• Journal of Nutrition and Health
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• v.17 no.1
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• pp.60-67
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• 1984

Inhibiting action of sulfur dioxide $(SO_{2})$ on enzymatic discoloration was investigated with crude enzyme preparations from homogenized tissues or sliced disks of raw potato tubers. $SO_{2}$ appeared to inhibit the formation of dopachrome in a competitive manner. At insufficient concentrations of $SO_{2}, the formation of dopachrome was reinitiated as time elapsed. The present results suggested that $SO_{2}$ would form an additional compounds with certain intermediate during the course of enzymatic browning and delay the enzymatic discoloration. To inhibit the production of dopachrome in sliced disks of raw potato tubers, much higher concentrations of $SO_{2}$ were required than homogenized tissues.

• Korean Journal of Food Science and Technology
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• v.17 no.3
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• pp.232-236
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• 1985

The inhibiting properties of sodium benzoate on the dopachrome formation were investigated with crude enzyme preparations from homogenized tissues of raw potato tubers. The % inhibition of dopachrome formation was increased with increasing concentrations of sodium benzoate and decreasing concentrations of substrate, 3,4-dihydroxyphenylalanine(DOPA). The inhibiting action was gradually reduced with increasing temperature. Dopachrome formation tended to be greatly inhibited in the range of pH 3-5, while it revealed a sharp increase above pH 6. The results suggested that sodium benzoate might compete with the substrate for the enzyme.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.16 no.2
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• pp.96-118
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• 2003

The effect of Glycyrrhizae Radix water extract, known as depigmenting agent, on melanin biosynthesis was investigated in cellular level by using B16 mouse melanoma cells. The inhibitory effect of Glycyrrhizae Radix water extract on melanogenesis was determined by mushroom tyrosinase assay traditionally using in vitro screening test. To determine whether Glycyrrhizae Radix water extract suppress melanin synthesis in cellular level, B16 mouse melanoma cells were cultured in the presence of different concentrations of Glycyrrhizae Radix water extract. Effects on cell proliferation, melanin biosynthesis, tyrosinase activity, DOPAchrome tautomerase activity, and expression level of mRNA for tyrosinase were examined. The maximum concentration of Glycyrrhizae Radix water extract that was not inhibitory to growth of the cells was 2 mgml. At that concentration, melanin synthesis was significantly inhibited without cytotoxicity after 5 days, compared with untreated cells. The treatment with Glycyrrhizae Radix water extract reduced tyrosinase and DOPAchrome tautomerase activity in a dose-dependent manner. However, the treatment with Glycyrrhizae Radix water extract did not affect significantly mRNA levels for tyrosinase. These results suggest that the inhibitory effect of Glycyrrhizae Radix water extract on melanogenesis is correlated with the suppression of tyrosinase and DOPAchrome tautomerase activity more than altering mRNA levels of tyrosinase.

• Biomolecules & Therapeutics
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• v.20 no.3
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• pp.340-345
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• 2012

Sarsasapogenin (SAR) is a steroidal sapogenin that is used as starting material for the industrial synthesis of steroids. It has various pharmacological benefits, such as antitumor and antidepressant activities. Since its effect on melanin biosynthesis has not been reported, we used murine melanocyte melan-a cells to investigate whether SAR influences melanogenesis. In this study, SAR significantly increased the melanin content of the melan-a cells from 1 to 10 ${\mu}M$. Based on an enzymatic activity assay using melan-a cell lysate, SAR had no effect on tyrosinase and DOPAchrome tautomerase activities. It also did not affect the protein expression of tyrosinase-related protein 1 and DOPAchrome tautomerase. However, protein levels of tyrosinase and microphthalmia-associated transcription factor were strongly stimulated by treatment with SAR. Therefore, our reports suggest that SAR treatment may induce melanogenesis through the stimulation of tyrosinase and microphthalmia-associated transcription factor expression in melan-a cells.

