Enhanced Delivery of 5-Aminolevulinic Acid by Lecithin Invasomes in 3D Melanoma Cancer Model.

Photodynamic therapy (PDT) is a noninvasive therapeutic approach for the treatment of skin cancer and diseases. 5-Aminolevulinic acid is a prodrug clinically approved for PDT. Once internalized by cancer cells, it is rapidly metabolized to the photosensitizer protoporphyrin IX, which under the proper light irradiation, stimulates the deleterious reactive oxygen species (ROS) production and leads to cell death. The high hydrophilicity of 5-aminolevulinic acid limits its capability to cross the epidermis. Lipophilic derivatives of 5-aminolevulinic acid only partly improved skin penetration, thus making its incorporation into nanocarriers necessary. Here we have developed and characterized 5-aminolevulinic acid loaded invasomes made of egg lecithin, either 1,2-dilauroyl-sn-glycero-3-phosphocholine or 1,2-dioleoyl-sn-glycero-3-phosphocholine, and the terpene limonene. The obtained invasomes are highly thermostable and display a spherical morphology with an average size of 150 nm and an encapsulation efficiency of 80%; moreover, the ex vivo epidermis diffusion tests established that nanovesicles containing the terpene led to a much higher skin penetration (up to 80% in 3 h) compared to those without limonene and to the free fluorescent tracer (less than 50%). Finally, in vitro studies with 2D and 3D human cell models of melanoma proved the biocompatibility of invasomes, the enhanced intracellular transport of 5-aminolevulinic acid, its ability to generate ROS upon irradiation, and consequently, its antiproliferative effect. A simplified scaffold-based 3D skin model containing melanoma spheroids was also prepared. Considering the results obtained, we conclude that the lecithin invasomes loaded with 5-aminolevulinic acid have a good therapeutic potential and may represent an efficient tool that can be considered a valid alternative in the topical treatment of melanoma and other skin diseases.

Preparation, characterisation and applications of bone char, a food waste-derived sustainable material: A review.

The production of increasing quantities of by-products is a key challenge for modern society; their valorisation - turning them into valuable compounds with technological applications - is the way forward, in line with circular economy principles. In this review, the conversion of bones (by-products of the agro-food industry) into bone char is described. Bone char is obtained with a process of pyrolysis, which converts the organic carbon into an inorganic graphitic one. Differently from standard biochar of plant origin, however, bone char also contains calcium phosphates, the main component of bone (often hydroxyapatite). The combination of calcium phosphate and graphitic carbon makes bone char a unique material, with different possible uses. Here bone chars' applications in environmental remediation, sustainable agriculture, catalysis and electrochemistry are discussed; several aspects are considered, including the bones used to prepare bone char, the preparation conditions, how these affect the properties of the materials (i.e. porosity, surface area) and its functional properties. The advantages and limitations of bone chars in comparison to traditional biochar are discussed, highlighting the directions the research should take for bone chars' performances to improve. Moreover, an analysis on the sustainability of bone chars' preparation and use is also included.

Calcium Phosphates from Fishery Byproducts as a Booster of the Sun Protection Factor in Sunscreens.

Calcium phosphates (CaPs) have been recently proposed as a bio- and eco-compatible alternative to UV filters in sunscreens, which are in the spotlight for being associated with health risks for both people and the environment. Here, natural CaPs extracted from fish bones have been tested as a booster of the sun protection factor (SPF), that is, as material working in synergy with UV filters to increase sunscreen UV-shielding efficiency, in combination with three of the most used UV filters, namely, octocrylene (OCR), octinoxate, and padimate-O, at different concentrations (10.0 and 20.0 wt %). The material obtained by calcination at 800 °C (CaP-N) was also enriched with Zn (CaP-Zn) or Mn (CaP-Mn) in an attempt to increase its SPF-boosting abilities. CaP-N and CaP-Zn consisted of a biphasic mixture of hydroxyapatite and beta tricalcium phosphate, while CaP-Mn presented a small quantity of Mn oxides. CaP-N was the most effective at increasing the SPF of the final emulsions, doubling the SPF of the formulation containing 20.0 wt % of OCR from 40.6 to 80.8. The results show that these CaPs, produced according to a circular economy approach, can be used as effective SPF boosters to decrease the concentration of UV filters used in sunscreen, while retaining high SPF values.

