Synthesis and evaluation of diverse thio avarol derivatives as potential UVB photoprotective candidates

Article abstract of DOI:10.1016/j.bmcl.2007.02.007

Semisynthesis of 13 new thio avarol derivatives (4-16) and in vitro evaluation on the photodamage response induced by UVB irradiation are described. Their ability to inhibit NF-κB activation and TNF-α generation in HaCaT cells as well as their antioxidant

Full text of DOI:10.1016/j.bmcl.2007.02.007

Bioorganic & Medicinal Chemistry Letters 17 (2007) 2561–2565
Synthesis and evaluation of diverse thio avarol derivatives
as potential UVB photoprotective candidates
a
a
a,
*
´
´
´
´
Marıa Amigo, Marıa C. Terencio, Miguel Paya,
Carmine Iodiceb and Salvatore De Rosab
aDepartamento de Farmacologı´a, Facultad de Farmacia, Universidad de Valencia, Av. Vicente Andre´s Estelle´s s/n,
46100 Burjasot, Valencia, Spain
bIstituto di Chimica Biomolecolare, CNR, Via Campi Flegrei, 34, 80078 Pozzuoli, Napoli, Italy
Received 19 December 2006; revised 1 February 2007; accepted 3 February 2007
Available online 7 February 2007
Abstract—Semisynthesis of 13 new thio avarol derivatives (4–16) and in vitro evaluation on the photodamage response induced by
UVB irradiation are described. Their ability to inhibit NF-jB activation and TNF-a generation in HaCaT cells as well as their anti-
oxidant capacity in human neutrophils has also been studied. Among them we have identified two monophenyl thio avarol deriv-
atives (4–5) lacking cytotoxicity which can be considered promising UVB photoprotective agents through the potent inhibition of
NF-jB activation with a mild antioxidant pharmacological profile.
ꢀ 2007 Elsevier Ltd. All rights reserved.
Ultraviolet radiation (UVR) is known for its negative
impact on health. The skin, situated at the interface be-
tween the body and its environment, directly suffers
from the deleterious effects of UV radiation. This UV
radiation jeopardizes the integrity of the skin that is crit-
ical for cellular homeostasis. The component of the solar
UV-light that reaches the earth’s surface and that is
most responsible for acute and long-term effects is
UVB (280–320 nm). Acute effects include erythema, tan-
ning, and inflammatory and immune modulatory chang-
es. Chronic exposure eventually leads to photoaging1
and photocarcinogenesis.2,3 In recent years, photo-
chemoprevention has matured into an accepted modali-
ty for controlling skin cancer. UVB mediated
inflammation that includes the release of growth factors,
proinflammatory cytokines, infiltration of inflammatory
cells, and radical oxygen species (ROS) production,4
plays an important role in skin cancer development. Dif-
ferent studies have demonstrated that UV-induced acti-
vation of NF-jB-dependent gene transactivation
pathways is a critical event for the subsequent develop-
ment of sunburn reactions in skin.5,6 Supplementation
of antioxidants a-lipoic acid, N-acetyl-L-cysteine
(NAC), and the flavonoid extract silymarin modulates
the activation of the transcription factor NF-jB in
HaCaT keratinocytes after exposure to a solar UV
simulator.7,8 These results indicate that some antioxi-
dants can efficiently amend the cellular response to UV
radiation through their selective action on NF-jB
activation.9,10 It is well established that NF-jB is activat-
ed upon UV irradiation and induces various genes includ-
ing IL-1 and TNF-a, which subsequently stimulate
the signal transduction pathway to activate NF-jB.11
Cutaneous alterations mediated by UV irradiation
through the NF-jB activation pathway could be likewise
effectively prevented by blocking NF-jB activation.
Keratinocytes are the major target of UVR and play a
central role in the inflammatory and immune modulato-
ry changes observed after UV exposure, at least partly
via the UV-induced release of cytokines.12
Avarol is a marine sesquiterpenoid hydroquinone with
interesting pharmacological properties13 including anti-
inflammatory,14 antitumor,15 antioxidant,16 antiplate-
let,17 anti-HIV,18 and antipsoriatic19 effects. Recently,
we reported that its derivative 30-(salicylthio) avarol
(3) inhibited superoxide generation in stimulated human
neutrophils and PGE2 release in the human HaCaT
keratinocyte cell line,20 offering interesting perspectives
as a possible anti-inflammatory or antipsoriatic drug.
