Efficacy of Vitamin B12 and Adenosine Triphosphate in Enhancing Skin Radiance: Unveiled with a Drug-Target Interaction Deep Learning-Based Model.

Skin radiance is crucial for enhancing facial attractiveness and is negatively affected by factors like hyperpigmentation and aging-related changes. Current treatments often lack comprehensive solutions for improving skin radiance. This study aimed to develop a cosmetic formula that enhances skin radiance by reducing hyperpigmentation and improving skin regeneration by targeting specific receptors-the endothelin receptor type B (EDNRB) for hyperpigmentation and the adiponectin receptor 1 (ADIPOR1) for sagging and wrinkles. To achieve this, we used artificial intelligence technologies to screen and select ingredients with an affinity for EDNRB and ADIPOR1. Vitamin B12 (VitB12) was identified as a molecule that targets EDNRB, which is involved in melanogenesis. Adenosine triphosphate (ATP) targets ADIPOR1, which is associated with skin regeneration. VitB12 successfully inhibited intracellular calcium elevation and melanogenesis induced by endothelin-1. In contrast, ATP increased the mRNA expression of collagen and elastin and promoted wound healing. Moreover, the VitB12 and ATP complex significantly increased the expression of hyaluronan synthases, which are crucial for skin hydration. Furthermore, in human participants, the application of the VitB12 and ATP complex to one-half of the face significantly improved skin radiance, elasticity, and texture. Our findings provide valuable insights for the development of skincare formulations.

Safety Assessment of Vinylpyrrolidone Polymers as Used in Cosmetics.

The Expert Panel for Cosmetic Ingredient Safety (Panel) reviewed the safety of 30 vinylpyrrolidone polymers as used in cosmetic products; most of these ingredients have the reported cosmetic function of film former in common. The Panel reviewed data relevant to the safety of these ingredients, and determined that 27 vinylpyrrolidone polymers are safe in cosmetics in the present practices of use and concentration described in the safety assessment. The Panel also concluded that the available data are insufficient to make a determination that 3 vinylpyrrolidone polymers (all urethanes) are safe under the intended conditions of use in cosmetic formulations.

Biologically Active Sheep Colostrum for Topical Treatment and Skin Care.

Colostrum is gaining popularity in cosmetic products. The present study compared the composition and selected biological properties of colostrum from Polish sheep (colostrum 1) and Swiss sheep (colostrum 2), particularly those that can affect healthy or diseased skin. The antioxidant activity of the colostrums was measured using ABTS and DPPH assays. The effect on the proliferation of human skin fibroblasts, neonatal epidermal keratinocytes, and human diabetic fibroblast (dHF) cells isolated from diabetic foot ulcers was also assayed in vitro by MTT and Presto Blue tests, respectively. The colostrum simulated dHF cell proliferation by up to 115.4%. The highest used concentration of colostrum 1 stimulated normal fibroblast proliferation by 191.2% (24 h) and 222.2% (48 h). Both colostrums inhibited epidermal keratinocyte viability. The influence of the colostrums on the expression of genes related to proliferation (Ki67) and immune response (IL-6, PTGS-2, TSG-6) in dHF cells were compared. Colostrum 1 increased the rate of wound closure (scar test). Analysis of total fat, protein and fatty acid content found the Polish colostrum to be a richer source of fat than the Swiss colostrum, which contained a larger amount of protein. Both colostrums exhibit properties that suggest they could be effective components in cosmetic or medicinal formulations for skin care, especially supporting its regeneration, rejuvenation, and wound healing.

Mass-Spectrometry-Based Research of Cosmetic Ingredients.