• Korean Journal of Medicinal Crop Science
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• v.15 no.6
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• pp.434-436
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• 2007

In the previous study, we reported the inhibitory effects of Rodgersia podophylla root extract on tyrosinase activity and melanin production in melan-a cells. However, mechanism of the inhibitory activity and in vivo assay were not yet examined. This study performed the examination of the effects of Rodgersia podophylla root extract on protein expression and in vivo depigmenting activity using melan-a cells and brown guinea pigs. As the results of western immunoblotting analysis, treatment of Rodgersia podophylla root extract reduced tyrosinase expression rates in 10 and 100 ppm concentrations, dose dependently. Moreover, Rodgersia podophylla root extract exhibited depigmenting activity on UV-B induced hyperpigmentation in brown guinea pig skin. These results suggested that Rodgersia podophylla root extract could act as whitening agent for the skin via not only direct tyrosinase activity inhibition but also reducing of tyrosinase expression.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.15 no.2
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• pp.20-32
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• 2002

The aim of this study was to investigate the effect of Codonopsis lanceolatae on the melanogenesis of HM3KO human melanoma cells biologically. The cells were treated for 3 days and 5 days with Codonopsis lanceolatae at several concentrations. The effects on tyrosinase activity and melanin contents were examined. And DOPAchrome tautomerase(TRP-2) activity was also examined to search a pathway influencing this inhibitory effect. 1. It was examined mushroom tyrosinase activity was significantly inhibited by Codonopsis lanceolatae. 2. Treatment with Codonopsis lanceolatae did not affect cell viability at the highest concentration, 1 ㎎／ml, and suppressed melanin contents as a time and dose dependent manner. 3. It was investigated treatment with Codonopsis lanceolatae inhibited tyrosinase, a key enzyme forming melanin, activity in a dose-dependent manner. 4. Treatment with Codonopsis lanceolatae did not affect DOPAchrome tautomerase(TRP-2) activity. 5. There was not a morphological change by Codonopsis lanceolatae microscopically. These results suggest that Codonopsis lanceolatae is a candidate for an efficient whitening agent which supresses melanogenesis by a Raper-Mason pathway.

• International Journal of Industrial Entomology and Biomaterials
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• v.9 no.1
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• pp.29-33
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• 2004

Low molecular weight silk fibroin (LMSF), which was prepared by hydrolysis of silk fibroin using high-temperature and high-pressure method, was found to inhibit the oxidation of L-3,4,-dihydroxyphenylalanine (L-DOPA) catalyzed by mushroom tyrosinase (EC 1.14.18.1). LMSF contained mostly free amino acids such as L-glycine, L-alanine, and L-serine and oligopeptides, mainly glycine-alanine dimer. As a result of analyzing the inhibition kinetics from Lineweaver-Burk plots, L-glycine and glycine-alanine dimer showed noncompetitive behavior while uncompetitive behavior was observed in L-alanine, and L-serine. When weight percent concentration of ${ID_50}$ was compared, L-glycine was most effective on the inhibition and LMSF was also good enough for the inhibition effect of tyrosinase activity. LMSF showed a mixed-type inhibition and the inhibitory mechanism of LMSF might be caused by free amino acids and oligopeptides. As a result of spectroscopic observation with time, initial rate of increase of DOPAchrome decreased remarkably and the time to reach maximum absorbance increased as an increase of the concentration of L-glycine, meaning that L-glycine made itself mainly responsible for the formation of chelate with ${Cu^2+}$ in tyrosinase. However, in case of L-alanine, L-serine, and especially glycine-alanine dimmer, the production of DOPAchrome after an arrival at maximum absorbance decreased, indicating the production of adducts through the reaction with DOPAquinone.

