Study of Tyrosinase inhibitors from natural sources Organic & Natural Product Chemistry Lab CMLS, Cell. Mol. Life Sci. 62 (2005) Presented by Hoi Young Kim

Contents Contents Organic & Natural Product Chemistry Lab Melanin Tyrosinase Tyrosinase inhibitors Applicaton Biosynthetic pathway of melanin Function of melanin Mechanism of tyrosinase action Natural and synthetic sources New Development of Tyrosianse inhibitors

Melanin Melanin Melanin is one of the most widely distributed pigments and is found in Melanin is one of the most widely distributed pigments and is found in bacteria, fungi, plants and animals. It is a heterogeneous polyphenol-like biopolymer with a complex structure and color varying from yellow to black. melanoine keranocyte melanocyte melanine Melanocyte Organic & Natural Product Chemistry Lab

Biosynthetic pathway of melanin Biosynthetic pathway of melanin EumelaninPheomelanin Mixed-melanins Organic & Natural Product Chemistry Lab

Melanin Function Melanin Function Several types Several types - Eumelanin (predominant in human) - Eumelanin (predominant in human) - Pheomelanin - Pheomelanin Functional bio-macromolecule found throughout nature: Functional bio-macromolecule found throughout nature: - Increases that live and competition - Increases that live and competition - protect the skin from ultraviolet(UV) - protect the skin from ultraviolet(UV) - Detoxification for cell toxicity substances - Detoxification for cell toxicity substances Melanin overproduction: Melanin overproduction: bring about freckles, hyperpigmentation and cancer bring about freckles, hyperpigmentation and cancer Melanin is formed through a series of oxidative reactions involving the Melanin is formed through a series of oxidative reactions involving the amino acid tyrosine in the presence of tyrosinase amino acid tyrosine in the presence of tyrosinase Organic & Natural Product Chemistry Lab

Monophenolase activity Monophenolase activity Diphenolase activity Diphenolase activity Tyrosinase (EC ) Tyrosinase (EC ) Phenolase, Monopheol oxidase, Catechol oxidase, etc. Phenolase, Monopheol oxidase, Catechol oxidase, etc. Copper-containing enzyme. Copper-containing enzyme. Catalyzes two distinct reactions of melanin synthesis Catalyzes two distinct reactions of melanin synthesis Organic & Natural Product Chemistry Lab

Classification and properties of tyrosinase Classification and properties of tyrosinase Table 1. Properties of Well-Characterized Tyrosinases source Number of subunits Molecular weight Of subunit (kDa) Spectroscopy of oxy form Absorption (nm) CD (nm) Streptomyces glaucescens (Eubacteria) Neurospora crassa (Fungi) Agaricus bisporus (Fungi; mushroom) Beta vulgaris (Plant; spinach-beet) Human melanocyte (Animal) a a No data available. Organic & Natural Product Chemistry Lab

Mechanism of tyrosinase action Mechanism of tyrosinase action Figure. Catalytic cycles of the hydroxylation of monophenol and oxidation of o-quinone by tyrosinase Monophenolase Catalytic Cycle Diphenolase Catalytic cycle Organic & Natural Product Chemistry Lab

Tyrosinase inhibitors from natural sources Tyrosinase inhibitors from natural sources Structure of tyrosinase inhibitors Organic & Natural Product Chemistry Lab

Tyrosinase inhibitors from natural sources Tyrosinase inhibitors from natural sources Mushroom Tyrosinase inhibitory activities of the Flavonoids Mushroom Tyrosinase inhibitory activities of the Flavonoids IC50 (μM) Compound No R 3 OH R 5 OH R 6 OH R 7 OH R 8 OH R 2' OH R 3' OH R 4' OH R 5' OH R 6' Organic & Natural Product Chemistry Lab

Tyrosianse inhibitors from synthetic sources Tyrosianse inhibitors from synthetic sources Structure of tyrosinase inhibitors Organic & Natural Product Chemistry Lab

Plant Source : Sophora flavescens Plant Source : Sophora flavescens Organic & Natural Product Chemistry Lab Plant taxonomy Plant taxonomy - Leguminous - Leguminous - Herbaceous perennial plant - Herbaceous perennial plant Pharmacological reserch Pharmacological reserch - Antipyretic - Antibiotic - Antipyretic - Antibiotic - Diuretic - Antifungal - Diuretic - Antifungal - Anthelmintic - Antitumor - Anthelmintic - Antitumor Main component Main component - Flavonoids - Flavonoids - Quinolizidine alkaloids - Quinolizidine alkaloids - Triterpenoid - Triterpenoid

Reported compounds from Sophora flavescens Organic & Natural Product Chemistry Lab

Isolated compounds from Sophora flavescens Organic & Natural Product Chemistry Lab Compound 1 Kushenol Compound 2 Sophoraflavanone G Compound 3 (2S)-2’-methoxykurarinone Compound 4 Kurarinone Compound 5 Kurarinol

5. Kurarinol Compound12345Arbutin Kojic acid IC 50 (μM) >302> K i (μM) Inhibition type -Noncompetitive-NoncompetitiveCompetitive-- Organic & Natural Product Chemistry Lab Tyrosinase inhibition compounds from Sophora flavescens 4. Kurarinone 2. Sophoraflavanone G

Enzyme kinetic analysis Enzyme kinetic analysis Organic & Natural Product Chemistry Lab Figure. Lineweaver-Burk plots for inhibition of Kurarinone on mushroom tyrosinase for catalysis of L-tyroine at 30 ℃, pH 6.8 Figure. Lineweaver-Burk plots for inhibition of Kurarinol on mushroom tyrosinase for catalysis of L-tyroine at 30 ℃, pH 6.8 Lineweaver-Burk plot

Organic & Natural Product Chemistry Lab Conclusion Tyrosinase inhibition mechanism stydy. Tyrosinase inhibition mechanism stydy. SAR (Structure Activity Relationship) from Flavonoids SAR (Structure Activity Relationship) from Flavonoids Isolated of potent tyrosinase inhibitors from Sophora flavescens Isolated of potent tyrosinase inhibitors from Sophora flavescens - Sophoraflavaone G (IC50 =2μM, noncompetitive inhibition) - Sophoraflavaone G (IC50 =2μM, noncompetitive inhibition) - Kurarinone (IC50 = 0.1μM, noncompetitive inhibition) - Kurarinone (IC50 = 0.1μM, noncompetitive inhibition) - Kurarinol (IC 50 = 0.05μM, competitive inhibition) - Kurarinol (IC 50 = 0.05μM, competitive inhibition) Active site - Hydroxyl groups at the R 7 and R 4' sites in the A and B rings. - Dihydroxyl group in the A and B rings. Development of Whitening agent