Comparative Study of High-throughput Screening Models for Anti-hyperpigmentation Compounds

HIGHLIGHTS

• It is the first time to compare mushroom tyrosinase model, B16 mouse melanoma cell model and zebrafish model.

• A high-throughput method for the quantification of melanin in zebrafish skin based on the Phyton for the first time.

Abstract

The mushroom tyrosinase assay, B16-F10 mouse melanoma cell model, and zebrafish model are frequently used for high-throughput screening and are widely used for developing anti-hyperpigmentation compounds, although these systems cannot be compared. We used each of these three systems to evaluate the seven anti-hyperpigmentation compounds. We investigated 1. tyrosinase activity using a mushroom tyrosinase assay, 2. viability, tyrosinase activity, and melanin content in B16-F10 cells, and 3. embryonic toxicity, tyrosinase activity, and melanin content in zebrafish. α-Arbutin, raspberry ketone (RK), raspberry ketone glucoside (RKG), glabridin (GLA), and 3-o-ethyl-ascorbic (EA), inhibited the activity of mushroom tyrosinase; dipotassium glycyrrhizinate (DG) did not inhibit mushroom tyrosinase activity, and glycyrrhetic acid (GA) promoted tyrosinase activity. Tyrosinase activity was inhibited by α-arbutin, GLA, GA and DG in B16-F10 cells; RK, RKG and EA did not inhibit tyrosinase activity. α-Arbutin, RK, RKG, EA, and GA, inhibited tyrosinase activity in zebrafish; GLA and DG did not inhibit tyrosinase activity. α-arbutin, RK, RKG, EA, GLA, and DG reduced melanin synthesis in B16-F10 cells in a dose-dependent manner without significant toxicity; GA did not inhibit melanin synthesis. α-arbutin, RK, RKG and GA significantly reduced melanin content on the zebrafish body surface. Mushroom tyrosinase analysis was the most practical assay among the three systems but had poor reliability. The B16-F10 mouse melanoma cell system was the most sensitive but had the worst stability. The zebrafish system had better reproducibility than other systems; however, most compounds were difficult to screen in this system.

Keywords:
Melanin; Zebrafish; B16-F10 mouse melanoma cell; Whitening ingredient.