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Life Sciences 144 (2016) 80–85

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Life Sciences

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Effect of apigenin-7-glucoside, genkwanin and naringenin on tyrosinase


activity and melanin synthesis in B16F10 melanoma cells
Nouha Nasr Bouzaiene a,b, Fadwa Chaabane a,b, Aicha Sassi a,b, Leila Chekir-Ghedira a,b,⁎, Kamel Ghedira a
a
Unit of Bioactive and Natural Substances and Biotechnology UR12ES12, Faculty of Pharmacy, University of Monastir, Avicenne Street, Monastir 5000, Tunisia
b
Laboratory of Cellular and Molecular Biology, Faculty of Dental Medicine, University of Monastir, Avicenne Street, Monastir 5000, Tunisia

a r t i c l e i n f o a b s t r a c t

Article history: Aims: In this study, we have investigated the effects of apigenin-7-glucoside, genkwanin and naringenin, on
Received 31 August 2015 mouse melanoma B16F10 cell proliferation. Influence of these natural products on percentage cell distribution
Received in revised form 23 November 2015 in cycle phases and melanogenesis was also studied.
Accepted 28 November 2015 Main methods: Cell viability was determined at various periods using the MTT assay, whereas effects of tested
Available online 30 November 2015
compounds on progression through the cell cycle were analyzed by flow cytometry. In addition, amounts of mel-
anin and tyrosinase were measured spectrophotometrically at 475 nm. Besides, the mechanism involved on the
Keywords:
B16F10 melanoma cells
death route induced by the tested molecules was evaluated using the bis-benzimide trihydrochloride coloration
Flavonoids method (Hoechst 33258).
Melanin synthesis Key findings: Apigenin-7-glucoside, genkwanin and naringenin exhibited significant anti-proliferative activity
Tyrosinase activity against B16F10 melanoma cells after 24 and 48 h of incubation. Furthermore, apigenin-7-glucoside, genkwanin
Apoptotic effect and naringenin provoked an increase of subG0/G1, S and G2/M phase cell proportion with a significant decrease
of cell proportion in G0/G1 phases. The results evaluated using Hoechst 33,258, confirm that the percentage of
B16F10 cells observed in the sub G0/G1 phase were undergoing apoptosis. Moreover, apigenin-7-glucoside
and naringenin revealed an ability to enhance melanogenesis synthesis and tyrosinase activity of B16F10 mela-
noma cells. Whereas genkwanin induces a decrease of melanin synthesis by inhibiting tyrosinase activity.
Significance: Our results promote the introduction of genkwanin in cosmetic preparations, as skin whitening
agent, whereas apigenin-7-glucoside and naringenin should be introduced into cosmetic products as natural tan-
ning agents.
© 2015 Elsevier Inc. All rights reserved.

1. Introduction (L-DOPA), while the second step involves oxidation of L-DOPA to L-


DOPA quinine through the enzymatic reaction of tyrosinase. Conse-
Malignant melanoma is the most invasive and deadly form of skin quently, red-orange pheomelanin and blackish-brown eumelanin are
cancer and its incidence is expected to rise over the next two decades formed in our living tissues [9].
[2,12]. The control of melanogenesis is a key approach for the treatment Indeed, traditional use of plants to treat skin diseases, especially for
of abnormal skin pigmentation disease or for the purpose of appearance cosmetic purposes, is a common practice in indigenous medicine of
beautification [20,35]. many cultures; thus, traditional herbal medicines may contain com-
Melanin is a pigment that is biosynthesized from L-tyrosine in mela- pounds with better anti-pigmentation or hyperpigmentation properties
nocytes and plays an important role in preventing skin cancer caused by [28]. Much effort has been devoted to isolate bioactive ingredients from
ultraviolent rays [26,38]. Decreased melanogenesis results in gray hair traditional herbs because of their safety and huge bioactive components
formation, sunburn, and mottling; therefore, controlling melanogenesis [39].
is important for maintaining human body to be beauty and healthy [13, Recently several reports have focused on the potential use of flavo-
36]. Tyrosinase is a key enzyme involved in melanin biosynthesis. This noids for preventing oxidative skin damage [29], such as apigenin-7-
enzyme contains copper ions and catalyzes two reactions in the melanin glucoside and genkwanin (flavones), and naringenin (flavanone).
biosynthetic pathway. The first key step of melanin biosynthesis in- They were reported to exhibited antioxidant, anti-tumoral and anti-
volves the hydroxylation of L-tyrosine to L-3,4-dihydroxyphenylalanine inflammatory activities [3,8,30]. In fact, a previous study, described the
efficacy of naringenin in suppressing UV-induced reactive oxygen spe-
⁎ Corresponding author at: Laboratory of Cellular and Molecular Biology, Faculty of
cies and apoptotic cell death in keratinocytes [22].
Dental Medicine, University of Monastir, Avicenne Street, Monastir 5000, Tunisia. In this study, we have investigated the potential of apigenin-7-glu-
E-mail address: leila.chekir@laposte.net (L. Chekir-Ghedira). coside, genkwanin and naringenin for anti-proliferative and differential

