Diphenolase Activity Research Articles

Continuous Fluorometric Method for Determining the Monophenolase Activity of Tyrosinase on L-Tyrosine, through Quenching L-DOPA Fluorescence by Borate.

Tyrosinase is the key enzyme in melanin biosynthesis and inherently involves both monophenolase activity and diphenolase activity. A continuous fluorometric assay method was developed for the first time to directly monitor the real monophenolase activity without the interference of diphenolase reactions through exclusively quenching the native fluorescence of DOPA by borate. Complexation with borate at pH 8.0 allowed for selective quantitation of tyrosine in a binary mixture of tyrosine and DOPA at 335 nm. The time course for consumption of tyrosine was established to measure the initial velocity by recording the tyrosine fluorescence intensity at discrete intervals. The assay worked in the monophenolase activity range from 0.13 to 2.01 U mL-1 with the limit of detection (LOD) of 0.10 U mL-1. The assay method exhibited a promising prospect in application in kinetics of monophenolase and high throughput screening for monophenolase inhibitors.

Discovery and characterization of tyrosinases from sea anemone pedal disc

The formation of underwater adhesion is a complicated physiological process and many different types of enzymes are found to be essential apart from structural proteins. Previous studies have shown that various tyrosinases were present in marine adhesives, but little information is available about the over-expression and enzymatic characterization of these enzymes. Specifically, this study first identified four significantly up-regulated tyrosinases in the pedal disc of Haliplanella luciae by means of multi-omics technology, and made preliminary bioinformatics predictions. Sequence alignment showed that the Tyr1_Hl contained six conserved His residues that bind to copper ions, of which a tyrosinase with diphenolase activity named as Tyr1_HlΔ, was expressed in Escherichia coli BL21 (DE3) cells and purified by affinity chromatography. Enzymatic characterization showed that the activity of Tyr1_HlΔ was Cu2+ dependent and maximum catalytic activities were in 20 mM Tris–HCl (pH 8.0) at 37 °C. In summary, we identified novel tyrosinases in the pedal disks of sea anemone for the first time and the Tyr1_HlΔ was successfully recombinant expressed. Our study will provide basis for future exploration of bio-adhesion mechanism and design of bio-adhesives derived from sea anemones.

Conversion of walnut tyrosinase into a catechol oxidase by site directed mutagenesis

Polyphenol oxidases (PPOs) comprise tyrosinases (TYRs) and catechol oxidases (COs), which catalyse the initial reactions in the biosynthesis of melanin. TYRs hydroxylate monophenolic (monophenolase activity) and oxidize diphenolic (diphenolase activity) substrates, whereas COs react only with diphenols. In order to elucidate the biochemical basis for the different reactions in PPOs, cDNA from walnut leaves was synthesized, the target gene encoding the latent walnut tyrosinase (jrPPO1) was cloned, and the enzyme was heterologously expressed in Escherichia coli. Mutations targeting the two activity controller residues (Asn240 and Leu244) as well as the gatekeeper residue (Phe260) were designed to impair monophenolase activity of jrPPO1. For the first time, monophenolase activity of jrPPO1 towards L-tyrosine was blocked in two double mutants (Asn240Lys/Leu244Arg and Asn240Thr/Leu244Arg) while its diphenolase activity was partially preserved, thereby converting jrPPO1 into a CO. Kinetic data show that recombinant jrPPO1 resembles the natural enzyme, and spectrophotometric investigations proved that the copper content remains unaffected by the mutations. The results presented herein provide experimental evidence that a precisely tuned interplay between the amino acids located around the active center controls the substrate specificity and therewith the mono- versus diphenolase activity in the type-III copper enzyme jrPPO1.

Simultaneous Determination of Six Isoflavones from Puerariae Lobatae Radix by CPE-HPLC and Effect of Puerarin on Tyrosinase Activity.

