Melanogenesis Inhibition by Homoisoflavavone Sappanone A from Caesalpinia sappan

Homoisoflavanone, sappanone A, was isolated from Caesalpinia sappan and proven to dose-dependently inhibit both melanogenesis and cellular tyrosinase activity via repressing tyrosinase gene expression in mouse B16 melanoma cells. To our knowledge, sappanone A is the first homoisoflavanone to be discovered with melanogenesis inhibitory activity. Our results give a new impetus to the future search for other homoisoflavanone melanogenesis inhibitors.     

美白化妆品添加剂效能的实验研究

利用体外,体内实验方法对美白添加剂及配方进行筛选和评估,通过观察美白添加剂对酪氨酸酶的抑制作用,对Ｂ１６细胞活力及黑素生成的影响,结合临床受试者实验,结果表明,根据黑素生成理论的新的认识所复配的美白添加剂效果较为理想。     

Antioxidative Characteristics of Anisomeles indica Extract and Inhibitory Effect of Ovatodiolide on Melanogenesis

The purpose of the study was to investigate the antioxidant characteristics of Anisomeles indica methanol extract and the inhibitory effect of ovatodiolide on melanogenesis. In the study, the antioxidant capacities of A. indica methanol extract such as DPPH assay, ABTS radical scavenging assay, reducing capacity and metal ion chelating capacity as well as total phenolic content of the extract were investigated. In addition, the inhibitory effects of ovatodiolide on mushroom tyrosinase, B16F10 intracellular tyrosinase and melanin content were determined spectrophotometrically. Our results revealed that the antioxidant capacities of A. indica methanol extract increased in a dose-dependent pattern. The purified ovatodiolide inhibited mushroom tyrosinase activity (IC(50) = 0.253 mM), the compound also effectively suppressed intracellular tyrosinase activity (IC(50) = 0.469 mM) and decreased the amount of melanin (IC(50) = 0.435 mM) in a dose-dependent manner in B16F10 cells. Our results concluded that A. indica methanol extract displays antioxidant capacities and ovatodiolide purified from the extract inhibited melanogenesis in B16F10 cells. Hence, A. indica methanol extract and ovatodiolide could be applied as a type of dermatological whitening agent in skin care products.     

Depigmenting effect of <Emphasis Type="Italic">Cinnamomum cassia</Emphasis> Presl in B16F10 melanoma cells

To find efficient depigmenting agents, we examined several Chinese herbs for melanogenesis inhibition and toxicity. Cinnamomum cassia Presl exhibited low cytotoxicity at even high concentration (200 μg/ml). The effects on melanogenesis of cultured B16 melanoma cells, mushroom tyrosinase activity, and free radical scavenging activity were further assessed. The methanol extracts of this plant showed the suppression of melanin synthesis. Melanin content was dose-dependently decreased by this herb extract as compared with control cells. It also showed good anti-oxidative activity (IC₅₀=3.7 μg/ml) but no inhibition of mushroom tyrosinase activity. This result showed that Cinnamomum cassia Presl extract might be useful and safe as a new whitening agent in cosmetics.     

Evaluation of Depigmenting Activity by 8-Hydroxydaidzein in Mouse B16 Melanoma Cells and Human Volunteers