http://dx.doi.org/10.1016/j.lfs.2015.11.030
0024-3205/© 2015 Elsevier Inc. All rights reserved.
N. Nasr Bouzaiene et al. / Life Sciences 144 (2016) 80–85 81

effects on B16F10 cells. To our knowledge, this is the first report on the room temperature and staining with 50 ml propidium iodide
effect of apigenin-7-glucoside, genkwanin and naringenin on (1 mg ml−1) for 10 min, cell cycle analysis was conducted using FACS
melanocytes. system (Beckman Coulter, Switzerland). Percentages of cells in each
phase of the cell cycle were calculated.
2. Material and methods
2.5. Quantitative fluorescence microscopy
2.1. Chemicals and reagents
Cells were seeded into 96-well microtiter plates and 24 h later, serial
The apigenin-7-glucoside, genkwanin and naringenin were pur- dilution of the test samples were added to dedicated wells. The plates
chased from Sigma Aldrich (Genay, France) unless otherwise noted were then incubated 48 h before the cells in each well were harvested,
and were of the highest available purity. All compounds were dissolved washed with PBS, re-suspended in 1 ml PBS containing 10 μg bis-
in DMSO first and then diluted with buffer (1:199, v/v). benzimide trihydrochloride (Hoechst 33,258), and incubated at room
Trypsin, penicillin, streptomycin, vitamins, sodium pyruvate, RPMI- temperature for 15 min. Aliquots of the cells (10 μl) were then placed
1640 medium, non-essential amino acids (NEA) and fetal bovine on glass slides, and triplicate samples of 100 cells each were counted
serum (FBS) were purchased from Gibco ⁄BRL (Scotland, UK). 3-(4,5- and scored for incidence of apoptotic chromatin condensation using a
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and fluorescent microscope (Zeiss, Oberkochen, Germany). Stained nuclei
propidium iodide (PI) were purchased from Sigma-Aldrich (Steinheim, with condensed chromatin (super-condensed chromatin at nuclear pe-
Germany). Ribonuclease A (RNase), Triton X-100, 3,4-dihydroxy-L- riphery) or nuclei that were fragmented into multiple smaller dense
phenylalanine (L-DOPA), bis-benzimide trihydrochloride (Hoechst bodies were considered as apoptotic. Nuclei with uncondensed and dis-
33258) and dimethyl sulphoxide (DMSO) were procured from Sigma persed chromatin were considered not apoptotic.
(St Louis, MO, USA).
2.6. Measurement of cellular melanin content
2.2. Cell line and culture conditions
Melanin release by cells was measured as described in Skandrani
The B16F10 melanoma line was obtained from American Type Cul- et al. [33]. Briefly, B16F10 cells (5 × 105) were seeded into a 50 cm2 cul-
ture Collection (ATCC, Manassas, VA). Cells were grown at 37 °C in a hu- ture dish with 10 ml of culture medium, and incubated for 24 h. Then,
midified atmosphere containing 5% CO2. The cells were culture in RPMI- cells were treated with different concentration of apigenin-7-glucoside,
1640 medium supplemented with 10% (v/v) fetal bovine serum, 2 mM genkwanin and naringenin for 48 h. After treatment, melanogenesis ac-
glutamine, 1% NEA (100 ×), 1% sodium pyruvate 100 mM (complete tivity (closely related to the amount of produced melanin), was estimat-
RPMI). ed from amount of melanin secreted into cultured medium
(extracellular melanin). Adherent cells were detached by incubation
2.3. Cell viability assay in trypsin (2.5%). Cells were then placed in tubes and solubilized in
1 ml of Triton ×100 (0.1%). Spectrophotometric absorbance of extracel-
Effects of tested compounds on viability of B16F10 cells were deter- lular melanin content was measured at 475 nm. Absorbance was com-
mined using the MTT assay. Tetrazolium salt has the property to be re- pared against a standard curve of known concentration of synthetic
