OBJECTIVETo confirm the inhibitory effect of Chinese herbs of acid taste on melanin synthesis.

METHODSActive ingredients of 22 kinds Chinese herbs of acid tastes were extracted by alkali extraction and acid precipitation, alcohol extraction, and water extraction, respectively, which was then dispensed into 25.00, 12.50, and 6.25 g/L suspension. Their effects on activities of tyrosinase were detected using mushroom-tyrosinase-DOPA speed oxidation. Their inhibition rates on activities of tyrosinase were respectively compared with inhibition rates of 1.0, 0.5, and 0.1 mmol/L arbutin.

RESULTSThe 22 kinds Chinese herbs of acid taste included Cornus Officinalis, Crataegus pinnatifida, dark plum fruit, Schisandra Chinensis, Chaenomeles sinensis Koehne, Reynoutria japonica Houtt, Achyranthes Bidentata, Sanguisorba officinalis L., Semen Ziziphi Spinosae, Herba Ecliptae, blueberry, immature bitter orange, submature bitter orange, Prunus mume Var, Hovenia acerba Lindl., Fructus Mori, Pomegranate Rind, white paeony root, Rosa laevigata Michx., Portulaca oleracea L, Terminalia chebula Retz, Rhus chinensis Mill. Their alkaline extractions showed inhibition to activities of tyrosinase to different degrees except Herba Ecliptae. Of them, the highest inhibition rate (88.49% ± 9.98%) was got by dark plum fruit at 25 g/L, while the lowest inhibition rate (11.22% ± 3.36%) was got by immature bitter orange at 6.25 g/L. Their alcohol extractions showed inhibition to activities of tyrosinase to different degrees except Herba Ecliptae. Of them, the highest inhibition rate (75.92% ± 5.57%) was got by Hovenia acerba Lindl. at 25 g/L, while the lowest inhibition rate (9.60% ± 1.15%) was got by submature bitter orange at 6.25 g/L. Their water extractions all had inhibition on activities of tyrosinase. Of them, the highest inhibition rate (54.23% ± 3.56%) was got by Fructus Mori at 25 g/L, while the lowest inhibition rate (10.25% ± 1.83%) was got by Semen Ziziphi Spinosae at 6.25 g/L. Compared with 1 mmol/L arbutin water solution, alkaline extractions of dark plum fruit, Schisandra Chinensis, Rhus chinensis Mill., Rosa laevigata Michx., blueberry, Chaenomeles sinensis Koehne, Portulaca oleracea L, Fructus Mori, Achyranthes Bidentata, Pomegranate Rind; alcohol extractions of dark plum fruit, Rhus chinensis Mill., Pomegranate Rind, Hovenia acerba Lindl., Crataegus pinnatifida, Achyranthes Bidentata; water extractions of Chaenomeles sinensis Koehne, blueberry, and Fructus Mori at 25 g/L got obviously higher inhibition rates (P < 0.05, P < 0.01). Compared with 0.5 mmol/L arbutin water solution, alcohol extraction of Chaenomeles sinensis Koehne and alcohol extraction of dark plum fruit at 12.5 g/L got obviously higher inhibition rates (P < 0.05, P < 0.01).

CONCLUSIONChinese herbs of acid taste could inhibit melanin synthesis, and its mechanism was related to inhibiting activities of tyrosinase.

Drugs, Chinese Herbal ; pharmacology ; Humans ; Melanins ; metabolism ; Monophenol Monooxygenase ; metabolism ; Oxidation-Reduction ; Schisandra ; Taste

AIMTo evaluate the effect of paeonol on the activity of tyrosinase and provide experimental evidence for the treatment of hyperpigmentation disorders.

METHODSTyrosinase activity was estimated by measuring the oxidation rate of L-3,4-dihydroxyphenylalanine (L-Dopa). The inhibitory effects of paeonol on the activity of mushroom tyrosinase and Michaelis-Menten kinetics were deduced from the Lineweaver-Burk plots.

RESULTSThe inhibitory concentration of paeonol leading to 50% enzyme activity lost (IC50) was estimated to be 0.60 mmol x L(-1). The inhibition constants for paeonol binding free enzyme, K(I), and substrate-enzyme, K(IS), are 0.084 and 0.12 mmol x L(-1), respectively.

CONCLUSIONPaeonol is a potential mixed inhibitor of mushroom tyrosinase. The mixed inhibition function may originate from its ability to form a Schiff base with a primary amino group and to chelate copper at the active site of tyrosinase.

