Mushroom tyrosinase

Manufactured by Merck Group

Sourced in United States, Germany, Italy, Singapore, Spain, Sao Tome and Principe

Mushroom tyrosinase is a laboratory enzyme derived from mushrooms. It catalyzes the conversion of tyrosine to melanin, a pigment. The core function of mushroom tyrosinase is to facilitate this biochemical reaction in a controlled laboratory setting.

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Lab products found in correlation

L dopa, by Merck Group (53 mentions) Kojic acid, by Merck Group (51 mentions) L tyrosine, by Merck Group (31 mentions) Dimethyl sulfoxide (dmso), by Merck Group (24 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (17 mentions) Arbutin, by Merck Group (14 mentions) Gallic acid, by Merck Group (13 mentions) 2 2 diphenyl 1 picrylhydrazyl (dpph), by Merck Group (13 mentions) Ascorbic acid, by Merck Group (12 mentions) L 3 4 dihydroxyphenylalanine l dopa, by Merck Group (10 mentions) α msh, by Merck Group (10 mentions) 3 4 dihydroxy l phenylalanine l dopa, by Merck Group (9 mentions) Sodium carbonate, by Merck Group (9 mentions) Quercetin, by Merck Group (9 mentions) Trolox, by Merck Group (8 mentions) Methanol, by Merck Group (8 mentions) Folin ciocalteu reagent, by Merck Group (7 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (7 mentions) Sodium hydroxide (naoh), by Merck Group (6 mentions) 3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide mtt, by Merck Group (6 mentions)

203 protocols using mushroom tyrosinase

Mushroom Tyrosinase Inhibition Assay

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The mushroom tyrosinase (Sigma Chemical, USA) inhibition was performed following our previously reported methods.^{38–40 (link)} In detail, 140 μL of phosphate buffer (20 mM, pH 6.8), 20 μL of mushroom tyrosinase (30 U mL⁻¹) and 20 μL of the inhibitor solution were placed in the wells of a 96-well microplate. After pre-incubation for 10 minutes at room temperature, 20 μL of l-DOPA (3,4-dihydroxyphenylalanine, Sigma Chemical, USA) (0.85 mM) was added and the assay plate was further incubated at 25 °C for 20 minutes. Afterward, the absorbance of dopachrome was measured at 475 nm using a microplate reader (OPTI Max, Tunable). Kojic acid was used as a reference inhibitor and phosphate buffer was used as a negative control. The amount of inhibition by the test compounds was expressed as the percentage of concentration necessary to achieve 50% inhibition (IC₅₀). Each concentration was analyzed in three independent experiments. IC₅₀ values were calculated by nonlinear regression using GraphPad Prism 5.0.
The % inhibition of tyrosinase was calculated as following: here, the B and S are the absorbance's for the blank and samples.

Khan F.M., Abbasi M.A., Rehman A.U., Siddiqui S.Z., Sadiq Butt A.R., Raza H., Hassan M., Ali Shah S.A., Shahid M, & Kim S.J. (2024). Design of potent tyrosinase inhibiting N-arylated-4-yl-benzamides bearing 2-aminothiazole-triazole bi-heterocycles: mechanistic insight through enzyme inhibition, kinetics and computational studies. RSC Advances, 14(23), 16546-16559.

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Mushroom Tyrosinase Inhibition Assay

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Mushroom tyrosinase (Sigma-Aldrich, St Louis, MO, USA) inhibition was performed following our previously reported methods.25 (link) In detail, 140 µL of phosphate buffer (20 mM, pH 6.8), 20 µL of Mushroom tyrosinase (30 U/mL) and 20 µL of the inhibitor solution were placed in the wells of a 96-well microplate. After preincubation for 10 min at room temperature, 20 µL of l-DOPA (3,4-dihydroxyphenylalanine, Sigma-Aldrich) (0.85 mM) was added and the assay plate was further incubated at 25°C for 20 min. Afterwards, the absorbance of dopachrome was measured at 475 nm using a microplate reader (Optimax Max Tunable, Sunnyvale, CA, USA). Kojic acid was used as a reference inhibitor and phosphate buffer was used as a negative control. The extent of inhibition by the test compounds was expressed as the percentage of concentration necessary to achieve 50% inhibition (IC₅₀). Each concentration was analyzed in three inde-pendent experiments. IC₅₀ values were determined by data analysis and by a graphing software, Origin 8.6, 64-bit.
Percent inhibition of tyrosinase was calculated using the following equation:

Inhibition (%) = [\frac{B - S}{B}] \times 100

Here, B and S are the absorbance of the blank and samples.

Abbas Q., Ashraf Z., Hassan M., Nadeem H., Latif M., Afzal S, & Seo S.Y. (2017). Development of highly potent melanogenesis inhibitor by in vitro, in vivo and computational studies. Drug Design, Development and Therapy, 11, 2029-2046.