Cosmetic products are chemical substances or mixtures used on the skin, hair, nails, teeth, and the mucous membranes of the oral cavity, whose use is intended to clean, protect, correct body odor, perfume, keep in good condition, or change appearance. The analysis of cosmetic ingredients is often challenging because of their huge complexity and their adulteration. Among various analytical tools, mass spectrometry (MS) has been largely used for compound detection, ingredient screening, quality control, detection of product authenticity, and health risk evaluation. This work is focused on the MS applications in detecting and quantification of some common cosmetic ingredients, i.e., preservatives, dyes, heavy metals, allergens, and bioconjugates in various matrices (leave-on or rinse-off cosmetic products). As a global view, MS-based analysis of bioconjugates is a narrow field, and LC- and GC/GC×GC-MS are widely used for the investigation of preservatives, dyes, and fragrances, while inductively coupled plasma (ICP)-MS is ideal for comprehensive analysis of heavy metals. Ambient ionization approaches and advanced separation methods (i.e., convergence chromatography (UPC2)) coupled to MS have been proven to be an excellent choice for the analysis of scented allergens. At the same time, the current paper explores the challenges of MS-based analysis for cosmetic safety studies.

Workshop report: Challenges faced in developing inhalation thresholds of Toxicological Concern (TTC) - State of the science and next steps.

A challenging step in human risk assessment of chemicals is the derivation of safe thresholds. The Threshold of Toxicological Concern (TTC) concept is one option which can be used for the safety evaluation of substances with a limited toxicity dataset, but for which exposure is sufficiently low. The application of the TTC is generally accepted for orally or dermally exposed cosmetic ingredients; however, these values cannot directly be applied to the inhalation route because of differences in exposure route versus oral and dermal. Various approaches of an inhalation TTC concept have been developed over recent years to address this. A virtual workshop organized by Cosmetics Europe, held in November 2020, shared the current state of the science regarding the applicability of existing inhalation TTC approaches to cosmetic ingredients. Key discussion points included the need for an inhalation TTC for local respiratory tract effects in addition to a systemic inhalation TTC, dose metrics, database building and quality of studies, definition of the chemical space and applicability domain, and classification of chemicals with different potencies. The progress made to date in deriving inhalation TTCs was highlighted, as well as the next steps envisaged to develop them further for regulatory acceptance and use.

Artificial Intelligence That Predicts Sensitizing Potential of Cosmetic Ingredients with Accuracy Comparable to Animal and In Vitro Tests-How Does the Infotechnomics Compare to Other "Omics" in the Cosmetics Safety Assessment?

The aim of the current study was to develop an in silico model to predict the sensitizing potential of cosmetic ingredients based on their physicochemical characteristics and to compare the predictions with historical animal data and results from "omics"-based in vitro studies. An in silico model was developed with the use of WEKA machine learning software fed with physicochemical and structural descriptors of haptens and trained with data from published epidemiological studies compiled into estimated odds ratio (eOR) and estimated attributable risk (eAR) indices. The outcome classification was compared to the results of animal studies and in vitro tests. Of all the models tested, the best results were obtained for the Naive Bayes classifier trained with 24 physicochemical descriptors and eAR, which yielded an accuracy of 86%, sensitivity of 80%, and specificity of 90%. This model was subsequently used to predict the sensitizing potential of 15 emerging and less-studied haptens, of which 7 were classified as sensitizers: cyclamen aldehyde, N,N-dimethylacrylamide, dimethylthiocarbamyl benzothiazole sulphide, geraniol hydroperoxide, isobornyl acrylate, neral, and prenyl caffeate. The best-performing model (NaiveBayes eAR, 24 parameters), along with an alternative model based on eOR (Random Comittee eOR, 17 parameters), are available for further tests by interested readers. In conclusion, the proposed infotechnomics approach allows for a prediction of the sensitizing potential of cosmetic ingredients (and possibly also other haptens) with accuracy comparable to historical animal tests and in vitro tests used nowadays. In silico models consume little resources, are free of ethical concerns, and can provide results for multiple chemicals almost instantly; therefore, the proposed approach seems useful in the safety assessment of cosmetics.

Study of the protective effects of cosmetic ingredients on the skin barrier, based on the expression of barrier-related genes and cytokines.

