Elastase Enzyme Research Articles

Chemical profiling and dermatological and anti-aging properties of Syzygium jambos L. (Alston): evidence from molecular docking, molecular dynamics, and in vitro experiments.

The phytoconstituents of the aqueous extract from Syzygium jambos L. (Alston) leaves were defined using HPLC-PDA-MS/MS and the antioxidant, anti-aging, antibacterial, and anti-biofilm activities of the extract were in silico and in vitro investigated. The antioxidant activities were performed using in vitro DPPH and FRAP assays as well as H2-DCFDA assay in HaCaT cells in which oxidative stress was induced by UVA radiation. Anti-aging activity was tested in vitro, using aging-related enzymes. The antibacterial, anti-biofilm and inhibitory effects on bacterial mobilities (swarming and swimming) were assessed against Pseudomonas aeruginosa. Results showed that S. jambos aqueous extract contained 28 phytochemicals belonging to different metabolite classes, mainly phenolic acids, gallic acid derivatives, flavonoids, and ellagitannins. Mineral content analysis showed that S. jambos leaves contained moderate amounts of nitrogen, potassium, manganese, magnesium, and zinc, relatively low amounts of phosphorus and copper, and high concentration of calcium and iron. The extract displayed strong antioxidant activities in vitro and inhibited UVA-induced oxidative stress in HaCaT cells. Docking the major compounds identified in the extract into the four main protein targets involved in skin aging revealed an appreciable inhibitory potential of these compounds against tyrosinase, elastase, hyaluronidase, and collagenase enzymes. Moreover, molecular dynamic simulations were adopted to confirm the binding affinity of some selected compounds towards the target enzymes. The extract exhibited pronounced in vitro anti-aging effects, compared to kojic acid and quercetin (the reference compounds). It also inhibited the growth of P. aeruginosa, counteracted its ability to form biofilm, and impeded its swarming and swimming mobilities. Altogether, these findings strongly propose S. jambos leaves as a promising source of bioactive metabolites for the development of natural cosmeceutical and dermatological agents.

IN SILICO STUDY OF ELASTASE ENZYME WITH NAPHTHOQUINONE DERIVATIVES AS LIGAND

We simulated 17 molecules classified as Naphthoquinones derivatives with the Enzyme Elastase to observe data regarding energies produced after bonding. These 17 Molecules were Eleutherin, Isoeleutherin, Elacanacin, Eleutherinone, Eleutherol A, Eleutherol B, Eleutherol C, Eleuthinones B, Eleuthinones C, Eleutherine A, Eleutherine B, Eleutherine C, Eleutherine D, Eleutherine E, Eleutherine F, Eleutherine G, Eleucanainones A. To prepare the ligand and protein for docking, we used the Discovery Studio application. For the molecular docking itself, we used the Pyrx application. Regarding interpreting the result, first, we chose the lowest rmsd/ub or rmsd/lb, and then we analyzed the energy result in which the lowest rmsd occurred. The docking results data indicated that all the ligand-enzyme bonding had negative binding affinity energy, but Eleucanainones A produced the lowest energy (Binding Affinity -7.7, mode 1, Rmsd/ub 1.787, Rmsd/lb 3.54), meaning it bound most easily with the enzyme Elastase. This study was only an initial or foundational step and further studies were highly needed for the development of the correlation between the ligands and the enzyme mentioned above.

Chemical annotation of the infusion of Jungia floribunda Less and its inhibitory potential on the elastase enzyme

Jungia floribunda Less. is a shrub belonging to the Asteraceae. The infusion of its leaves has been used, in folk medicine of several South American countries, as anti-inflammatory and hypoglycaemic agent. In the present study, the infusion of leaves from J. floribunda was obtained and its chemical composition was determined by UHPLC-MS associated with molecular network allowing the annotation of flavonoids, sesquiterpene lactones, coumarins, and chlorogenic acid derivatives. Besides, in vitro elastase activity assay was carried out with the infusion. As observed, elastase was inhibited at concentrations ranging from 15 to 240 µg/mL, reaching to 71% of inhibition at the maximum of evaluated concentration. Given that species of plants are promising sources for the discovery of new drugs, these results corroborate the infusion of J. floribunda as a potential source of bioactive compounds for the discovery of new inhibitors for elastase, besides its ethnopharmacological aspects.