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Mushroom Tyrosinase Inhibition Assay

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Mushroom tyrosinase (EC 1.14.18.1, Sigma-Aldrich) was dissolved in 1/15 M PBS (pH 6.8) to a concentration of 276 units/mL and used. About 150 μL of Mushroom tyrosinase (13.8 units/mL) was reacted with L-tyrosine (T−3754, Sigma-Aldrich) and cycloheterophyllin (10, 1, 0.1 ug/mL) for 5 min, and the absorbance was measured at a wavelength of 475 nm. Each experimental group was independently tested three times, and the tyrosinase enzyme inhibition rate (%) of the material was calculated by applying the following calculation formula. As a positive control, kojic acid (K3125, Sigma-Aldrich) at a concentration of 100 μg/mL was used.
Tyrosinase inhibition ratio (%) = (1 − absorbance of treatment/absorbance of non-treatment) × 100

, & Shim J.H. (2021). Inhibitory Effects of Cycloheterophyllin on Melanin Synthesis. Molecules, 26(9), 2526.

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Mushroom Tyrosinase Inhibition Assay

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The measurement of Mushroom tyrosinase activity was modified from the published method [65 (link),66 (link)]. l-Tyrosine (Bio Basic, Ontario, Canada) or l-DOPA (Sigma Chemical, St. Louis, MO, USA) were used as substrates for monophenolase and diphenolase activity, respectively. Each sample was diluted to a series of concentrations (0.039, 0.078, 0.156, 0.3125, 0.625, 1.25, 2.5, and 5 mg/mL). Mushroom tyrosinase (Sigma Chemical, St. Louis, MO, USA) was dissolved in 0.1 M phosphate buffer (pH = 6.8) to obtain the final concentration of 100 units/mL. The reaction was started by adding 40 µL of sample solution, 80 µL of phosphate buffer, 40 µL of Mushroom tyrosinase, and 40 µL of substrates (1.5 mM l-tyrosine or 2.5 mM l-DOPA solution). After incubation at room temperature for 15 min, the l-dopachrome formation was measured at 475 nm. The percentage of Mushroom tyrosinase inhibition was calculated using the following equation:
where A = vehicle control, B: = vehicle control without Mushroom tyrosinase, C = sample mixed with Mushroom tyrosinase, and D = sample without Mushroom tyrosinase. The results were expressed as IC₅₀ values (the concentration that caused 50% Mushroom tyrosinase inhibition).

Linsaenkart P., Ruksiriwanich W., Jantrawut P., Chittasupho C., Rachtanapun P., Jantanasakulwong K., Sommano S.R., Prom-u-thai C., Jamjod S., Arjin C., Sringarm K, & Barba F.J. (2023). Natural Melanogenesis Inhibitor, Antioxidant, and Collagen Biosynthesis Stimulator of Phytochemicals in Rice Bran and Husk Extracts from Purple Glutinous Rice (Oryza sativa L. cv. Pieisu 1 CMU) for Cosmetic Application. Plants, 12(4), 970.

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Tyrosinase Inhibition by Dienols

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Small molecule dienols’ (4a–e) tyrosinase inhibition activity were assayed using a modified protocol by Chen et al. and compared to that of free KA in vitro.^{9 (link)} Mushroom tyrosinase inhibition was determined by adding dienols in sample media (25 µL) to 96-well plates containing phosphate buffer (80 µL, pH = 6.8) and 125 µL substrate (0.5 mM L-DOPA) and incubated for 5 min at room temperature.^{9 (link)} Mushroom tyrosinase (20 µL, 1250 U/mL, Sigma, Milwaukee, WI) in phosphate buffer (pH = 6.8) was added and incubated an additional 5 min at room temperature. The amount of dopachrome produced was measured using a microplate reader (Coulter, Boulevard Brea, CA) at 475 nm. Dienol (4a–e) tyrosinase inhibition was expressed as a function of the dienol concentration and inhibitory concentration 50 (IC₅₀) values calculated and compared to KA (positive control).

Faig J.J., Moretti A., Joseph L.B., Zhang Y., Nova M.J., Smith K, & Uhrich K.E. (2017). Biodegradable Kojic Acid-Based Polymers: Controlled Delivery of Bioactives for Melanogenesis Inhibition. Biomacromolecules, 18(2), 363-373.

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Mushroom Tyrosinase Inhibition Assay

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mushroom tyrosinase Inhibitory Activities The reac-tion mixture for mushroom tyrosinase (EC 1.14.18.1, Sigma) activity consisted of 150 µL of 0.1 M phosphate buffer (pH 6.5), 3 µL of sample solution, 8 µL of mushroom tyrosinase (2100 unit/mL, 0.05 M phosphate buffer at pH 6.5), and 36 µL of 1.5 mM L-tyrosine. Tyrosinase activity was determined by reading the optical density at 475 nm on a microplate reader (Bio-Rad 3550, Richmond, CA, U.S.A.) after incubation at 37°C for 20 min. The inhibitory activity of the sample was expressed as the concentration that inhibits 50% of the enzyme activity (IC 50 ).