Acetophenones ; isolation & purification ; pharmacology ; Enzyme Inhibitors ; pharmacology ; Kinetics ; Levodopa ; metabolism ; Monophenol Monooxygenase ; metabolism ; Paeonia ; chemistry ; Plants, Medicinal ; chemistry

Tea contains polyphenols and is one of the most popular beverages consumed worldwide. Because most tyrosinase inhibitors that regulate melanogenesis are phenol/catechol derivatives, this study investigated the inhibitory effects of Camellia sinensis water extracts (CSWEs), including black tea, green tea, and white tea extracts, on melanogenesis using immortalized melanocytes. CSWEs inhibited melanin accumulation and melanin synthesis along with tyrosinase activity in a concentration-dependent manner. These inhibitory effects were superior to those of arbutin, a well-known depigmenting agent. The anti-melanogenic activity of black (fermented) tea was higher than that of a predominant tea catecholamine, epigallocatechin gallate. CSWEs, especially black tea extract, decreased tyrosinase protein levels in a concentration-dependent manner. These results suggest that the anti-melanogenic effect of CSWEs is mediated by a decrease in both tyrosinase activity and protein expression, and may be augmented by fermentation. Thus, CSWEs could be useful skin-whitening agents in the cosmetic industry.
Animals ; Catechin/analogs & derivatives/metabolism ; Cell Line ; Melanins/*metabolism ; Melanocytes/enzymology/*metabolism ; Mice ; Monophenol Monooxygenase/*metabolism ; Plant Extracts/*pharmacology ; Plant Leaves/chemistry ; Tea/*chemistry

OBJECTIVETo study the effects of aloesin and arbutin on normal cultured human melanocytes in synergetic method.

METHODSBuilding up the system of cultured human melanocytes. The cultured melanocytes in vitro were treated with the mixture of aloesin and arbutin. The cell viability and tyrosinase activity was measured by MTT assay, utilization of L-Dopa as the substrate respectively; melanin content was measured by image analysis system. Furthermore, the effects of the mixture on melanocytes were compared with that of aloesin and arbutin.

RESULTSThe mixture of aloesin and arbutin showed an inhibition on tyrosinase activity of human melanocytes and reduced significantly melanin content. Between the mixture and the single use of aloesin or arbutin, there is significant difference (P < 0.05). On the other hand, the mixture has little influence on melanocytes viability and there is negative significance.

CONCLUSIONThe mixture of aloesin and arbutin can significantly inhibit the tyrosinase activity and melanogenesis of cultured human melanocytes. It showed the effects of aloesin and arbutin in a synergistic manner. It is worth to give farther study later.

Arbutin ; pharmacology ; Cells, Cultured ; Chromones ; pharmacology ; Dose-Response Relationship, Drug ; Drug Synergism ; Glucosides ; pharmacology ; Humans ; Melanocytes ; cytology ; drug effects ; metabolism ; Monophenol Monooxygenase ; drug effects ; metabolism

OBJECTIVETo evaluate the transfect results of recombinant adenovirus vector carrying tyrosinase gene (Ad-tyr) in vitro by magnetic resonance imaging (MRI) after the Ad-tyr was transfected into HepG2 cell.

METHODSThe Ad-tyr which carried the full-length cDNA of tyrosinase gene was transfected into HepG2 cell. The transfected cells were scan by MRI sequences of T1 weighted image (T1WI) , T2 weighted image (T2WI) , and short time inversion recovery (STIR) to observe the MRI signals of expressed melanin. Masson-Fontana staining was performed to search for melanin granules in transfected cells. Real-time PCR method was used to search for cDNA of tyrosinase gene.

RESULTSAd-tyr was transfected into HepG2 cells and synthesized a large amount of melanin inside. The synthesized melanin of 1 x 10(6) cells which had been transfected by Ad-tyr with the 50, 150, and 300 multiplicity of infection separately were all sufficient to be detected by MRI and showed high signals in MRI T1WI, T2WI, and STIR sequences. The signal intensities of MRI were positively correlated to the amounts of transfected Ad-tyr. The melanin granules were found in HepG2 cells in Masson-Fontana staining. The cDNA amount of tyrosinase gene in transfected HepG2 cells, which was detected by real-time PCR, was remarkably higher than that in nontransfected cells.

CONCLUSIONThe synthesized melanin of HepG2 cells, which controlled by expression of exogenous gene, can be detected by MRI, indicating that the adenovirus vector can efficiently carry the tyrosinase gene into HepG2 cells.