In our previous study, 8-hydroxydaidzein (8-OHDe) was demonstrated to be a potent and unique suicide substrate of mushroom tyrosinase. In this study, the compound was evaluated for in vitro cellular tyrosinase and melanogenesis inhibitory activities in mouse B16 melanoma cells and for in vivo skin-whitening activity in human volunteers. Tyrosinase activity and melanogenesis in the cell culture incubated with 10 μM of 8-OHDe were decreased to 20.1% and 51.8% of control, respectively, while no obvious cytotoxicity was observed in this concentration. In contrast, a standard tyrosinase inhibitor, kojic acid, showed 69.9% and 71.3% of control in cellular tyrosinase and melanogenesis activity, respectively, at a concentration as high as 100 μM. Hence, 8-OHDe exhibited more than an inhibitory effects on melanin production in B16 cells 10-fold stronger than kojic acid. In addition, when a cream containing 4% 8-OHDe was applied to human skin in an in vivo study, significant increases in the dL*-values were observed after three weeks. Moreover, the increase in the dL*-values after 8-week treatment with 4% 8-OHDe (from −0.57 to 1.94) is stronger than those of 2% 8-OHDe treatment (from 0.26 to 0.94) and 2% ascorbic acid-2-glucoside treatment (from 0.07 to 1.54). From the results of the study, it was concluded that 8-OHDe, the potent suicide substrate of mushroom tyrosinase, has depigmenting activities in both mouse melanoma cells and in human volunteers. Thus, the compound has significant potential for use in cosmetics as a skin-whitening ingredient.     

Protocatechuic Aldehyde Inhibits α-MSH-Induced Melanogenesis in B16F10 Melanoma Cells via PKA/CREB-Associated MITF Downregulation

Protocatechuic aldehyde (PA) is a naturally occurring phenolic compound that is a potent inhibitor of mushroom tyrosinase. However, the molecular mechanisms of the anti-melanogenesis activity of PA have not yet been reported. The aim of the current study was to clarify the melanogenesis inhibitory effects of PA and its molecular mechanisms in murine melanoma cells (B16F10). We first predicted the 3D structure of tyrosinase and used a molecular docking algorithm to simulate binding between tyrosinase and PA. These molecular modeling studies calculated a binding energy of −527.42 kcal/mol and indicated that PA interacts with Cu400 and 401, Val283, and His263. Furthermore, PA significantly decreased α-MSH-induced intracellular tyrosinase activity and melanin content in a dose-dependent manner. PA also inhibited key melanogenic proteins such as tyrosinase, tyrosinase-related protein 1 (TRP-1), and TRP-2 in α-MSH-stimulated B16F10 cells. In addition, PA decreased MITF expression levels by inhibiting phosphorylation of cAMP response element-binding protein (CREB) and cAMP-dependent protein kinase A (PKA). These results demonstrate that PA can effectively suppress melanin synthesis in melanoma cells. Taken together, our results show that PA could serve as a potential inhibitor of melanogenesis, and hence could be explored as a possible skin-lightening agent.     

Inhibition of purified pig and human liver retinyl ester hydrolase by pharmacologic agents

Identification of inhibitors of retinyl ester hydrolase (RFH) would help to elucidate its role in vitamin A metabolism in vivo. By using standard incubation conditions, the effects of 215 drugs as potential inhibitors of purified pig and human liver REH when acting on micellar substrate retinyl palmitate were evaluated at 16.7, 167, and 1670 μM. Out of the compounds tested, 103 were inhibitors of the pig liver enzyme. The most potent compounds, in order of decreasing activity, were chloral hydrate, lovastatin, phytomenadione, alimemazine, physostigmine, thioridazine, phenoxybenzamine, probucol, cinnarizine, cyclandelate, amiodarone, flupenthixol, and naftidrofuryl; this order is roughly similar to that of their inhibition of human liver REH. Of the 10 tricyclic ring-containing drugs tested, alimemazine was the most potent enzyme inhibitor. The concentrations necessary for 50% enzyme inhibition ranged from <2.6 up to >540 μM. Moreover, inhibitory kinetic studies showed that at least two pharmaceutica's, chloral hydrate and amiodarone, are potent REH inhibitors at therapeutically achievable serum concentrations. First-pass metabolites were inactive as REH inhibitors compared to that of the parent compounds, in the cases of chloral hydrate, lovastatin, and cyclandelate.     