In this paper, the effects of 3 and 13 new thio avarol
Keywords: Keratinocytes; TNF-a; NF-jB; UVB; Thio avarol
derivatives.
*
Corresponding author. Tel.: +34 963544946; fax: +34 963544943;
e-mail: Miguel.paya@uv.es
0960-894X/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.02.007

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M. Amigo et al. / Bioorg. Med. Chem. Lett. 17 (2007) 2561–2565
2562
derivatives (4–16) are investigated, with respect to the
response induced by UVB irradiation in HaCaT cells.
Their ability to inhibit in vitro NF-jB activation and
TNF-a generation, as well as their antioxidant capacity
in human neutrophils, has also been studied.
production, and NF-jB activation. Cells were cultured
in DMEM supplemented with 100 U/mL penicillin,
100 lg/mL streptomycin, and 10% fetal bovine serum
at 37 ꢁC in 5% CO2. For TNF-a experiment cells were
seeded in 96-well plate at 2 · 105 cells/mL and in 6-well
plate at 4 · 105 cells/mL for NF-jB activation experi-
ment. HaCaT cells were incubated overnight until con-
fluence was reached. Two hours before irradiation, the
cells were washed twice in phosphate-buffered saline
(PBS) and the compounds were added dissolved in eth-
anol. Cells were irradiated through a thin film of PBS
with a single sublethal UVB dose (40–50 mJ/cm2) using
a LuzChem LZC-5 photoreactor calibrated for UVB
irradiation (280–320 nm) with an emission peak cen-
tered at 313 nm. After the UVB exposure, fresh culture
medium was added to the cells and media were harvest-
ed at the appropriated time points. Control conditions
consisted of cells pretreated with identical concentra-
tions of ethanol. Final ethanol concentration did not
exceed 0.1% and did not affect the experiments. The
mitochondrial-dependent reduction of 3-(4,5-dimethyl-
thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)
to formazan25 was used to assess the possible cytotoxic
effects of compounds on HaCaT cells after 24 h of
irradiation. The concentration of TNF-a in the
culture supernatants derived from HaCaT cells 24 h
after UVB irradiation26 was determined by time-re-
solved fluoroimmunoassay.27 NF-jB activation was
performed 18 h after irradiation26 by the electrophoretic
mobility shift assay (EMSA).28 Human neutrophils
(2.5 · 106 cells/mL) stimulated with 12-O-tetradecanoyl
phorbol 13-acetate (TPA) were used to evaluate the
inhibitory effect on ROS of the thio avarol derivatives
studied.29
Avarol (1) was isolated from the sponge Dysidea
avara,21 collected in the Bay of Naples, Italy. Avarone
(2) was obtained by Ag2O oxidation of avarol, in etha-
nol, as previously reported.21 The compound 3 was
obtained by adding thiosalicylic acid to a solution of
avarone (2) in ethanol, as previously reported.20 Thio
derivatives (4–16) (Fig. 1) were generally obtained by
slowly adding the corresponding thio compound dis-
solved in ethanol to a solution of avarone in ethanol.22
On the one hand, for the thioglycerol and thioglycolic
acid two isomers were obtained with substitution at 30
(13 and 15) and 40 (14 and 16) of the benzoquinone ring.
On the other hand, for the thiophenol, p-thiocresol, and
thioglycol three isomers were obtained with substitution
at 30 (4, 7, and 10), 40 (5, 8, and 11), and in both 30 and 40
(6, 9, and 12). The position of the substituent was
1
determined by the analysis of H NMR spectra. Signals
of protons in the benzoquinone ring are doublets in
30-substituted compounds and singlets in 40-substituted
compounds, whereas 30,40-disubstituted compounds
show only one signal of proton as singlet. Only chemical
shifts of hydroquinone and thio residues are reported
because all other signals belonging to the sesquiterpe-
noid portion were earlier reported.20,23
Biological activities of the thio avarol derivatives were
evaluated in spontaneously immortalized human kerati-
nocyte cell line (HaCaT)24 for cellular viability, TNF-a
R1
O
HO
R2
O
H
OH
H
Avarone (2)
1 R1 = R2 = H (avarol)
3 R2 = H, R1 =
9 R1 = R2 =
S
S
10 R2 = H, R1 = CH2OH-CH2S
11 R1 = H, R2 = CH2OH-CH2S
COOH
S
4 R2 = H, R1 =
5 R1 = H, R2 =
12 R1 = R2 = CH2OH-CH2S
S
S
13 R2 = H, R1 = CH2OH-CHOH-CH2S
6 R1 = R2 =
14 R1 = H, R2 = CH2OH-CHOH-CH2S
15 R2 = H, R1 = HOOC-CH2S
7 R2 = H, R1 =
S
8 R1 = H, R2 =
S
16 R1 = H, R2 = HOOC-CH2S
Figure 1. Chemical structures of the thio avarol derivatives.