BACKGROUND: Sensitive skin is the result of a complex process that is closely linked to the damage of the skin barrier. There are no recognized methods for evaluating the efficacy of anti-allergy products. METHODS: In this study, a model of skin barrier damage was created by treating HaCaT cells with 60 µg/ml of sodium dodecyl sulfate for 48 h. The protective effects of nine cosmetic ingredients, including oat extract (S1), on the skin barrier were investigated based on the gene expression levels of aquaporin3 (AQP3), filaggrin (FLG), caspase-14 (CASP14), and human tissue kallikrein7 (KLK7), as well as those of various interleukins (IL) and vascular endothelial growth factor (VEGF). RESULTS: Among the nine ingredients, S1 had a good protective effect on the function of the skin barrier. It promoted the expression of AQP3, FLG, and CASP14, while inhibiting the expression of KLK7 in HaCaT cells, at a concentration of 0.06%. It also maintained IL-6, IL-8, and VEGF at appropriate levels while promoting the proliferation and differentiation of HaCaT cells. CONCLUSIONS: The above indicators allow for the preliminary establishment of a method to evaluate the efficacy of the barrier protection ability of sensitive skin.

Natural Compounds and Products from an Anti-Aging Perspective.

Aging is a very complex process that is accompanied by a degenerative impairment in many of the major functions of the human body over time. This inevitable process is influenced by hereditary factors, lifestyle, and environmental influences such as xenobiotic pollution, infectious agents, UV radiation, diet-borne toxins, and so on. Many external and internal signs and symptoms are related with the aging process and senescence, including skin dryness and wrinkles, atherosclerosis, diabetes, neurodegenerative disorders, cancer, etc. Oxidative stress, a consequence of the imbalance between pro- and antioxidants, is one of the main provoking factors causing aging-related damages and concerns, due to the generation of highly reactive byproducts such as reactive oxygen and nitrogen species during the metabolism, which result in cellular damage and apoptosis. Antioxidants can prevent these processes and extend healthy longevity due to the ability to inhibit the formation of free radicals or interrupt their propagation, thereby lowering the level of oxidative stress. This review focuses on supporting the antioxidant system of the organism by balancing the diet through the consumption of the necessary amount of natural ingredients, including vitamins, minerals, polyunsaturated fatty acids (PUFA), essential amino acids, probiotics, plants' fibers, nutritional supplements, polyphenols, some phytoextracts, and drinking water.

Extraction, Encapsulation into Lipid Vesicular Systems, and Biological Activity of Rosa canina L. Bioactive Compounds for Dermocosmetic Use.

Valorization of wild plants to obtain botanical ingredients could be a strategy for sustainable production of cosmetics. This study aimed to select the rosehip extract containing the greatest amounts of bioactive compounds and to encapsulate it in vesicular systems capable of protecting their own antioxidant activity. Chemical analysis of Rosa canina L. extracts was performed by LC-DAD-MS/MS and 1H-NMR and vitamins, phenolic compounds, sugars, and organic acids were detected as the main compounds of the extracts. Liposomes, prepared by the film hydration method, together with hyalurosomes and ethosomes, obtained by the ethanol injection method, were characterized in terms of vesicle size, polydispersity index, entrapment efficiency, zeta potential, in vitro release and biocompatibility on WS1 fibroblasts. Among all types of vesicular systems, ethosomes proved to be the most promising nanocarriers showing nanometric size (196 ± 1 nm), narrow polydispersity (0.20 ± 0.02), good entrapment efficiency (92.30 ± 0.02%), and negative zeta potential (-37.36 ± 0.55 mV). Moreover, ethosomes showed good stability over time, a slow release of polyphenols compared with free extract, and they were not cytotoxic. In conclusion, ethosomes could be innovative carriers for the encapsulation of rosehip extract.

Comprehensive characterization of the structure and properties of human stratum corneum relating to barrier function and skin hydration: modulation by a moisturizer formulation.

