prevails.
The first description of the 6,8-rearrangement was reported by
Jurd for the compound 4’,5,7,8-tetrahydroxyflavylium.[10] He
observed that at pH=2.6 the isomer bearing hydroxyl in 6-
position gives almost quantitatively the one in 8-position after
about seven hours, while in 1% aqueous hydrochloric acid at
room temperature, was much slower, circa three days. Andersen
Scheme 3. Retrosynthesis of 8-alkylated 3-deoxyanthocyanidins
and co-workers published a complete study of flavylium cations
6,8-rearrangement derived from C-Glycosyl-3-
deoxyanthocyanidins family and arrived to similar conclusions.[11]
Recently we reported that in the case of substitution of Br in 6/8-
position, it was possible to separate both flavylium isomers by
HPLC and prove that they are converted in a mixture of circa
50% of each isomer at pH=1.0.[9]
In order to obtain more information regarding the role of the
substituents in 6-position (or 8-) of the flavylium cation, two kinds
of 8-substituted-5,7-dihydroxyflavylium were synthesized. For
this purpose, we established a new synthetic route (Scheme 3).
Among the previous reports, synthetic routes in Scheme 4-A[12]
and B[13] should afford a mixture of regioisomers due to involving
the cyclization of C-ring. In the other classical route by reduction
of per-O-acetylflavanone followed by oxidation of quinone
(Scheme 4-C)[14], there are many difficulties on alkylation of
flavanone skeleton because flavanone may be readily ring-
opened under acidic and basic condition.[15] Furthermore, the
yield of quinone oxidation step is low and the purification from
quinone oxidant is tedious.[14] To overcome these problems, we
planned a synthetic strategy using stable flavone which can be
alkylated by Suzuki-Miyaura coupling reaction.[16] To this end, an
efficient transformation reaction from flavone to 3-
deoxyanthocyanidin should be required. Based on our reductive
synthesis of per-O-methylcyanidins using LiAlH4 (Scheme 5),[17]
we developed a novel synthetic route of unprotected 3-
deoxyanthocyanidin and synthesized chrysinidin (1) and two 8-
alkylated 3-deoxyanthocyanidins 2 and 3 (Scheme 4-D).
Using
those
synthetic
compounds,
8-phenyl-5,7-
dihydroxyflavylium (2) and 8-methyl-5,7-dihydroxyflavylium (3),
we studied their 6,8-rearrangement. The effect of a modified β-
cyclodextrin was also used as external stimuli to improve the
photochemical response of the multistate and confirm the 6,8-
rearrangement.
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trihydroxyacetophenone (4)[20] followed by condensation
between the resulting methylated compound and benzaldehyde
to furnish a trans-chalcone.[16a,c,19] Cyclization of the trans-
chalcone with catalytic amount of iodine afforded
permethylchrysin 5 in good yield.[16a,c,19] Demethylation of 5 with
BBr3 in CH2Cl2 gave chrisin (6) in 89% yield. Based on our
previous reports (Scheme 5),[17] the reductive transformation of
chrisin (6) by LiAlH4 was examined. However, this reaction did
not give chrysinidin (1). Thus, the phenolic hydroxyl groups in 6
was protected by MOM group, then, the obtained 7 was reduced
by using LiAlH4 (4 eq.) in THF at room temperature. The reaction
mixture was analyzed by HPLC and Mass (Supporting
Information). After 30 min 7 was converted to MOM protected 3-
deoxyanthocyanidin 8 (HR-ESI-MS of 8: calcd for C19H19O5 [M]+
327.1227, found
Scheme 4. Synthesis of unprotected 3-deoxyanthocyanidin
Scheme 6. Synthesis of chrysinidin (1)
Scheme 5. Synthesis of 8-aryl-per-O-methylcyanidin by LiAlH4
Results and Discussion
Our retrosynthetic analysis is outlined in Scheme 4. The
reductive transformation of 8-alkylated flavone to the
corresponding 3-deoxyanthocyanidins by LiAlH4 was planned as
a key reaction at the final step.[17] 8-alkylated flavone could be
Scheme 7. Transformation of flavone 7 to chrysidin (1).
obtained by Suzuki-Miyaura reaction[16] of 8-iodochrysin (8).[18]
8
327.1231) and the successive deprotection of
8 using
could be prepared from phloroacetophenone (4) by the
previously reported method.[16a,c,18,19]
hydrochloric acid (HCl) in MeOH afforded chrysinidin (1) in 45%
yield (2 steps)
At first, we focused our attention to the development of synthetic
route of chrysinidin (1) as shown in Scheme 6. The synthesis of
chrysinidin (1) commenced with a methylation of 2',4',6'-
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are quinoidal base, by far the major species, and trans-chalcone,
minor one. The synthesis of 8-phenyl-5,7-dihydroxyflavylium (2)
leads to the pure 8-isomer, and at pH=1.0 we did not observed
significant spectral variations after 1 week.