Dual Bioactivities of Essential Oil Extracted from the Leaves of Artemisia argyi as an Antimelanogenic versus Antioxidant Agent and Chemical Composition Analysis by GC/MS

The study was aimed at investigating the antimelanogenic and antioxidant properties of essential oil when extracted from the leaves of Artemisia argyi, then analyzing the chemical composition of the essential oil. The inhibitory effect of the essential oil on melanogenesis was evaluated by a mushroom tyrosinase activity assay and B16F10 melanoma cell model. The antioxidant capacity of the essential oil was assayed by spectrophotometric analysis, and the volatile chemical composition of the essential oil was analyzed with gas chromatography-mass spectrometry (GC/MS). The results revealed that the essential oil significantly inhibits mushroom tyrosinase activity (IC₅₀ = 19.16 mg/mL), down-regulates B16F10 intracellular tyrosinase activity and decreases the amount of melanin content in a dose-dependent pattern. Furthermore, the essential oil significantly scavenged 2,2-diphenyl-1-picryl-hydrazyl (DPPH) and 2,2′-azino-bis (3-ethylbenzthiazoline- 6-sulphonic acid) ABTS radicals, showed an apparent reduction power as compared with metal-ion chelating activities. The chemicals constituents in the essential oil are ether (23.66%), alcohols (16.72%), sesquiterpenes (15.21%), esters (11.78%), monoterpenes (11.63%), ketones (6.09%), aromatic compounds (5.01%), and account for a 90.10% analysis of its chemical composition. It is predicted that eucalyptol and the other constituents, except for alcohols, in the essential oil may contribute to its antioxidant activities. The results indicated that essential oil extracted from A. argyi leaves decreased melanin production in B16F10 cells and showed potent antioxidant activity. The essential oil can thereby be applied as an inhibitor of melanogenesis and could also act as a natural antioxidant in skin care products.     

The melanogenesis-inhibitory, anti-inflammatory, and chemopreventive effects of limonoids in n-hexane extract of Azadirachta indica A. Juss. (neem) seeds

Seventeen limonoids (tetranortriterpenoids 1-17) were isolated from the n-hexane extract of Azadirachta indica (neem) seeds. The previously unidentified compound 16 was established by spectroscopy to be 17-defurano-17-oxosalannin. The effects of six compounds, 6 and 11-15, on melanogenesis in B16 melanoma cells was evaluated; 2 compounds, salannin (13) and 3-deacetylsalannin (15), exhibited marked inhibitory effects (70-74% reduction of melanin content at 25 μg/mL) with only minor cytotoxicity (79-85% of cell viability). Eleven compounds, 2, 3, 5, 6, and 9-15, were evaluated for inhibitory activity against 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammation (1.7 nmol/ear) in mice; all exhibited marked anti-inflammatory activity (ID(50) values 0.22-0.57 μmol/ear). In addition, compounds 6 and 11-16 exerted moderate inhibition (IC(50) values of 410-471 mol ratio/32 pmol TPA) of TPA-induced Epstein-Barr virus early antigen (EBV-EA) activation in Raji cells. The triacylglycerol fraction of the n-hexane extract contained oleic acid (50.2%) as the most predominant fatty acid constituent.     

Discovery of 3‐Hydroxy‐3‐phenacyloxindole Analogues of Isatin as Potential Monoamine Oxidase Inhibitors

A series of 3‐hydroxy‐3‐phenacyloxindole analogues of isatin were designed, synthesized, and evaluated in vitro for their inhibitory activity toward monoamine oxidase (MAO) A and B. Most of the synthesized compounds proved to be potent and selective inhibitors of MAO‐A rather than MAO‐B. 1‐Benzyl‐3‐hydroxy‐3‐(4′‐hydroxyphenacyl)oxindole (compound 18 ) showed the highest MAO‐A inhibitory activity (IC₅₀: 0.009±0.001 μm , K_i: 3.69±0.003 nm ) and good selectivity (selectivity index: 60.44). Kinetic studies revealed that compounds 18 and 16 (1‐benzyl‐3‐hydroxy‐3‐(4′‐bromophenacyl)oxindole) exhibit competitive inhibition against MAO‐A and MAO‐B, respectively. Structure–activity relationship studies suggested that the 3‐hydroxy group is an essential feature for these analogues to exhibit potent MAO‐A inhibitory activity. Computational studies revealed the possible molecular interactions between the inhibitors and MAO isozymes. The computational data obtained are congruent with experimental results. Further studies on the lead inhibitors, including co‐crystallization of inhibitor–MAO complexes and in vivo evaluations, are essential for their development as potential therapeutic agents for the treatment of MAO‐associated neurological disorders.     