The stratum corneum (SC) is key in the maintenance of the biomechanical barrier and hydration of skin. Our previous investigations showed beneficial effects of a combination of emollients on water capture and retention and protein and lipid organization, all of which are linked to dryness and dry skin damage. Here, we show how a formulation containing an emollient combination ("Trio") and its basal formulation (placebo) impacted the descriptors of SC hydration in SC layers. Only the Trio formulation-not its placebo formulation-modified SC biomechanical drying stress behaviour and imparted a high capacity to protect it from dehydration. This was in accordance with findings at the molecular level using Raman analyses and at the structural level using cryo-scanning electron microscopy (SEM). After topical application, only the Trio formulation profoundly increased lateral packing of lipids and their compactness. Cryo-SEM showed that, unlike the placebo formulation, the Trio formulation prevented the water loss when applied before the dehydration process. In conclusion, these studies demonstrate that stresses in the SC due to dehydration can be alleviated using a formulation containing emollients that interact with the SC lipid components.

Current EU regulatory requirements for the assessment of chemicals and cosmetic products: challenges and opportunities for introducing new approach methodologies.

The EU Directive 2010/63/EU   on the protection of animals used for scientific purposes and other EU regulations, such as REACH and the Cosmetic Products Regulation advocate for a change in the way toxicity testing is conducted. Whilst the Cosmetic Products Regulation bans animal testing altogether, REACH aims for a progressive shift from in vivo testing towards quantitative in vitro and computational approaches. Several endpoints can already be addressed using non-animal approaches including skin corrosion and irritation, serious eye damage and irritation, skin sensitisation, and mutagenicity and genotoxicity. However, for systemic effects such as acute toxicity, repeated dose toxicity and reproductive and developmental toxicity, evaluation of chemicals under REACH still heavily relies on animal tests. Here we summarise current EU regulatory requirements for the human health assessment of chemicals under REACH and the Cosmetic Products Regulation, considering the more critical endpoints and identifying the main challenges in introducing alternative methods into regulatory testing practice. This supports a recent initiative taken by the International Cooperation on Alternative Test Methods (ICATM) to summarise current regulatory requirements specific for the assessment of chemicals and cosmetic products for several human health-related endpoints, with the aim of comparing different jurisdictions and coordinating the promotion and ultimately the implementation of non-animal approaches worldwide. Recent initiatives undertaken at European level to promote the 3Rs and the use of alternative methods in current regulatory practice are also discussed.

Analytical method development and percutaneous absorption of propylidene phthalide, a cosmetic ingredient.

Propylidene phthalide (PP) is a cosmetic ingredient used in the fragrance industry and regulated for the limited content of 0.01% in cosmetic products in Korea. The aim of this study was to determine PP dermal absorption rate according to the Korea Ministry of Food and Drug Safety (MFDS) guidelines using in vitro Franz diffusion system. An analytical method in assessing PP was developed through method validation using LC-MS/MS. Linearity, precision, and accuracy were acceptable based upon MFDS guidelines. The stability of PP in receptor fluid (50% ethanol) at 32°C was sufficient up to 24 hr. Cream formulation (o/w) was topically applied to excised rat skin at a dose of 113 mg/cm2 containing 0.7% PP. The time points for receptor fluid collection were set at 0, 1, 2, 4, 8, 12, and 24 hr. After 24 hr, the remaining formulation on the skin and stratum corneum (SC) were collected through swabbing with an alcohol cotton and tape stripping, respectively. The collected samples (swabbed-remained formulation, SC, and skin) were extracted using acetonitrile for 24 hr. Total dermal absorption rate of PP was approximately 24% in cream formulation. These findings may be used for further exposure evaluation of PP in human consumers.

The Threshold of Toxicological Concern (TTC) is a pragmatic tool for the safety assessment: Case studies of cosmetic ingredients with low consumer exposure.