Tyrosinase inhibitors with excellent inhibitory activities and lower side effects have promising applications in the fields of medicine, agriculture, food sciences and cosmetics. In this study, a method for simultaneous separation and determination of six target compounds (puerarin, daidzin, genistein, daidzein, genistin, and formononetin) in Puerariae Lobatae Radix was established by cloud point extraction (CPE) and concentration combined with high performance liquid chromatography (HPLC). To achieve high extraction yields, an ultrasound-assisted extraction method was developed based on a salt-modified Triton X-100 system. The optimal extraction conditions are: surfactant Triton X-100 concentration 0.07 g/mL, liquid-solid ratio 80:1 (mL/g), NaCl addition amount 0.6 g, equilibrium time 40 min, equilibrium temperature 70 °C. Under the optimal conditions, the total maximum extraction yield of the six target isoflavones reached 8.92 mg/g. Using l-tyrosine and l-dopa as substrates, the effects of puerarin on the monophenolase and diphenolase activity of tyrosinase activity were investigated by the enzyme kinetics method. The results showed that puerarin inhibited monophenolase activity with an IC50 of 0.537 mg/mL and activated diphenolase activity. The inhibition type of puerarin on monophenolase and the activation type of puerarin on diphenolase were analyzed by Lineweaver-Burk plots which show that puerarin showed mixed inhibition on monophenolase and mixed activation on diphenolase. Therefore, puerarin can be used as both a tyrosinase inhibitor and a tyrosinase activator.

Thermodynamic, kinetic and docking studies of some unsaturated fatty acids-quercetin derivatives as inhibitors of mushroom tyrosinase

Inhibition of activity and stability structure of mushroom tyrosinase (MT) is highly important, since it is a key enzyme of melanogenesis playing various roles in organisms. In this study, thermodynamic stability and diphenolase activities were investigated in the presence of quercetin-7-linoleate (ligand I) and quercetin-7-oleate (ligand II) on mushroom tyrosinase by experimental and computational methods. Kinetic analyses showed that the inhibition mechanism of these ligands is reversible and competitive manner. The inhibition constants values (KiI = 0.31 and KiII = 0.43 mM) and the half maximal inhibitory concentration (IC50 = 0.58 and 0.71 mM) were determined for ligand I and ligand II respectively. Thermal denaturation for the sole and modified enzyme were performed by using fluorescence spectroscopy to obtain the thermodynamic parameters of denaturation. Type of interactions and orientation of ligands were determined by molecular docking simulations. The binding affinities of the MT–ligand complexes during docking were calculated. In the computational studies performed using the MT (PDBID: 2Y9X) from which tropolone was removed, we showed that the ligands occupied different pockets in MT other than the active site. The best binding energies with values of −9 and −7.9 kcal/mol were calculated and the MolDock scores of the best poses with the lowest root mean square deviation (RMSD) were obtained as −172.70 and −165.75 kcal/mol for complexes of MT–ligand I and MT–ligand II, respectively. Computational simulations and experimental analysis demonstrated that the ligands increased the mushroom tyrosinase stability by reducing the activity of enzyme. In this regard, ligand I showed the potent inhibitory and played an important role in enzyme stability.

Monophenolase and catecholase activity of Aspergillus oryzae catechol oxidase: insights from hybrid QM/MM calculations.

Catechol oxidase from Aspergillus oryzae (AoCO4) can not only catalyze oxidation of o-diphenols to o-quinones, but can also catalyze monooxygenation of small phenolics. To gain insight into the catecholase and monophenolase activities of AoCO4, the reaction mechanism of catechol oxidation was investigated by means of hybrid quantum mechanical/molecular mechanical (QM/MM) calculations. The oxy-form of AoCO4 was found to be a μ-η2:η2 side-on peroxo dicopper(ii) complex, which can undergo a proton coupled electron transfer from the substrate rather than a proton transfer from the nearby Ser302 residue to generate a hydroperoxide. The μ-1,1-OOH Cu2(i,ii) complex is thermodynamically more stable than the μ-η1:η2 hydroperoxide. Moreover, the cleavage of the O-O bond in the μ-1,1-OOH Cu2(i,ii) intermediate has a much lower barrier than that in the μ-η1:η2 hydroperoxide species. In both cases, the O-O bond cleavage is the rate-limiting step, generating the reactive (μ-O˙)(μ-OH) dicopper(ii) complex. In addition, our results demonstrated that the oxidation of catechol to quinone is much more preferred than the hydroxylation reaction. These findings may provide useful information for understanding the reactivity of the Cu2O2 active site of coupled binuclear copper enzymes.

Inhibitory mechanism of Penicillin V on mushroom tyrosinase.

