Inhibition of Tyrosinase Activity by N,N-Unsubstituted Selenourea Derivatives

Abstract

Melanin formation is the most important determinant of mammalian skin color. Melanin is secreted by melanocyte cells, which are distributed in the basal layer of the dermis. 1) One of the roles of melanin is to protect the skin and underlying tissues from UV-induced skin injury. However, excessive melanin production in the skin has negative hyperpigmenting effects such as melasma, freckles, and senile lentigines. The synthesis of melanin starts with the conversion of the amino acid L-tyrosine to L-dopa (3,4-dihydroxyphenylalanine), the subsequent oxidation of L-dopa then yields dopaquinone. Tyrosinase is the key enzyme in the biosynthesis of melanin, and participates in the oxidation of tyrosine to L-dopa, and of dopa to dopaquinone. 2,3) Therefore, tyrosinase inhibitors are accepted as important constituents of cosmetics and as depigmenting agents in cases of hyperpigmentation. 4) In this study, we investigated inhibitory effects of N,N-unsubstituted selenourea derivatives on mushroom tyrosinase and their depigmenting effect in melan-a cells. MATERIALS AND METHODS General MethodsMelting points were determined using a Yanagimoto micromelting point apparatus. IR spectra were obtained using a Perkin-Elmer 1600 spectrometer, and 1 H-and 13 C-NMR spectra were recorded on a JEOL-JNMa400 (400 MHz) spectrometer. Mass spectra were obtained using a Shimadzu 9020-DF mass spectrometer, and UV spectra using a Molecular Devices E09090 microplate reader.Materials Mushroom tyrosinase, L-dopa (3-(3,4-dihydroxyphenyl)-L-alanine), and kojic acid (5-hydroxy-2-(hydroxymethyl)-4H-pyran-4one) were purchased from Aldrich Chemical; Inc. (U.S.A.). Solvents used for organic syntheses were redistilled. All other chemicals and solvents were of analytical grade and used without further purification.Typical procedure for preparation of N,N-unsubstituted selenourea derivatives, 1 M HCl (2 eq) in diethyl ether was added to N,N-dimethylcyanamide (1 eq) of THF solution. The reaction mixture became milky white suspension from colorless solution in less than 30 s and then was stirred at 0°C for 2 h. The compound was formed in situ. Subsequently, LiAlHSeH (1 eq) 5) was added to the reaction mixture. The reaction mixture was further stirred at 0°C for 3 h. Assay of Tyrosinase Activity Tyrosinase activity was measured by determining its dopa oxidase activity using a modification of the method reported by Shono et al. 6) Test substances were dissolved in MeOH to 1 mM, 500 mM, 100 mM, or 10 mM. 120 ml of L-dopa (8 mM, dissolved in 67 mM phosphate buffer, pH 6.8) and 40 ml of either the same buffer or test sample were added to each well of a 96-well microplate, and then 40 ml of mushroom tyrosinase (125 U) This study investigated inhibitory effects of N,N-unsubstituted selenourea derivatives on tyrosinase activity. Three types of N,N-unsubstituted selenoureas derivatives exhibited inhibitory effect on dopa (3,4-dihydroxyphenylalanine) oxidase activity of mushroom tyrosinase. Compound D at a concentration of 200 m mM exhibited 55.5% of inhibition on dop...

Inhibitory Effects of Bakuchiol, Bavachin, and Isobavachalcone Isolated fromPiper longumon Melanin Production in B16 Mouse Melanoma Cells

“…1) However, excessive melanin formation and accumulation of melanin in the skin causes hyperpigmentation skin disorders such as melasma, freckles, and geriatric pigment spots. 2,3) In mammals, -melanocyte-stimulating hormone (-MSH) is required for the development of pigmentation. Notably, -MSH binds to its specific melanocortin-1 receptor (MC1R) and increases cyclic AMP (cAMP), which induces melanogenesis by strongly up-regulating the expression of melanogenic enzymes such as tyrosinase.…”

“…We found that an EtOH extract of the fruits of the Indian long pepper Piper longum had a potent inhibitory effect against -MSH-induced melanin production in B16 cells. We isolated the known prenylated phenolic compounds bakuchiol (1), bavachin (2), and isobavachalcone (3) from the fruits of P. longum, and found that they inhibited melanin production in a dose-dependent manner without showing strong cytotoxicity (Fig. 1).…”

“…In conclusion, we searched to identify new plant components that inhibit melanin production in -MSHinduced B16 mouse melanoma cells, and isolated prenylated phenolic compounds bakuchiol (1), bavachin (2), and isobavachalcone (3) from fruits of the Indian long pepper P. longum. These compounds were to inhibit melanin biosynthesis in a dose-dependent manner without showing strong cytotoxicity.…”

Purification and Determination of the Chemical Structure of the Tyrosinase Inhibitor Produced by Trichoderma viride Strain H1-7 from a Marine Environment

“…[12][13][14][15][16] In recent years, attention has been focused on bioactive compound-producing marine microorganisms and a number of novel compounds have been isolated. 17) Strain H1-7 was also isolated from a marine sediment sample at a depth of 100 m off the Izu Islands, and it produced at least 4 kinds of mushroom tyrosinase inhibitors (Fig.…”

Assessment of the Effect of Arbutin Isomers and Kojic Acid on Melanin Production, Tyrosinase Activity, and Tyrosinase Expression in B16-4A5 and HMV-II Melanoma Cells

“…See Supporting Information for detailed protocols. Tyrosinase inhibition assay: Mushroom tyrosinase activity was measured by determining its DOPA-oxidase activity using a modification of the method of Ha et al [15]. First, 120 µL of L-DOPA (8 mM, dissolved in 50 mM phosphate buffer, pH 6.8) and 40 µL of the sample were mixed.…”