The Threshold of Toxicological Concern (TTC) is an internationally accepted pragmatic and conservative tool for the safety assessment of substances, which is used in a wide range of regulatory contexts. The TTC approach produces human exposure threshold values (TTC values) originally derived by Munro from oral toxicity data on cancer and non-cancer toxicity endpoints. This database has been recently substantially enlarged by the COSMOS database, an enhanced oral non-cancer TTC dataset on a larger chemical domain, thereby resulting in a new, transparent and public TTC database also including 552 cosmetics-related chemicals. The 5th percentile point of departure value for each Cramer Class was determined, from which human exposure TTC values have been derived. The combined COSMOS/Munro dataset provided TTC values of 46, 6.2 and 2.3 µg/kg bw/day for Cramer Classes I, II or III, respectively. In order to demonstrate the diverse scope and successful application of the TTC concept to cosmetic ingredients including hair dyes, fragrances and plant-derived ingredients, Cosmetics Europe has prepared several case studies. Overall, the TTC concept is not only useful to replace animal testing but can also successfully be applied to the safety evaluation of cosmetic ingredients in the marketed formulas with low human exposure.

The SCCS Notes of Guidance for the testing of cosmetic ingredients and their safety evaluation, 11th revision, 30-31 March 2021, SCCS/1628/21.

The "SCCS Notes of Guidance for the Testing of Cosmetic Ingredients and Their Safety Evaluation, 11 th Revision" (SCCS/1628/21) contains relevant and updated information on the different aspects of testing and safety evaluation of cosmetic substances in Europe. The emphasis is on cosmetic ingredients for which a concern has been expressed for human health. Indirectly, the Guidance also provides some advice on the safety of finished products. A general aim is to improve harmonised compliance with the current cosmetic EU legislation, Regulation (EC) No 1223/2009, for which animal testing and marketing bans fully apply from 2013 onwards. This means that no in vivo testing of ingredients or finished products is allowed in Europe for the purpose of cosmetics. For this reason, the SCCS has closely followed the progress made in regard to the development and validation of alternative replacement methods, also referred to as new approach methodology (NAM). The "SCCS Notes of Guidance" are regularly revised and updated in order to incorporate progress made and experience gained over time, in particular on the use of NAMs, and the new methods and data that became available since previous revision (SCCS/1602/18) formed the basis of the current (11 th) Revision.

Confocal Raman Spectroscopic Imaging for Evaluation of Distribution of Nano-Formulated Hydrophobic Active Cosmetic Ingredients in Hydrophilic Films.

Film-forming systems are highly relevant to the topical administration of active ingredients (AI) to the body. Enhanced contact with the skin can increase the efficacy of delivery and penetration during prolonged exposure. However, after the evaporation of volatile solvents to form a thin film, the distribution of the ingredient should remain homogenous in order to ensure the effectiveness of the formula. This is especially critical for the use of hydrophobic molecules that have poor solubility in hydrophilic films. In order to address this concern, hydroxyphenethyl esters (PHE) of Punica granatum seed oil were prepared as a nanosuspension stabilised by poloxamers (NanoPHE). NanoPHE was then added to a formulation containing polyvinyl alcohol (PVA) as a film forming agent, Glycerol as a plasticiser and an antimicrobial agent, SepicideTM HB. Despite their reliability, reference methods such as high-performance liquid chromatography are increasingly challenged due to the need for consumables and solvents, which is contrary to current concerns about green industry in the cosmetics field. Moreover, such methods fail to provide spatially resolved chemical information. In order to investigate the distribution of ingredients in the dried film, Confocal Raman imaging (CRI) coupled to Non-negatively Constrained Least Squares (NCLS) analysis was used. The reconstructed heat maps from a range of films containing systematically varying PHE concentrations highlighted the changes in spectral contribution from each of the ingredients. First, using NCLS scores it was demonstrated that the distributions of PVA, Glycerol, SepicideTM HB and PHE were homogenous, with respective relative standard deviations (RSD) of 3.33%, 2.48%, 2.72% and 6.27%. Second, the respective relationships between ingredient concentrations in the films and their Raman responses, and the spectral abundance were established. Finally, a model for absolute quantification for PHE was be constructed using the percentage of spectral abundance. The prepared %w/w concentrations regressed against predicted %w/w concentrations, displaying high correlation (R2 = 0.995), while the Root Mean Squared Error (0.0869% w/w PHE) confirmed the precision of the analysis. The mean percent relative error of 3.75% indicates the accuracy to which the concentration in dried films could be determined, further supporting the suitability of CRI for analysis of composite solid film matrix. Ultimately, it was demonstrated that nanoformulation of hydrophobic PHE provides homogenous distribution in PVA based film-forming systems independent of the concentration of NanoPHE used in the formula.

