Study on the inhibitory effect of oleuropein on tyrosinase
Tyrosinase (EC. 1.14.18.1) is widely present in microorganisms, animals, plants, and humans, and is an important rate limiting enzyme involved in the process of melanin synthesis. Its expression level and activity determine the speed and yield of melanin synthesis. Inhibiting the activity of tyrosinase can improve the tyrosinase metabolism of pigment cells in the skin, prevent the formation of pigmentation, and play a role in preventing the formation of freckles, melasma, age spots, and whitening. The commonly used tyrosinase inhibitors, such as quercetin and hydroquinone, have good inhibitory abilities, but can cause side effects such as dermatitis, skin allergies, and even skin cancer. Therefore, the search for safe, effective, and non-toxic tyrosinase inhibitors is currently a hot research topic. At present, research has shown that phenols, flavonoids and their derivatives, polysaccharides, volatile oils, organic acids, coumarins, and other compounds in many plant extracts are good sources of tyrosinase inhibitors.
Oleuropein is a iridoid glycoside compound formed by hydroxytyrosol and olive skeleton, distributed throughout the olive tree, with the highest content in olive leaves, followed by olive fruit. In recent years, with the continuous deepening of research on oleuropein by domestic and foreign scholars, it has been found that it has various biological activities, such as antiviral, anti-tumor, antibacterial, antioxidant, hypoglycemic, hypotensive, anti Alzheimer’s disease, skin wound protection, etc. However, there are few reports on the effect of oleuropein on tyrosinase activity. This article aims to provide theoretical basis for the development and utilization of oleuropein as a whitening ingredient by studying its effect on tyrosinase activity and its mechanism of action.

In enzymatic reactions, the rate of enzymatic reaction is expressed by measuring the increment of products per unit time. The initial velocity refers to the velocity within a certain period of time at the beginning of the enzymatic reaction, at which the slope remains almost unchanged. At a certain substrate concentration, it is obtained from the slope of the zero crossing tangent of the product time relationship curve. The method of measuring the initial rate of enzymatic reaction is generally used to determine enzyme activity, with fewer interfering factors. In this experiment, the A475nm Time curve showed an approximately linear relationship from 0 to 5.3 minutes of reaction. As time increased, the curve gradually flattened, the slope decreased, and the reaction rate also decreased. At this time, the measured enzyme activity could not represent the true enzyme activity, so the reaction rate at 5.3 minutes of reaction was taken as the initial rate.
The experiment of inhibiting tyrosinase activity in vitro is divided into cell experiments and biochemical experiments. Chan et al. believe that cells can simulate real physiological environments, and measuring tyrosinase activity through cell experiments is more reliable than directly measuring tyrosinase activity, and can better reflect its inhibitory activity. In this study, there were differences in the inhibitory activity of oleuropein on B16 cell tyrosinase and mushroom tyrosinase. At the cellular level, oleuropein inhibited tyrosinase activity better than the positive control quercetin. This is consistent with the research results of Li et al.
When human skin is exposed to ultraviolet radiation, it produces a large amount of reactive oxygen species (ROS) and free radicals, which accelerate skin aging, pigmentation, and deepening wrinkles. Research has shown that elevated ROS activates tyrosinase by mobilizing alpha melanocyte stimulating hormone in the epidermis, ultimately stimulating melanocytes to produce melanin. Antioxidants promote the normal metabolism of the body by clearing reactive oxygen species and free radicals, improving the condition of the skin, gradually dissipating pigments, and thus playing a whitening role. Wang et al. found that antioxidants such as glycyrrhizic acid, resveratrol, oxidized resveratrol, and phenethyl resorcinol have a synergistic effect between antioxidant activity and tyrosinase inhibition when using L-dopa as a substrate, greatly inhibiting tyrosinase activity. Olive bitter glycoside has strong antioxidant activity. Whether its inhibition of tyrosinase activity is related to antioxidant activity and whether there is a synergistic effect between olive bitter glycoside and antioxidants deserves further exploration.
Tyrosinase is a key enzyme in the synthesis of melanin. By inhibiting the activity of tyrosinase, the production of melanin can be suppressed, thereby achieving the effect of whitening. The results of this study indicate that oleuropein has a significant inhibitory effect on tyrosinase in B16 cells and mushroom tyrosinase, showing a good dose-dependent relationship. Through kinetic analysis, the type of inhibitory effect of oleuropein on mushroom tyrosinase is reversible non competitive inhibition, that is, the binding of oleuropein to tyrosinase is on an essential group outside the substrate binding site. Oleuropein does not reduce the affinity of tyrosinase to the substrate (L-dopa), but rather inhibits enzyme activity by blocking the catalytic action of the enzyme, reducing Vm. Olive bitter glycoside has a wide range of sources, high safety, and good antioxidant activity. As a potential raw material for whitening products, it has broad application prospects.