Skin aging is a complex process influenced by both internal factors like genetics and external factors such as UV radiation and accumulation of reactive oxygen species, leading to visible signs like wrinkles, fine lines, loss of elasticity, and sagging. A significant contributor to the appearance of age is a lack of skin plumpness, emphasizing the need to maintain adequate production of skin layer components and reduce their degradation. Key elements like fibrillin-1, a protein crucial for elastic fibers at the dermal-epidermal junction, are degraded by photoaging, contributing to wrinkles and loss of elasticity. Similarly, extracellular matrix proteins like collagen and elastin are degraded by matrix metalloproteinases (MMPs), with MMP-1 being a key player in photoaging. Wrinkle formation also involves the thinning of the subcutaneous fat layer. Additionally, hyaluronic acid (HA), vital for skin hydration and elasticity, is found abundantly in the dermis and is regulated by enzymes like hyaluronic acid synthase (HAS2). Strategies to combat skin aging include suppressing the degradation of fibrillin proteins and MMP activation, while facilitating the synthesis of collagen fibers, subcutaneous fat, and HA.
Traditional medicine offers potential sources for novel anti-aging cosmetic ingredients. Gongjin-dan, an oriental medicine historically used as a restorative tonic for longevity in Korea and China, has scientifically demonstrated effects like improving chronic fatigue and modulating the immune system. Inspired by this traditional medicine, researchers explored a modified formula comprising Angelica gigas, Cornus officinalis, Ganoderma lucidum, Thymus vulgaris, and Asparagus cochinchinensis, ingredients with documented anti-aging potential. The goal was to investigate if a fraction enriched in glycopeptides and glycoproteins from this modified formula could offer skin anti-aging benefits, potentially leveraging important anti-aging constituents beyond the typical low-molecular-weight compounds found in extracts.
Methods
The study involved processing five raw herbal materials—Angelica gigas, Cornus officinalis, Ganoderma lucidum, Thymus vulgaris, and Asparagus cochinchinensis—by drying, cutting, and pulverizing them. A glycopeptide- and glycoprotein-enriched fraction (GEF) was obtained using an enzyme-based extraction process with commercial proteases (Protamex and Alcalase) followed by purification via ethanol precipitation. Chemical analysis was performed to identify glycopeptides and glycoproteins using ammonium sulfate precipitation and a carbohydrate estimation kit, and to determine total carbohydrate and protein contents using phenol-sulfuric acid and modified Lowry methods, respectively. Amino acid and monosaccharide compositions were analyzed using HPLC and HPAEC-PAD after hydrolysis. The in vitro skin anti-aging properties of GEF were evaluated on human fibroblasts (HS68), keratinocytes (HaCaT), and adipose-derived mesenchymal stem cells (ADMSCs) by assessing effects on fibrillin-1 synthesis via immunocytochemistry, gene expression of MMP-1, COL1A1, COL4A1, and HAS2 via RT-PCR, and adipogenesis via oil red O staining and absorbance measurement.
Key Findings
•GEF is a peptide and protein fraction with significant binding of oligosaccharide chains, confirmed by carbohydrate estimation showing higher absorbance (13.1) than standard glycoproteins like fetuin (5.6) and α1-acid glycoprotein (5.3) at the same concentration
•GEF exhibits a high proportion of conditionally essential amino acids (CEAAs) at 51.0% and acidic/basic amino acids at 67.7% of its composition. Aspartic acid (25.5%), glutamic acid (20.7%), and arginine (17.5%) were particularly abundant.
•Five monosaccharides—glucose (65.7%), fructose (16.6%), fucose (12.3%), galactose (3.3%), and arabinose (2.1%)—were identified in GEF.
•GEF demonstrated inhibitory effects on MMP-1 gene expression induced by UV irradiation in HS68 fibroblasts. At concentrations of 5 ppm and 50 ppm, MMP-1 mRNA expression was decreased by 18.3% and 28.4%, respectively, compared to the UV-treated negative control.
•GEF showed a restoration effect on fibrillin-1 protein loss caused by UV irradiation in HS68 fibroblasts. The amount of fibrillin-1 recovered by 37.2% at 5 ppm, and by 22.5% and 23.3% at 0.5 ppm and 50 ppm, respectively, compared to the UV-treated negative control.
•GEF facilitated COL1A1 gene expression in HS68 fibroblasts, with an increase of up to 25% observed at the optimal concentration of 5 ppm.
•GEF facilitated COL4A1 gene expression in HS68 fibroblasts, with an increase of up to 35% observed at the optimal concentration of 5 ppm.
•GEF significantly facilitated HAS2 gene expression in HaCaT keratinocytes. Expression increased 1.2-fold at 0.5 ppm and 3.1-fold at 5 ppm compared to the untreated control.
•GEF facilitated adipogenesis in ADMSCs, leading to lipid accumulation. An increase in lipids of about 10–15% was observed at tested concentrations, considered significant compared to previous studies with commercial peptide mixtures and a positive control.
This study successfully extracted a glycopeptide- and glycoprotein-enriched fraction (GEF) from a cosmetic variation of the traditional longevity medicine, Gongjin-dan, by reinterpreting the formulation to exclude animal-derived ingredients and modifying the preparation method to obtain high-molecular-weight components. The novelty of this research lies in scientifically demonstrating the skin anti-aging properties of this specific enriched fraction derived from a modified traditional herbal formula, highlighting its unique chemical characteristics including the abundance of glycopeptides/glycoproteins and high proportions of skin-essential amino acids like CEAAs.
The in vitro results provide scientific evidence supporting the potential of GEF as a raw material for skin anti-aging cosmetics. The findings indicate a dual mechanism of action: inhibiting the breakdown of skin components (like MMP-1 expression and fibrillin-1 loss) and facilitating the synthesis of crucial elements (like COL1A1, COL4A1, HAS2, and subcutaneous fat).
The future implications of this research are significant for the cosmetic industry, suggesting GEF as a promising plant-derived ingredient for anti-aging formulations. Further research is recommended to explore the effects of purified glycopeptides or glycoproteins from this fraction and to gain a deeper understanding of their specific mechanisms of action in skin cells. This study serves as a valuable example of scientifically validating traditional medicine for modern cosmetic applications.
Link to the study: https://www.mdpi.com/2079-9284/12/3/91