Measurement of the penetration of 56 cosmetic relevant chemicals into and through human skin using a standardized protocol.

OECD test guideline 428 compliant protocol using human skin was used to test the penetration of 56 cosmetic-relevant chemicals. The penetration of finite doses (10 µL/cm2 ) of chemicals was measured over 24 hours. The dermal delivery (DD) (amount in the epidermis, dermis and receptor fluid [RF]) ranged between 0.03 ± 0.02 and 72.61 ± 8.89 µg/cm2 . The DD of seven chemicals was comparable with in vivo values. The DD was mainly accounted for by the amount in the RF, although there were some exceptions, particularly of low DD chemicals. While there was some variability due to cell outliers and donor variation, the overall reproducibility was very good. As six chemicals had to be applied in 100% ethanol due to low aqueous solubility, we compared the penetration of four chemicals with similar physicochemical properties applied in ethanol and phosphate-buffered saline. Of these, the DD of hydrocortisone was the same in both solvents, while the DD of propylparaben, geraniol and benzophenone was lower in ethanol. Some chemicals displayed an infinite dose kinetic profile; whereas, the cumulative absorption of others into the RF reflected the finite dosing profile, possibly due to chemical volatility, total absorption, chemical precipitation through vehicle evaporation or protein binding (or a combination of these). These investigations provide a substantial and consistent set of skin penetration data that can help improve the understanding of skin penetration, as well as improve the prediction capacity of in silico skin penetration models.

Chitin and Chitosans: Characteristics, Eco-Friendly Processes, and Applications in Cosmetic Science.

Huge amounts of chitin and chitosans can be found in the biosphere as important constituents of the exoskeleton of many organisms and as waste by worldwide seafood companies. Presently, politicians, environmentalists, and industrialists encourage the use of these marine polysaccharides as a renewable source developed by alternative eco-friendly processes, especially in the production of regular cosmetics. The aim of this review is to outline the physicochemical and biological properties and the different bioextraction methods of chitin and chitosan sources, focusing on enzymatic deproteinization, bacteria fermentation, and enzymatic deacetylation methods. Thanks to their biodegradability, non-toxicity, biocompatibility, and bioactivity, the applications of these marine polymers are widely used in the contemporary manufacturing of biomedical and pharmaceutical products. In the end, advanced cosmetics based on chitin and chitosans are presented, analyzing different therapeutic aspects regarding skin, hair, nail, and oral care. The innovative formulations described can be considered excellent candidates for the prevention and treatment of several diseases associated with different body anatomical sectors.

Granulated cork as biosorbent for the removal of phenol derivatives and emerging contaminants.