The spectral variations between the pH dependent absorption
spectra of 8-phenyl-5,7-dihydroxyflavylium (2) taken immediately
after a direct pH jump, Figure 1a, and at the equilibrium, Figure
1b, are similar. In Figure 1a the spectral variations are due to the
equilibrium between flavylium cation and quinoidal base,
pKa=3.25, because proton transfer takes place during the mixing
time and evidences of other reactions were not observed. At the
equilibrium, besides quinoidal base absorption, a relatively small
increasing of the absorption in the spectral region around 360 nm
(compared with Figure 1a) can be attributed to the trans-
chalcone. 20% of trans-chalcone at the equilibrium can be
roughly calculated from the difference between K’a and Ka.[23] The
existence of an equilibrium between flavylium cation and a
mixture of quinoidal base (major) and trans-chalcone (minor)
was previously reported for other deoxanthocyanidins.[24]
Similar
observations
were
made
for
8-methyl-5,7-
dihydroxyflavylium (3) and the results are reported in the
Supporting Information (SI). For this compound further
evidences of the presence of trans-chalcone as minor species
was obtained from photochemical experiments (see SI).
Scheme 8. Synthesis of 8-alkylated 3-deoxyanthocyanidins (2 and 3)
In Figure 2 the absorption spectra of the flavylium cation and the
apparent conjugated base CB are reported in the presence of 50
mM of captisol, immediately after a direct pH jump and at the
equilibrium. Comparing Figure 2a with Figure 1a it can be
observed that the pKa does not change significantly in the
absence and presence of the host and the spectra are
compatible with an equilibrium between flavylium cation and
quinoidal base. However, at the equilibrium not only pK’a
becomes significantly lower, but also a clear disappearance of
the quinoidal base to give a significant absorption of the trans-
chalcone is observed, Figure 2b. This is the result of the already
reported higher affinity of the β-cyclodextrin and derivatives for
the trans-chalcone when compared with other species of the
respective multistate.[25]
Having developed
a
robust route to unprotected 3-
deoxyanthocyanidin from flavone, we focused on synthesis of 8-
alkylated 3-deoxyanthocyanidin 2 and 3 (Scheme 8). For the
introduction of alkyl groups at 8-position, permethylflavone 5 was
converted into 8-iodochrysin (9) by using NIS (N-
iodosuccinimide).[18] Based on our previous reported ligand-free
condition,[17a] the Suzuki-Miyaura coupling of
9
with
phenylboronic acid in the presence of Pd(OAc)2 and K3PO4 in
toluene at 100 ºC afforded 8-phenylflavone 10 in 92%. In this
reaction, the temperature should be controlled higher than 100
ºC, otherwise the yield decreased. Next, the Suzuki-Miyaura
coupling of 9 with methylboronic acid in the presence of
Pd(PPh3)4 was carried out according to Park’s report.[16d]
However, this coupling reaction afforded only a trace amount of
the desired 8-methylflavone with a significant amount of 5, which
is the reduced compound of 9. Furthermore, we tried the
coupling reaction with MeBF3K in the presence of PdCl2(dppf)
was examined.[21] However, the desired 8-methylflavone could
not be obtained. Finally, the Suzuki-Miyaura coupling with
DABAL-Me3 in the presence of Pd2(dba)3 and XPhos in
toluene[22] afforded the 8-methylflavone 11 in 80% yield involving
demethylation at 5-position. Demethylation of 10 and 11 by BBr3
followed by MOM protection afforded 14 and 15 in high yield.
Reductive transformation of 14 and 15 by LiAlH4 (4 eq.) in THF
at room temperature followed by deprotection with HCl afforded
the desired 8-alkylated 3-deoxyanthocyanidin 2 and 3 in 47 and
41% yield, respectively (2 steps).
Notably, for the compound 8-methyl-5,7-dihydroxyflavylium (3)
the selective complexation of the trans-chalcone with the host is
smaller than for 2 and, as a result, the fraction of this species at
the equilibrium is also smaller (see SI).
