Synthesis and melanin biosynthesis inhibitory activity of (±)-terrein produced by Penicillium sp. 20135

Article abstract of DOI:10.1016/j.bmcl.2004.10.057

Terrein was isolated from Penicillium sp. 20135, prepared by a practical synthetic way, and evaluated first time for its melanin biosynthesis inhibitory activity. Terrein was isolated from Penicillium sp. 20135, prepared by a practical synthetic way, and evaluated first time for its melanin biosynthesis inhibitory activity.

Full text of DOI:10.1016/j.bmcl.2004.10.057

Bioorganic & Medicinal Chemistry Letters 15 (2005) 471–473
Synthesis and melanin biosynthesis inhibitory activity of
( )-terrein produced by Penicillium sp. 20135
Sangku Lee,a Won-Gon Kim,a Eungsoo Kim,a In-Ja Ryoo,a Hyeong Kyu Lee,a
Jae Nyoung Kim,b Sang-Hun Jungc and Ick-Dong Yooa,*
aKorea Research Institute of Bioscience and Biotechnology, 52 Oun, Yusong, Taejon 305-333, Republic of Korea
bDepartment of Chemistry and Institute of Basic Science, Chonnam National University, Gwangju 500-757, Republic of Korea
cCollege of Pharmacy, Chungnam National University, Taejon 305-764, Republic of Korea
Received 14 July 2004; accepted 12 October 2004
Available online 6 November 2004
Abstract—Terrein was isolated from Penicillium sp. 20135, prepared by a practical synthetic way, and evaluated first time for its
melanin biosynthesis inhibitory activity.
Ó 2004 Elsevier Ltd. All rights reserved.
Melanin pigments are known to be biosynthesized in the
melanosomes of melanocytes, moved to kerotinocytes,
and stored in the epidermis.1 Hyperpigmentation such
as chloasma, colouration or freckles increased abnor-
mally the amounts of melanin in the epidermis. Melano-
genesis consists of two rate-limiting reactions catalyzed
by tyrosinase, that is, the hydroxylation of tyrosine
to 3,4-dihydroxyphenylalanine (DOPA) and the
subsequent oxidation of DOPA to DOPA-quinone.2
Thus melanin biosynthesis inhibitors, such as inhibitors
of tyrosinase or regulators of tyrosinase expression and
activation, may be useful as a skin-whitening agent in
cosmetics.
Mel-Ab, terrein (1) have been newly isolated from Pen-
icillium sp. 20135 as a racemic form with an optical rota-
tion of zero (c 0.1, MeOH)6 (Fig. 1). Although terrein
was discovered nearly seventy years ago,7 there have
been only a few reports about its biological activities.8,9
Terrein was reported to show plant growth inhibition8
and antibacterial activities.9 We found that terrein
inhibited effectively melanin formation in melanocyte
Mel-Ab cells. As we have been interested in further bio-
logical studies including in vivo activities, we needed a
practical synthetic approach to terrein, thereby also ena-
bling access to terrein analogues for examining struc-
ture–activity relationships. Herein we describe
a
practical synthesis of terrein and its melanin biosynthe-
sis inhibitory activity.
Fungi are a source of many important biomedical sub-
stances. Kojic acid currently used as a cosmetic agent
for the purpose of skin whitening was isolated from
Aspergillus oryzae.3 Stronger and safer skin whitening
agents, however, are needed because of a carcinogenic
potential as well as a weak whitening effect of kojic
acid.4 For exploring a new class of cosmetic agents, we
have screened melanin biosynthesis inhibitors from fun-
gal metabolites and reported strong tyrosinase inhibi-
tors, melanocins A–C, produced by Eupenicillium
shearii.5 In the course of our continuous screening for
melanin biosynthesis inhibitors using melanocyte cells
The synthesis of terrein was reported by several research
groups. Auerbach and Weinreb synthesized racemic ter-
rein from cis-1,4-bisbenzyloxy-2,3-epoxycyclopentane in
a multistep route and in low overall yield.10 Barton and
Hulshoff prepared ( )-terrein by performing photo-
chemical ring contraction of 5-hydroxy-4-pyrone.11
O
OH
OH
Keywords: Fungal metabolite; Inhibition; Melanin; Cosmetics.
*
Corresponding author. Tel.: +82 42 860 4330; fax: +82 42 860
4595; e-mail: idyoo@kribb.re.kr
Figure 1. Structure of terrein (1).
0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2004.10.057