Anti-Aging Properties of Temulawak Extract (Curcuma xanthorrhiza L.) by In Vitro Assay

Highlights:1. Elastase, hyaluronidase, and tyrosinase can all be inhibited by temulawak extract.2. Temulawak extract has strong anti-aging ability and can be used as a candidate for cosmetic applications. AbstractThe process of aging is the gradual loss of a tissue's capacity to heal and maintain normal or physiological form and function. Elastase, hyaluronidase, and tyrosinase are the enzymes that contribute to skin aging. The anti-aging effect is connected to the inhibition of these enzymes' activity. Many medicinal plants with active metabolite have been utilized extensively to treat aging. Active compounds of temulawak (Curcuma xanthorrhiza L.) such as phenolic, curcuminoid and xanthorrhizol have potential as antiaging. This study aims to determine whether temulawak extract (TE), a potential candidate for cosmeceutical agent, has anti-aging characteristics by inhibiting the elastase, hyaluronidase, and tyrosinase enzymes. TE was processedxby PT FAST. Stock solution of TE was made by dissolving 20 mg of extract with 1 ml of DMSO 100%, the stock solution was then diluted to achieve the concentrations level 31.25 - 1000 μg/ml working solution of TE. The antiaging activity assays were carried out at PT Aretha Medika Utama. In vitro antiaging assay of TE includes elastase, hyaluronidase, and tyrosinase enzyme inhibition at 7 level of various concentrations of 2.08-66.67 μg/ml for anti-elastase assay, 5.21-166.67 μg/ml for anti-hyaluronidase assay, and 3.125-100 μg/ml for anti-tyrosinase assay with three replications. SPSS software was used to conduct the statistical analysis (p<0,05 is considered significant). TE exhibited highest inhibition percentage of 82.72%, 89.41%, and 94.17% for anti-tyrosinase, anti-elastase, and anti-hyaluronidase, respectively. The median inhibitory concentration (IC50) for elastase inhibitory activity was 10.66 μg/ml, hyaluronidase 70.39 μg/ml, and tyrosinase 55.87 μg/ml. According to this study, TE can be used as cosmeceu

Advances in Sardinian Withania somnifera (L.) Dunal crops through phytochemical and biological approaches

Withania somnifera (L.) Dunal is widely used in the Indian traditional system of medicine to promote general health, wellness, and longevity. Its pharmacological properties are attributed to a group of molecules called withanolides, among which Withaferin A holds great interest for its anti-carcinogenic action. For this reason, numerous studies in recent years have focused on different metabolic or genetic engineering solutions to increase its yield. Here, we present the Sardinian chemotype of Withania somnifera as a potential crop for the extraction of Withaferin A. W. somnifera was cultivated from Sardinian wild germplasm collected in the northeast of the island. After 18 months, the leaves and the roots were collected and their methanolic extract was analyzed by HPLC. 0.3 mg/g DW of Withanolide A (WA), 1.0 mg/g DW of Withanolide B (WB) and 17.7 mg/g DW of Withaferin A (WF) were detected in the leaf sample, while lower values were detected in the roots (0.1 mg/g WF, 0.3 WA mg/g, 0.1 mg/g WB, 0.2 mg/g WO). This research not only confirms the high Withaferin A content found in the wild population leaves, but shows how they are reproducible in cultivated specimens, highlighting Sardinian W. somnifera leaves as a potential source of high-content Withaferin A products. Finally, we focused on the leaves extract by characterizing the phenolic and flavonoid content, as well as the in-vitro antioxidant capacity by DPPH and ABTS assays, revealing a significant amount of phenolic compounds and a related free radical scavenging activity. The leaves extract was further characterized for its anti-aging properties for potential cosmetic application, by the inhibition of tyrosinase, elastase, and collagenase enzymes.

