Effects of tetrahydropterines on the generation of quinones catalyzed by tyrosinase

Article abstract of DOI:10.1271/bbb.90924

Tetrahydrobiopterine (6BH⁴) can diminish the oxidative stress undergone by keratinocytes and melanocytes by reducing the o-quinones generated by the oxidation of the corresponding o-diphenols. We found that 6BH⁴ and their analogs reduced all the o-quinones studied. The formal potentials of different quinone/diphenol pairs indicate that the o-quinones with withdrawing groups are more potent oxidants than those with donating groups.

Full text of DOI:10.1271/bbb.90924

Biosci. Biotechnol. Biochem., 74 (5), 1108–1109, 2010
Note
Effects of Tetrahydropterines on the Generation of Quinones Catalyzed
by Tyrosinase
1
1
2
Francis GARCIA-MOLINA, Joseph Louis MUNOZ-MUNOZ, Francis MARTINEZ-ORTIZ,
1
1;y
1
Joseph TUDELA, Francis GARC´IA-CA´NOVAS, and Joseph Neptune RODRIGUEZ-LOPEZ
1
GENZ: Investigative Group for Enzymology, Department of Biochemistry and Molecular Biology A,
Faculty of Biology, University of Murcia, Espinardo, Murcia E-30100, Spain
2
Investigative Group for Theoretical and Applied Electrochemistry, Department of Physical Chemistry,
Faculty of Chemistry, University of Murcia, Espinardo, Murcia E-30100, Spain
Received December 11, 2009; Accepted February 4, 2010; Online Publication, May 7, 2010
[doi:10.1271/bbb.90924]
Tetrahydrobiopterine (6BH4) can diminish the oxi-
tetrahydropterin). All the o-quinones were assayed,
including 1,2-benzoquinone, oxidise BH4, and its ana-
logs MBH4 and DMBH4 (Scheme 1). In addition we
studied the influence of withdrawing and donating
groups in C-1 on the redox potential of the o-quinones.
First, the formal potentials of the different compounds
under study were determined by square wave voltammo-
grams. From these results, it can be seen that the
oxidation/reduction potential was greater for com-
pounds (o-quinones) that contained a withdrawing group
in C-1 (see below). The potential of p-benzoquinone was
dative stress undergone by keratinocytes and melano-
cytes by reducing the o-quinones generated by the
oxidation of the corresponding o-diphenols. We found
that 6BH4 and their analogs reduced all the o-quinones
studied. The formal potentials of different quinone/
diphenol pairs indicate that the o-quinones with with-
drawing groups are more potent oxidants than those
with donating groups.
Key words: tetrahydropterines; quinones; o-diphenols;
voltammograms; tyrosinase
near that of BH and its derivatives, which means that
4
they are not oxidised by the quinone.
The oxidation of diphenols to quinones in keratino-
cytes and melanocytes involves an increase in oxidative
stress in these cells, since H2O2 is generated at the same
time.1 However, topical applications of formulations
containing high concentrations (mM) of coenzyme Q10-
quinol, hydroquinone (HQ), or estrogens can contribute
to increasing oxidative stress in the human epidermis.
A reduction in quinones can contribute to decreasing
oxidative stress. A possible mechanism for o-quinone
reduction is the thioredoxin reductase/thioredoxin (TR/
T) system.3 In human keratinocytes and melanocytes,
the presence of the TR isoform has been demonstrated,4
while 6BH4 can reduce o-quinone L-dopaquinone back
to L-dopa.5
Of the o-diphenols studied, catechol, L-dopa, and
4-tert-butylcatechol (TBC) were easily oxidised by
tyrosinase, generating the corresponding o-quinones. In
this study, we oxidized them with tyrosinase or with a
deficiency of periodate. The results obtained with
catechol are shown in Figs. 1 and 2.
,2)
1,2-Benzoquinone was generated enzymatically by
the action of tyrosinase on catechol. In the presence of
6BH , a lag period in the accumulation of 1,2-benzo-
4
quinone was observed (Fig. 1). Similar results were
obtained in the case of MBH4 (Fig. 1, inset A) and
DMBH4 (Fig. 1, inset B). Note that in all cases the
o-quinone generated by the enzyme oxidized to tetrahy-
dropterines and the system returned to the initial steady
state (in the absence of a reductant). Compounds such as
L-dopa and TBC, which are easily oxidized by tyrosi-
nase, showed the same behavior (results not shown).
Another experimental focus involved the generation
of o-quinone from catechol by oxidation in a deficiency
)
)
)
It has been found that keratinocytes and melanocytes
have a capacity for de novo synthesis, recycling, and
regulation of 6-tetrahydrobiopterin (6BH4). TR reduces
p-quinone 1,4 benzoquinone and coenzyme Q10-
6
)
quinone back to HQ and coenzyme Q10-quinol. On
the other hand, 6BH , can reduce coenzyme Q10-
of NaIO . Successive addition of tetrahydropterines led
4
4
quinone back to coenzyme Q10-quinol, while 1,4
benzoquinone is not reduced to HQ. However, there is
controversy concerning the reduction of o-quinones by
BH4, and it has been proposed that BH4 does not reduce
to a reduction in o-quinone (Fig. 2 and Fig. 2, insets A
and B). However, when the quinone was 1,4 benzoqui-
none, the addition of tetrahydropterin caused no reduc-
tion (results not shown).
1
,4 benzoquinone or 1,2 benzoquinone. It has been
With respect to the influence of withdrawing or
donating groups in the para position on the o-quinone
redox potential, the oxidation capacity increased in the
presence of withdrawing groups but not of donating
groups, as described in ref. 6. All the above was
demonstrated by generating o-quinones of TBC (which
has a donating group), of 4-nitrocatechol (which has a
suggested that the presence of electron-donating sub-
tituents in the quinone structure is needed before
reduction by 6BH4 can occur.
The objective of this work was to demonstrate that
quinones are reduced by 6BH and their analogs MBH
6
)
4
4
(
6-methyltetrahydropterin) and DMBH (6,7-dimethyl-
4
y
To whom correspondence should be addressed. Fax: +34-868-883963; E-mail: canovasf@um.es