Penicillin V is a bacteriolytic β-lactam antibiotic drug. In the present work, we investigated the inhibitory effect of Penicillin V on the activity of mushroom tyrosinase for the first time. The molecular mechanism for the inhibition of tyrosinase by Penicillin V was investigated by means of kinetics analysis, fluorescence quenching and molecular docking techniques. The results showed that Penicillin V could inhibit both monophenolase and diphenolase activities with IC50 of 16.6 ± 0.5 and 11.0 ± 0.2mmol/L, respectively. The inhibitory type of Penicillin V on mushroom was mixed type, and the values of KI and KIS were 13.46 and 17.26mmol/L, respectively. The fluorescence quenching and molecular docking showed that Penicillin V could form static interaction near the catalytic pocket of the enzyme to hinder the transportation of substrate to the active site, as well as reduce the copper plasticity for catalysis. Our results contributed to the usage of Penicillin V as a novel tyrosinase inhibitor with dual effect in field of antimicrobial and food preservation and could also provide guidance for the design of novel tyrosinase inhibitors.

A New Natural Antioxidant Biomaterial from Cinnamomum osmophloeum Kanehira Leaves Represses Melanogenesis and Protects against DNA Damage.

Cinnamomoum osmophloeum Kanehira (COK) is an indigenous tree species in Taiwan. Chemical compositions, antioxidant activity, mushroom tyrosinase inhibition, melanin synthesis repression, and protection against DNA damage of hydrosol from the COK leaves by steam distillation were examined. We performed 1,1-diphenyl-2-picrylhydrazyl radical scavenging, metal ion chelating, reducing power, and Trolox equivalent antioxidant capacity (TEAC) assays and determined the correlations between total phenolic contents and antioxidant activities. The findings showed that the anti-oxidative properties of COK hydrosol are closely correlated with their phenol contents. Additionally, the major constituents of hydrosol, i.e., cinnamaldehyde and benzaldehyde, had dose-dependent anti-tyrosinase effects against both monophenolase and diphenolase activities. GC-MS analysis revealed that the major bioactive components of hydrosol were trans-cinnamaldehyde (87.7%), benzaldehyde (7.0%), and cinnamyl acetate (5.3%). Moreover, we found that the hydrosol with the presence of benzaldehyde is more potent than pure cinnamaldehyde, and enhances the tyrosinase inhibitory activity of hydrosol. In kinetic analyses, Lineweaver–Burk plots and replots showed that COK hydrosol is a mixed-type inhibitor. Additionally, we found that very low doses of COK hydrosol repressed α-melanocyte-stimulating hormone-induced synthesis of microphthalmia-associated transcription factor, leading to decreased melanin synthesis in B16-F10 melanoma cells. These results demonstrated that production of hydrosol from COK leaves using steam distillation may provide a safe and efficacious source of skin-whitening agents for cosmetic and pharmaceutical applications, with antioxidant, anti-tyrosinase, anti-melanogenesis, and DNA protective activities.

Antityrosinase, Antioxidant, and Cytotoxic Activities of Phytochemical Constituents from Manilkara zapota L. Bark.

Hyperpigmentation is considered by many to be a beauty problem and is responsible for photoaging. To treat this skin condition, medicinal cosmetics containing tyrosinase inhibitors are used, resulting in skin whitening. In this study, taraxerol methyl ether (1), spinasterol (2), 6-hydroxyflavanone (3), (+)-dihydrokaempferol (4), 3,4-dihydroxybenzoic acid (5), taraxerol (6), taraxerone (7), and lupeol acetate (8) were isolated from Manilkara zapota bark. Their chemical structures were elucidated by analysis of their nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) data, and by comparing them with data found in the literature. The in vitro antityrosinase, antioxidant, and cytotoxic activities of the isolated compounds (1–8) were evaluated. (+)-Dihydrokaempferol (4) exhibited higher monophenolase inhibitory activity than both kojic acid and α-arbutin. However, it showed diphenolase inhibitory activity similar to kojic acid. (+)-Dihydrokaempferol (4) was a competitive inhibitor of both monophenolase and diphenolase activities. It exhibited the strongest 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS), and ferric reducing antioxidant power (FRAP) activities of the isolated compounds. Furthermore, (+)-dihydrokaempferol (4) also demonstrated potent cytotoxicity in breast carcinoma cell line (BT474), lung bronchus carcinoma cell line (Chago-K1), liver carcinoma cell line (HepG2), gastric carcinoma cell line (KATO-III), and colon carcinoma cell line (SW620). These results suggest that M. zapota bark might be a good potential source of antioxidants and tyrosinase inhibitors for applications in cosmeceutical products.