Synthesis of enamine derivatives bearing of 2H-pyran-2,4(3H)-dione moiety from dehydroacetic acid: A promising frontier in skin protection and anti-aging, in-vitro, and in-silico insights

In aging research, the most crucial objective is to find longevity-enhancing drugs and analyze their subsequent mode of action. Search for longevity-enhancing drugs and their targets is, however, very challenging. The current study aimed to synthesize novel enamine-based derivatives as anti-aging products. The structures of these derivatives were characterized using multinuclear nmR (1H, 13C, and DEPT-135), mass spectrometry, single crystal XRD, and elemental analysis. Almost all the derivatives 3a-g showed good to excellent activities against elastase enzyme more than the reference drug. The IC50 value of compounds 3e and 3f (0.8 ± 0.2 µM and 0.7 ± 0.4 µM resp.) make them the most potent derivatives against elastase enzyme. Compound 3f exhibited prominent free radical scavenging activity (95.7%) and showed a zone of inhibition (24 mM/100 µL) against candida albicans. The docking studies showed that derivative 3b has the lowest binding affinity (-8.8 kcal/mol) and the best fitting orientation in the active pocket of the elastase enzyme. We infer from the biological activity results that some derivatives can be skin-protecting agents.

Design and development of a hepatic lyo-dECM powder as a biomimetic component for 3D-printable hybrid hydrogels

Bioprinting offers new opportunities to obtain reliable 3D in vitro models of the liver for testing new drugs and studying pathophysiological mechanisms, thanks to its main feature in controlling the spatial deposition of cell-laden hydrogels. In this context, decellularized extracellular matrix (dECM)-based hydrogels have caught more and more attention over the last years because of their characteristic to closely mimic the tissue-specific microenvironment from a biological point of view. In this work, we describe a new concept of designing dECM-based hydrogels; in particular, we set up an alternative and more practical protocol to develop a hepatic lyophilized dECM (lyo-dECM) powder as an ‘off-the-shelf’ and free soluble product to be incorporated as a biomimetic component in the design of 3D-printable hybrid hydrogels. To this aim, the powder was first characterized in terms of cytocompatibility on human and porcine mesenchymal stem cells (MSCs), and the optimal powder concentration (i.e. 3.75 mg ml−1) to use in the hydrogel formulation was identified. Moreover, its non-immunogenicity and capacity to reactivate the elastase enzyme potency was proved. Afterward, as a proof-of-concept, the powder was added to a sodium alginate/gelatin blend, and the so-defined multi-component hydrogel was studied from a rheological point of view, demonstrating that adding the lyo-dECM powder at the selected concentration did not alter the viscoelastic properties of the original material. Then, a printing assessment was performed with the support of computational simulations, which were useful to define a priori the hydrogel printing parameters as window of printability and its post-printing mechanical collapse. Finally, the proposed multi-component hydrogel was bioprinted with cells inside, and its post-printing cell viability for up to 7 d was successfully demonstrated.

Comparison of Biological Activities and Protective Effects on PAH-Induced Oxidative Damage of Different Coffee Cherry Pulp Extracts.

Polycyclic aromatic hydrocarbons (PAHs) are the main toxic components of ambient air particulate matter (PM), causing oxidative damage to the skin and ultimately resulting in skin aging. This study was conducted to determine the anti-oxidant, anti-aging properties and protective effects of the extracts of coffee cherry pulp (Coffea arabica L.), which is a by-product of the coffee industry, against the oxidative damage induced by PAH exposure in human epidermal keratinocytes (HaCaT). Three different techniques were used to extract the coffee cherry pulp: maceration, Soxhlet and ultrasonication to obtain CCM, CCS and CCU extract, respectively, which were then compared to investigate the total phenolic content (TPC) and total flavonoid content (TFC). The chemical compositions were identified and quantified using high-performance liquid chromatography (HPLC). The results demonstrated that Soxhlet could extract the highest content of chlorogenic acid, caffeine and theophylline. CCS showed the significantly highest TPC (324.6 ± 1.2 mg GAE/g extract), TFC (296.8 ± 1.2 mg QE/g extract), anti-radical activity against DPPH free radicals (98.2 ± 0.8 µM Trolox/g extract) and lipid peroxidation inhibition (136.6 ± 6.2 µM Trolox/g extract). CCS also showed the strongest anti-aging effects based on collagenase, elastase, hyaluronidase and tyrosinase inhibitory enzymes. In addition, CCS can protect human keratinocyte cells from PAH toxicity by increasing the cellular anti-oxidant capacity. This study suggests that CCS has the potential to be used as a cosmetic material that helps alleviate skin damage caused by air pollution.