Monodisperse and Nanometric-Sized Calcium Carbonate Particles Synthesis Optimization.

Calcium carbonate (CaCO3) particles represent an appealing choice as a drug delivery system due to their biocompatibility, biodegradability, simplicity and cost-effectiveness of manufacturing, and stimulus-responsiveness. Despite this, the synthesis of CaCO3 particles with controlled size in the nanometer range via a scalable manufacturing method remains a major challenge. Here, by using a co-precipitation technique, we investigated the impact on the particle size of different synthesis parameters, such as the salt concentration, reaction time, stirring speed, and temperature. Among them, the salt concentration and temperature resulted in having a remarkable effect on the particle size, enabling the preparation of well-dispersed spherical nanoparticles with a size below 200 nm. Upon identification of optimized synthesis conditions, the encapsulation of the antitumoral agent resveratrol into CaCO3 nanoparticles, without significantly impacting the overall size and morphology, has been successfully achieved.

Poly(l-lactide-co-caprolactone-co-glycolide)-Based Nanoparticles as Delivery Platform: Effect of the Surfactants on Characteristics and Delivery Efficiency.

Polymeric nanoparticles made of the copolymer Poly(L-lactide-co-caprolactone-co-glycolide) were prepared using the solvent evaporation method. Two different surfactants, polyvinyl alcohol and dextran, and a mixture of the two were employed. The three types of nanoparticles were used as hosting carriers of two chemotherapeutic drugs, the hydrophilic doxorubicin and the hydrophobic SN-38. The morphostructural characterization showed similar features for the three types of nanoparticles, while the drug encapsulation efficiency indicated that the dextran-based systems are the most effective with both drugs. Cellular studies with breast cancer cells were performed to compare the delivery capability and the cytotoxicity profile of the three nanosystems. The results show that the unloaded nanoparticles are highly biocompatible at the administered concentrations and confirmed that dextran-coated nanoparticles are the most efficient vectors to release the two drugs, exerting cytotoxic activity. PVA, on the other hand, shows limited drug release in vitro, probably due to strong interactions with both drugs. Data also show the release is more efficient for doxorubicin than for SN-38; indeed, the doxorubicin IC50 value for the dextran-coated nanoparticles was about 35% lower than the free drug. This indicates that these nanocarriers are suitable candidates to deliver hydrophilic drugs while needing further modification to host hydrophobic molecules.

Valorization of porcine by-products: a combined process for protein hydrolysates and hydroxyapatite production.

The meat industry generates large amounts of by-products that are costly to be treated and discarded ecologically; moreover, they could be used to extract high added-value compounds. In this work, we present an innovative combined process which allowed the parallel extraction of both organic and mineral compounds; more specifically protein hydrolysates and single-phase hydroxyapatite were obtained. The protein hydrolysates, extracted through an enzymatic hydrolysis with alcalase, showed a degree of hydrolysis of 53.3 ± 5.1%; moreover, they had a high protein content with peptides with molecular weight lower than 1.2 kDa. Their antioxidant activities, measured with ABTS and ORAC tests, were 21.1 ± 0.5 mg ascorbic acid equivalent/g of dry extract and 87.7 ± 6.3 mg Trolox equivalent/g of dry extract, respectively. Single-phase hydroxyapatite, obtained with a simple calcination at 700 °C on the residues of the hydrolysis process, showed a Ca/P ratio close to the stoichiometric one (1.65 vs. 1.67) and presented a nanometric structure. This study reports a simple and feasible process for the valorization of porcine by-products in a large-scale up generating products with potential applications for environment remediation, biomedicine, nutrition and catalysis/bioenergy.