Adenoviridae ; genetics ; metabolism ; Gene Transfer Techniques ; Genetic Vectors ; genetics ; Hep G2 Cells ; Humans ; Magnetic Resonance Imaging ; methods ; Melanins ; analysis ; genetics ; Monophenol Monooxygenase ; biosynthesis ; genetics ; Transfection

Increasing evidence suggests that stress may induce changes in hair color, with the underlying mechanism incompletely understood. In this study, female C57BL/6 mice subjected to electric foot shock combined with restraint stress were used to build chronic stress mouse model. The melanin contents and tyrosinase activity were measured in mouse skin and B16F10 melanoma cells. The enzyme-linked immunosorbent assay (ELISA) was used to determine the content of tumor necrosis factor α (TNF-α), interleukin- 1β (IL-1β) and interleukin-6 (IL-6) in the mouse skin. The content of nuclear factor κB (NFκB)/p65 subunit in mouse skins was valued by immunofluorescence staining. The results demonstrated that under chronic stress, the fur color turned from dark to brown in C57BL/6 mice due to the decrease of follicle melanocytes and tyrosinase activity in C57BL/6 mouse skin. Simultaneously, inflammatory responses in skins were detected as shown by increased NFκB activity and TNF-α expression in stressed mouse skin. In cultured B16F10 melanoma cells, TNF-α reduced the melanogenesis and tyrosinase activity in a dose-dependent manner. These findings indicate that chronic stress induces fur color change by decreasing follicle melanocytes and tyrosinase activity in female C57BL/6 mice, and TNF-α may play an important role in stress-induced hair color change.
Animal Fur ; Animals ; Color ; Female ; Melanins ; Melanocytes ; enzymology ; Melanoma, Experimental ; Mice ; Mice, Inbred C57BL ; Monophenol Monooxygenase ; metabolism ; Pigmentation ; Skin ; physiopathology ; Stress, Physiological

Zinc oxide quantum dots (ZnO QDs) were synthesized by gel-sol method and employed as the transdermal aloesin (Alo) carriers. ZnO QDs were surface-functionalized with amino using aminopropyltriethoxysilane (APTES). Alo was covalently bonded on the surface of ZnO QDs via N,N'-carbonyldiimidazole to obtain Alo NPs, which were characterized by transmission electron microscope (TEM), dynamic light scattering (DLS), Fourier transform infrared spectroscopy (FTIR) and thermal gravimetric analyzer (TGA). TEM images showed that ZnO QDs were analogously sphere and monodisperse with a reasonably narrow size distribution, of which was around 4 nm. The size of Alo NPs increased to around 8 nm due to the surface modification. The intense bands at around 3 400 cm and 1 200 cm in the FTIR spectrum of Alo NPs from the vibration of -OH indicated the linkage of Alo on the surface of ZnO QDs. The results of TGA analysis showed that the mass ratio of ZnO QDs and Alo were 39.27% and 35.14%, respectively. The penetration of Alo NPs was much higher than raw Alo according to the passive penetration experiments with Franz-type diffusion cells instrument using full-thickness cavy skin, which manifested the improvement of the penetration for Alo delivered by ZnO QDs. The pH-controlled drug release behavior was investigated. At pH 7.4, only a small amount of Alo (1.45% ± 0.21%) had been released after 2 h. In contrast, as incubation at pH 5.0 of which pH was similar to endosomal environment, Alo was released very fast (87.63% ± 0.46% in 2 h) from Alo NPs, confirming that Alo NPs could response to the pH and realize the intracellular drug release. The inhibitory effect of Alo NPs on tyrosinase was in a dose dependent manner. When the concentration of Alo NPs was 12.5 μg/mL, the inhibition rate was up to 40.32% ± 1.57%. All the results show that the Alo NPs hold a great potential in transdermal tyrosinase inhibition.
Administration, Cutaneous ; Animals ; Chromones ; administration & dosage ; Drug Delivery Systems ; Glucosides ; administration & dosage ; Guinea Pigs ; Monophenol Monooxygenase ; metabolism ; Nanoparticles ; Quantum Dots ; Zinc Oxide