The Wound-Healing Potential of the Endemic Plant Helianthemum canum (L.) Baumg: Preclinical Studies Supported with Phytochemical Profiling.

The study's objective is to clarify the probable mechanisms underlying the wound-healing properties of Helianthemum canum L. (Cistaceae), a traditional anti-inflammatory and wound-healing medicine. LC/MS-MS was used to perform phytochemical analyses on a 70 % methanol extract of the plant's aerial parts. In vivo, linear incision and circular excision models were used to evaluate the wound healing activity. For anti-inflammatory effect, in vivo acetic acid capillary permeability assay and in vitro Interleukin 1, Interleukin 6, and Interferon ɣ levels in LPS-induced FR skin fibroblast cell line were also evaluated. The extract significantly improved wound healing in experimental models, with tensile strength values of 27.8 % and a contraction value of 35.09 %. Histopathological examinations, hydroxyproline estimation, hyaluronidase, collagenase, and elastase enzyme inhibitory assays confirmed wound healing potential. Inflammatory cytokines were significantly inhibited in the LPS-induced FR cell line, with the highest effect seen on IL-6 (34.5±2.12 pg/mL). This study offered the first concrete proof that H. canum can be used to treat wounds by suggesting that the myricetin and quinic acid content identified by LCMS-MS analysis may be accountable for the effect of H. canum on wound contraction and hydroxyproline production.

Eucalyptus-derived essential oils alleviate microbes and modulate inflammation by suppressing superoxide and elastase release

The Eucalyptus tree, belonging to the myrtle family, grows all over the world for its pharmaceutical and industrial benefits. In this article, we present a comparative analysis of the chemical composition of the hydrodistilled oils obtained from three different Eucalyptus species growing in Egypt viz. E. citriodora, E. camaldulensis, and E. ficifolia. Gas Chromatography-Mass Spectrometric guided analysis resulted in the identification of a total of 20 metabolites in E. citriodora oil with citronellal (54.9%) and citronellol (25.4%) being the most dominant components. β-cymene (12.7%) and 1,8-cineole (11.7%) were the major volatile constituents identified in E. camaldulensis oil, while trans-β-ocimene (22.4%), 1,8-cineole (13.5%), and L-trans-pinocarveol (12.5%) were the dominating components in the oil of E. ficifolia. The essential oils of the studied species were evaluated for their in vitro anti-inflammatory, antiviral including anti-SARS-CoV-2 (severe acute respiratory syndrome corona virus 2), antibacterial, and antifungal activities. E. citriodora oil displayed the highest inhibitory activity on the release of the superoxide radical (32%) and elastase enzyme (31%) in human neutrophils, while E. ficifolia oil had enhancing effects on elastase. The latter showed significant antiviral effects against hepatitis A, herpes simplex, and coxsackie viruses with IC50 values at 2.1, 2.5, and 5.6 μg/mL, respectively. Moderate antibacterial and antifungal activities were observed for Eucalyptus oils with Staphylococcus aureus being the most susceptible bacterial strain. E. ficifolia oil, similarly, displayed the best antibacterial activity with minimum inhibitory concentration (MIC) value at ca. 25 μg/mL (for S. aureus). On the contrary, E. camaldulensis oil was the most active against Candida albicans with an MIC value at 45 μg/mL. In silico studies were performed with a number of macromolecular drug targets for confirming the biological activities of the identified compounds and for interpreting their ADME (absorption-distribution-metabolism-elimination) parameters.