Role of Herba Pyrolae Essential Oil in Skin Aging and Their Inhibition of Free Radicals

Essential oil of Herba pyrolae, a Chinese herbal medicine, has been suggested to exhibit anti-aging effects. To understand underlying mechanism, we have examined its effect on inhibition of two free radicals (2,2-diphenyl-1-picrylhydrazyl and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) and tyrosinase. The half maximal inhibitory concentration (IC50) of the essential oil was 0.1045 mg/mL for DPPH, while that for ABTS was 0.3961 mg/mL. In addition, the essential oil inhibited both monophenolase and diphenolase activities of tyrosinase. Further examination of the tyrosinase inhibition mechanism and kinetics showed that the diphenolase activity inhibition by H. pyrolae essential oil was a non-competitive reversible process. These data suggest possible mechanism(s) underlying anti-aging effects of H. pyrolae essential oil.

Antityrosinase and antioxidant activities of guanidine compounds and effect of guanylthiourea on melanogenesis

Abstract In this study, the antityrosinase and antioxidant activities of four guanidine compounds (GCs), including guanylthiourea (GT), guanidine thiocyanate (GTC), phenylguanidine (PG), and phenethyldiguanide (PTDG), were investigated. The results showed that the GCs reduced the monophenolase and diphenolase activities of tyrosinase. The IC50 values of GT, GTC, PG, and PTDG were 0.27, 3.53, 6.68, and 5.18 mM for monophenolase and 0.23, 3.24, 3.97 and 2.50 mM for diphenolase, respectively. The inhibition of diphenolase activity was reversible for the four GCs. The inhibition types of GT and PTDG were competitive, while those of GTC and PG were mixed. FRAP, ABTS, DPPH, and DNA damage protection assays revealed that only GT possessed significant antioxidant activity. Furthermore, GT was found to decrease the melanin content and expression of TYR, MITF and DCT in both B16F10 cells and zebrafish. Overall, our results indicated that GT can inhibit tyrosinase activity in vitro, which suggests that these GCs are potential candidates for novel small-molecule tyrosinase inhibitors and that GT could have broader application prospects.

Melanogenic inhibitory effects of Triangularin in B16F0 melanoma cells, in vitro and molecular docking studies

The lack of secure therapies for hyperpigmentation disorders, without serious adverse effects, and the latest reports relating melanogenic disorders with development of neurodegenerative diseases, encourage the continuing search for new drugs for the treatment of such disorders. In this sense, the plant kingdom is an important source of bioactive natural products with great potential for the research and development of new therapeutics. The present study evaluated the anti-melanogenic activity of the natural methoxylated chalcone, 2′,6′-dihydroxy-4′-methoxy-3′-methylchalcone (Triangularin, T), on diphenolase activity from mushroom tyrosinase and on murine B16F0 melanoma cell model. In addition, molecular modelling studies were carried out in order to understand the inhibitory activity observed. T showed a potent anti-melanogenic activity being more active than kojic acid (KA) on tyrosinase isolated of both sources and on intracellular tyrosinase. Molecular docking studies displayed important interactions between T and the active site of tyrosinase. Our results suggest that T may be useful for the treatment of hyperpigmentary disorders.

Catechol Oxidase versus Tyrosinase Classification Revisited by Site‐Directed Mutagenesis Studies

AbstractCatechol oxidases (COs) and tyrosinases (TYRs) are both polyphenol oxidases (PPOs) that catalyze the oxidation of ortho‐diphenols to the corresponding quinones. By the official classification, only TYRs can also catalyze the hydroxylation of monophenols to ortho‐diphenols. Researchers have been trying to find the molecular reason for the mono‐/diphenolase specificity for decades. However, the hypotheses for the lack of monophenolase activity of plant COs are only based on crystal structures so far. To test these hypotheses, we performed site‐directed mutagenesis studies and phylogenetic analyses with dandelion PPOs offering high phylogenetic diversity, the results of which refute the structure‐based hypotheses. While plant PPOs of phylogenetic group 2 solely exhibit diphenolase activity, plant PPOs of phylogenetic group 1 unexpectedly also show monophenolase activity. This finding sheds new light upon the molecular basis for mono‐/diphenol substrate specificity and challenges the current practice of generally naming plant PPOs as COs.

Catechol Oxidase versus Tyrosinase Classification Revisited by Site-Directed Mutagenesis Studies.