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M. Amigo et al. / Bioorg. Med. Chem. Lett. 17 (2007) 2561–2565
2563
Table 1. Inhibitory effect of thio avarol derivatives on cellular viability and TNF-a production in UVB irradiated HaCaT cells
Compound
Viability
TNF-a
% I (10 lM)
% I (5 lM)
% I (10 lM)
% I (5 lM)
IC50 (lM)
3
15.7 1.9*
0.0 0.0
0.0 0.0
0.6 0.3
0.0 0.0
0.0 0.0
0.0 0.0
0.0 0.0
0.0 0.0
0.0 0.0
20.6 2.6*
21.4 1.3*
18.1 1.2*
13.6 0.2
12.6 3.8
18.3 1.2*
12.4 2.9
0.0 0.0
ND
76.3 6.4**
55.4 3.7**
53.6 4.6**
0.0 0.0
0.0 0.0
0.0 0.0
0.0 0.0
ND
3.3 (2.3–5.0)
3.7 (2.5–4.9)
4.2 (2.1–5.6)
4
92.7 2.5**
74.3 10.7**
0.0 0.0
0.0 0.0
0.0 0.0
0.0 0.0
ND
5
6
0.0 0.0
0.0 0.0
7
8
0.0 0.0
0.0 0.0
9
10
22.5 2.0*
48.4 1.8**
28.5 3.0**
14.0 2.3
18.6 1.5*
28.5 3.5**
13.3 1.1
3.4 2.1
11
12
ND
ND
ND
ND
13
14
ND
ND
ND
ND
15
16
ND
ND
68.1 6.6**
ND
ND
58.6 3.7**
Fraxetin
2.8 (1.4–8.7)
Data were calculated as inhibitory percentages (% I) by taking the values of maximal viability or TNF-a production in drug-free samples as 100%.
Values represent means SEM (n = 6). IC50 values and 95% confidence intervals were calculated from at least four significant concentrations for
those compounds that reached 50% of inhibition at 10 lM. **P < 0.01 and *P < 0.05 with respect to the control group.
UVB
Table 1 shows the inhibitory effect of thio avarol deriv-
5 µM
3
10 µM
5
atives (3–16) on cellular viability and TNF-a production
in UVB-irradiated HaCaT cells. The glycyl thio (10–12),
glyceryl thio (13–14), and glycolyl thio (15–16) avarol
derivatives exerted a significant cytotoxicity at both dos-
es tested, whereas phenyl thio (4–6) and cresyl thio (7–9)
avarol derivatives showed absence of cytotoxicity. TNF-
a production was only determined at concentrations
which showed an inhibition of cell viability lower than
10%. Among the noncytotoxic compounds only the
monophenyl thio (4–5) and the salicyl thio (3) avarol
derivatives were able to inhibit TNF-a production with
IC50 values very close to the reference compound fraxe-
tin, antioxidant capable to inhibit TNF-a release.30
Then, we investigated whether the changes in TNF-a
production produced by the thio avarol derivatives (3–
5) correlated with changes in NF-jB activation. Nuclear
protein extracts from UVB-irradiated HaCaT cells were
analyzed for NF-jB-DNA binding activity using a radi-
olabeled NF-jB-specific oligonucleotide, either in the
presence or absence of compounds (3–5) (Fig. 2). The
inhibitory effect of compounds (3–5) was more potent
than proteasome inhibitor MG132 at 10 lM.
B
C
4
MG
NF-κB
Figure 2. Inhibitory effect of compounds 3–5 on NF-jB activation in
nuclear extracts of UVB-irradiated (40–50 mJ/cm2) HaCaT cells. B,
non-UVB-irradiated cells. C, UVB irradiated cells. MG, MG132. The
figure is representative of three independent experiments.
90
**
80
**
70
**
60
50
40
30
20
10
0
**
*
**
In the assay of ROS generation in stimulated human neu-
trophils, all of these compounds inhibited the generation
of oxygen-derived species in a concentration-dependent
manner, being the salicyl thio avarol derivative the most
potent antioxidant compound (Fig. 3).