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S. Lee et al. / Bioorg. Med. Chem. Lett. 15 (2005) 471–473
O
O
O
a
b
O
c
OH
HO
O
O
OBu-t
OBu-t
OAc
5
2
4
3
O
O
HO
OBu-t
OTBDMS
OBu-t
f
e
OTBDMS
d
OTBDMS
OBu-t
7
8
6
O
O
g
i
h
OH
OBu-t
1
OTBDMS
OTBDMS
9
10
Scheme 1. Reagents and conditions: (a) NBS, NaHCO3, Ac2O, THF/H2O, 0°C to rt, 18h, 55%; (b) t-BuOH, SnCl4, CH2Cl2, rt, 4h, 93%; (c) Et3N,
DMF, 70°C, 24h, 47%; (d) TBDMSCl, imidazole, CH2Cl2, rt, 2h, 92%; (e) allylmagnesium bromide, Et2O, À78°C to rt, 16h, 97%; (f) PCC, Celite,
CH2Cl2, rt, 18h, 60%; (g) NaOH, MeOH, rt, 15min, 77%; (h) TiCl4, CH2Cl2, 0°C, 20min, 92%; (i) HCl, EtOH/H2O, rt, 2h, 62%.
The approach has some drawbacks in the construction
of the E-double bond and the trans-dihydroxy moieties
to be nonstereoselective. Zwanenburg and co-workers
completed the synthesis of ( )-terrein via flash vacuum
pyrolysis of functionalized tricyclo[5.2.1.0]decenone
epoxide to cyclopentadienone epoxide.12 Recently, Kolb
and Hoffmann prepared terrein utilizing the ring con-
traction of a pyranone to a cyclopentenone,13 however
their synthetic route involves expensive reagents such
as 2-(trimethylsilyl)ethanol and E-1-propenyllithium.
all aspects including IR, 1H NMR, 13C NMR and
TLC in three different solvent systems. Also, the syn-
thetic compound 1 showed identical melanin biosynthe-
sis inhibitory activities to naturally occurring ( )-
terrein.
Inhibitory activities of terrein on melanin formation
were evaluated by measurement of the amounts of mel-
anin produced by melanocyte Mel-Ab cells.20 After ter-
rein was treated at the concentrations of 5–50lM for 4
days, terrein-treated cells were much less pigmented
than the untreated cells (Fig. 2). In agreement with the
microscopic observations, melanin levels were strongly
reduced in a dose-dependent manner with an IC50 value
of 18.8lM. Terrein showed above 10 times stronger
activity than kojic acid, as a control, which exhibited
Caddick et al. reported a base-mediated isomerization of
pyranones to functionalized cyclopentenones.14 We have
envisioned that the application of the isomerization
reaction of pyranone to a dihydroxylated cyclopent-
enone would provide a practical synthetic approach to
terrein. To this end, acetoxy pyranone 3 was prepared
from furfuryl alcohol (2) by treatment with N-bromo-
succinimide and acetic anhydride in aqueous THF
(Scheme 1).15 Acetate 3 was converted to acetal 4 by
treatment with tert-butyl alcohol in the presence of
SnCl4.16 Isomerization of pyranone 4 with triethylamine
in DMF at 70°C yielded five-membered cyclopentenone
5 in a stereospecific fashion.15b,17 Silylation of secondary
alcohol 5 followed by addition of allyl magnesium bro-
mide provided tertiary alcohol 7. Oxidation of tertiary
allylic alcohol 7 using PCC18 afforded enone 8, which
upon exposure to a methanolic NaOH solution afforded
fully conjugated cyclopentadienone 9. Finally, cleavage
120
100
80
60
40
20
0
19
of tertiary-butyl ether in 9 utilizing TiCl4 and subse-
quent desilylation gave ( )-1. This synthetic pathway
provided a practical route to be easily amenable to large
scale and produced 10g quantities of ( )-1 starting from
115g of furfuryl alcohol (2). The synthetic compound 1
was in good accord with a natural authentic sample in
0
5
12.5
25
37.5
50
Terrrein (µm)
Figure 2. Inhibitory activity of terrein (1) on melanin biosynthesis.