duced to blue crystals of formazan by mitochondrial succinate melanin and amounts estimated.
dehydrogenase.
This enzyme, which plays an important role in the Krebs cycle, cata- 2.7. Tyrosinase activity assay
lyzes dehydrogenation of succinate to fumarate. It is a flavoprotein lo-
cated on internal mitochondrial membranes of viable cells. Activity of Tyrosinase enzyme activity was estimated by measuring rate of L-
this enzyme is measured by reduction of the MTT. Toxicity of doses to DOPA oxidation, as described previously [33] with slight modification.
a given cell population is determined by spectrophotometry. Absor- Briefly, cells (106) were treated with different levels of apigenin-7-glu-
bance is directly related to the activity of succinate dehydrogenase, itself coside, genkwanin and naringenin for 48 h, the cells were then solubi-
related to cell toxicity or to number of viable cells. lized in phosphate buffer (0.1 M; pH 6.8) containing 0.1% Triton ×100.
Cells were seeded into 96-well microtiter plates and 24 h later, the Lysate was clarified by centrifugation at 17,500 g for 10 min at 4 °C;
test samples were added in serial dilutions before incubating the plates 400 μl of supernatant was mixed with 400 μl of L-DOPA (0.15%), and ab-
for an additional 24 and 48 h. At each time point, dedicated sets of cells sorbance was followed spectrophotometrically at 475 nm, every minute
were washed once with phosphate-buffered saline (PBS, pH 7.4) before for 10 min, since the substrate added to the reaction mixture.
adding 10 μl PBS containing 2 mg/ml MTT. After 2 h of incubation, the
medium was discarded, and the formazan blue in cells was dissolved 2.8. Statistical analysis
by adding 100 μl DMSO in ethanol (1:1). Negative control cells without
test compound were prepared in the same manner. After incubation at All tests were carried out in triplicate and results were presented as
37 °C for 15 min, the optical density (OD) in each well was measured at mean ± SD (Standard deviation). Statistical analyses were performed
570 nm in a Thermo Scientific (Vantaa, Finland) plate reader. Cytotoxic- by the means of Student test to compare data of control (untreated
ity was expressed as IC50, the concentration that reduced the absor- cells) with those of cells treated with different extracts. Statistical differ-
bance of treated cells by 50% with reference to the control (untreated ences were determined at P b 0.05, P b 0.01 and P b 0.0001.
cells). Actual IC50 values were extrapolated from dose–response curves.
3. Results
2.4. Cell cycle analysis using flow cytometry
3.1. Cell viability assay
Mouse melanoma cells (B16F10 5 × 105 cells) were seeded into a
50 cm2 culture dish and incubated for 24 h. Cells were treated with dif- As illustrated in Fig. 1, incubation of each compound at concentra-
ferent concentrations of compounds for 48 h, trypsinized and washed tions between 10 and 50 μM (apigenin-7-glucoside and genkwanin)
twice in PBS (pH = 7.4). Cells were harvested and incubated for and 10–100 μM (naringenin) with B16F10 cell lines, inhibited cell pro-
15 min at room temperature and washed twice in cold PBS (pH = liferation in a dose-dependent manner after 24 as well as 48 h of
7.4). After treatment with Ribonuclease A (10 mg ml−1) for 30 min at incubation.
82 N. Nasr Bouzaiene et al. / Life Sciences 144 (2016) 80–85

Fig. 1. Inhibitory effect of apigenin-7-glucoside, genkwanin and naringenin on B16F10 melanoma cells viability. Values represent the mean ± SD of three separate experiments. The sta-
tistical significance of results was evaluated by the Student's t-test. *P b 0.05, **P b 0.01 and ***P b 0.001 means significant difference between negative control and treated sample.