3
4
5
3
4
5
3
4
5
10 μM
5 μM
1 μM
The importance of the skin as an immunologic organ is
now recognized. In addition to being a mechanical barri-
er, the skin is critically involved in the regulation of im-
mune responses that can affect the health of the entire
organism.31 Keratinocytes are the major constituent of
the epidermis and can synthesize, store, and release differ-
ent cytokines, including TNF-a.31,32 The skin is continu-
ally exposed to oxidants such as UVR, air pollutants, and
chemicals. In order to prevent deleterious oxidative reac-
tions, endogenous antioxidants (both enzymatic and non-
Figure 3. Inhibitory effect of compounds 3–5 on radical oxygen species
in TPA stimulated human neutrophils. Results show percentages of
inhibition. Statistical evaluation included one-way analysis of variance
followed by Dunnett’s t test. Data represent means SEM (n = 6).
**P < 0.01 and *P < 0.05 with respect to the control group.
enzymatic) are present in the skin. UVB is considered to
be the main component of the solar UV irradiation acti-
vating NF-jB in HaCaT cells,8 being the activation of
NF-jB significantly enhanced 18 h after UVB irradia-

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M. Amigo et al. / Bioorg. Med. Chem. Lett. 17 (2007) 2561–2565
2564
tion.26 Topical application of antioxidants has been used
to prevent skin antioxidant loss and photooxidative dam-
age. However, in addition to the capacity of antioxidants
to prevent this damage in the skin, growing evidence indi-
cates that they also play a role in modulating the expres-
sion of genes whose products are involved in
inflammation, aging, and carcinogenesis.7,8,33
5. Abeyama, K.; Eng, W.; Jester, J. V.; Vink, A. A.;
Edelbaum, D.; Cockerell, C. J.; Bergstresser, P. R.;
Takashima, A. J. Clin. Invest. 2000, 105, 1751.
6. Fisher, G. J.; Datta, S. C.; Talwar, H. S.; Wang, Z. Q.;
Varani, J.; Kang, S.; Voorhees, J. J. Nature 1996, 379, 335.
7. Larsson, P.; Ollinger, K.; Rosdahl, I. Br. J. Dermatol.
2006, 155, 292.
8. Saliou, C.; Kitazawa, M.; McLaughlin, L.; Yang, J. P.;
Lodge, J. K.; Tetsuka, T.; Iwasaki, K.; Cillard, J.;Okamoto,
T.; Packer, L. Free Radical Biol. Med. 1999, 26, 174.
9. Mantena, S. K.; Katiyar, S. K. Free Radical Biol. Med.
2006, 40, 1603.
It is reported that NF-jB activation and translocation is
previous to TNF-a production26 and that not all antiox-
idants are equally effective inhibitors in UVB-irradiated
keratinocytes.34 The critical event for UVB erythema-
tous damage seems to be the NF-jB-dependent gene
transactivation.10
10. Afaq, F.; Adhami, V. M.; Mukhtar, H. Mutat. Res. 2005,
571, 153.
11. Tanaka, K.; Hasegawa, J.; Asamitsu, K.; Okamoto, T.
J. Pharmacol. Exp. Ther. 2005, 315, 624.
12. Pupe, A.; Moison, R.; De Haes, P.; van Henegouwen, G.
B.; Rhodes, L.; Degreef, H.; Garmyn, M. J. Invest.
Dermatol. 2002, 118, 692.
13. De Rosa, S. In Natural Products in the New Millennium:
Prospects and Industrial Application; Rauter, A. P., Palma,
F. B., Justino, J., Arau´jo, M. E., Santos, S. P., Eds.; Kluwer
Academic Publishers: Dordrecht, 2002; pp 441–461.
We have identified several thio derivatives of avarol, a
well-reputed antipsoriatic drug, as effective UVB photo-
protective candidates acting mainly through the potent
inhibition of NF-jB activation. In addition, these prod-
ucts clearly reduced TNF-a generation in keratinocytes
and demonstrated antioxidant properties. These results
are according to other studies indicating that ROS are
implicated in the induction of TNF-a expression by
UVBviaNF-jB-dependent pathway.10,26,34 Inthisregard
it is interesting to note that the best antioxidant properties
of compound 3 correlate with the more potent inhibition
of NF-jB activation showed by this compound.
´
´
14. Ferrandiz, M. L.; Sanz, M. J.; Bustos, G.; Paya, M.;
Alcaraz, M. J.; De Rosa, S. Eur. J. Pharmacol. 1994, 253, 75.