These results were confirmed by 1H NMR. An aqueous solution
of 8-phenyl-5,7-dihydroxyflavylium (2) in the presence of captisol
50 mM at pH= 4, was kept 1 day in the dark, extracted with ethyl
acetate and dried in vacuum. The respective residue was
dissolved in CD3OD and the 1H NMR spectrum acquired
immediately, Figure 3a. This spectrum can be attributed to a
mixture of the trans-chalcone identified by the large coupling
constant (16.0 Hz) between proton 3 and 4 and a small fraction
of other species, probably quinoidal bases. Figure 3b shows the
1H NMR spectrum of the trans-chalcone solution acidified to DCl
0.1 M. The spectrum shows signals attributed to the two flavylium
compounds 8-phenyl-5,7-dihydroxyflavylium (3) and 6-phenyl-
Similarly to other 3-deoxyanthocyanidins, the “basic” species at
the equilibrium with flavylium cation in the present compounds
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(a)
(b)
Figure 1. (a) Absorption spectra of the compound 8-phenyl-5,7-dihydroxyflavylium (2) in water:ethanol (1:1) as a function of pH taken immediately after a direct pH
jump pKa=3.25; (b) The same at the equilibrium, pKa=3.15.
(a)
(b)
Figure 2. Absorption spectra of flavylium cation and its apparent conjugated base CB, in the presence of captisol 50 mM, immediately after a direct pH jump (a)
and at the equilibrium (b).
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Figure 3. 1H NMR spectra acquired for an equilibrated solution (95:5 water:methanol) of 8-phenyl-5,7-dihydroxyflavylium (2) at pH = 4 in the presence of captisol
50 mM after extraction with ethyl acetate and vacuum concentration. (a) immediately after dissolving the solid residue in the CD3OD (540 L), (b) immediately after
addition of 60 µL of DCl 1M, (c) after 30 minutes, (d) after ca. 24 hours, (e) after one week. (f) 1H NMR spectrum of 8-phenyl-5,7-dihydroxyflavylium (2) in CD3OD:D2O
1:9 with DCl 0.1 M.
Figure 4. Absorption spectra of the compound 8-phenyl-5,7-dihydroxyflavylium (2) in the presence of the host 50 mM: black traced line- equilibrium at pH=1 AH+
;
8
black full line- equilibrium at pH=5.0; red line- photostationary state; blue line upon addition of acid to the photostationary state.
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Figure 5. 1H NMR spectra acquired for an equilibrated solution (95:5 water:methanol) of 8-methyl-5,7-dihydroxyflavylium (3) at pH = 4.5 in the presence of captisol
50 mM after extraction with ethyl acetate and vacuum concentration. (a) immediately after dissolving the solid residue in the CD3OD (540 µL), (b) after addition of
60 µL of DCl 1M, (c) after ca. 24 hours, (d) after one week, (e) after two weeks, (f) after four weeks.
leads to the blue absorption spectra. Taking into account that the
5,7-dihydroxyflavylium and trans-chalcone. After 30 minutes
(Figure 3c) the signals corresponding to the last species
disappear and only those corresponding to the mixture of
flavylium compounds could be observed. After one day the
relative fraction of 6-Ph decreased considerably (Figure 3d) and
after one week in the dark the mixture evolved to the total
conversion of 6-Ph into 8-Ph. This experiment shows
unequivocally that cis-chalcone is able to close the ring through
the hydroxyl in 2-position or 5-position,[26] leading to the two
flavylium cations, as shown in Scheme 2.
proton transfer to transform the quinoidal bases into the
respective flavylium cation is extremely fast (microseconds
range) the blue spectrum should be attributed to the respective
flavylium cations. The fact that the obtained absorption it is not
coincident with the one of the equilibrium, black traced line (AH+ ),
8
is an indication that some quinoidal base 6 was formed at the
photostationary state and converted into the respective flavylium
cation (AH+ ) at pH=1.0. Moreover, it can be concluded that the
6
absorption of AH+6 is blue shifted in comparison with AH+ .