3.2. Cell cycle analysis 3.4. Effect of apigenin-7-glucoside, genkwanin and naringenin on melanin
synthesis and tyrosinase activity
B16F10 cell cycle distribution was studied after exposure to different
concentrations of apigenin-7-glucoside, genkwanin and naringenin. To investigate the effect of apigenin-7-glucoside, genkwanin and
B16F10 cells were treated with 70, 60 and 50 μM of apigenin-7-gluco- naringenin compounds on melanin synthesis, B16F10 melanoma cells
side and genkwanin, and 100, 50 and 25 μM of naringenin. Cell cycle dis- were exposed to different concentrations of apigenin-7-glucoside,
tribution was examined after 48 h treatment. genkwanin and naringenin. After 48 h incubation at 37 °C, 5% CO2, mel-
The data listed in Table 1 were obtained by separation of cell cycle anin content of cells were measured.
phases by setting adjacent cursors without deconvolution of overlap- It appears that both apigenin-7-glucoside and naringenin stimulated
ping G0/G1, S and G2/M phases. significantly the production of intracellular melanin in a dose depen-
In comparison with the control group (without treatment), treated dent manner (42.12 μg per 106 cells and 43.03 μg per 106 cells, in the
cells showed a significant increase in the proportion of cells in the presence of the highest tested dose, respectively) when compared to
subG0/G1, S and G2/M phases accompanied by a significant decrease the rate of melanin synthesis in untreated cells (23.93 μg per 106
in the G0/G1 phase (Table 1). cells). Whereas genkwanin decreased significantly melanin production
in a dose dependent manner (Fig. 3).
As far as melanin synthesis pathway involved a rate-limiting regula-
3.3. Apigenin-7-glucoside, genkwanin and naringenin induce apoptosis on tory melanogenic enzyme [14], which is the tyrosinase, we attempted
B16F10 cells the assessing of the aforementioned protein in cells incubated with dif-
ferent doses of the tested molecules. Or results revealed that apigenin-
To evaluate whether apigenin-7-glucoside, genkwanin and 7-glucoside and naringenin increased tyrosinase activity in a dose and
naringenin decreased cell viability through apoptosis, morphological time dependent manner, whereas genkwanin decreased tyrosinase ac-
changes characteristic of apoptotic cells (condensed chromatin) were tivity in a dose and time dependent manner (Fig. 4).
analyzed and quantified by fluorescent microscopy (Fig. 2A and B). Eval-
uation of nuclear morphology indicated that the percentage of apoptotic 4. Discussion
cells increased from 5% (control) to 43% in apigenin-7-glucoside, 52% in
genkwanin and 33% in naringenin treated cells after 48 h of exposure at It is well-known that many compounds from natural plants have
the highest concentrations (Fig. 2A). chemopreventative and chemotherapeutic efficacy in human cancers

Table 1
Cell cycle distribution of B16F10 cells after treatment with apigenin-7-glucoside, genkwanin and naringenin.

Sub-G0/G1 (apoptosis) (%) G0/G1 (%) S (%) G2/M (%)

Control 1 ± 0.01 66 ± 0.08 18 ± 0.02 15 ± 0.02

50 μM 7.31 ± 0.17⁎ 45.61 ± 0.20⁎ 28.97 ± 0.22⁎ 18.11 ± 1.23


Apigenin-7-glucoside 60 μM 11.40 ± 0.1⁎⁎ 41.7 ± 0.25⁎ 28.51 ± 0.25⁎ 18.40 ± 0.23⁎
70 μM 16 ± 0.02⁎⁎⁎ 38.90 ± 0.1⁎⁎ 27.10 ± 0.30⁎ 18 ± 1.18
50 μM 3.97 ± 1.02 48.01 ± 0.13⁎ 31.70 ± 0.11⁎⁎ 16.31 ± 1.20
Genkwanin 60 μM 6.73 ± 0.21⁎ 45.41 ± 0.12⁎ 31.13 ± 0.22⁎ 16.71 ± 1.25
70 μM 10.77 ± 0.17⁎⁎ 40.09 ± 0.15⁎ 31.23 ± 0.23⁎ 17.89 ± 1.16
25 μM 2.80 ± 1.01 48.05 ± 0.17⁎ 30.30 ± 0.28⁎ 18.85 ± 0.30⁎
Naringenin 50 μM 3.45 ± 1.05 47.61 ± 0.15⁎ 29.91 ± 0.24⁎ 19.01 ± 0.25⁎
100 μM 6.90 ± 0.17⁎ 46.8 ± 0.15⁎ 29.10 ± 0.21⁎ 17.20 ± 1.36
⁎P b 0.05 and ⁎⁎P b 0.01 compared to control groups, using the Student's t-test.
N. Nasr Bouzaiene et al. / Life Sciences 144 (2016) 80–85 83

