Ultraviolet radiation from sunlight induces overproduction of reactive oxygen species (ROS) resulting in skin photoaging and hyperpigmentation disorders. Novel whitening and anti-wrinkle compounds from natural products have recently become of increasing interest. The purpose of this study was to find products that reduce ROS in 14 Thai plant extracts. To determine total phenolic and flavonoid content, antioxidant activity, anti-tyrosinase activity and anti-collagenase activity, we compared extracts of 14 Thai plants prepared using different solvents (petroleum ether, dichloromethane and ethanol). Antioxidant activities were determined by DPPH and ABTS assays. Total phenolic content of the 14 Thai plants extracts was found at the highest levels in ethanol followed by dichloromethane and petroleum ether extracts, respectively, while flavonoid content was normally found in the dichloromethane fraction. Scavenging activity ranged from 7 to 99% scavenging as assessed by DPPH and ABTS assays. The ethanol leaf extract of Ardisia elliptica Thunb. had the highest phenolic content, antioxidant activity and collagenase inhibition, while Cassia alata (L.) Roxb. extract had the richest flavonoid content. Interestingly, three plants extracts, which were the ethanolic fractions of Annona squamosa L., Ardisia elliptica Thunb. and Senna alata (L.) Roxb., had high antioxidant content and activity, and significantly inhibited both tyrosinase and collagenase. Our finding show that the ethanol fractions of Annona squamosa L., Ardisia elliptica Thunb. and Senna alata (L.) Roxb. show promise as potential ingredients for cosmetic products such as anti-wrinkle agents and skin whitening products.

Background: Ultraviolet radiation (UVR) is the major cause for hyperpigmentation, and to prevent this natural products are increasingly being explored as potential skin whitening agents. The aim of this study was to determine the total phenolic and flavonoid content, free radical scavenging activity, anti-tyrosinase activity and the inhibition of melanin content in α-melanocyte stimulating hormone-induced B16F10 melanoma cells of an aqueous extract of Garcinia atroviridis Griff. ex. T. Anderson fruit pericarps. Methods: The aqueous extract was prepared by extraction with distilled water at 105 o C for 60 min. Total phenolic and flavonoid content were determined using the Folin-Ciocalteau and aluminium chloride methods, respectively. Scavenging activity was assessed using 2,2-Diphennyl-1-picrylhydrazyl (DPPH) and 2,2′-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS). Tyrosinase activity and melanin content were determined spectrophotometrically. Results: The results showed that the aqueous extract of Garcinia atroviridis fruit pericarps had a phenolic (26.33 ± 0.77 mg GAE/g plant extract) and flavonoid content (9.31 ± 0.40 mg QE/g plant extract). The aqueous extract of Garcinia atroviridis significantly inhibited mushroom tyrosinase activity (IC 50 of 40.72 ± 1.83 µg/mL) and cellular tyrosinase activity (at a concentration of 125 µg/mL) in α-melanocyte stimulating hormone-induced B16F10 melanoma cells. The Garcinia atroviridis extract also suppressed melanin content at concentrations of 31.25-125 µg/mL. Correlations of mushroom tyrosinase inhibition with DPPH and ABTS scavenging activities were 0.8673 and 0.9468, respectively. Conclusion: Our findings show that an aqueous extract of Garcinia atroviridis fruit pericarps is a source of natural compounds and antioxidant capacity which can inhibit tyrosinase activity and melanin content. Thus, aqueous extracts of Garcinia atroviridis may be a potential source of skin whitening agents for hyperpigmentation.

Croton roxburghii and Croton sublyratus have been used as skin treatments in traditional medicine. The objective of the present study was to investigate the antimelanogenic effect of ethanol extracts of Croton roxburghii (CRE) and Croton sublyratus (CSE) leaves on cellular melanin content and cellular tyrosinase activity as mediated by the action of microthalmia transcription factor (MITF) and melanogenic enzymes. Croton roxburghii and Croton sublyratus leaves were extracted by petroleum ether, dichloromethane and absolute ethanol, sequentially. The ethanolic crude extracts were examined for antimelanogenic activity by their ability to decrease melanin content and cellular tyrosinase activity in alpha-melanocyte-stimulating hormone-stimulated B16F10 melanoma cells. In addition, the extracts were evaluated to determine a plausible mechanism of melanogenesis suppression through determining the activation of MITF transcription factor and melanogenic proteins (tyrosinase, tyrosinase-related protein 1 or TRP-1 and tyrosinase-related protein 2 or TRP-2) at the transcriptional and translation levels in α-MSH-induced B16F10 cells. Upon treatment with CRE and CSE, the cells showed significant decreases in melanin content and cellular tyrosinase activity. CRE and CSE also suppressed MITF, tyrosinase, TRP-1 and TRP-2 at the transcription and translation levels in α-MSH-stimulated melanin biosynthesis in B16F10 cells. Our finding shows that CRE and CSE inhibit melanin content and cellular tyrosinase activity through suppressing MITF and melanogenic enzymes. CRE and CSE may be useful to combine with skin whitening agents for cosmetic uses.