15. Shen, Y. C.; Lu, C. H.; Chakraborty, R.; Kuo, Y. H. Nat.
Prod. Res. 2003, 17, 83.
16. Belisario, M. A.; Maturo, M.; Pecce, R.; De Rosa, S.;
Villani, G. R. Toxicology 1992, 72, 221.
17. Belisario, M. A.; Maturo, M.; Avagnale, G.; De Rosa, S.;
Scopacasa, F.; De Caterina, M. Pharmacol. Toxicol. 1996,
79, 300.
18. Sarin, P. S.; Sun, D.; Thornton, A.; Muller, W. E. G.
J. Natl. Cancer Inst. 1987, 78, 663.
19. Sipkema, D.; Osinga, R.; Schatton, W.; Mendola, D.;
Tramper, J.; Wijffels, R. H. Biotechnol. Bioeng. 2005, 90, 201.
Results obtained in the present study show some clear
structure–activity relationships. In this regard, monophe-
nyl thio avarol derivatives (4–5), as well as salicyl thio ava-
rol derivative (3), are the only ones that exert a potent
UVB photoprotective profile. The disubstituted phenyl
thio avarol derivative (6) presents a total reduction in
activity probably related with the higher steric hindrance
of the bulk. The methylation of phenyl thiol moiety in
order to get the respective cresyl thio derivatives (7–9)
produces a lack of inhibitory activity. In addition, the
presence of glycyl thio/glyceryl thio and glycolyl thio
moieties seems to increase the cytotoxicity (10–16).
´
´
20. Amigo, M.; Terencio, M. C.; Mitova, M.; Iodice, C.; Paya,
M.; De Rosa, S. J. Nat. Prod. 2004, 67, 1459.
21. Minale, L.; Riccio, R.; Sodano, G. Tetrahedron Lett. 1974,
15, 3401.
22. Synthesis of 30-(phenylthio)avarol (4), 40-(phenylthio)ava-
rol (5), and 30,40-(phenylthio)avarol (6). Thiophenol
(100 lL) dissolved in EtOH (5 mL) was added to a
solution of avarone (2) (100 mg) in EtOH (10 mL) and
stirred for 5 min at room temperature. After evaporation
of EtOH, the residue was chromatographed on a Si gel
column and eluted with petroleum ether/Et2O (4:1), to give
30-(phenylthio)avarol (4) as the more polar component
In summary, salicyl thio avarol and monophenyl thio
avarol derivatives can be considered promising UVB
photoprotective agents through the potent inhibition
of NF-jB activation and the interference with cellular
processes mediated by ROS generation.
25
(65 mg; yield 48.0%): amorphous solid; ½aꢀD 7.1ꢁ (c 0.20,
CHCl3); UV (MeOH) kmax (loge) 313 (3.36), 433 (1.71); 1H
NMR (CDCl3) d 7.20 (2H, dd, J = 7.7 and 7.3 Hz, H-300
and H-500), 7.14 (H, d, J = 7.7 Hz, H-400), 7.06 (2H, d,
J = 7.3 Hz, H-200 and H-600), 6.88 (1H, d, J = 2.9 Hz, H-40),
6.73 (1H, d, J = 2.9 Hz, H-60); EIMS m/z 424 [M+2]+
(0.4), 422 [M]+ (12), 233 (32), 192 (12), 189 (8), 107 (35), 95
(100); HREIMS m/z 422.2281 (Calcd for C27H34O2S,
422.2279), 40-(phenylthio)avarol (5) (13 mg; yield 9.6%):
Acknowledgments
M.A. was the recipient of a Research Fellowship from
´
the FPU program of Spanish Ministerio de Educacion
25
y Ciencia. This work was supported partly by the Grant
UV-AE-20060243 from the University of Valencia and
by FIS-PI051659.
amorphous solid; ½aꢀD 28.7ꢁ (c 0.10, CHCl3); UV (MeOH)
k
max (loge) 312 (3.43), 430 (2.32); 1H NMR (CDCl3) d 7.22
(2H, dd, J = 7.7 and 7.3 Hz, H-300 and H-500), 7.15 (H, d,
J = 7.7 Hz, H-400), 7.09 (2H, d, J = 7.3 Hz, H-200 and H-
600), 6.88 (1H, s, H-40), 6.82 (1H, s, H-60); EIMS m/z 424
[M+2]+ (0.5), 422 [M]+ (10), 233 (30), 192 (15), 189 (10),
107 (35), 95 (100); HREIMS m/z 422.2281 (Calcd for
C27H34O2S, 422.2279), and 30,40-(phenylthio)avarol (6) as
the less polar component (6 mg; yield 3.5%): amorphous
References and notes
1. Rittie, L.; Fisher, G. J. Ageing Res. Rev. 2002, 1, 705.
2. Hussein, M. R. J. Cutan. Pathol. 2005, 32, 191.
3. Matsumura, Y.; Ananthaswamy, H. N. Toxicol. Appl.
Pharmacol. 2004, 195, 298.