Calcium Phosphate Particles Coated with Humic Substances: A Potential Plant Biostimulant from Circular Economy.

Nowadays, the use of biostimulants to reduce agrochemical input is a major trend in agriculture. In this work, we report on calcium phosphate particles (CaP) recovered from the circular economy, combined with natural humic substances (HSs), to produce a plant biostimulant. CaPs were obtained by the thermal treatment of Salmo salar bones and were subsequently functionalized with HSs by soaking in a HS water solution. The obtained materials were characterized, showing that the functionalization with HS did not sort any effect on the bulk physicochemical properties of CaP, with the exception of the surface charge that was found to get more negative. Finally, the effect of the materials on nutrient uptake and translocation in the early stages of development (up to 20 days) of two model species of interest for horticulture, Valerianella locusta and Diplotaxis tenuifolia, was assessed. Both species exhibited a similar tendency to accumulate Ca and P in hypogeal tissues, but showed different reactions to the treatments in terms of translocation to the leaves. CaP and CaP-HS treatments lead to an increase of P accumulation in the leaves of D. tenuifolia, while the treatment with HS was found to increase only the concentration of Ca in V. locusta leaves. A low biostimulating effect on both plants' growth was observed, and was mainly scribed to the low concentration of HS in the tested materials. In the end, the obtained material showed promising results in virtue of its potential to elicit phosphorous uptake and foliar translocation by plants.

Lipid-Based Nanovesicles for Simultaneous Intracellular Delivery of Hydrophobic, Hydrophilic, and Amphiphilic Species.

Lipid nanovesicles (NVs) are the first nanoformulation that entered the clinical use in oncology for the treatment of solid tumors. They are indeed versatile systems which can be loaded with either hydrophobic or hydrophilic molecules, for both imaging and drug delivery, and with high biocompatibility, and limited immunogenicity. In the present work, NVs with a lipid composition resembling that of natural vesicles were prepared using the ultrasonication method. The NVs were successfully loaded with fluorophores molecules (DOP-F-DS and a fluorescent protein), inorganic nanoparticles (quantum dots and magnetic nanoparticles), and anti-cancer drugs (SN-38 and doxorubicin). The encapsulation of such different molecules showed the versatility of the developed systems. The size of the vesicles varied from 100 up to 300 nm depending on the type of loaded species, which were accommodated either into the lipid bilayer or into the aqueous core according to their hydrophobic or hydrophilic nature. Viability assays were performed on cellular models of breast cancer (MCF-7 and MDA-MB-231). Results showed that NVs with encapsulated both drugs simultaneously led to a significant reduction of the cellular activity (up to 22%) compared to the free drugs or to the NVs encapsulated with only one drug. Lipidomic analysis suggested that the mechanism of action of the drugs is the same, whether they are free or encapsulated, but administration of the drugs by means of nanovesicles is more efficient in inducing cellular damage, likely because of a quicker internalization and a sustained release. This study confirms the versatility and the potential of lipid NVs for cancer treatment, as well as the validity of the ultrasound preparation method for their preparation.

Mussel Shell-Derived Macroporous 3D Scaffold: Characterization and Optimization Study of a Bioceramic from the Circular Economy.