Catechol oxidases (COs) and tyrosinases (TYRs) are both polyphenol oxidases (PPOs) that catalyze the oxidation of ortho-diphenols to the corresponding quinones. By the official classification, only TYRs can also catalyze the hydroxylation of monophenols to ortho-diphenols. Researchers have been trying to find the molecular reason for the mono-/diphenolase specificity for decades. However, the hypotheses for the lack of monophenolase activity of plant COs are only based on crystal structures so far. To test these hypotheses, we performed site-directed mutagenesis studies and phylogenetic analyses with dandelion PPOs offering high phylogenetic diversity, the results of which refute the structure-based hypotheses. While plant PPOs of phylogenetic group 2 solely exhibit diphenolase activity, plant PPOs of phylogenetic group 1 unexpectedly also show monophenolase activity. This finding sheds new light upon the molecular basis for mono-/diphenol substrate specificity and challenges the current practice of generally naming plant PPOs as COs.

Enzyme Modification and Oxidative Cross‐linking Using Carboxylate‐, Phenol‐ and Catechol‐Conjugated 1,8‐Naphthalimides

The ground- and excited-state interactions of β-alanine, tyrosine and l-dopa substituted 1,8 naphthalimides (NI-Ala, NI-Tyr and NI-Dopa) with lysozyme and mushroom tyrosinase were evaluated to understand the mechanism of oxidative modification. Photooxidative cross-linking of lysozyme was observed for all three conjugates. The yield was significantly reduced for NI-Tyr and NI-Dopa due to intramolecular electron transfer to the excited singlet state of the 1,8-naphthalimide. Incubation of NI-Tyr and NI-Dopa with mushroom tyrosinase resulted in an increased fluorescence from the naphthalimide, suggesting that the phenol and catechol portion of the conjugates are oxidized by the enzyme. This result demonstrates that the compounds bind in the active site of mushroom tyrosinase. The catalytic activity of mushroom tyrosinase to oxidize both tyrosine (monophenolase) and l-dopa (diphenolase) was modified by NI-Tyr and NI-Dopa. Monophenolase activity was inhibited, and the diphenolase activity was enhanced in the presence of these conjugates. Detailed Michaelis-Menten studies show that both Vmax and Km are modified, consistent with a mixed inhibition mechanism. Collectively, the results show that the compounds interact in the enzyme's active site, but also modify the distribution of the enzyme's oxidation states that are responsible for catalysis.

Keggin-type polyoxotungstates as mushroom tyrosinase inhibitors - A speciation study

Mushroom tyrosinase abPPO4 is a commercially relevant polyphenol oxidase and has been being targeted for numerous inhibition studies including polyoxometalates (POMs). In the present work, its diphenolase activity was inhibited at pH 6.8 by a series of structurally related polyoxotungstates (POTs) of the α-Keggin archetype, exhibiting the general formula [Xn+W12O40](8−n)− in order to elucidate charge-dependent activity correlations. Kinetic data were obtained from the dopachrome assay and 183W NMR was applied to obtain crucial insights into the actual Keggin POT speciation in solution, facilitating a straightforward assignment of inhibition effects to the identified POT species. While [PW12O40]3− was completely hydrolyzed to its moderately active lacunary form Hx[PW11O39](7−x)− (Ki = 25.6 mM), [SiW12O40]4− showed the most pronounced inhibition effects with a Ki of 4.7 mM despite of partial hydrolysis to its ineffective lacunary form Hx[SiW11O39](8−x)−. More negative Keggin cluster charges of 5− and 6− generally resulted in preclusion of inhibitory efficacy as well as hydrolysis, but with the Ni-substituted cluster [PW11O39{Ni(H2O)}]5− enzymatic inhibition was clearly restored (Ki = 9.7 mM). The inhibitory capacity of the structurally intact Keggin POTs was found to be inversely correlated to their net charge. The here applied speciation strategy is of utmost importance for any biological POM application to identify the actually active POM species.

Isolation of phenylpropanoid sucrose esters from the roots of Persicaria orientalis and their potential as inhibitors of melanogenesis

Cosmetically, hyperpigmentation is an important issue and the most prominent target for inhibiting hyperpigmentation is, tyrosinase, the rate-limiting enzyme in melanogenesis. Therefore, in this research, we have screened several medicinal plants collected from Bangladesh for their anti-tyrosinase activity and found that roots of Persicaria orientalis have potent inhibitory activity. Nine compounds, including five phenylpropanoid sucrose esters (1–5), were isolated from the EtOH extract of P. orientalis roots. The chemical structures were determined based on spectroscopic methods. All the tested compounds (1–5), significantly reduced extracellular melanin formation in B16 melanoma cells and inhibited tyrosinase monophenolase and diphenolase activity in a dose dependent manner. Compound 4 was twice as effective as kojic acid, when l-DOPA was used as the substrate. Compound 1 and 4 showed better inhibitory activity (>59%) on melanin synthesis at a treated concentration of 50 µM compare with arbutin (730 µM). At 6.25 µM concentration, compounds 2 and 5, inhibited extracellular melanin production by 53.1% and 40.2%, respectively. Structure-activity-relationship suggested that both the feruloyl (C-6′) and acetyl (C-2′/ C-4′) groups in sucrose moiety is essential for cytotoxicity. Our finding indicated the roots of P. orientalis is a potential source of natural compounds that could be used in dermatological and cosmetological research. To the best of our knowledge, this is the first study to report the potential melanogenesis inhibitory activity of phenylpropanoid sucrose esters.