25
solid; ½aꢀD 7.3ꢁ (c 0.05, CHCl3); UV (MeOH) kmax (loge)
332 (3.52), 433 (1.70), 492 (1.21); 1H NMR (CDCl3) d 7.16
(6H, m, H-300, H-400 and H-500), 6.97 (4H, m, H-200 and H-
4. Halliday, G. M. Mutat. Res. 2005, 571, 107.

´
M. Amigo et al. / Bioorg. Med. Chem. Lett. 17 (2007) 2561–2565
2565
600), 6.65 (1H, s, H-60); EIMS m/z 532 [M+2]+ (0.6), 530
[M]+ (12), 233 (30), 192 (12), 189 (8), 107 (35), 95 (100);
HREIMS m/z 530.2310 (Calcd for C33H38O2S2, 530.2313).
Synthesis of 30-(cresylthio)avarol (7), 40-(cresylthio)ava-
rol (8), and 30,40-(cresylthio)avarol (9). p-Thiocresol
(100 lL) dissolved in EtOH (5 mL) was added to a
solution of avarone (2) (100 mg) in EtOH (10 mL) and
stirred for 5 min at room temperature. After evaporation
of EtOH, the residue was chromatographed on a Si gel
column and eluted with petroleum ether/Et2O (4:1), to give
30-(cresylthio)avarol (7) as the more polar component
(100 mg) in EtOH (10 mL) and stirred for 30 min at room
temperature. After evaporation of EtOH, the residue was
chromatographed on a Si gel column and eluted with ethyl
acetate, to give 30-(glycerylthio)avarol (13) as the less polar
component (45 mg; yield 33.4 %): amorphous solid;
25
½aꢀD 8.6ꢁ (c 0.20, CHCl3); UV (MeOH) kmax (loge) 310
(3.18), 433 (2.32); 1H NMR (CDCl3) d 6.85 (1H, d,
J = 2.7 Hz, H-40), 6.63 (1H, d, J = 2.7 Hz, H-60), 3.67 (2H,
m, H-200 and H-3a00); 3.50 (1H, m, H-3b00), 2.77 (1H, dd,
J = 13.9 and 6.4 Hz, H-1a00), 2.59 (1H, dd, J = 13.9 and
7.6 Hz, H-1b00); EIMS m/z 422 [M+2]+ (0.4), 420 [M]+ (10),
258 (8), 229 (35), 191 (25), 155 (15), 107 (60), 95 (100);
HREIMS m/z 420.2330 (Calcd for C24H36O4S, 420.2334),
and 40-(glycerylthio)avarol (14) as the less polar component
25
(60 mg; yield 43.0 %): amorphous solid; ½aꢀD 9.2ꢁ (c 0.14,
CHCl3); UV (MeOH) kmax (loge) 312 (3.35), 433 (2.40); 1H
NMR (CDCl3) d 7.01 (2H, d, J = 8.3 Hz, H-300 and H-500),
6.98 (2H, d, J = 8.3 Hz, H-200 and H-600), 6.86 (1H, d,
J = 2.9 Hz, H-40), 6.70 (1H, d, J = 2.9 Hz, H-60), 2.29 (3H,
s, H-700); EIMS m/z 438 [M+2]+ (0.4), 436 [M]+ (10), 232
(98), 191 (25), 107 (40), 95 (100); HREIMS m/z 436.2445
(Calcd for C28H36O2S, 436.2442), 40-(cresylthio)avarol (8)
25
(40 mg; yield 29.7 %): amorphous solid; ½aꢀD 32.7ꢁ (c 0.20,
CHCl3); UV (MeOH) kmax (loge) 309 (3.86), 433 (3.07); 1H
NMR (CDCl3) d 6.86 (1H, s, H-30), 6.73 (1H, s, H-60), 3.72
(2H, m, H-200 and H-3a00); 3.51 (1H, dd, J = 11.2 and
6.30 Hz, H-3b00), 2.87 (1H, dd, J = 13.8 and 4.1 Hz, H-1a00),
2.74 (1H, dd, J = 13.8 and 8.3 Hz, H-1b00); EIMS m/z 422
[M+2]+ (0.4), 420 [M]+ (12), 258 (8), 229 (30), 191 (20), 155
(20), 107 (50), 95 (100); HREIMS m/z 420.2330 (Calcd for
C24H36O4S, 420.2334).
