Photoprotection and Antiaging Activity of Extracts from Honeybush (Cyclopia sp.)-In Vitro Wound Healing and Inhibition of the Skin Extracellular Matrix Enzymes: Tyrosinase, Collagenase, Elastase and Hyaluronidase.

Cyclopia sp. (honeybush) is an African shrub known as a rich source of polyphenols. The biological effects of fermented honeybush extracts were investigated. The influence of honeybush extracts on extracellular matrix (ECM) enzymes responsible for the skin malfunction and aging process-collagenase, elastase, tyrosinase and hyaluronidase-was analysed. The research also included assessment of the in vitro photoprotection efficiency of honeybush extracts and their contribution to the wound healing process. Antioxidant properties of the prepared extracts were evaluated, and quantification of the main compounds in the extracts was achieved. The research showed that the analysed extracts had a significant ability to inhibit collagenase, tyrosinase and hyaluronidase and a weak influence on elastase activity. Tyrosinase was inhibited effectively by honeybush acetone (IC50 26.18 ± 1.45 µg/mL), ethanol (IC50 45.99 ± 0.76 µg/mL) and water (IC50 67.42 ± 1.75 µg/mL) extracts. Significant hyaluronidase inhibition was observed for ethanol, acetone and water extracts (IC50 were 10.99 ± 1.56, 13.21 ± 0.39 and 14.62 ± 0.21µg/mL, respectively). Collagenase activity was inhibited effectively by honeybush acetone extract (IC50 42.5 ± 1.05 µg/mL). The wound healing properties of the honeybush extracts, estimated in vitro in human keratinocytes (HaCaTs), were indicated for water and ethanol extracts. In vitro sun protection factor (SPF in vitro) showed medium photoprotection potential for all the honeybush extracts. The quantity of polyphenolic compounds was estimated with the use of high-performance liquid chromatography equipped with diode-array detection (HPLC-DAD), indicating the highest mangiferin contents in ethanol, acetone and n-butanol extracts, while in the water extract hesperidin was the dominant compound. The antioxidant properties of the honeybush extracts were estimated with FRAP (2,4,6-Tris(2-pyridyl)-s-triazine) and DPPH (2,2-diphenyl-1-picrylhydrazyl) tests, indicating their strong antioxidant activity, similar to ascorbic acid for the acetone extract in both tests. The wound healing abilities, estimation of SPF in vitro and the direct influence on selected enzymes (elastase, tyrosinase, collagenase and hyaluronidase) of the tested honeybush extracts were analysed for the first time, indicating a high potential of these well-known herbal tea for antiaging, anti-inflammation, regeneration and protection of the skin.

Mangiferin Affects Melanin Synthesis by an Influence on Tyrosinase: Inhibition, Mechanism of Action and Molecular Docking Studies.

Mangiferin is a strong antioxidant that presents a wide range of biological activities. The aim of this study was to evaluate, for the first time, the influence of mangiferin on tyrosinase, an enzyme responsible for melanin synthesis and the unwanted browning process of food. The research included both the kinetics and molecular interactions between tyrosinase and mangiferin. The research proved that mangiferin inhibits tyrosinase activity in a dose-dependent manner with IC50 290 +/- 6.04 µM, which was found comparable with the standard kojic acid (IC50 217.45 +/- 2.54 µM). The mechanism of inhibition was described as mixed inhibition. The interaction between tyrosinase enzyme and mangiferin was confirmed with capillary electrophoresis (CE). The analysis indicated the formation of two main, and four less significant complexes. These results have also been supported by the molecular docking studies. It was indicated that mangiferin binds to tyrosinase, similarly to L-DOPA molecule, both in the active center and peripheral site. As it was presented in molecular docking studies, mangiferin and L-DOPA molecules can interact in a similar way with surrounding amino acid residues of tyrosinase. Additionally, hydroxyl groups of mangiferin may interact with amino acids on the tyrosinase external surface causing non-specific interaction.