This study evaluated the ability of cork to adsorb a broad range of phenolic, pharmaceutical and cosmetic compounds: phenol, 2-chlorophenol, 2-nitrophenol, 2,4-dichlorophenol, pentachlorophenol carbamazepine, naproxen, ketoprofen, diclofenac, triclosan, and methyl paraben. The effect of variables such as the compound concentrations and the amount of cork were studied resulting in a highly pH dependence in the case of phenolic compounds. Maximum removal percentages and uptake values of 75% (1.61 mg/g) for 2,4-dichlorophenol, 55% (1.25 mg/g) for 2-nitrophenol, 45% (1.47 mg/g) for 2-chlorophenol, 20% (0.63 mg/g) for phenol, and 100% for pentachlorophenol, were obtained for a 30 mg L-1 solution at pH 6, showing that the adsorption process increased with greater electronegativity of the phenolic substituting group. Removal percentages and uptakes of 82% (3.56 mg/g) for naproxen, 57% (2.31 mg/g) for ketoprofen, 50% (1.84 mg/g) for carbamazepine, 50% (1.78 mg/g) for methyl paraben, 100% for sodium diclofenac, and 100% for triclosan, were obtained using 5 mg of cork and a 1 mg L-1 solution of each compound. The adsorption process was almost complete after 30 min for all the micropollutants. Experimental equilibrium data were analysed by Freundlich and Langmuir adsorption models. Cork has proved to be an effective sorbent for the removal of phenols and emerging contaminants from contaminated waters and is a readily available material that can be acquired at minimal or no cost in cork-producing areas.

Comparison of the Skin Penetration of 3 Metabolically Stable Chemicals Using Fresh and Frozen Human Skin.

BACKGROUND: The Cosmetics Europe ADME Task Force is developing in vitro and in silico tools for predicting skin and systemic concentrations after topical application of cosmetic ingredients. There are conflicting reports as to whether the freezing process affects the penetration of chemicals; therefore, we evaluated whether the storage of human skin used in our studies (8-12 weeks at -20°C) affected the penetration of model chemicals. METHODS: Finite doses of trans-cinnamic acid (TCA), benzoic acid (BA), and 6-methylcoumarin (6MC) (non-volatile, non-protein reactive and metabolically stable in skin) were applied to fresh and thawed frozen skin from the same donors. The amounts of chemicals in different skin compartments were analysed after 24 h. RESULTS: Although there were some statistical differences in some parameters for 1 or 2 donors, the penetration of TCA, BA, and 6MC was essentially the same in fresh and frozen skin, i.e., there were no biologically relevant differences in penetration values. Statistical differences that were evident indicated that penetration was marginally lower in frozen than in fresh skin, indicating that the barrier function of the skin was not lost. CONCLUSION: The penetration of the 3 chemicals was essentially unaffected by freezing the skin at -20°C for up to 12 weeks.

Preparation, Characterization, and Biological Activities of Topical Anti-Aging Ingredients in a Citrus junos Callus Extract.

In this study, we prepared and characterized a callus extract from Citrus junos and assessed its utility as a source of topical anti-aging ingredients. Callus extract was produced by aqueous extraction from Citrus junos grown on Murashige and Skoog medium with picloram as a growth regulator. After measuring the total phenolic and flavonoid contents, the major phenolic compound in calli was identified as p-hydroxycinnamoylmalic acid (1) by spectroscopic analysis. The total phenol content in the extract was determined to be 24.50 ± 0.43 mg/g of gallic acid equivalents; however, the total flavonoid content of the extract was not determined. The biological activities of the callus extract, in terms of skin anti-aging, were assessed by measuring the anti-tyrosinase activity in, and melanogenesis by, melanoma cells; and proliferation of, and procollagen synthesis by, human fibroblasts. The callus extract was incorporated into nanoliposomes (NLs) to improve its percutaneous absorption. Addition of the callus extract resulted in a 1.85-fold decrease in the melanin content of melanocytes compared with that with arbutin. The extract (500 µg/mL) significantly promoted the proliferation of, and procollagen synthesis by, fibroblasts (by 154% and 176%, respectively). In addition, the flux through the human epidermis of Citrus junos callus extract incorporated into NLs was 17.67-fold higher than that of the callus extract alone. These findings suggest that Citrus junos callus extract-loaded NLs have promise as an anti-aging cosmetic, as well as having a skin-lightening effect.