Tetrahydropterines and Quinones
1109
O
O
N
H
N
R1
R2
N
N
R1
R2
OH
OH
O
N
HN
+
+
H2N
N
H2N
N
H
O
Tetrahydropterin
1,2-Benzoquinone
Dihydropterin
Catechol
Scheme 1. Oxidation/Reduction Reaction of Tetrahydropterines by 1,2-Benzoquinone to Dihydropterines and Catechol.
The tetrahydropterines used were 5,6,7,8-tetrahydrobiopterine (R1 = CH2OH–CH2OH–CH3, R2 = H), 6-methyl-5,6,7,8-tetrahydropterin
R1 = CH3, R2 = H), and 6,7-dimethyl-5,6,7,8-tetrahydropterin (R1 = CH3, R2 = CH3).
(
0
0
0
0
.12
.08
.04
.00
0.20
0.10
0
.10
0
0
0
0
.06
.04
.02
.00
0.06
0.04
0.02
0.00
A
B
DMBH4
(a)
A
B
DMBH4
MBH4
0
0
.08
.06
0.08
0.06
0.04
MBH
4
(
a)
(f)
(f)
0
0
0
.15
.10
.05
0.04
0.02
0
.02
τ
τ
0
.00
0.00
150
0
50
100
t (s)
150
200
0
20
40
60
80
0
20
40
60
80
0
50
100
t (s)
200
t (s)
t (s)
6
BH4
(a)
(f)
(a)
6
BH4
(b)
τ
0.00
0
50
100
150
200
0
20
40
60
80
t (s)
t (s)
Fig. 1. Catechol Oxidation by Tyrosinase in the Presence of
Tetrahydropterines.
Fig. 2. Reduction of 1,2-Benzoquinone by Tetrahydropterines at
ꢀ
¼ 475 nm.
Spectrophotometric recording at ꢀ ¼ 475 nm of the oxidation of
Catechol (1 mM) was oxidized in a deficiency of sodium periodate
0.4 mM), and the evolution of o-quinone was followed spectropho-
0
.5 mM catechol by 1 nM tyrosinase in the presence of 6BH4 (mM):
(
tometrically, recording (a). At the times indicated by arrows, 6BH4
(a) 0, (b) 100, (c) 125, (d) 150, (e) 175, and (f) 200. The buffer used
was 30 mM sodium phosphate (pH 7.0), 25 C. Inset A, the same
ꢁ
(
100 mM) was added, recording (b). The buffer used was 30 mM
conditions as in Fig. 1, but in the presence of MBH4 (mM); (a) 0,
ꢁ
sodium phosphate (pH 7.0), 25 C. Inset A, exactly the same as in
Fig. 2, but with the addition of MBH4 (100 mM). Inset B, exactly the
same as in Fig. 2, but with the addition of DMBH4 (100 mM).
(
b) 100, (c) 125, (d) 150, (e) 175, and (f) 200. Inset B, the same
conditions as in Fig. 1, but in the presence of DMBH4 (mM); (a) 0,
b) 100, (c) 125, (d) 150, (e) 175, and (f) 200.
(
withdrawing group), and of 3,4-dihydroxybenzaldehyde
which has a withdrawing group) by oxidation in a
deficiency of NaIO4, and in these cases, the tetrahy-
dropterines were also oxidized by these o-quinones
Educaci o´ n (CARM, Murcia, Spain) BIO-BMC 06/01-
0004. JLMM has a fellowship from the Ministerio
de Educaci o´ n y Ciencia (Madrid, Spain), reference
(
AP2005-4721. FGM has
a fellowship from the
(results not shown). Taking into account the redox
potentials of the tetrahydropterine oxidized/reduced
Fundaci o´ n Caja Murcia (Murcia, Spain).
pairs (ꢀ0:099, ꢀ0:105, and ꢀ0:110 for BH4, DMBH4
References
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4
o-diphenol pairs (0.192, 0.163, 0.156, 0.152, 0.114, and
0
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Ministerio de Educaci o´ n y Ciencia (Madrid, Spain)
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