• Microbiology and Biotechnology Letters
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• v.23 no.6
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• pp.641-646
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• 1995

During the screening of inhibitors of melanin biosynthesis from microbial secondary metabolites, a fungal strain MR304 which was capable of producing high level of an inhibitor was selected. Based on taxonomic studies, this fungus could be classified as Trichoderma harzianum. The active compound (MR304-1) was purified from culture broth by Diaion HP-20 column chromatography, ethylacetate extraction, Sephadex LH-20 column chromatographv and HPLC. The inhibitor was identified as 3-(1,5-dihvdroxy-3-isocyanocyclopent-(E)-3-envl)prop-2-enoate by spectroscopic methods of UV, ESIMS, $^{1}$H-NMR, $^{13}$C-NMR, NOE, HMQC and HMBC. MR304-1 showed strong mushroom tyrosinase inhibitory activity with IC$_{50}$ value of 0.25 $\mu $g/ml. It inhibited melanin biosynthesis with 15 mm inhibition zone at 30 $\mu $g/paper disc in Streptomyces bikiniensis, a bacterium used as an indicator organism in this work. It also inhibited melanin biosynthesis in B16 melanoma cells with a niinimum inhibitory concentration of 0.05 $\mu $g/ml.

• Herbal Formula Science
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• v.28 no.1
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• pp.81-90
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• 2020

Objectives : This study was conducted to investigate the effects of major constituents of Epimedium koreanum Nakai (Icariin, epimedium A, epimedium B, and epimedium C) on melanin synthesis. Methods : We measured melanin contents, tyrosinase activity, and expression of Rab27a in B16F10 cells cultured with Epimedium koreanum Nakai ethanol extract (EKN) and their major constituents. After treatment with H89 and dibutyryl cAMP, which inhibit or promote the activation of PKA, we observed changes in melanin synthesis and tyrosinase activity stimulated by EKN. Results : Among them, EKN and icariin enhanced tyrosinase activity and melanin contents. We confirmed that EKN augmented melanin synthesis via cAMP/PKA pathway. Icariin-induced tyrosinase activity and melanin content were attenuated by PKA inhibitor H89, while melanogenic effect of icariin was further augmented by cAMP analog, dbc AMP. However, icariin did not affect the expression of small GTPase Rab27a involved in melanosome transport. Conclusions : These results suggest that icariin promotes melanogenesis through cAMP/PKA pathway but does not affect small GTPase Rab27a.

• Journal of Microbiology and Biotechnology
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• v.16 no.5
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• pp.667-672
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• 2006

Tyrosinase is an enzyme that catalyzes the biosynthetic pathway of melanin pigments participating in the coloring of skin, hair, and eyes, and is widely distributed in nature. The inhibitory compounds of tyrosinase have been extensively used as a cosmetic agent with a skin-whitening effect. In this paper, several plant extracts were screened using Melan-a cells for the melanin biosynthesis inhibition activity, and Idesia polycarpa was selected. A melanin biosynthesis inhibitor was isolated from I. polycarpa fruits by activity-guided fractionation, and the inhibitor was identified as 6-hydroxy-2-[[[(1-hydroxy-6-oxo-2-cyclohexenl-yl)carbonyl]oxy]methyl]phenyl$\beta$-D-glucopyranoside (idescrapin) by comparing it with reported spectral data. Idescarpin $(IC_{50}=8{\mu}g/ml)$ reduced melanin content compared with the vehicle. In addition, the inhibitory activity of idescarpin for melanin synthesis is mediated by decreasing tyrosinase protein rather than directly inhibiting the tyrosinase activity. These results suggest that idescarpin isolated from I. polycarpa fruits may be used as a skin-whitening agent.