Comparison of biochemical properties of membrane-bound and soluble polyphenol oxidase from Granny Smith apple (Malus × domestica Borkh.)

Polyphenol oxidase from Granny Smith apples was purified and characterized in both its soluble form (sPPO) and its membrane-bound form (mPPO). Both forms were purified by temperature-induced phase partitioning, precipitation with ammonium sulfate, and ion exchange chromatography. The specific activity of mPPO was 19.17 times that of sPPO. The optimum pH and temperature for both forms were 7.0 and 35 °C when catechol was the substrate. The Michaelis constant and maximum reaction rate for sPPO were 34.1 mM and 500 U/mL/min, whereas those for mPPO were 53 mM and 10,000 U/mL/min, respectively. The enzymes exhibited diphenolase activity, and their affinity was highest for catechol (sPPO) and 4-methylcatechol (mPPO). Inhibitors of sPPO and mPPO included ascorbic acid, glutathione, and l-cysteine. However, ethylenediaminetetraacetic acid increased the activity of mPPO. Purified sPPO was dimeric with a molecular weight of 31 kDa, whereas mPPO was monomeric with an estimated molecular weight of 65 kDa.

Inhibitory effects of compounds isolated from Lepechinia meyenii on tyrosinase

To contribute enzymatic browning inhibitors to the food industry and also extend knowledge about the phytochemical profile of the anti-tyrosinase plant Lepechinia meyenii, its ethanol extract was subjected to bioguided fractionation. Three hydroxycinnamic acids, p-coumaric acid (1), caffeic acid (2) and rosmarinic acid (3), were isolated as mainly responsible for its activity. Compounds 1, 2 and 3 showed themselves highly effective for inhibiting tyrosinase with IC50 values of 0.30, 1.50 and 4.14 μM, respectively, for monophenolase activity and 0.62, 2.30 and 8.59 μM, respectively for diphenolase activity. This is the first report describing the isolation of the compounds causing the tyrosinase inhibitory activity of L. meyenii extract. The inhibitory kinetics of 1–3 using both L-tyrosine and L-DOPA as substrates was investigated and the results obtained were discussed at molecular level by docking analysis. The resulting compounds 1–3 and a phenolic-enriched fraction of the extract, 2.9-fold more active than the starting material, may be suitable as non-toxic and inexpensive alternatives for the control of deleterious enzymatic darkening.

Anti-melanization effects and inhibitory kinetics of tyrosinase of bird's nest fern (Asplenium australasicum) frond extracts on melanoma and human skin

Some bioactive properties of p-coumaric acid and fucose-rich polysaccharide in skin health have been studied, including melanogenesis inhibition of the phenolic acid and growth inhibitory effects of the polysaccharide on melanoma. The dermatological benefits of bird's nest fern extracts (BNFE), containing both substantial fucose-rich polysaccharide and p-coumaric acid, like promoting collagen production and growth of fibroblast cell and further improving the elasticity and dryness of human skins have been demonstrated in our previous study. Besides, the anti-melanization effects of various BNFE on B16-F10 melanoma and human skin were first studied here. The promising extracts revealed that the main phenolic acid, p-coumaric acid, in BNFE resulted in suppression against tyrosinase activity from melanogenesis. The inhibitory kinetics on the diphenolase activity indicated that AE40 was a noncompetitive inhibitor of mushroom tyrosinase. On the other hand, the fucose-rich mucilage of BNFE showed pronouncedly suppressing effect on B16-F10 melanoma viability. Clinical trial was performed by recruiting 46 female volunteers and the results indicated that the lotions with 1% of BNFE was non-irritant and reduced effectively the pigmentation on human skin after 7-14 days of continuous application. It was suggested that the fucose-rich mucilage and p-coumaric acid in BNFE may have potential for nutricosmetics and phytotherapy applications as a natural hypopigmenting agent.