Lee D.Y., Jeong S.C., Jeong Y.T., Lee M.K., Seo K.H., Lee J.W., Kim G.S., Lee S.E., Baek N.I, & Kim J.H. (2015). Antimelanogenic Effects of Picrionoside A Isolated from the Leaves of Korean Ginseng. Biological & pharmaceutical bulletin, 38(10).

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Melanin Production Inhibition Assay

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B16F10 cells were treated with Fsk (20 µM) for 48 h and recovered in 2 mL tubes (approximately 5 × 10⁶ cells). The cells were suspended in 500 µL of PBS (50 mM, pH 6.8) and lysed by sonication for 10 s. Ten µL of the cell lysate with 83 µL of PBS and 2 µL of GIF-2209 or GIF-2216 in DMSO (maximum 100 µM) were mixed in 96-well plates [18 (link)]. The reaction was initiated by the addition of l-DOPA (5 µL, final concentration 900 µM), and the monitored using a spectrophotometer (GloMax-Multi Detection System, Promega) at OD450. The reliability of the assay was verified with mushroom tyrosinase (4.15U; Merck) in PBS (Figure 8).

Watanabe M., Kawaguchi K., Nakamura Y., Furuta K, & Takemori H. (2021). GIF-2209, an Oxindole Derivative, Accelerates Melanogenesis and Melanosome Secretion via the Modification of Lysosomes in B16F10 Mouse Melanoma Cells. Molecules, 27(1), 177.

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Mushroom Tyrosinase Antioxidant Assay

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Mushroom tyrosinase, butylated hydroxytoluene (BHT), α-tocopherol, 6-hydroxy-2,5,7,8-tetramethylcroman-2-carboxylic acid (Trolox), trichloroacetic acid (TCA), gallic acid, potassium ferricyanide, ferric chloride, ferrous chloride, ferrozine, 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2′-azino-bis-3-ethylbenzthiazoline-6- sulphonic acid (ABTS), Folin–Ciocalteu reagent, L-tyrosine, and L-3,4-dihydroxyphenylalanine (L-DOPA) were obtained from Merck Co. (Darmstadt, Germany). All the chemicals and solvents used in the study were of analytical grade or of high-performance liquid chromatography. Antibodies against microphthalmia-associated transcription factor (MITF) and β-actin were obtained from Cell Signalling Technology (Danvers, MA, U.S.A.) and Sigma Chemical Co. (St. Louis, U.S.A.), respectively.

Ho Y.S., Wu J.Y, & Chang C.Y. (2019). A New Natural Antioxidant Biomaterial from Cinnamomum osmophloeum Kanehira Leaves Represses Melanogenesis and Protects against DNA Damage. Antioxidants, 8(10), 474.

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Enzymatic Assays for Bioactive Compounds

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All chemicals and reagents, solvents, α-amylase (EC 3.2.1.1), α-glucosidase (EC 3.2.1.20) and mushroom tyrosinase (EC 1.14.18.1) were purchased from Merck (Milan, Italy).

Mirabile S., D’Angelo V., Germanò M.P., Pouramin Arabi S., Parisi V., Raimondo F.M, & Rosa E. (2023). Chemical Profile and Health-Promoting Activities of Crataegus laciniata (Rosaceae) Flowers. Plants, 13(1), 34.

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Development and Characterization of Copper Tripeptide Formulations

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The copper tripeptide (SpecPed-GCu11P, Spec-Chem Industry Inc., Nanjing, China) was of cosmetic grade and was kindly supplied by Alfa Sagittarius (Kraków, Poland). Hydrogenated lecithin (Emulmetik 950, Lucas Meyer Cosmetics, Massy, France) was kindly supplied by Naturallia Sp. z o.o. (Brzeg, Poland). Dicetyl phosphate, N-Succinyl-Ala-Ala-Ala-p-nitroanilide, elastase from porcine pancreas, mushroom tyrosinase and L-DOPA were all purchased from Sigma-Aldrich (Merck KGaA, Darmstadt, Germany). Stearylamine (97%) and cholesterol (95%) were purchased from Alfa Aesar (Thermo Fisher (Kandel) GmbH, Kandel, Germany). From Pol-Aura (Zawroty, Poland), 0.1 M Tris-HCl buffer was purchased. All other chemicals, such as potassium dihydrogen phosphate (POCh, Gliwice, Poland), sodium hydrogen phosphate (POCh), methanol (Chempur, Piekary Śląskie, Poland) and acetonitrile (Chempur), were of analytical grade. Spectra/Por 1 regenerated cellulose membrane with a 6–8 kDa molecular weight cut-off (MWCO), which was used in the microdialysis tests, was purchased from Spectrum Laboratories Inc., (DG Breda, The Netherlands). The deionized water used in all formulations was additionally filtered through a Milli-Q system.

Dymek M., Olechowska K., Hąc-Wydro K, & Sikora E. (2023). Liposomes as Carriers of GHK-Cu Tripeptide for Cosmetic Application. Pharmaceutics, 15(10), 2485.

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