• Microbiology and Biotechnology Letters
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• v.24 no.2
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• pp.227-233
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• 1996

Tyrosinase is an enzyme which catalyzes an enzymatic browning of some foods and in vivo synthesis of melanin. In order to produce natural and edible inhibitor of the enzyme which is expected to have whitening effect on melanogenesis, a microorganism was selected from fermented foods. It was named as NU-7, and cultured in mushroom (Lentinus edodes, Shiitake) media. Optimal media to produce tyrosinase inhibitor was formulated by varing nitrogen or carbon content. If glucose content was in a range of 3-20% and ammonium sulfate was in a range of 0-0.25%, production of inhibitor was independent of cell mass. Addition of ammonium sulfate as a nitrogen source had little effect on inhibitor production. Production of inhibitor (Y) was proportionally related to shiitake content (X) with a regression equation of Y= -0.96X$^{2}$ + 13.07X + 14.43 (R = 0.96). These results indicate that shiitake and glucose are necessary for the production of tyrosinase inhibitor. In the analysis of mycotoxin in culture broth, aflatoxin was not detected, suggesting that it would be probably edible.

• YAKHAK HOEJI
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• v.57 no.1
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• pp.24-31
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• 2013

Although statins, inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, have been shown to increase melanin synthesis, the exact mechanism of this action is not fully understood. In this study we investigated the possible involvement of intracellular $Ca^{2+}$ signal in the mechanism of stimulation of melanin synthesis induced by lovastatin in B16 cells. Lovastatin stimulated the production of melanin in a dose-dependent manner in the cells. Treatment with mevalonate, FPP and GGPP, precursors of cholesterol, did not significantly suppress the lovastatin-induced melanin production, suggesting that inhibition of cholesterol synthesis may not be involved in the mechanism of the action of lovastatin. In addition, lovastatin did not significantly alter the cAMP concentration and the stimulated production of melanin by lovastatin was not significantly changed by treatment with H89, a potent inhibitor of protein kinase A, which demonstrates that cAMP pathway may not be involved. However, lovastatin increased intracellular $Ca^{2+}$ concentration in a dose-related fashion. Treatment with EGTA, an extracellular $Ca^{2+}$ chelator did not significantly alter the lovastatin-induced intracellular $Ca^{2+}$ increase and melanin synthesis, whereas intracellular $Ca^{2+}$ reduction with BAPTA/AM and intracellular $Ca^{2+}$ release blockers (dantrolene and TMB-8) completely blunted these actions of lovastatin. Taken together, these results suggest that the intracellular $Ca^{2+}$ release may play an important role in the lovastatin-induced stimulation of melanin synthesis in B16 cells. These results further suggest that lovastatin may be useful for the treatment of hypopigmentation disorders, such as vitiligo.

• Journal of Applied Biological Chemistry
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• v.65 no.2
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• pp.93-99
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• 2022

Lysosome organelle extract (LOE) was derived from egg whites. The lysosome is an intracellular organelle that contains several hydrolysis enzymes. Previous studies have reported that LOE performs important functions, such as melanin de-colorization and anti-melanin production in B16F10 melanoma cells. However, its principal molecular and cellular mechanisms have not been elucidated till date. In non-cytotoxic conditions, LOE significantly inhibited α-MSH induced melanin synthesis of murine B16F10 cells. The anti-melanogenic activity of LOE was mediated by suppressing the mRNA expression of tyrosinase enzyme, tyrosinase related protein-1/2 (TRP-1/2), and microphthalmia-associated transcription factor (MITF) genes. By performing western blot analysis, we found that LOE significantly attenuated melanogenesis. In this case, LOE helped in increasing extracellular receptor kinase (ERK) phosphorylation in α-MSH induced B16F10 cells. Furthermore, MITF is found to be a key regulatory transcription factor in melanin synthesis; it was down-regulated by LOE through ERK phosphorylation. In this experiment, PD98059 (MEK inhibitor) was used to check whether LOE directly regulated the activity of ERK. Although LOE exerted inhibitory effect on melanin synthesis, we could not observe this effect in PD98059-treated α-MSH induced B16F10. These results strongly indicate that LOE is related to ERK activation and MITF degradation in anti-skin pigmentation. Hence, LOE should be utilized as a whitening agent of skin in the near future.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.47 no.2
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• pp.107-121
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• 2021