25
(25 mg; yield 18.0%): amorphous solid; ½aꢀD 21.7ꢁ (c 0.10,
CHCl3); UV (MeOH) kmax (loge) 312 (3.76), 430 (2.73); 1H
NMR (CDCl3) d 7.02 (2H, d, J = 8.3 Hz, H-300 and H-500),
6.98 (2H, d, J = 8.3 Hz, H-200 and H-600), 6.85 (1H, s, H-
30), 6.80 (1H, s, H-60), 2.30 (3H, s, H-700); EIMS m/z 438
[M+2]+ (0.4), 436 [M]+ (12), 232 (95), 191 (30), 107 (45), 95
(100); HREIMS m/z 436.2445 (Calcd for C28H36O2S,
436.2442), and 30,40-(cresylthio)avarol (9) as the less polar
Synthesis of 30-(glycolylthio)avarol (15) and 40-(glycolyl-
thio)avarol (16). Thioglycolic acid (200 lL) dissolved in
EtOH (5 mL) was added to a solution of avarone (2)
(100 mg) in EtOH (10 mL) and stirred for 3 h at room
temperature. After evaporation of EtOH, the residue was
chromatographed on a Si gel column and eluted with
petroleum ether–Et2O–HOAc (7:3:0.1), to give 30-(glycol-
ylthio)avarol (15) as the less polar component (34 mg; yield
25
component (15 mg; yield 8.4%): amorphous solid; ½aꢀD
9.9ꢁ (c 0.10, CHCl3); UV (MeOH) kmax (loge) 332 (3.57),
433 (1.98), 441 (1.99); 1H NMR (CDCl3) d 6.98 (4H, d,
J = 8.3 Hz, H-300 and H-500), 6.87 (4H, d, J = 8.3 Hz, H-200
and H-600), 6.68 (1H, s, H-60), 2.26 (6H, s, H-700); EIMS m/
z 560 [M+2]+ (0.7), 558 [M]+ (10), 232 (55), 191 (25), 107
(40), 95 (100); HREIMS m/z 558.2630 (Calcd for
C35H42O2S2, 558.2626).
25
26.3%): amorphous solid; ½aꢀD 14.4ꢁ (c 0.15, CHCl3); UV
(MeOH) kmax (loge) 312 (4.35), 433 (2.93); 1H NMR
(CDCl3) d 6.88 (1H, d, J = 2.9 Hz, H-40), 6.65 (1H, d,
J = 2.9 Hz, H-60), 3.47 (2H, s, H-100); EIMS m/z 406 [M+2]+
(0.4), 404 [M]+ (12), 229 12(35), 191 (25), 107 (60), 95 (100);
HREIMS m/z 404.2017 (Calcd for C23H32O4S, 404.2021),
and 40-(glycolylthio)avarol (16) as the less polar component
Synthesis of 30-(glycylthio)avarol (10), 40-(glycylthio)ava-
rol (11), and 30,40-(glycylthio)avarol (12). Thioglycol
(200 lL) dissolved in EtOH (5 mL) was added to a solution
of avarone (2) (100 mg) in EtOH (10 mL) and stirred for
5 min at room temperature. After evaporation of EtOH, the
residue was chromatographed on a Si gel column and eluted
with petroleum ether/Et2O/HOAc (7:3:0.1), to give 30-
(glycylthio)avarol (10) as the less polar component (25 mg;
25
(28 mg; yield 21.6%): amorphous solid; ½aꢀD 14.0ꢁ (c 0.15,
CHCl3); UV (MeOH) kmax(loge) 311 (3.52), 433 (2.20); 1H
NMR (CDCl3) d 6.81 (1H, s, H-30), 6.65 (1H, s, H-60), 3.43
(2H, s, H-100); EIMS m/z 406 [M+2]+ (0.4), 404 [M]+ (8), 229
(25), 191 (20), 107 (50), 95 (100); HREIMS m/z 404.2025
(Calcd for C23H32O4S, 404.2021).
25
yield 20.0%): amorphous solid; ½aꢀD 7.1ꢁ (c 0.15, CHCl3);
1
UV (MeOH) kmax (loge) 312 (3.54), 433 (2.45); H NMR
23. De Rosa, S.; Minale, L.; Riccio, R.; Sodano, G. J. Chem.
Soc., Perkin Trans. 1 1976, 1408.
