Applications of Panax ginseng leaves-mediated gold nanoparticles in cosmetics relation to antioxidant, moisture retention, and whitening effect on B16BL6 cells

Abstract

Background: Bioactive compounds in plant extracts are able to reduce metal ions to nanoparticles through the process of green synthesis. Panax ginseng is an oriental medicinal herb and an adaptogen which has been historically used to cure various diseases. In addition, the P. ginseng leaves-mediated gold nanoparticles are the value-added novel materials. Its potential as a cosmetic ingredient is still unexplored. The aim of this study was to evaluate the antioxidant, moisture retention and whitening properties of gold nanoparticles (PgAuNPs) in cosmetic applications.

Methods: Cell-free experiments were performed to evaluate PgAuNP's antioxidant and moisture retention properties and inhibition activity on mushroom tyrosinase. Furthermore, in vitro cell cytotoxicity was evaluated using normal human dermal fibroblast and murine B16BL6 melanoma cells (B16) after treatment with increasing concentrations of PgAuNPs for 24 h, 48 h, and 72 h. Finally, in vitro cell assays on B16 cells were performed to evaluate the whitening effect of PgAuNPs through reduction of cellular melanin content and tyrosinase activity.

Results: In vitro DPPH radical scavenging assay results revealed that PgAuNPs exhibited antioxidant activity in a dose-dependent manner. PgAuNPs exhibited moisture retention capacity and effectively inhibited mushroom tyrosinase. In addition, 3-(4,5-dimethyl-thiazol-2yl)-2,5-diphenyl tetrazolium bromide results revealed that PgAuNPs were not toxic to human dermal fibroblast and B16 cells; in addition, they significantly reduced melanin content, tyrosinase activity, and mRNA expression of melanogenesis-associated transcription factor and tyrosinase in B16 cells.

Conclusion: Our study is the first report to provide evidence supporting that P. ginseng leaves-capped gold nanoparticles could be used as multifunctional ingredients in cosmetics.

Keywords: Panax ginseng leaves; antioxidant; antityrosinase activity; gold nanoparticles; moisture retention.

Fig. 1

Increasing 2,2-diphenyl-1-picrylhydrazyl (DPPH) free-radical-scavenging activity of different concentrations (1–150 μg/mL) PgAuNPs. Results are expressed as mean ± SD of three separate experiments. For DPPH assay, ascorbic acid (1–150 μg/mL) was used as reference standard. PgAuNPs, Panax ginseng leaves-capped gold nanoparticles; SD, standard deviation.

Fig. 2

Moisture retentions of PgAuNP solution (1–10%). Results are expressed as mean ± SD of three separate experiments. For moisture retention assay, glycerin solution (10%) was used as reference standard. PgAuNPs, Panax ginseng leaves-capped gold nanoparticles; SD, standard deviation.

Fig. 3

Cell viability of HDF and B16BL6 cells after treatment with PgAuNPs at different time points: (A) 24 h. (B) 48 h. (C) 72 h. Cell viability of HDF and B16BL6 cells after treatment with RA for different time intervals: (D) 24 h. (E) 48 h. (F) 72 h. Cells (1 × 10⁵ cells/well) were incubated with various concentrations (0.1–100 μg/mL) of PgAuNPs or RA. Cell viability was determined by 3-(4,5-dimethyl-thiazol-2yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Note that samples did not show any toxicity effect up to 100 μg/mL for PgAuNPs. Results are expressed as a percentage of sample-treated control and presented as mean ± SD of three separate experiments. ***p < 0.001 versus control by Student t test. HDF, human dermal fibroblast; PgAuNPs, Panax ginseng leaves-capped gold nanoparticles; RA, retinoic acid; SD, standard deviation.

Fig. 4

Inhibition effect of PgAuNPs on melanin content in B16 cells. Cells (2 × 10⁵ cells/well) were incubated with different concentrations (1–100 μg/mL) of PgAuNPs or arbutin (100 μg/mL) in the presence of 100nM of α-MSH for 72 h. (A) Extracellular melanin content, the melanin levels contained in 100 μL of the medium containing the cells were determined reading the OD value of each sample. Below is a picture of the color intensity of extracellular melanin production in media. (B) Intracellular melanin content was determined as described in Materials and methods. Below is a picture of B16 cell pellet after centrifugation. Results are expressed as a percentage of α-MSH-treated control and presented as mean ± SD of three separate experiments. *p < 0.05, **p < 0.01, ***p < 0.001 versus α-MSH treated control by Student t test. α-MSH, α-melanocyte stimulating hormone; OD, optical density; PgAuNPs, Panax ginseng leaves-capped gold nanoparticles; SD, standard deviation. C stands as for assay control and ^###P 0.001 versus assay control (C).

Fig. 5

Inhibitory effect of melanogenesis of PgAuNPs through inhibition of tyrosinase activity. B16 cells (4 × 10⁵ cells/well) were incubated with various concentrations of (1–100 μg/mL) PgAuNPs or arbutin (100 μg/mL) in the presence of 100nM of α-MSH for 72 h. Tyrosinase activity in cellular lysates was determined as described in Material and methods. Results are expressed as a percentage of α-MSH-treated control and presented as mean ± SD of three separate experiments. **p < 0.01, ***p < 0.001 versus α-MSH-treated control by Student t test. α-MSH, α-melanocyte stimulating hormone; PgAuNPs, Panax ginseng leaves-capped gold nanoparticles; SD, standard deviation. C stands as for assay control and ^###P 0.001 versus assay control (C).

Fig. 6

Effect of on PgAuNPs mRNA expression of melanogenesis-related genes. Cells (4 × 10⁵ cells/well) were treated with various concentrations of (1–100 μg/mL) PgAuNPs or arbutin (100 μg/mL) in the presence of 100nM of α-MSH for 72 h. Then cells were harvested and total RNA was extracted. mRNA expression was visualized by RT-PCR. (A) Tyrosinase (TYR). (B) Microphthalmia-associated transcription factor (MITF). The size of amplified gene products was 528 bp for GAPDG. (C) Tyrosinase, 563 bp.(D) MITF, 216 bp. Results as a percentage of α-MSH-treated control and presented as mean ± SD of three separate experiments. *p < 0.05, **p < 0.01, ***p < 0.001 versus α-MSH-treated control by Student t test. α-MSH, α-melanocyte stimulating hormone; PgAuNPs, Panax ginseng leaves-capped gold nanoparticles; RT-PCR, reverse transcription-polymerase chain reaction; SD, standard deviation. C stands as for assay control and ^###P 0.001 versus assay control (C).