8
In the case of the compound 8-methyl-5,7-dihydroxyflavylium (3)
the behavior is similar but, for the sake of brevity, the complete
UV-Vis study is reported in the Supporting Information. NMR
experiments similar to those described above were conducted
for 3 to demonstrate the formation of the trans-chalcone and both
regioisomers (8-methyl-5,7-dihydroxyflavylium and 6-methyl-
5,7-dihydroxyflavylium) after equilibration in presence of captisol
and solvent extraction (Figure 5). The results show the initial
The 6,8-rearrangement was also unveiled by a sequence of pH
jumps and light irradiation in the presence of the host captisol,
Figure 4. In this figure the absorption spectra at the equilibrium
for pH=1.0 (AH+ , black traced line) and pH=5.0 (black full line)
8
are shown. At pH=5.0 trans-chalcone formation is favored by its
interaction with the host. Light irradiation of the solution at
pH=5.0 at 365 nm where the trans-chalcone absorbs leads to a
photostationary state (PSS) represented by the red line. The
PSS shows an increasing of the absorption bands of the
quinoidal bases at the expenses of the trans-chalcone
disappearance, as expected from the photochromic system.
Immediate acidification of the photostationary solution to pH=1.0
formation of
a
mixture of 6-methyl and 8-methyl-5,7-
dihydroxyflavylium in a 38:62 mole ratio immediately after
acidification and a slow conversion of the 6-methyl into the more
stable 8-methyl isomer to a 14:86 mole ration after 4 weeks.
These experiments suggest a slower interconversion for 3 in
comparison with 2
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the trans-chalcone, the pivot species of the 6,8-rearrangement,
is greatly increased.
Experimental Section
General Procedures for Synthesis: Melting points were measured on a
Yanaco Mp-S3 micro melting point apparatus and are uncorrected.
Infrared (IR) spectra were recorded on
a JASCO FT/IR 6100
spectrometer, ultraviolet-visible (UV-Vis) absorption spectra on a JASCO
V-560 spectrophotometer (cell length: 10 mm), and 1H and 13C NMR
spectra on a JEOL JNM-ECA-500 spectrometer. Chemical shifts are
reported in ppm on the δ scale relative to CDCl3 (δ = 7.26 for 1H NMR),
CDCl3 (δ = 77.0 for 13C NMR), DMSO-d6 (δ = 2.49 for 1H NMR), and
DMSO-d6 (δ = 39.5 for 13C NMR), CD3OD (δ = 3.31 for 1H NMR), and
CD3OD (δ = 49.0 for 13C NMR) as internal references. Signal patterns are
indicated as s, singlet; d, doublet; m, multiplet. High-resolution mass
.
Scheme 9. Energy level diagram to account for the pH jumps and the
photochemistry of the 6,8-rearrangement of the compound 8-phenyl-5,7-
dihydroxyflavylium (2) in the presence of the cyclodextrin host. Traced energy
levels are tentatively positioned. The arrow of the excitation energy is not in
scale.
spectra (HRMS) were recorded on
a Bruker micrOTOF-QII (ESI)
spectrometer. Analytical thin-layer chromatography (TLC) was performed
on Merck precoated analytical plates, 0.25 mm thick, silica gel 60 F254.
High performance liquid chromatography (HPLC) was performed with a
system equipped with JASCO PU-1580 pumps, an MD-1515 detector,
HG-1580-32 mixer, and DG-580-35 degasser, JASCO. An ODS column
(SunShell C18, 2.6 μm, 2.1 i.d × 100 mm, ChromaNik Technologies) was
eluted at 40 °C with linear gradient elution (0.2 mL/min) over 20 min from
10% to 90% aq. MeCN solution containing 0.5% trifluoroacetic acid (TFA).
Flash silica gel chromatography was performed using silica gel PSQ60B
(Fuji Silysia Chemical). All reagents were purchased from commercial
sources. All reactions were performed under an argon (Ar) atmosphere.
Chrysinidin (1).
