Quality-by-Design Approach for Biological API Encapsulation into Polymersomes Using "Off-the-Shelf" Materials: a Study on L-Asparaginase.

Polymersomes are versatile nanostructures for protein delivery with hydrophilic core suitable for large biomolecule encapsulation and protective stable corona. Nonetheless, pharmaceutical products based on polymersomes are not available in the market, yet. Here, using commercially available copolymers, we investigated the encapsulation of the active pharmaceutical ingredient (API) L-asparaginase, an enzyme used to treat acute lymphoblastic leukemia, in polymersomes through a quality-by-design (QbD) approach. This allows for streamlining of processes required for improved bioavailability and pharmaceutical activity. Polymersomes were prepared by bottom-up (temperature switch) and top-down (film hydration) methods employing the diblock copolymers poly(ethylene oxide)-poly(lactic acid) (PEG45-PLA69, PEG114-PLA153, and PEG114-PLA180) and the triblock Pluronic® L-121 (poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide), PEG5-PPO68-PEG5). Quality Target Product Profile (QTPP), Critical Quality Attributes (CQAs), Critical Process Parameters (CPPs), and the risk assessment were discussed for the early phase of polymersome development. An Ishikawa diagram was elaborated focusing on analytical methods, raw materials, and processes for polymersome preparation and L-asparaginase encapsulation. PEG-PLA resulted in diluted polymersomes systems. Nonetheless, a much higher yield of Pluronic® L-121 polymersomes of 200 nm were produced by temperature switch, reaching 5% encapsulation efficiency. Based on these results, a risk estimation matrix was created for an initial risk assessment, which can help in the future development of other polymersome systems with biological APIs nanoencapsulated.

Another Reason for Using Caffeine in Dermocosmetics: Sunscreen Adjuvant.

The excessive exposure to ultraviolet (UV) radiation is the main cause of skin cancer, the most commonly diagnosed cancer in the world. In this context, the development of innovative and more effective sunscreens, with bioactive compounds like caffeine, displaying antioxidant and anticancer potential, is required. This research work assessed in vitro and in vivo the efficacy and safety of topical sunscreen formulations containing caffeine as an adjuvant of the UV filters. Sunscreens were prepared with 2.5% w/w caffeine or in the absence of this compound. In order to evaluate the safety of these formulations, stratum corneum hydration, skin barrier and colorimetry were assessed in vivo in healthy subjects before and after skin treatment with the samples. The efficacy of the sunscreens was assessed in vitro, using PMMA plates and a spectrophotometer equipped with an integrating sphere; and in vivo by the determination of the sun protection factor (SPF). None of the formulations caused erythema or impaired the skin barrier function. The in vitro functional characterization showed higher SPF values for the caffeine formulation. The in vivo studies also confirmed the higher SPF value of the formulation combining caffeine with the filters, compared to the caffeine-free sample. This improvement contributed to an increase of, approximately, 25% in the in vivo anti-UVB protection. In conclusion, caffeine was well tolerated by the skin and increased the photoprotective activity, being a new alternative adjuvant in sunscreens formulation.

SPF enhancement provided by rutin in a multifunctional sunscreen.

Unprotected chronic exposure to solar radiation can contribute to premature skin cancer and sunscreens are a key factor to avoid those detrimental effects. Currently, there is a growing interest in the photoprotector and antioxidant potential of bioactive substances, such as rutin, that could increase the sun protection factor (SPF) value and, also, donate multifunctional characteristics to sunscreens. Recent in vitro findings indicated that rutin, when incorporated into sunscreens, can provide antioxidant activity and SPF improvement. However, clinical studies are fundamental to determine this activity, due to the lack of repeatability of in vitro methodology and low correlation with the in vivo data. We aimed at evaluating the clinical safety and in vivo SPF of rutin by comparing sunscreen formulations containing 0.1% (w/w) rutin, 3.0% (w/w) butyl methoxydibenzoylmethane and 8.0% (w/w) octyl dimethyl PABA (2-ethylhexyl 4-(dimethylamino)benzoato) with a similar bioactive-free preparation. Additionally, skin hydration, in vitro SPF and in vitro antioxidant activity of rutin, in association with the ultraviolet (UV) filters, were investigated. The safety profile of the formulations under sun-exposed skin conditions qualified the formulas for clinical efficacy assays. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) test confirmed the antioxidant properties of rutin, revealing around 40% increase in radical scavenging potential when the bioactive compound was present. Rutin in combination with the UV filters robustly elevated the clinical SPF around 70%, when compared with the bioactive-free formulation. To date, this is the first report in the specialized literature of an in vivo SPF measurement of a rutin-containing photoprotective preparation, supporting the claim that rutin is an effective and safe bioactive compound to be used in multifunctional sunscreens.