• Journal of Physiology & Pathology in Korean Medicine
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• v.16 no.6
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• pp.1230-1235
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• 2002

This study was conducted to evaluate the effects of Glycyrrhizae Radix water extract, known as depigmenting agent, on melanin biosynthesis in cellular level. The inhibitory effect of Glycyrrhizae Radix water extract on melanogenesis was identified by mushroom tyrosinase assay, To determine whether Glycyrrhizae Radix water extract suppress melanin synthesis in cellular level, B16 mouse melanoma cells were cultured in the presence of different concentrations of Glycyrrhizae Radix water extract. The maximum concentration of Glycyrrhizae Radix water extract that was not inhibitory to growth of the cells was 2 mg/ml. At that concentration, melanin synthesis was significantly inhibited without cytotoxicity after 5 days, compared with untreated cells. The treatment with Glycyrrhizae Radix water extract reduced tyrosinase and DOPAchrome tautomerase activity in a dose-dependent manner. These results suggest that the inhibitory effect of Glycyrrhizae Radix water extract on melanogenesis is due to the suppression of tyrosinase and DOPAchrome tautomerase activity.

• Journal of the Society of Cosmetic Scientists of Korea
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• v.33 no.2
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• pp.69-77
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• 2007

We examined the depigmentation effect on Korean traditional citrus 17 species. With B16 melanoma cells, we have seen inhibition of the tyrosinase and melanin formation, which eventually were dose dependently decreased by three citrus fruits, immature Citrus unshiu, Citrus hassaku, and Citrus sinensis ${\times}$ reticulata as compared with positive control. Also, we examined expression of tyrosinase, DOPAchrome tautomerase (TRP-2), and DHICA oxidase (TRP-1) which affect melanin synthesis. Especially, immature Citrus unshiu decreased the protein levels of tyrosinase and TRP-1. In conclusion, immature Citrus unshue showed the strongest activity in all the experiments mentioned above and we expect that it can be used for preventing UV-induced pigmentation.

• The Korean Journal of Zoology
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• v.39 no.1
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• pp.36-46
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• 1996

An endogenous phenoloxidase (EPO) from earthworm, Lumbricus rubellus, has been purified and characterized. The purified EPO using ammonium sulfate fractionation, Blue-2, Phenyl-, and Q-sepharose chromatography steps was revealed in SDS-PAGE as a single protein banri with Mr. of 59 kl)a. A native strudure of the enzyme was examined with an in situ staining of a nondenatudng-PAGE using DL-dopa as a substrate. The result showed that a single band due to the EPO activity was located siighdy above a standard polypeptide with Mr. of 210 kl)a. These fads indicate that the EPO is an oligomeric enzyme. The presence of a monophenolase activity of the purified EPO, which hydroxylates tyrosine to dopa, was confirmed by observing dopachrome accumulation at 475 nm at PH 8.0 with a typical lag phase during 60 mm. of meausrement. A series of inhibition study has been performed for the enzyme with several divalent cation chelators such as phenyithiourea (Flu), 1, lO-phenanthroline, EDTA, and EGTA. Among them, only V'flj inhibited the enzyme with 1C0.5 of 65 MM, which indicated that copper was critical for the catalysis of EPO. The enzyme was maximally active at 35'C and pH 8.0 when L-dopa to dopachrome conversion was spectrophotometricaily monitored at 475 nm. The apparent Km values of P0 for L-opa were obtained as 1.86 mM and 13.8 mM at pH 6.5 and 8.0, respectively. The catalytic efficiencies at both pH were almost identical [(kat/Km)pH8.0/(kcat/Km)pH6.5 = O.92] while the Vmax at p11 8.0 was 6.6-fold higher than that at pH 6.5. This fact may indicate that pH affeds the catalysis at substrate and/or enzyme-substrate complex level rather than the enzyme itself. Taken together, the EPO was an oligomeric enzyme which did not require proteolysis for its activation. These results also indicated that the enzyme can exist, at least, in part as a latent form In vivo, which might be distinct from the prophenoloxidase activating system. Therefore, it is pertinent to consider that there must be certain regulatory molecules or phenomena in L. rubellus which make the 1,0 in a latent form in vivo before the foreign invasions.