To a solution of 5,7-di-O-methoxymethylchrysin (7) (50 mg, 146 µmol) in
anhydrous THF (3 mL) was added LiAlH4 (0.6 mL, 1.0 M solution in THF)
at room temperature. After stirring for 30 min at room temperature,
anhydrous MeOH (3 mL) and 5 N HCl/MeOH (2.0 mL) were added to the
reaction mixture at 0 ºC. After stirring for 17 h at room temperature,
distilled water (10 mL) was added to the reaction mixture. The solvent
was concentrated under reduced pressure and the residue was purified
by flash silica gel chromatography (1% MeOH/AcOEt with 0.5% TFA 
5% MeOH/AcOEt with 0.5% TFA  7% MeOH/AcOEt with 0.5% TFA 
10% MeOH/AcOEt with 0.5% TFA 20% MeOH/AcOEt with 0.5% TFA)
to give 1 (23 mg, 45%) as a TFA salt (dark red solid). mp: > 300ºC, UV-
Vis (0.1% HCl-MeOH) λmax nm (ε): 392 nm (13,740), 464 nm (10,040);
IR (KBr) 3424, 1650, 1535, 1340, 1206 cm–1;1H NMR (10% TFA-d-CDCl3,
500 MHz) δ 6.72 (1H, d, J = 2.0 Hz), 7.01 (1H, d, J = 2.0 Hz), 7.70 (2H, t,
J = 7.5 Hz), 7.79 (1H, t, J = 7.5 Hz), 8.18 (1H, d, J = 7.5 Hz), 8.35 (2H, d,
J = 7.5 Hz), 9.26 (1H, d, J = 7.5 Hz); 13C NMR (10%TFA-d-CD3OD, 125
MHz) δ 96.3, 103.8, 111.1, 116.2, 129.7, 130.7, 131.1, 136.5, 151.0,
160.9, 161.3, 171.8, 174.5; ESI-TOF HRMS Calcd. For C15H11O3 [M]+,
239.0703; Found: 239.0710.
A further information was achieved by carrying out a HPLC
separation of the solution labeled b in Figure 6, as reported in
the Supporting Information (SI). LC-MS experiments (see SI)
confirm the presence of both isomers after extraction and
acidification. This experiment demonstrates the existence of two
species with the same molecular mass (compatible with flavylium
3) and different retention times. The isomer with higher retention
time was assigned to 8-methyl-5,7-dihydroxyflavylium (3) based
on the UV-Vis spectrum and the one with lower affinity for
stationary phase assigned to 6-methyl-5,7-dihydroxyflavylium by
exclusion.
The results can be visualized by means of an energy level
diagram obtained as reported elsewhere, Scheme 9.[27] The
trans-chalcone (and a fraction of quinoidal bases) are extracted
and dissolve in methanol, as in the NMR experiments. After
acidification the flavylium cations become the more stable
species and are formed from the quinoidal bases (within mixing
time) and chalcone, in circa 30 minutes. The mixture of the two
flavylium isomers is a kinetic product that slowly evolves to the
equilibrium situation which privileges the 8-isomer. On the other
hand, light absorption by the fraction of trans-chalcone, at pH=5
for example, leading to both quinoidal bases. These bases can
be converted in the respective flavylium cations by addition of
acid, once more giving a kinetic mixture.
8-phenylchrysinidin (2)
To a solution of 5,7-di-O-methoxymethyl-8-C-phenylchrysin (14) (177 mg,
0.42 mmol) in anhydrous THF (4.2 mL) was added LiAlH4 (1.7 mL, 1.0 M
solution in THF) at room temperature. After stirring for 40 min at room
temperature, distilled water (3 mL) and 15% TFA aqueous solution (6 mL)
were added to the mixture at 0 ºC. After stirring for 1 h at room
temperature, 4 mL of 5 N HCl/MeOH was added to the resulting mixture.
After stirring for 2.5 h at room temperature, distilled water (30 mL) was
added to the reaction mixture. The solvent was concentrated under
reduced pressure and the residue was purified by ODS HPLC (Develosil
ODS-HG-5, Nomura Chemicals) with 30% CH3CN aqueous solution
containing 0.5% TFA to give 2 (82 mg, 47%) as a TFA salt (dark red solid).
mp: 194–196 ºC; UV-Vis (0.1% HCl-MeOH) λmax nm (ε): 397 nm
Conclusions
In spite of the fact that in 8-phenyl-5,7-dihydroxyflavylium (2) and
8-methyl-5,7-dihydroxyflavylium (3) the 8-isomer is stable at
pH=1.0 and the 6,8-rearrangement is not observed, extending
these studies to the moderately acid region proved unequivocally
that in these compounds the 6,8-rearrangement takes place. The
6,8-rearrangement is amplified in the presence of a modified β-
cyclodextrin (captisol) because the mole fraction distribution of
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10.1002/ejoc.201701009
European Journal of Organic Chemistry
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(10,390), 476 nm (5,450); IR (KBr) 3476, 1651, 1531, 1332, 1195 cm–1
1H NMR (10%TFA-d-CD3OD, 500 MHz) δ 6.95 (1H, s), 7.51–7.60 (7H,
m), 7.71 (1H, t, J = 7.5 Hz), 8.01 (2H, d, J = 7.5 Hz), 8.21 (1H, d, J = 8.0
Hz), 9.33 (1H, d, J = 8.0 Hz); 13C NMR (10%TFA-d-CD3OD, 125 MHz) δ
103.3, 110.8, 111.3, 116.1, 129.5, 129.6, 130.6, 130.9, 131.1, 132.0,
136.4, 151.1, 157.2, 160.1, 171.5, 171.7; ESI-TOF HRMS Calcd. For
C21H15O3 [M]+, 315.1016; Found: 315.1011.