Fig. 2. Evaluation of apoptotic effects of apigenin-7-glucoside, genkwanin and naringenin against B16F10 cells. (A) Cells were stained with bis-benzimide trihydrochloride (Hoechst
33,258) after treatment with different concentrations of each compound for 48 h, and the presence of apoptotic cells was evaluated by fluorescence microscopy. Values shown are
mean (±SD). The histogram is representative of two independent experiments (each performed in duplicate). *P b 0.05, **P b 0.01 and ***P b 0.001 means significant difference between
negative control and treated group. (B) Photomicrographs of representative fields of B16F10 cells, treated with 60 μM of genkwanin, to evaluate nuclear chromatin condensation (i.e.
apoptosis).

[10]. The discovery of phytomedicinal plants as well as elucidations of through inhibition of tumor cell proliferation, and enhancing of antiox-
their underlying mechanism in anticancer activity is important. idant potential [19,24].
Apigenin-7-glucoside, genkwanin and naringenin, included among The effect of apigenin-7-glucoside, genkwanin and naringenin on
the major representative of flavonoids family compounds, are present melanoma cancer cell (B16F10) proliferation, was evaluated after 24
in many natural plants, and have been shown to suppress tumor growth and 48 h of treatment period. Maximum growth inhibition was obtain-
ed on the second day of treatment at the highest tested concentration.
Such inhibitory effect has been previously observed on human cancer
cell lines (HL-60, SMMC-7721, A-549, MCF-7, and SW480), [24]. These
inhibitory effects were reported to be mediated by the suppression of
DNA synthesis [31].
Further, to investigate the anti-proliferative mechanism of apigenin-
7-glucoside, genkwanin and naringenin, we analyzed changes in
B16F10 cell cycle progression induced by the aforementioned mole-
cules, using flow cytometry.
In comparison with the control group (without treatment), treated
cells showed a significant increase of cell proportion in the sub G0/G1,
S and G2/M phases accompanied by a significant decrease in the G0/
G1 phase (Table 1). Several studies show that cell cycle disorders are
often associated with neoplastic growth, so the knowledge of these
mechanisms plays a key role in the development of effective therapies
against cancer. It was reported that cell cycle arrest activates several
mechanisms of DNA repair, [25,37] however, if DNA repair does not
occur properly, p53 can induce cell apoptosis [11,17]. In the presence
Fig. 3. Effect of apigenin-7-glucoside, genkwanin and naringenin on melanin content in
of genomic damage, a cell three choices: repair the damage, go into ap-
B16F10 cells after 48 h incubation. Values represent the mean ± SD of three separate ex-
periments. The statistical significance of results was evaluated by the Student's t-test.
optosis, or die by necrosis. If the damage is extensive enough to deplete
*P b 0.05, **P b 0.01 and ***P b 0.001 means significant difference between negative con- the concentration of ATP or inactivate the caspases, apoptosis becomes
trol (untreated cells) and treated sample. impossible and the cell dies by necrosis. If the damage is more
84 N. Nasr Bouzaiene et al. / Life Sciences 144 (2016) 80–85

Fig. 4. Effect of apigenin-7-glucoside, genkwanin and naringenin on tyrosinase activity in B16F10 cells after 48 h of incubation. Values represent the mean ± SD of three separate
experiments.