Depigmenting effect of <Emphasis Type="Italic">Sterculia lynchnophera</Emphasis> on B16F10 melanoma and C57BL/6 melan-a cells

To develop a novel skin depigmenting agent from natural sources, the inhibition of melanogenesis by the Chinese herb, Sterculia lynchnophera (SL), was evaluated. Treatment of B16F10 melanoma cells and melan-a cells with SL exhibited a 32.9% and 68.2% inhibition of melanin synthesis without cytotoxicity at a concentration of 200 μg/ml, respectively. This herb possessed a high free radical scavenging activity with IC₅₀=11.02 μM. The methanol extract of SL slightly inhibited in vitro mushroom tyrosinase activity (23.4% at a concentration of 200 μg/ml) and had a significant inhibitory effect on cellular tyrosinase activivity (48.65% and 88.56% inhibition at the concentration 200 μg/ml in B16F10 cells and C57BL/6 melan-a cells, respectively). From the western blotting results, SL inhibited the expression of tyrosinase and tyrosinase related protein 1 (TRP-1). Taken together, we suggest that SL may be a safe and effective depigmentation agent.     

Investigation of the Anti-Melanogenic and Antioxidant Characteristics of Eucalyptus camaldulensis Flower Essential Oil and Determination of Its Chemical Composition

The effects of essential oil from Eucalyptus camaldulensis flowers oil on melanogenesis and the oil’s antioxidant characteristics were investigated. Assays of mushroom and cellular tyrosinase activities and melanin content of mouse melanoma cells were performed spectrophotometrically, and the expression of melanogenesis-related proteins was determined by Western blotting. The possible signaling pathways involved in essential oil-mediated depigmentation were also investigated using specific protein kinase inhibitors. The results revealed that E. camaldulensis flower essential oil effectively suppresses intracellular tyrosinase activity and decreases melanin amount in B16F10 mouse melanoma cells. The essential oil also exhibits antioxidant properties and effectively decreases intracellular reactive oxygen species (ROS) levels. The volatile chemical composition of the essential oil was analyzed with gas chromatography–mass spectrometry (GC/MS). The chemical constituents in the essential oil are predominately oxygenated monoterpenes (34.9%), followed by oxygenated sesquiterpenes (31.8%), monoterpene hydrocarbons (29.0%) and sesquiterpene hydrocarbons (4.3%). Our results indicated that E. camaldulensis flower essential oil inhibits melanogenesis through its antioxidant properties and by down-regulating both mitogen-activated protein kinases (MAPK) and protein kinase A (PKA) signaling pathways. The present study indicates that the essential oil has the potential to be developed into a skin care product.     

新型功能性植物盐美白功效的体外评价

对松盐、梅盐与竹盐的美白功效进行了体外评价,分别研究其对体外酪氨酸酶的单酚酶和二酚酶活性的抑制作用,并以熊果苷为阳性对照,采用体外培养的B16黑素瘤细胞模型探究其对胞内酪氨酸酶活性、细胞增殖及迁移的影响。结果表明,植物盐元素组成丰富,且水溶液呈碱性,松盐、竹盐、梅盐对酪氨酸酶活性表现出极显著的浓度依赖的抑制作用(p0.01),当这3种植物盐水溶液的质量浓度为50.0 g/L时,它们对酪氨酸酶单酚酶抑制率分别为97.1%、94.8%、48.2%,对二酚酶的抑制率均高于92.0%;与原盐相比,5.0g/L的植物盐对B16细胞生长抑制作用不明显,但会抑制其横向迁移;植物盐对B16细胞胞内酪氨酸酶活性抑制作用优于熊果苷,5.0g/L的梅盐组的细胞内酪氨酸酶活性仅有8.76%,抑制效果好于松盐、竹盐。所研究的植物盐可以抑制酪氨酸酶的活性,且对B16细胞无附加毒性,显示出作为一种无机酪氨酸酶抑制剂的潜力,在人体美白方面有较好的潜在应用价值。     