Ferulic acid photoprotective properties in association with UV filters: multifunctional sunscreen with improved SPF and UVA-PF.

Ultraviolet (UV) radiation stimulates several injurious biological effects on cutaneous tissue, causing, for instance, photocarcinogenesis. Sunscreens are topical products designed to protect the skin against these harmful effects and their use must be encouraged. The addition of antioxidants, as ferulic acid (FA), a phenolic compound from the class of the hydroxycinnamic acids, in sunscreens could improve their sun protection factor (SPF) and prevent inflammatory reactions. Here, the clinical safety and efficacy of an association of ethylhexyl triazone and bis-ethylhexyloxyphenol methoxyphenyl triazine (UV filters) with ferulic acid were assessed. Samples had good skin biocompatibility and presented satisfactory safety profile, even in a sun-exposed condition. A synergic effect between the natural polyphenol and the UV filters was evidenced, as well as, FA increased in vivo SPF in 37% and the UVA protection factor (UVA-PF) in 26%. The in vivo data indicated that FA reinforced the broad-spectrum characteristic of the photoprotective formulations. Additionally, according to the results from the ex vivo antioxidant test, it is plausible to recommend adjustments on the ex vivo protocol to explicitly determine the positive effects of topical antioxidant ingredients applied over the skin. These results provided a new perspective for the development of multifunctional bioactive sunscreens using FA as a new platform.

Safety and Antioxidant Efficacy Profiles of Rutin-Loaded Ethosomes for Topical Application.

Topical application of dermocosmetics containing antioxidant and/or the intake of antioxidants through diet or supplementation are remarkable tools in an attempt to slow down some of the harmful effects of free radicals. Rutin is a strong antioxidant compound used in food and pharmaceutical industries. It was established that rutin presents a low skin permeation rate, a property that could be considered an inconvenience to the satisfactory action for a dermocosmetic formulation to perform its antioxidant activity onto the skin. Therefore, it is indispensable to improve its delivery, aiming at increasing its antioxidant capacity in deeper layers of the epidermis, being a possibility to associate the rutin to liposomal vesicles, such as ethosomes. Thus, in this work, the pre-clinical safety of rutin-loaded ethosomes was investigated employing an in vitro method, and the clinical safety and efficacy were also assessed. Rutin-loaded ethosomes were efficaciously obtained in a nanoscale dimension with a relevant bioactive compound loading (80.2%) and provided antioxidant in vitro activity in comparison with the blank sample. Pre-clinical and clinical safety assays assured the innocuous profile of the rutin-loaded ethosomes. The ethosomes containing the bioactive compound accomplished a more functional delivery system profile, since in the tape stripping assay, the deeper layers presented higher rutin amounts than the active delivered in its free state. However, the ex vivo antioxidant efficacy test detected no positive antioxidant activity from the rutin-loaded ethosomes, even though the in vitro assay demonstrated an affirmative antioxidant action.

Functional photostability and cutaneous compatibility of bioactive UVA sun care products.

Sunscreens are the most-established approach for photoprotection. The strategy of providing antioxidant properties to sun care products by addition of natural and potent anti-free radical compounds has led to the development of bioactive sunscreens, able to neutralize the harmful effects of ultraviolet (UV) radiation. UVA filters, such as benzophenone-3 (BP) and butyl methoxydibenzoylmethane (BMDBM), can exhibit photodegradation which limits the development of broad spectrum sunscreens. Previous research verified that rutin interacts with filters incorporated in sunscreens. In this work, we focused on the development and evaluation of the efficacy of the sunscreens containing either BP or BMDBM with and without rutin. The addition of rutin to the UVA filters afforded antioxidant properties to the formulations and they were considered safe for human use. Additionally, rutin in combination with either BP or BMDBM increased the antioxidant activity about 40 times when compared with the UVA filters alone. Remarkably, the addition of rutin 0.1% (w/w) to BP 6.0% (w/w) raised the SPF from 24.3±1.53 to 33.3±2.89. In conclusion, these findings demonstrated that the addition of rutin into sunscreens can markedly improve the antioxidant properties of the formulation as well as photostabilize some of the UVA filters.