Mangiferin and Hesperidin Transdermal Distribution and Permeability through the Skin from Solutions and Honeybush Extracts (Cyclopia sp.)-A Comparison Ex Vivo Study.

Polyphenolic compounds-mangiferin and hesperidin-are, among others, the most important secondary metabolites of African shrub Cyclopia sp. (honeybush). The aim of this study was to compare the percutaneous absorption of mangiferin and hesperidin from solutions (water, ethanol 50%, (v/v)) and extracts obtained from green and fermented honeybush (water, ethanol 50%, (v/v)). Research was performed with the Bronaugh cells, on human dorsal skin. The mangiferin and hesperidin distributions in skin layers (stratum corneum, epidermis, and dermis) and in acceptor fluid (in every 2, 4, 6, and 24 h) were evaluated by HPLC-Photodiode Array Coulometric and Coulometric Electrochemical Array Detection. The transdermal distribution of hesperidin was also demonstrated by fluorescence microscopy. Results indicated that mangiferin and hesperidin were able to cross the stratum corneum and penetrate into the epidermis and dermis. An advantage of hesperidin penetration into the skin from the water over ethanol solution was observed (451.02 ± 14.50 vs. 357.39 ± 4.51 ng/cm2), as well as in the mangiferin study (127.56 ± 9.49 vs. 97.23 ± 2.92 ng/cm2). Furthermore, mangiferin penetration was more evident from nonfermented honeybush ethanol extract (189.85 ± 4.11 ng/cm2) than from solutions. The permeation of mangiferin and hesperidin through the skin to the acceptor fluid was observed regardless of whether the solution or the honeybush extract was applied. The highest ability to permeate the skin was demonstrated for the water solution of hesperidin (250.92 ± 16.01 ng/cm2), while the hesperidin occurring in the extracts permeated in a very low capacity. Mangiferin from nonfermented honeybush ethanol extract had the highest ability to permeate to the acceptor fluid within 24 h (152.36 ± 8.57 ng/cm2).

Comparison of the in vitro cytotoxicity among phospholipid-based parenteral drug delivery systems: Emulsions, liposomes and aqueous lecithin dispersions (WLDs).

Lecithin and isolated phospholipids (mainly phosphatidylcholine) have been used for years as pharmaceutical excipients in parenteral formulations: submicron emulsions, liposomes and mixed micelles. Under development are also other lecithin-based drug delivery systems, e.g. aqueous lecithin dispersions (WLDs). The aim of the study was to investigate the properties and potential cytotoxicity of 7 different phospholipid-based dispersions intended for parenteral administration: emulsions, liposomes and WLDs. Each formulation contained egg phosphatidylcholine (PC) in the concentration range of 0.6-5.0%, and to some formulations other surfactants, such as polysorbate 80 (P80), Solutol HS 15 (HS) and cholesterol (Ch) were added. Particles in all dispersions were homogenous (PDI < 0.26) and submicron in size (Z-average in the range of approx. 100-260 nm). The cytotoxicity of all tested formulations was evaluated by means of 3 independent methods: a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, a real-time xCELLigence (RTCA) system, and a flow cytometry analysis, using two cell lines: human embryonic kidney 293 (HEK-293) and human promyelocytic leukaemia (HL-60). The results indicated that regardless of the test method and cell line type, the cytotoxicity of all formulations was low, especially when dispersions diluted to concentrations of =10% were tested. A more pronounced cytotoxic effect was noticed only for the following formulations: E-P80 (emulsion containing P80), WLD (unbuffered aqueous lecithin dispersion) and L-Ch (liposomes containing Ch), tested as less diluted (concentration 10% or 25%). IC50 values measured for these dispersions (on HL-60 cells) amounted to: 10.4 ±â€¯0.5% (v/v), 14.4 ±â€¯0.2% (v/v) and 24.2 ±â€¯0.6% (v/v), respectively. Our investigation confirmed the biocompatibility of all tested phospholipid-based formulations: emulsions, liposomes and also newly-developed WLDs, which can be considered as safe parenteral drug carriers.