moderate, the cell can induce p53 protein and repair the damage during Structure-function analysis of flavonoids suggests that flavonoids
the cell cycle arrest, or deliberately induce apoptosis. In some situations, with an α-keto group exhibit potent tyrosinase inhibition effect due to
the cell can initiate an apoptosis process and then, with ATP depletion the similarity between the dihydroxyphenyl group of DOPA and the
and inactivation of caspases, progress into necrosis [23,37]. Therefore, α-keto containing flavonoids [21].
cell cycle arrest and the induction of apoptosis in cancer cells become In addition, melanin plays an important role in protecting human
the major indicators of anticancer effects. Inhibitors or modulators of skin from the harmful effects of UV radiations by absorbing UV sunlight
tumor cell cycle are of great interest as novel therapeutic agents in can- [34]. Thus, we can suggest a protective effect of apigenin-7-glucoside,
cer. Besides, checkpoints at G0/G1 of the cell cycle in cultured cancer cell genkwanin and naringenin against skin irritations induced by UV sun-
lines have been found to be perturbed by flavonoids, such as quercitin, light by enhancing melanogenesis. Tyrosinase, which is a multifunction-
luteolin, kaempferol, apigenin and epigallocatechin 3-gallate [5,7]. al copper-containing enzyme, is responsible of melanin synthesis that
To elucidate the death mechanism of B16F10 cells induced by starts with the hydroxylation of L-tyrosine to L-
apigenin-7-glucoside, genkwanin and naringenin, we investigated dihydroxyphenylalanine (DOPA) and is followed by the oxidation of
whether they could induce apoptosis or not by observing morphological DOPA to DOPA quinine [14,16]. These quinines spontaneously polymer-
changes, which are described to be characteristic of apoptotic cells (con- ize to high molecular weight brown pigmented components, known as
densed chromatin) [32]. Evaluation of nuclear morphology indicated melanin.
that the percentage of apoptotic cells increase among cells treated HOWEVER, it appears that genkwanin inhibits the melanogenesis
with apigenin-7-glucoside, genkwanin and naringenin. These results and decreased tyrosinase activity after 48 h treatment in a dose depen-
confirm those obtained by flow cytometry, and revealing an increase dent manner (Figs. 3 and 4). Many studies have reported that flavonoids
of sub G0/G1 cell number. They are also compatible with results obtain- have hypopigmenting effects, as they can directly inhibit tyrosinase and
ed when assessing cell viability by MTT assay (Fig. 1). also can act on the distal part of the melanogenesis oxidative pathway
On the other hand, as endogenous pigmentation is associated with such as nobiletin (5,6,7,8,3′,4′-hexamethoxyflavone), naringin (5,7,4′-
markedly reduced risk of skin cancer, agents that enhance skin pigmen- trihydroxyflavanone), and neohesperidin (5,7,3′-trihydroxy-4′-
tation and have the potential to reduce both photo damage and skin methoxyflavone) [6].
cancer incidence took much attention. Since genkwanin is a flavone, several studies showed that flavones
Thus, evaluations of topically applied substances that stimulate the exhibited tyrosinase inhibitory activities such as norartocarpetin by
natural pigmentation process become the target of many studies [33]. competitive inhibition mode.
Alesiani et al. [1] demonstrated that melanoma population growth re- Many authors [4,27] have demonstrated that during the melanogen-
duction was linked to differentiation processes detected by monitoring esis, there is a production of H2O2 and other ROS by human melanoma
some specific markers (i) morphological changes with the development cells, which put the skin cells under high grade oxidative stress. Hydro-
of dendrite-like projections from the cell surface; and we observed so gen peroxide, which is a main source of reactive oxygen species (ROS),
with B16F10 cells treated with apigenin-7-glucoside and naringenin; has an important role in the pathogenesis of a wide range of diseases in-
(ii) melanin synthesis: and our results indicate that apigenin-7-gluco- cluding cancers [18].
side and naringenin exposure stimulate tyrosinase activity and
melanogenesis. 5. Conclusion
We believe that apigenin-7-glucoside and naringenin are able to en-
hance melanin synthesis in B16 mouse melanoma cells, by activating In summary, this study suggests that apigenin-7-glucoside and
the cyclic AMP-protein kinase A pathway and upregulating the expres- naringenin have a potential to be used as a natural product for tanning
sion and the activity of the melanogenesis-related enzyme tyrosinase, in cosmetic applications. We also report that genkwanin might be a use-
and the microphthalmia-associated transcription factor, a transacting ful therapeutic agent in the treatment of hyperpigmentation and pro-
factor that regulates the gene transcription of tyrosinase [15]. vides effective component in skin-whitening cosmetics, since there is
The bioavailability, metabolism, and biological activity of flavo- a concerted effort to search for naturally occurring tyrosinase inhibitors
noids depend upon the configuration, total number of hydroxyl from plant because plants constitute a rich source of bioactive chemicals
groups, and substitution of functional groups about their nuclear and many of them are largely free from harmful adverse effect.
structure. Moreover, apigenin-7-glucoside, genkwanin and naringenin
In Fact, active compounds isolated from plants, such as apigenin-7- inhibited population growth of mouse melanoma B16F10 cells in a
glucoside, genkwanin and naringenin effect melanogenesis without dose-dependent manner. Further investigations are required to under-
melanocytotoxicity by different mechanisms [40]. stand molecular mechanisms by which apigenin-7-glucoside,
N. Nasr Bouzaiene et al. / Life Sciences 144 (2016) 80–85 85

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