Fish industry by-products constitute an interesting platform for the extraction and recovery of valuable compounds in a circular economy approach. Among them, mussel shells could provide a calcium-rich source for the synthesis of hydroxyapatite (HA) bioceramics. In this work, HA nanoparticles have been successfully synthesized starting from mussel shells (Mytilus edulis) with a two steps process based on thermal treatment to convert CaCO3 in CaO and subsequent wet precipitation with a phosphorus source. Several parameters were studied, such as the temperature and gaseous atmosphere of the thermal treatment as well as the use of two different phosphorus-containing reagents in the wet precipitation. Data have revealed that the characteristics of the powders can be tailored, changing the conditions of the process. In particular, the use of (NH4)2HPO4 as the phosphorus source led to HA nanoparticles with a high crystallinity degree, while smaller nanoparticles with a higher surface area were obtained when H3PO4 was employed. Further, a selected HA sample was synthesized at the pilot scale; then, it was employed to fabricate porous 3D scaffolds using the direct foaming method. A highly porous scaffold with open and interconnected porosity associated with good mechanical properties (i.e., porosity in the range 87-89%, pore size in the range 50-300 µm, and a compressive strength σ = 0.51 ± 0.14 MPa) suitable for bone replacement was achieved. These results suggest that mussel shell by-products are effectively usable for the development of compounds of high added value in the biomedical field.

Films of chitosan and natural modified hydroxyapatite as effective UV-protecting, biocompatible and antibacterial wound dressings.

Chitosan is a natural polysaccharide widely used in biomedicine, for instance for wound dressing. Hydroxyapatite is a very bioactive calcium phosphate which, if modified with an appropriate element (iron Fe), can also have UV-absorbing properties. In this work, we report the study of films of chitosan incorporated with iron-modified hydroxyapatite of natural origin (from cod fish bones); this combination led to an innovative chitosan-based material with excellent and advanced functional properties. The films showed very high UV absorption (Ultraviolet Protection Factor (UPF) value higher than 50). This is the first time that a chitosan-based material has shown such high UV protection properties. The films also showed to be non-cytotoxic, and possessed antimicrobial activity towards both Gram-positive and negative strains. Their mechanical properties, optimised with an experimental design approach, confirmed their potential use as multifunctional wound dressing, capable of reducing bacterial infections and, at the same time, protecting from UV light.

Nanoheterostructures (NHS) and Their Applications in Nanomedicine: Focusing on In Vivo Studies.

Inorganic nanoparticles have great potential for application in many fields, including nanomedicine. Within this class of materials, inorganic nanoheterostructures (NHS) look particularly promising as they can be formulated as the combination of different domains; this can lead to nanosystems with different functional properties, which, therefore, can perform different functions at the same time. This review reports on the latest development in the synthesis of advanced NHS for biomedicine and on the tests of their functional properties in in vivo studies. The literature discussed here focuses on the diagnostic and therapeutic applications with special emphasis on cancer. Considering the diagnostics, a description of the NHS for cancer imaging and multimodal imaging is reported; more specifically, NHS for magnetic resonance, computed tomography and luminescence imaging are considered. As for the therapeutics, NHS employed in magnetic hyperthermia or photothermal therapies are reported. Examples of NHS for cancer theranostics are also presented, emphasizing their dual usability in vivo, as imaging and therapeutic tools. Overall, NHS show a great potential for biomedicine application; further studies, however, are necessary regarding the safety associated to their use.

Safety of Yam-Derived (Dioscorea rotundata) Foodstuffs-Chips, Flakes and Flour: Effect of Processing and Post-Processing Conditions.

The production of yam-derived (Dioscorea rotundata) foodstuffs is mainly performed by small and medium scale processors that employ old traditional methods. This can lead to differences in quality from processor to processor, and from location to location, with consequent safety concerns. As such, the effects of processing and post-processing phases (i.e., storage, transport, etc.) on the safety of some yam-derived foodstuffs-namely chips, flakes, and flour-has been evaluated, with a focus on bacterial and fungal contamination, aflatoxins, pesticides, and heavy metals (Pb, Ni, Cd and Hg). Yams harvested and processed in Nigeria were screened, being that the country is the largest producer of the tuber, with 70⁻75% of the world production. Results highlighted no presence of pesticides, however, many samples showed high levels of bacterial and fungal contamination, together with heavy metal concentrations above the recommended safety levels. No trend was observed between the items considered; it was noticed, however, that samples purchased from the markets showed higher contamination levels than those freshly produced, especially regarding bacterial and aflatoxins presence. The processing stage was identified as the most critical, especially drying. Nonetheless, post-processing steps such as storage and handling at the point of sale also contributed for chemical contamination, such as aflatoxin and heavy metals. The results suggested that both the processing and post-processing phases have an impact on the safety of yam chips, flakes, and flour.