S. Lee et al. / Bioorg. Med. Chem. Lett. 15 (2005) 471–473
473
compound was checked to be over than 99% by high-
performance liquid chromatography with a ODS column
(YMC C18) eluted with MeOH–H2O (30:70).
7. Raistrick, H.; Smith, G. Biochem. J. 1935, 29, 606.
8. Kamata, S.; Sakai, H.; Hirota, A. Agric. Biol. Chem. 1983,
47, 2637.
9. Qureshi, I. H.; Begum, T.; Noorani, R. Pakistan J. Sci.
Ind. Res. 1976, 19, 120.
10. Auerbach, J.; Weinreb, S. M. J. Chem. Soc., Chem.
Commun. 1974, 298.
inhibition of melanin synthesis by 20% at 100lM. Inter-
estingly, terrein showed no inhibitory activity against
tyrosinase even at 200lM concentration.
In conclusion, terrein, isolated from Penicillium sp.
20135, was prepared by a practical synthetic way that
would bring a large quantity of 1 and its melanin bio-
synthesis inhibitory activity was first evaluated. Further
studies on mechanistic aspects for melanin formation
inhibition of terrein are under investigation.
11. Barton, D. H. R.; Hulshof, L. A. J. Chem. Soc., Perkin
Trans. 1 1977, 1103.
12. Klunder, A. J. H.; Bos, W.; Zwanenburg, B. Tetrahedron
Lett. 1981, 22, 4557.
13. Kolb, H. C.; Hoffmann, M. R. Tetrahedron: Asymmetry
1990, 1, 237.
14. Caddick, S.; Cheung, S.; Frost, L. M.; Khan, S.;
Pairaudeau, G. Tetrahedron Lett. 2000, 41, 6879.
15. (a) Achmatowicz, O., Jr.; Bukowski, P.; Szechner, B.;
Zwierzchowska, Z.; Zamoiski, A. Tetrahedron 1971, 27,
1973; (b) Caddick, S.; Khan, S.; Frost, L. M.; Smith,
N. J.; Cheung, S.; Pairaudeau, G. Tetrahedron 2000, 56,
8953.
Acknowledgements
This work was supported by a grant from the National
Research Laboratory Program funded by the Ministry
of Science and Technology, Korea.
References and notes
16. Grynkiewicz, G.; Barszazak, B.; Zamojski, A. Synthesis
1979, 364.
17. Caddick, S.; Delisser, V. M.; Doyle, V. E.; Khan, S.;
Avent, A. G.; Vile, S. Tetrahedron 1999, 55, 2737.
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682.
1. (a) Schroder, J. J. Invest. Dermatol. 1995, 105, 20; (b)
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19. Caddick, S.; Delisser, V. M. Tetrahedron Lett. 1997, 38,
2355, and references cited therein.
4. (a) Fujimoto, N.; Watanabe, H.; Nakatami, T.; Roy, G.;
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6. Isolation of terrein from the Penicillium culture: The
culture broth (1L) was extracted with 80% acetone and the
extract was concentrated in vacuo to an aqueous solution,
which was then extracted with an equal volume of EtOAc
three times. EtOAc extract was concentrated in vacuo to
dryness. The crude extract was subjected to SiO2 (Merck
Art No. 7734.9025) column chromatography followed by
stepwise elution with CHCl3–MeOH (50:1, 20:1, 10:1).
The active fractions eluted with CHCl3–MeOH (20:1) were
pooled and concentrated in vacuo. The residue was finally
applied to a Sephadex LH-20 and then eluted with MeOH
to afford the purified compound (15mg). The purity of the
20. Melanin synthesis inhibition assay: The Mel-Ab is a
mouse-derived spontaneously immortalized melanocyte
cell line that produces large amounts of melanin.21 Mel-
Ab cells were cultured in DMEM supplemented with 10%
fetal bovine serum (FBS), 100nM TPA, 1nM CT, 50lg/
mL streptomycin and 50U/mL penicillin at 37°C in 5%
CO2. Melanin contents were measured as described
previously22 with slight modification. Briefly, cells were
treated with terrein at various concentrations for 4 days.
Cell pellets were dissolved in 1mL of 1N NaOH at 100 °C
for 30min and centrifuged for 20min at 16,000g. Optical
densities (OD) of the supernatants were measured at
400nm using an ELISA reader.
21. Dooley, T. P.; Gadwood, R. C.; Kilgore, K.; Thomasco,
L. M. Skin Pharmacol. 1994, 7, 188.
22. Tsuboi, T.; Kondoh, H.; Hiratsuka, J.; Mishima, Y.
Pigment Cell Res. 1998, 11, 275.