Proteolitic Activity of Marine Strain Bacillus sp. 051

The main interest in the study of marine microorganisms is due to their ability to produce a wide range of unique enzymes, including peptidases with different specificities. In recent years, interest has increased in peptidases that are able to cleave elastin as a specific substrate. Streptomyces fradiae and Bacillus thermoproteolyticus elastases are among the most potent elastolytic proteinases discovered to date because they are 4-8-fold more effective than pancreatic elastases. The disadvantages of these producers include the fact that most of them are pathogenic for humans, and the elastase enzyme secreted from them is directly involved in the initiation of the pathogenetic process. All this significantly limits the scope of their practical application. Therefore, the search for new, more effective, safe for humans’ producers continues to be an urgent question, taking into account the fact that there are no highly active elastase producers in Ukraine. Previously we found elastase activity in only 4 of the 10 studied isolates of bacteria from the Black Sea. Since among them, the elastase activity of the Bacillus sp. 051 was the highest, the aim of this work was to study the physicochemical properties and substrate specificity of the enzyme. Methods. We used methods of determining proteolytic (caseinolytic, elastolytic, fibrinolytic, fibrinogenolytic) activity. Protein concentration was determined by the Lowry method. The study of the effect of temperature on the enzymatic activity was carried out in the range from 4 to 70 °C and pH values from 2.0 to 12.0, created using 0.01 M phosphate-citrate buffer. Results. It has been shown that the growing temperature of 12°C is the most optimal for biosynthesis of enzyme by the culture of Bacillus sp. 051. The complex enzyme preparation capable of hydrolyzing elastin, casein and fibrinogen. The enzyme showed maximum activity in relation to elastin (3.65 U/mg). The optimum pH of the enzyme action is 8.0, the thermal optimum is 40°C. The rate of casein hydrolysis compared to elastin was 2.7 times lower and amounted to 1.35 U/mg. The complex enzyme preparation also hydrolyzed fibrinogen (1.16 U/mg). Conclusions. According to its physicochemical and catalytic properties, the representative of the Black Sea, Bacillus sp. 051 is promising for further research as an enzyme producer with elastolytic activity.

Evaluation of Physiological Activity of Traditional Welsh Onion (Allium fistulosum L.)

Purpose: This study was conducted to evaluate physiological activities, such as antioxidant activity and enzyme activity, of hot water extract of the four parts (flower, leaf and pseudostem, stem, and root) of traditional Welsh onion (Allium fistulosum L.) and then to provide basic data on traditional Welsh onion to support future research.Methods: Four parts of traditional Welsh onion plant were extracted with water heated at 80℃. Antioxidative and enzymatic activities, such as total polyphenol content, total flavonoid content, 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, 2,2'-azino-bis-3-ethylbenzothiazoline–6-sulfonic acid (ABTS) radical scavenging activity, and the inhibitory activity on α-glucosidase, tyrosinase, and elastase enzymes, were measured.Results: Out of the hot water extract of the different parts, Allium fistulosum flower (AFF) extract had the highest content of antioxidants. In the Allium fistulosum leaf and pseudostem (AFLP), Allium fistulosum stem (AFS), and Allium fistulosum root (AFR) extracts, the total polyphenol content and DPPH radical scavenging activity of the root were high, and AFS had high total flavonoid content and ABTS radical scavenging activity. Regarding enzyme activity, α-glucosidase and elastase inhibitory activities were the highest in AFR, and tyrosinase inhibitory activity was the highest in AFS.Conclusion: We confirmed the physiological activity of the hot water extract of four parts of traditional Welsh onion. Our findings provide basic data for research into traditional Welsh onion, and can facilitate its future development as a functional material.

