The following are excerpts of a study reported in Scientific Reports volume 5, Article number: 7995 (2015)
Abstract
Tyrosinase is involved in melanin biosynthesis and the abnormal accumulation of melanin pigments leading to hyperpigmentation disorders that can be treated with depigmenting agents. A natural product T1, bis(4-hydroxybenzyl)sulfide, isolated from the Chinese herbal plant, Gastrodia elata, is a strong competitive inhibitor against mushroom tyrosinase (IC50 = 0.53 μM, Ki = 58 ± 6 nM), outperforms than kojic acid. The cell viability and melanin quantification assay demonstrate that 50 μM of T1 apparently attenuates 20% melanin content of human normal melanocytes without significant cell toxicity. Moreover, the zebrafish in vivo assay reveals that T1 effectively reduces melanogenesis with no adverse side effects. The acute oral toxicity study evidently confirms that T1 molecule is free of discernable cytotoxicity in mice. Furthermore, the molecular modeling demonstrates that the sulfur atom of T1 coordinating with the copper ions in the active site of tyrosinase is essential for mushroom tyrosinase inhibition and the ability of diminishing the human melanin synthesis. These results evident that T1 isolated from Gastrodia elata is a promising candidate in developing pharmacological and cosmetic agents of great potency in skin-whitening.
Introduction
Melanin is the major cellular component commonly observed in bacteria, fungi, plants, and animals responsible for skin color1. It presents as a complex, heterogeneous polyphenol-like biopolymer structure and colors vary from yellow to black2, secreted by melanocyte cells in the basal layer of the dermis3. Normal melanin pigmentation is able to shield from UV radiation, inhibit photocarcinogenesis and affect the synthesis of vitamin D34. In contrast, the abnormal pigmentation, such as senile lentigines, freckles, melasma, and other forms of melanin hyperpigmentation, causes serious esthetic problems5,6. The oxidative reactions of the tyrosine catalyzed by tyrosinase mainly contributes to the melanin biosynthesis7.
As a binuclear copper enzyme, tyrosinase (monophenol monooxygenase EC 1.14.14.1) catalyzes two distinct reactions of melanin biosynthesis. It catalyzes phenols to catechols and further oxidizes catechols to quinones8. The tyrosinase contains two copper ions, coordinating with histidine residues in the active site. The two copper ions are critical for the catalytic activities of this enzyme9 and exist in different tyrosinases regardless of their source10,11. Since tyrosinase-catalyzed reaction is highly associated with local hyperpigmentaiton such as ephelide, melasma, and lentigo5, discovering of tyrosinase inhibitors are of great importance in cosmetic and medicinal products for the prevention of pigmentation disorders12. Recently, significant efforts have been made to search for the tyrosinase inhibitors with copper chelator ability as whitening and anti-hyperpigment agents13,14,15,16,17,18, and several tyrosinase inhibitors have also been used as depigmentation ingredients of medical products19,20,21,22.
Many tyrosinase inhibitors, such as hydroquinone23,24,25,26, kojic acid20, azelaic acid27,28, electron-rich phenols29, and arbutin have been tested in pharmaceuticals and cosmetics for their capability of preventing overproduction of melanin30,31. Meanwhile, their structure-activity relationship (SAR) analysis have been widely discussed32 (Supplementary Table S1). Hydroquinone is one of the most frequently prescribed ingredients among the conventional skin-whitening agents. However, hydroquinone causes skin irritation33, and it is thought to be mutagenic to mammalian cells34 and cytotoxic to melanocytes. This leads to the use of kojic acid and arbutin as alternative agents, but these agents show poor efficacy in vivo. In addition, due to their adverse side effects, low formulation stability, and poor skin penetration35, their use is still limited. The Japanese officials also spurred to ban the use of kojic acid in skin treatment due to its carcinogenicity36. Thus, it is in great need of developing new tyrosinase inhibitors from different sources. In this regard, the natural products have been extensively utilized in the cosmetics industry37,38,39 because of lower adverse side effects and high safety.
Recently, the extracts and isolated compounds of a number of natural sources in particular botanical sources were well characterized with anti-tyrosinase activities40,41,42,43,44,45,46,47,48,49,50,51 and have been accepted as popular skin lightening agents52,53,54,55,56. In this study, we have screened 78 different medicine herbal plants and identified that rhizome of Gastrodia elata is of great potency for tyrosinase inhibition. The rhizome of Gastrodia elata has been applied in for the treatment of dizziness, headaches, vertigo, and convulsive illnesses57. The studies of how Gastrodia elata prevents the neuronal damage have been performed as well58,59,60,61,62,63, but its efficacy on tyrosinase inhibition and melanin biosynthesis has not been thoroughly investigated. Thus, here we aim to isolate the functional components from the rhizome of Gastrodia elata and investigate its inhibitory effect on the mushroom tyrosinase and human melanogenesis. The natural compounds, T1 and T2 extracted from Gastrodia elata and their derivatives T3–T5, exert profound mushroom tyrosinase inhibitory abilities. The bioactive natural product T1, bis(4-hydroxybenzyl)sulfide, with the most striking inhibitory potency against tyrosinase, was chosen as the target compound to characterize its biological effects in tyrosinase inhibition, cell viability, in vitro and in vivo melanin biosynthesis, and acute oral toxicity in mice.
Conclusion
This study screened 78 different Chinese herbal plants and identified the potent and functional components useful for tyrosinase inhibition, melanogenic inhibition, and depigmentation. The natural component T1, isolated from Gastrodia elata, is a strong competitive inhibitor of mushroom tyrosinase (IC50 = 0.53 μM, Ki = 58 ± 6 nM), which is superior to kojic acid (IC50 = 40.69 μM) and more effective than other known natural products. The in vitro assay demonstrates that 50 μM of T1 apparently attenuates 20% melanin content of human normal melanocytes and shows no significant cell toxicity. The in vivo assay reveals that T1 effectively reduces melanogenesis in zebrafish without any adverse side effects. The acute oral toxicity study evidently confirms that the T1 molecule is free of discernable cytotoxicity in mice. The molecular models reveals that the sulfur atom of T1 interacting with the copper ions in the active site of tyrosinase is crucial for mushroom tyrosinase inhibition and the ability of disturbing the human melanin synthesis. In conclusion, T1 isolated from Gastrodia elata is a potential candidate in developing safe cosmetic and pharmacological agents, which is of great potency in skin-whitening.