;
Acknowledgements
The financial support by the Associated Laboratory for
Sustainable Chemistry-Clean Processes and Technologies-
LAQV is acknowledged. The latter is financed by national funds
from FCT/ MEC (UID/QUI/50006/2013) and co-financed by the
ERDF under the PT2020 Partnership Agreement (POCI-01-
0145- FEDER-007265). The Portuguese FCT/MEC is also
acknowledged through the National Portuguese NMR Network
(RECI/BBB-BQB/0230/2012). N.B. is recipient of FCT
postdoctoral grant (SFRH/BPD/84805/2012). Luís Cruz
gratefully acknowledges the research assistant contract
(NORTE-01-0145-FEDER-000011). We acknowledge Prof. T.
Kondo for his discussions.
8-C-methylchrysinidin (3).
To a solution of 5,7-di-O-methoxymethyl-8-C-methylchrysin (15) (36 mg,
0.1 mmol) in anhydrous THF (1.0 mL) was added LiAlH4 (0.4 mL, 1.0 M
solution in THF) at room temperature. After stirring for 1 h at room
temperature, the reaction mixture was filtered through a pad of Celite®
and eluted with AcOEt. After the solvent was removed under reduced
pressure, the residue was dissolved with MeOH (0.5 mL) and then 1.0
mL of 7.5 N HCl/MeOH was added to this mixture. After stirring for 3 h at
room temperature, distilled water (1 mL) was added to the resulting
mixture. The solvent was concentrated under reduced pressure and the
residue was purified by flash silica gel chromatography (33%
AcOEt/Hexane with 0.5% TFA  67% AcOEt/Hexane with 1% TFA) to
give 3 (15 mg, 41%) as a TFA salt (dark red solid). mp: > 300 ºC; UV-Vis
(0.5% HCl-MeOH) λmax (ε): 393 nm (13,440), 488 nm (6,430); IR (KBr)
3424, 1661, 1541, 1458, 1363, 1204 cm–1; 1H NMR (10%TFA-d-CD3OD,
500 MHz) δ 2.47 (3H, s), 6.82 (1H, s), 7.70 (2H, t, J = 7.5 Hz), 7.78 (1H,
t, J = 7.5 Hz), 8.12 (1H, d, J = 8.5 Hz), 8.34 (2H, d, J = 8.5 Hz), 9.24 (1H,
d, J = 8.5 Hz); 13C NMR (10%TFA-d-CD3OD, 125 MHz) δ 7.4, 103.0,
105.9, 110.2, 116.4, 129.5, 131.0, 131.1, 136.3, 150.6, 157.7, 159.4,
171.5, 172.8; ESI-TOF HRMS Calcd. For C16H13O3 [M]+, 253.0859;
Found: 253.0865.
Keywords: 8-substituted-3-deoxyanthocyanidin • 6,8-
rearrangement • β-cyclodextrin • Suzuki-Miyaura reaction •
photochemistry
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Entry for the Table of Contents (Please choose one layout)
Layout 2:
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Nuno Basílio, João Carlos Lima, Luís
Cruz, Victor de Freitas, Fernando Pina,*
Hiroki Ando, Yuki Kimura, Kin-Ichi
Oyama, Kumi Yoshida,*
Page No. – Page No.
6,8-rearrangement of the compounds 8-phenyl-5,7-dihydroxyflavylium and 8-
methyl-5,7-dihydroxyflavylium is not observed at pH=1. However, the 6,8-
rearrangement could be investigated by extending these studies to the moderately
acidic region in particular in the presence of a β-cyclodextrin derivative (captisol).
Unveiling the 6,8-rearrangement in 8-
phenyl-5,7-dihydroxyflavylium and 8-
methyl-5,7-dihydroxyflavylium
through host-guest complexation
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