Nutritional Composition, Capsaicin Content and Enzyme Inhibitory Activities from “Bang Chang” Thai Cultivar Chili Pepper (Capsicum annuum Var. acuminatum) after Drying Process

Chili pepper cultivars come in a broad variety of tastes, colors, sizes, and spiciness levels due to various growth conditions. The significance of different chili peppers and other spices in Thai cuisine may be seen in dishes. This study evaluated the nutritional composition and capsaicin content from “Bang Chang” Thai cultivar chili pepper after drying, and the inhibition of lipase, tyrosinase and elastase enzymes from its oil and ethanol extracts. The nutritional composition of sun-dried chili pepper was high dietary fiber and beta-carotene, which were significantly high amount portion of Thai recommended dietary intake (RDI). According to calculations and definitions, “Bang Chang chili pepper” is a non-pungent Capsicum (0-700 SHU) since it contains a low amount of capsaicin. Total phenolic content was more abundant in ethanol extract (2.50 ± 0.13 mg GAE/g) than oil extract (1.05 ± 0.05 mg GAE/g). Compared to the positive control orlistat (IC50 = 3.26 ± 0.28 mg/mL), an anti-lipase drug, ethanol extract was 2.0 times more lipase inhibitory. In addition, ethanol extract was mildly anti-tyrosinase, while oil extract was non-activity. The anti-lipase activity of ethanol extract was due to phenolic content rather than capsaicin contained and this enzyme inhibition may act on active site. We found that ethanol extract of “Bang Chang” cultivar chili pepper yielded higher TPC content and more effective anti-lipase than oil extract. The finding provided benefits on applying this chili pepper on anti-lipase use and weight management.

Identification of polyphenols using UPLC-QTOF MS/MS, in-vitro photoprotective and antiaging activities of brown macroalga Padina tetrastromatica

Brown algae produce several bioactive compounds including unique secondary metabolites like polyphenols, which are known for various pharmacological activities. The present study focused on the cosmeceutical properties of polyphenol-rich fraction from brown macroalga (Padina tetrastromatica). Two fractions (ethyl acetate and aqueous) were obtained by liquid-liquid purification. Total phenolic and phlorotannin contents were assessed by in-vitro methods (Folin-Ciocalteu test and 2,4-dimethoxybenzaldehyde assay). Antioxidant properties were assessed through 2,2-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity, nitric oxide radical scavenging activity, and ferric-reducing antioxidant power assay. Ethyl acetate fraction showed high radical scavenging activity and was used for in-vitro photoprotective assays, cell viability tests, and anti-aging related enzyme inhibition tests (anti-tyrosinase, anti-elastase, anti-collagenase). Ethyl acetate fraction showed potential ultraviolet A absorption and a strong sun protection factor (SPF) value and was not toxic to the human skin cell line up to a concentration of 100 μg mL−1. Interestingly, this fraction also showed a strong tyrosinase inhibition and a significant inhibition of elastase and collagenase enzymes. Major polyphenols including phlorotannins present in the fraction were identified using Fourier transform infrared (FTIR) spectroscopy, Ultra pressure liquid chromatography (UPLC)-quadrupole-time of flight (QTOF) mass spectrometry (MS) and NMR spectroscopy. The findings suggest the ethyl acetate fraction of P. tetrastromatica as a promising source for cosmetic use with photoprotection and anti-aging properties.

Synthesis, DFT and molecular docking of novel (Z)-4-bromo-N-(4-butyl-3 (quinolin-3-yl)thiazol-2(3H)-ylidene)benzamide as elastase inhibitor