Discovery of a Novel Scaffold as an Indoleamine 2,3‐Dioxygenase 1 (IDO1) Inhibitor Based on the Pyrrolopiperazinone Alkaloid,Longamide B

Indoleamine 2,3‐dioxygenase 1 (IDO1) has emerged as a key target for cancer therapy, as IDO1 plays a critical role in the capacity of tumor cells to evade the immune system. The pyrrolopiperazinone alkaloid longamide B and its derivatives were identified as novel IDO1 inhibitors based on docking studies and small library synthesis. The thioamide derivative showed higher IDO1 inhibitory activity than longamide B, and displayed an activity similar to that of a representative IDO1 inhibitor, 1‐methyl‐tryptophan. These results suggest that the pyrrolopiperazinone scaffold of longamide B could be used in the development of IDO1 inhibitors.     

Proteasome Inhibitors with Photocontrolled Activity

Proteasome inhibitors are widely used in cancer treatment as chemotherapeutic agents. However, their employment often results in severe side effects, due to their non‐specific cytotoxicity towards healthy tissue. This problem might be overcome by using a photopharmacological approach, that is, by attaining external, dynamic, spatiotemporal photocontrol over the activity of a cytotoxic agent, achieved by the introduction of a photoswitchable moiety into its molecular structure. Here we describe the design, synthesis, and activity of photoswitchable proteasome inhibitors. Substantial differences in proteasome inhibitory activity in cell extracts were observed before and after irradiation with light. The presented results show potential for the development of chemotherapeutic agents that can be switched on and off with light, constituting a new strategy for spatiotemporally modulating proteasomal activity.     

Inhibition of melanogenesis and melanin transportation by <Emphasis Type="Italic">Gynostemma pentaphyllum</Emphasis>

The extract of Gynostemma pentaphyllum was tested to control the melanogenesis in B16 melanoma. Cytotoxic effect by the extract was observed when the dose concentration was higher than 2 mg/L. Most of the inhibitory effect was obtained by the reduced accumulation of extra-cellular melanin. When the extract was dosed as 2 mg/L, the extra-cellular melanin produced was only 24% of the control. Proteome analysis with 2-D PAGE showed that various proteins involved in melanogenesis were down-regulated by Gynostemma pentaphyllum. In addition to other proteins related to the intra-cellular melanogenesis, Rab-27b and Rab-38 could explain the remarkable decrease in extra-cellular melanin accumulation by reduced melanin transfer to keratinocyte.     

Partial protection against Botulinum B neurotoxin-induced blocking of exocytosis by a potent inhibitor of its metallopeptidase activity

Clostridium botulinum neurotoxins (BoNTs) cause botulism, which is characterized by a flaccid paralysis, through inhibition of acetylcholine release by peripheral cholinergic nerve terminals. This is due to the zinc metallopeptidase activity of the neurotoxin, cleaving one component (synaptobrevin for BoNT/B) of the exocytosis machinery. Yet, there are no specific agents able to control the peptidase-related effects of BoNT/B. We recently developed the first compounds to inhibit this enzymatic activity in the nanomolar range. Here we report that two of our best inhibitors prevent the BoNT/B-induced cleavage of native synaptobrevin on synaptic vesicles, and partially inhibit the suppression of [3H]noradrenaline release from synaptosomes that is caused by BoNT/B. These results were obtained at micromolar concentrations, consistent with the measured inhibitory potency of these inhibitors on the native toxin. These compounds provide a new way to possibly prevent and/or to control the neurotoxin effects of botulinum.