Questions about which reactive oxygen species (ROS) or reactive nitrogen species (RNS) can escape from the mitochondria and activate signals must be addressed. In this study, two parameters, the calculated dipole moment (debye, D) and permeability coefficient (Pm) (cm s⁻¹), are listed for hydrogen peroxide (H₂O₂), hydroxyl radical (•OH), superoxide (O₂^•−), hydroperoxyl radical (HO₂•), nitric oxide (•NO), nitrogen dioxide (•NO₂), peroxynitrite (ONOO⁻), and peroxynitrous acid (ONOOH) in comparison to those for water (H₂O). O₂^•− is generated from the mitochondrial electron transport chain (ETC), and several other ROS and RNS can be generated subsequently. The candidates which pass through the mitochondrial membrane include ROS with a small number of dipoles, i.e., H₂O₂, HO₂•, ONOOH, •OH, and •NO. The results show that the dipole moment of •NO₂ is 0.35 D, indicating permeability; however, •NO₂ can be eliminated quickly. The dipole moments of •OH (1.67 D) and ONOOH (1.77 D) indicate that they might be permeable. This study also suggests that the mitochondria play a central role in protecting against further oxidative stress in cells. The amounts, the long half-life, the diffusion distance, the Pm, the one-electron reduction potential, the pK_a, and the rate constants for the reaction with ascorbate and glutathione are listed for various ROS/RNS, •OH, singlet oxygen (¹O₂), H₂O₂, O₂^•−, HO₂•, •NO, •NO₂, ONOO⁻, and ONOOH, and compared with those for H₂O and oxygen (O₂). Molecules with negative electrical charges cannot directly diffuse through the phospholipid bilayer of the mitochondrial membranes. Short-lived molecules, such as •OH, would be difficult to contribute to intracellular signaling. Finally, HO₂• and ONOOH were selected as candidates for the ROS/RNS that pass through the mitochondrial membrane.

Melanin, the pigment responsible for human skin color, increases susceptibility to UV radiation, leading to excessive melanin production and hyperpigmentation disorders. This study investigated the ethanolic extract of Garcinia atroviridis fruits for its phenolic and flavonoid contents, antioxidant activity, and impact on melanogenesis pathways using qRT-PCR and Western blot analysis. Utilizing network pharmacology, molecular docking, and dynamics simulations, researchers explored G. atroviridis fruit extract’s active compounds, targets, and pharmacological effects on hyperpigmentation. G. atroviridis fruit extract exhibited antioxidant properties, scavenging DPPH^• and ABTS^•+ radicals radicals and chelating copper. It inhibited cellular tyrosinase activity and melanin content in stimulated B16F10 cells, downregulating TYR, TRP-1, phosphorylated CREB, CREB, and MITF proteins along with transcription levels of MITF, TYR, and TRP-2. LC-MS analysis identified thirty-three metabolites, with seventeen compounds selected for further investigation. Network pharmacology revealed 41 hyperpigmentation-associated genes and identified significant GO terms and KEGG pathways, including cancer-related pathways. Kaempferol-3-O-α-L-rhamnoside exhibited high binding affinity against MAPK3/ERK1, potentially regulating melanogenesis by inhibiting tyrosinase activity. Stable ligand–protein interactions in molecular dynamics simulations supported these findings. Overall, this study suggests that the ethanolic extract of G. atroviridis fruits possesses significant antioxidant, tyrosinase inhibitory, and anti-melanogenic properties mediated through key molecular targets and pathways.

Recently, paper has gained traction in the biotechnology research field due to its ability to be a substrate for 3D cell culture. In this work, we demonstrate the application of paper-based 3D cell culture for rapid and easy screening of the effect of natural compounds on melanin production. Whatman No. 1 filter paper was used as the substrate for B16F10 melanoma cell culture. The use of paper is beneficial for supporting the 3D structure of cells, which makes the result more reliable due to the similarity to in vivo conditions. Furthermore, paper is beneficial for melanin observation due to melanin's black color, which is easily in situ visualized after it is cultured on white paper. Matrigel was used to encapsulate cells before being pipetted onto the paper to prevent the passing of cells through paper pores. The intensity of melanin can then be observed with the naked eye and analyzed by scanning the paper. The analysis process took only 20 minutes, which is faster than that of the conventional absorbance spectroscopy, owing to the elimination of centrifugation, melanin solubilization, and the absorbance measurement step. The color intensity on the paper showed a direct proportion with increased α-MSH concentrations, confirming that the color on the paper was melanin. The 3D structure of cells was confirmed by using a scanning electron microscope. To demonstrate the application of the paper-based scaffold, paper-based 3D cell culture was used for screening the effects of Kojic acid and Arbutin on melanin production, which showed increased anti-melanogenesis effects with increased concentrations of natural compounds. High cell viability was observed over 120 hours. In conclusion, the developed paper-based scaffold can be used for screening the effect of natural compounds on melanin production, as a rapid and simple method with low cost.