A new compound, C23H20BrN3OS, containing a quinoline-based iminothiazoline with a thiazoline ring, was synthesized and its crystal and molecular structures were analyzed through single crystal X-ray analysis. The compound belongs to the triclinic system P − 1 space group, with dimensions of a = 9.2304 (6) Å, b = 11.1780 (8) Å, c = 11.3006 (6) Å, α = 107.146 (5)°, β = 93.701 (5)°, γ = 110.435 (6)°, Z = 2 and V = 1025.61 (12) Å3. The crystal structure showed that C–H···N and C–H···O hydrogen bond linkages, forming infinite double chains along the b-axis direction, and enclosing R22(14) and R22(16) ring motifs. The Hirshfeld surface analysis revealed that H…H (44.1%) and H…C/C…H (15.3%) interactions made the most significant contribution. The newly synthesized (Z)-4-bromo-N-(4-butyl-3 (quinolin-3-yl)thiazol-2(3H)-ylidene)benzamide, in comparison to oleanolic acid, exhibited more strong potential against elastase with an inhibition value of 1.21 µM. Additionally, the derivative was evaluated using molecular docking and molecular dynamics simulation studies, which showed that the quinoline based iminothiazoline derivative has the potential to be a novel inhibitor of elastase enzyme. Both theoretical and experimental findings suggested that this compound could have a number of biological activities.

Ferulic acid-loaded aspasomes: A new approach to enhance the skin permeation, anti-aging and antioxidant effects

In this study, an innovative approach to enhance the skin permeation, anti-aging and antioxidant effects of ferulic acid (FA)-loaded aspasomal formulations were developed and evaluated in vitro and ex vivo. The chemical interactions between FA and excipients in the formulation were characterized by Differential scanning calorimetry (DSC) and Fourier-transformed infrared (FTIR). Aspasomes (ASPs) were produced using the film hydration method with different components and were evaluated according to in vitro characterization parameters (particle size (PS), polydispersity index (PDI), zeta potential (ZP), scanning electron microscopy (SEM), drug entrapment efficiency (EE%) and drug release profile), and the optimized formulation was determined. The 3-(4,5-dimethyldiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to test the cytotoxicity of the ASPs and different FA solutions on the L929 cell line. In the stability studies, changes in characterization parameters and FA content of optimum ASPs stored in both suspended and lyophilized dry powder form in two different conditions over 3 months were monitored. Ex-vivo skin permeation study was done using mouse abdominal skin and the amount of penetrated FA was measured. Also, skin depth and density of blank and FA-loaded ASPs were evaluated by fluorescence microscopy. At the end of the permeation study, the antioxidant capacity of FA reaching the dermis layer and its inhibitory effect on collagenase and elastase enzymes were assessed. FTIR spectra and DSC thermograms assured chemical and physical compatibility. The selected optimum formulation (F2 coded) showed PS of 384 ± 1.52 nm, PDI of 0.015 ± 0.011, ZP of −39.2 ± 3.45 mV, EE of 58 ± 2.01% and drug release of 77.3 ± 0.554% and was used for permeation study. The lyophilized ASPs were physically stable, and the FA content of ASPs did not decrease significantly during 3 months of storage at 4 °C. The skin permeation experimental results indicated positive results that successfully developed FA-loaded ASPs have an effective FA permeability rate and better antioxidant and anti-aging effects than the free drug and are useful and effective for improving skin permeation and slowing skin aging with topical applications.

Phytocompounds and in vitro antiaging activity of ethanolic extract and fractions of Rubus fraxinifolius Poir. leaves

Context: The leading cause of skin sunburn is exposure to ultraviolet radiation (UVR), which can also cause reactive oxygen species (ROS), which accelerates aging. Rubus fraxinifolius Poir. leaf ethanolic extract may have antiaging properties by preventing elastase enzyme activity. Aims: To compare the antioxidant activity and elastase enzyme inhibition from old and young leaves using Soxhlet extraction and evaluate the antioxidant activity and elastase enzyme inhibition utilizing a variety of extraction techniques, including conventional Soxhlet extraction and unconventional methods like microwave-assisted extraction (MAE) and ultrasonic-assisted extraction (UAE). Methods: Using various extraction methods, samples were extracted with 96% ethanol (Soxhlet, MAE, and UAE). The active extract was then transformed into n-hexane, ethyl acetate, and distilled water fraction. Following the total phenolic and total flavonoid content analysis, the antioxidant activity was evaluated using the radical scavenging tests of 1,2-diphenyl-2-picrylhydrazyl (DPPH), ferric reducing antioxidant power assay (FRAP), and ABTS radicals, or 2,2'-azino-bisphenol S (2,2'-ABPS) (3-ethylbenzothiazoline-6-sulphonic acid). The antiaging potential of skin breakdown enzymes was examined using elastase. Results: According to the R. fraxinifolius leaf's optimization type, extract from older leaves performs better than from younger leaves in terms of antioxidant activity and elastase enzyme inhibitory activity. In terms of optimization extraction, UAE approaches to produce the best outcomes than Soxhlet and MAE. Additionally, using the chosen UAE extraction method, multilayer fractionation was done using n-hexane, ethyl acetate, and distilled water solvents, with distilled water fraction producing the best results. Conclusions: The distilled water fraction from R. fraxinifolius leaf’s shows the potential for future research to support its efficacy as an active component in antiaging cosmetics.