Glycyrrhizin (GR), triterpenoid saponin composed of one glycyrrhetinic acid (GA) and two glucuronic acids, is a main constituent of the hydrophilic fraction of licorice (Glycyrrhiza glabra) extracts and is known to have a wide range of pharmacological actions. In this study, we investigated the mechanism of GR effect on melanogenesis in B16 murine melanoma cells. The cellular levels of tyrosinase mRNA, protein, enzyme activities and melanin contents were increased by GR in a dose dependent manner. Expression of tyrosinase-related protein-2 (TRP-2) mRNA was also increased by GR, however, no significant change was observed on TRP-1. No cytotoxicity was observed at the effective concentration range of GR. GA showed no effect on melanogenesis at the equivalent nontoxic concentrations, indicating that glycoside structure is important in the stimulatory effect of GR on melanogenesis. These results indicate that GR-induced stimulation of melanogenesis is likely to occur through the transcriptional activation.
Animal ; Blotting, Western ; Glycyrrhetinic Acid/pharmacology ; Glycyrrhizic Acid/*pharmacology ; Intramolecular Oxidoreductases/genetics/metabolism ; Melanins/*biosynthesis ; Melanoma, Experimental/enzymology/*metabolism ; Mice ; Monophenol Monooxygenase/genetics/metabolism ; Proteins/genetics/metabolism ; Reverse Transcriptase Polymerase Chain Reaction

In this study, the essential oil from lotus flower extract, including petals and stamens, was assessed with regard to its effects on melanogenesis in human melanocytes. The lotus flower essential oil was shown to stimulate melanin synthesis and tyrosinase activity in a dose-dependent manner. The lotus flower essential oil induced the expression of tyrosinase, microphthalmia-associated transcription factor M (MITF-M), and tyrosinase-related proten-2 (TRP-2) proteins, but not tyrosinase mRNA. Moreover, it increased the phosphorylation of ERK and cAMP response element binding protein (CREB). In order to verify the effective components of the lotus flower oil, its lipid composition was assessed. It was found to be comprised of palmitic acid methyl ester (22.66%), linoleic acid methyl ester (11.16%), palmitoleic acid methyl ester (7.55%) and linolenic acid methyl ester (5.16%). Among these components, palmitic acid methyl ester clearly induced melanogenesis as the result of increased tyrosinase expression, thereby indicating that it may play a role in the regulation of melanin content. Thus, our results indicate that lotus flower oil may prove useful in the development of gray hair prevention agents or tanning reagents.
Blotting, Western ; Cell Proliferation ; Cyclic AMP/metabolism ; Cyclic AMP Response Element-Binding Protein/genetics/metabolism ; Flowers/*chemistry ; Gas Chromatography-Mass Spectrometry ; Humans ; Intramolecular Oxidoreductases/genetics/metabolism ; Lotus/*chemistry ; Melanins/*biosynthesis ; Melanocytes/*drug effects/metabolism ; Microphthalmia-Associated Transcription Factor/genetics/metabolism ; Monophenol Monooxygenase/genetics/metabolism ; Phosphorylation ; Plant Oils/*pharmacology ; RNA, Messenger/genetics/metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Skin/cytology/drug effects/metabolism

The anti-melanogenesis effect of glyceollins was examined by melanin synthesis, tyrosinase activity assay in zebrafish embryos and in B16F10 melanoma cells. When developing zebrafish embryos were treated with glyceollins, pigmentation of the embryos, melanin synthesis and tyrosinase activity were all decreased compared with control zebrafish embryos. In situ expression of a pigment cell-specific gene, Sox10, was dramatically decreased by glyceollin treatment in the neural tubes of the trunk region of the embryos. Stem cell factor (SCF)/c-kit signaling pathways as well as expression of microphthalmia-associated transcription factor (MITF) were determined by western blot analysis. Glyceollins inhibited melanin synthesis, as well as the expression and activity of tyrosinase induced by SCF, in a dose-dependent manner in B16F10 melanoma cells. Pretreatment of B16F10 cells with glyceollins dose-dependently inhibited SCF-induced c-kit and Akt phosphorylation. Glyceollins significantly impaired the expression and activity of MITF. An additional inhibitory function of glyceollins was to effectively downregulate intracellular cyclic AMP levels stimulated by SCF in B16F10 cells. Glyceollins have a depigmentation/whitening activity in vitro and in vivo, and that this effect may be due to the inhibition of SCF-induced c-kit and tyrosinase activity through the blockade of downstream signaling pathway.
Animals ; Embryo, Nonmammalian/drug effects ; Melanins/*biosynthesis ; Melanoma, Experimental/metabolism/pathology ; Mice ; Monophenol Monooxygenase/metabolism ; Phosphorylation/drug effects ; Pigmentation/drug effects ; Proto-Oncogene Proteins c-kit/*metabolism ; Pterocarpans/chemistry/*pharmacology ; SOXE Transcription Factors/metabolism ; Sesquiterpenes/chemistry/*pharmacology ; Signal Transduction/*drug effects ; Soybeans/*chemistry ; Stem Cell Factor/*pharmacology ; Zebrafish/embryology/metabolism