24. Boukamp, P.;Petrussevska, R. T.; Breitkreutz, D.;Hornung,
J.; Markham, A.; Fusenig, N. E. J. Cell Biol. 1988, 106, 761.
25. Borenfreund, E.; Babick, H.; Martin-Alguacil, N. Toxicol.
In Vitro 1988, 2, 1.
26. Lan, C. C.; Yu, H. S.; Wu, C. S.; Kuo, H. Y.; Chai, C. Y.;
Chen, G. S. Br. J. Dermatol. 2005, 153, 725.
27. Pennanen, N.; Lapinjoki, S.; Palander, A.; Urtti, A.;
Monkkonen, J. Int. J. Immunopharmacol. 1995, 17, 475.
(CDCl3) d 6.85 (1H, d, J = 2.9 Hz, H-40), 6.63 (1H, d,
J = 2.9 Hz, H-60), 3.67 (2H, t, J = 5.9 Hz, H-200); 2.86 (2H, t,
J = 5.9 Hz, H-100); EIMS m/z 392 [M+2]+ (0.4), 390 [M]+
(10), 343 (5), 258 (8), 196 (18), 189 (28), 107 (30), 95 (100);
HREIMS m/z 390.2230 (calcd for C23H34O3S, 390.2228), 40-
(glycylthio)avarol (11) (40 mg; yield 32.0%): amorphous
25
solid; ½aꢀD 11.0ꢁ (c 0.20, CHCl3); UV (MeOH) kmax (loge)
310 (3.61), 433(2.39); 1H NMR(CDCl3)d 6.86 (1H, s, H-30),
6.73 (1H, s, H-60), 3.73 (2H, t, J = 5.9 Hz, H-200); 2.89 (2H, t,
J = 5.9 Hz, H-100); EIMS m/z 392 [M+2]+ (0.4), 390 [M]+
(12), 343 (5), 258 (10), 196 (15), 189 (25), 107 (35), 95 (100);
HREIMS m/z 390.2230 (Calcd for C23H34O3S, 390.2228),
and 30,40-(glycylthio)avarol (12) as the more polar compo-
nent (6 mg; yield 4.0 %): amorphous solid; ½aꢀ2D5 20.4ꢁ (c 0.05,
CHCl3); UV (MeOH) kmax (loge) 327 (3.94), 433 (2.28); 1H
NMR (CDCl3) d 6.85 (1H, s, H-60), 3.66 (2H, t, J = 5.9 Hz,
H-200), 3.60 (2H, t, J = 5.9 Hz, H-200), 2.96 (2H, t, J = 5.9 Hz,
H-100), 2.92 (2H, t, J = 5.9 Hz, H-100); EIMS m/z 468 [M+2]+
(0.6), 466 [M]+ (10), 343 (3), 258 (5), 196 (15), 189 (20), 107
(30), 95 (100); HREIMS m/z 466.2214 (Calcd for
C25H38O4S2, 466.2211).
´
28. Amigo, M.; Schalkwijk, J.; Olthuis, D.; De Rosa, S.; Paya,
M.; Terencio, M. C.; Lamme, E. Life Sci. 2006, 79, 2395.
´
´
´
29. Bustos, G.; Ferrandiz, M. L.; Sanz, M. J.; Paya, M.;
Alcaraz, M. J. Naunyn-Schmiedeberg’s Arch. Pharmacol.
1995, 351, 298.
30. Kuo, P. L.; Huang, Y. T.; Chang, C. H.; Chang, J. K. Int.
J. Immunopharmacol. 2006, 6, 1167.
31. Oberyszyn, T. M.; Tober, K. L.; Ross, M. S.; Robertson,
F. M. Mol. Carcinog. 1998, 22, 16.
32. Banno, T.; Gazel, A.; Blumenberg, M. J. Biol. Chem.
2004, 279, 32633.
33. Jurkiewicz, B. A.; Bissett, D. L.; Buettner, G. R. J. Invest.
Dermatol. 1995, 104, 484.
Synthesis of 30-(glycerylthio)avarol (13) and 40-(glyceryl-
thio)avarol (14). 1-Thioglycerol (100 lL) dissolved in
EtOH (5 mL) was added to a solution of avarone (2)
34. Pupe, A.; Degreef, H.; Garmyn, M. Photochem. Photobiol.
2003, 78, 68.