Skin hypopigmentation, which is observed in albinism or vitiligo, occurs when melanin synthesis is decreased by genetic, epigenetic, and other factors. To identify drug candidates that can promote melanin synthesis in cells, we screened an epigenetic modulator library consisting of 141 cell-permeable, small molecule drugs. B16/F10 murine melanoma cells were treated with each drug at 0.1 𝜇M and melanin synthesis and cell viability were subsequently monitored. As a result, (-)-neplanocin A, 3-deazaneplanocin A (DZNep), and DZNep hydrochloride were found to increase cellular melanin synthesis without causing cytotoxicity. Because these three structurally related drugs exhibited similar dose-dependent effects on melanin synthesis and cell viability, DZNep was selected as a representative drug for additional experiments. DZNep increased intracellular melanin content and tyrosinase (TYR) activity. DZNep also induced the expression of TYR, tyrosinase-related protein 1 (TYRP1), and dopachrome tautomerase (DCT) at the mRNA and protein levels. DZNep also induced the mRNA and protein expression of microphthalmia-associated transcription factor (MITF), a key regulator of melanin synthesis. DZNep is a specific inhibitor of S-adenosylhomocysteine hydrolase and it caused the accumulation of S-adenosylhomocysteine that inhibits histone methyltransferases in cells. This study suggests that melanogenesis can be modulated by targeting S-adenosylhomocysteine hydrolase in certain cellular contexts.

• The Plant Pathology Journal
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• v.16 no.3
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• pp.125-129
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• 2000

A rapid and efficient assay to determine melanin biosynthesis inhibition of Magnaporthe grisea, a causal agent of the rice blast, by chemicals was developed. Wells in 24-well plates were loaded with spore suspension of the fungus and three known melanin biosynthesis inhibitors of KC10017, tricyclazole, and carpropamid. Subsequent color changes of mycelia and culture media in the wells were observed 7 days after incubation. The wells treated with KC10017 (an inhibitor of polyketide synthesis step and/or pentaketide cyclization step) became colorless, whereas tricyclazole (an inhibitor of 1, 3, 8-trihydroxynaphthalene reductase) or carpropamid (an inhibitor of scytalone dehydratase)-treated wells exhibited red color. They did not show any inhibitory effect on fungal growth. The inhibition of reaction steps prior to 1, 3, 6, 8-tetrahydroxynaphthalene formation was easily determined by colorless medium and mycelia. However, it was impossible to distinguish between inhibition of reduction steps and inhibition of dehydration steps by colors of the cultures. It was accomplished through HPLC analysis of the melanin biosynthesis-involving pentaketide metabolites accumulated by the inhibitors. Through screening of a number of synthetic chemicals using the in vitro assay, we could find a novel chemical group of melanin biosynthesis inhibitor.

• ALGAE
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• v.30 no.2
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• pp.163-170
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• 2015

Tyrosinase inhibitors are an important component of cosmetic products. Our previous studies have proposed that eckol isolated from the brown alga Ecklonia cava, can be explored as a tyrosinase inhibitor. However, cellular activities and mechanism of action of eckol remain unknown. Therefore, the current study analyzed the eckol binding modes using the crystal structure of Bacillus megaterium tyrosinase. The effects of eckol on melanin synthesis induced by ${\alpha}$-melanocyte stimulating hormone in B16F10 melanoma cells were also investigated. We predicted the 3D structure of tyrosinase and used a docking algorithm to simulate binding between tyrosinase and eckol. These molecular modeling studies were successful (calculated binding energy value, $-115.84kcal\;mol^{-1}$) and indicated that eckol interacts with Asn205, His208, and Arg209. Furthermore, eckol markedly inhibited tyrosinase activity and melanin synthesis in B16F10 melanoma cells. We also found that eckol decreased the expression of tyrosinase, tyrosinase-related protein (TRP) 1, and TRP2. These results indicate that eckol is a potent inhibitor of melanogenesis, and this finding may be useful for the development of novel pharmaceutical and cosmetic agents.

• Proceedings of the PSK Conference
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• 2002.10a
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• pp.271.3-272
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• 2002

This study shows that sphingosine-1-phosphate (SPP) significantly inhibits melanin synthesis in a concentration-dependent manner, and that the activity of tyrosinase was also reduced in SPP-treated cells. In contrast. a specific extracellular signal-regulated protein kinase (ERK) pathway inhibitor, PD98059 increased tyrosinase activity and melanin production, and PD98059 restored the reduced tyrosinase activity and pigmentation induced by SPP. (omitted)