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.

Exploring the Therapeutic Potential of Elastase Inhibition in Age-Related Macular Degeneration in Mouse and Human.

Abnormal turnover of the extracellular matrix (ECM) protein elastin has been linked to AMD pathology. Elastin is a critical component of Bruch's membrane (BrM), an ECM layer that separates the retinal pigment epithelium (RPE) from the underlying choriocapillaris. Reduced integrity of BrM's elastin layer corresponds to areas of choroidal neovascularization (CNV) in wet AMD. Serum levels of elastin-derived peptides and anti-elastin antibodies are significantly elevated in AMD patients along with the prevalence of polymorphisms of genes regulating elastin turnover. Despite these results indicating significant associations between abnormal elastin turnover and AMD, very little is known about its exact role in AMD pathogenesis. Here we report on results that suggest that elastase enzymes could play a direct role in the pathogenesis of AMD. We found significantly increased elastase activity in the retinas and RPE cells of AMD mouse models, and AMD patient-iPSC-derived RPE cells. A1AT, a protease inhibitor that inactivates elastase, reduced CNV lesion sizes in mouse models. A1AT completely inhibited elastase-induced VEGFA expression and secretion, and restored RPE monolayer integrity in ARPE-19 monolayers. A1AT also mitigated RPE thickening, an early AMD phenotype, in HTRA1 overexpressing mice, HTRA1 being a serine protease with elastase activity. Finally, in an exploratory study, examining archival records from large patient data sets, we identified an association between A1AT use, age and AMD risk. Our results suggest that repurposing A1AT may have therapeutic potential in modifying the progression to AMD.

Carotenoids from female Grapsus albolineatus as potential anti-ageing compounds

Introduction: Grapsus albolineatus is thought to have a high concentration of carotenoid pigments. Although studies on male G. albolineatus have been conducted, no studies on pigment extraction from female G. albolineatus have been carried out. Carotenoids have a high ability to fight free radicals. Previous research has shown that carotenoids can fight free radicals that cause premature skin-aging. Aim: The purpose of this research was to find out what kinds of carotenoids are found in G. albolineatus and whether they can act as natural inhibitors of proteins that cause premature aging, such as glucogenase, elastase, and hyaluronidase enzymes. Methods: Carotenoids were extracted from G. albolineatus using column chromatography and high-performance liquid chromatography (HPLC); molecular docking and visualization were done with Autodock 4.2 and Discovery Studio/Biovia, respectively. Results: According to HPLC data, there are carotenoid pigments such as didehydroastaxanthin, tetrahydroastaxanthin, dihydroastaxanthin, diatoxanthin, astaxanthin, and adonixanthin. According to molecular docking experiments, pigment carotenoids from G. albolineatus are efficient inhibitors of protein elastase and hyaluronidase with binding energy range -7.58 kcal/mol - -9.03 kcal/mol and -6.16 kcal/mol - -7.71 kcal/mol, respectively. Conclusions: G. albolineatus carotenoids have the potential to be anti-aging since they are more effective as protein elastase and hyaluronidase inhibitors than their native inhibitors.