Photocatalytic Degradation of Diclofenac by Hydroxyapatite⁻TiO2 Composite Material: Identification of Transformation Products and Assessment of Toxicity.

Diclofenac (DCF) is one of the most detected pharmaceuticals in environmental water matrices and is known to be recalcitrant to conventional wastewater treatment plants. In this study, degradation of DCF was performed in water by photolysis and photocatalysis using a new synthetized photocatalyst based on hydroxyapatite and TiO2 (HApTi). A degradation of 95% of the target compound was achieved in 24 h by a photocatalytic treatment employing the HApTi catalyst in comparison to only 60% removal by the photolytic process. The investigation of photo-transformation products was performed by means of UPLC-QTOF/MS/MS, and for 14 detected compounds in samples collected during treatment with HApTi, the chemical structure was proposed. The determination of transformation product (TP) toxicity was performed by using different assays: Daphnia magna acute toxicity test, Toxi-ChromoTest, and Lactuca sativa and Solanum lycopersicum germination inhibition test. Overall, the toxicity of the samples obtained from the photocatalytic experiment with HApTi decreased at the end of the treatment, showing the potential applicability of the catalyst for the removal of diclofenac and the detoxification of water matrices.

Biodegradation of Diclofenac by the bacterial strain Labrys portucalensis F11.

Diclofenac (DCF) is a widely used non-steroidal anti-inflammatory pharmaceutical which is detected in the environment at concentrations which can pose a threat to living organisms. In this study, biodegradation of DCF was assessed using the bacterial strain Labrys portucalensis F11. Biotransformation of 70% of DCF (1.7-34 µM), supplied as the sole carbon source, was achieved in 30 days. Complete degradation was reached via co-metabolism with acetate, over a period of 6 days for 1.7 µM and 25 days for 34 µM of DCF. The detection and identification of biodegradation intermediates was performed by UPLC-QTOF/MS/MS. The chemical structure of 12 metabolites is proposed. DCF degradation by strain F11 proceeds mainly by hydroxylation reactions; the formation of benzoquinone imine species seems to be a central step in the degradation pathway. Moreover, this is the first report that identified conjugated metabolites, resulting from sulfation reactions of DCF by bacteria. Stoichiometric liberation of chlorine and no detection of metabolites at the end of the experiments are strong indications of complete degradation of DCF by strain F11. To the best of our knowledge this is the first report that points to complete degradation of DCF by a single bacterial strain isolated from the environment.

Cassava (Manihot esculenta Crantz) and Yam (Dioscorea spp.) Crops and Their Derived Foodstuffs: Safety, Security and Nutritional Value.

Cassava (Manihot esculenta Crantz) and yam (Dioscorea spp.) are tropical crops consumed by ca. 2 billion people and represent the main source of carbohydrate and energy for the approximately 700 million people living in the tropical and sub-tropical areas. They are a guarantee of food security for developing countries. The production of these crops and the transformation into food-derived commodities is increasing, it represents a profitable business and farmers generate substantial income from their market. However, there are some important concerns related to the food safety and food security. The high post-harvest losses, mainly for yam, the contamination by endogenous toxic compounds, mainly for cassava, and the contamination by external agents (such as micotoxins, pesticides, and heavy metal) represent a depletion of economic value and income. The loss in the raw crops or the impossibility to market the derived foodstuffs, due to incompliance with food regulations, can seriously limit all yam tubers and the cassava roots processors, from farmers to household, from small-medium to large enterprises. One of the greatest challenges to overcome those concerns is the transformation of traditional or indigenous processing methods into modern industrial operations, from the crop storage to the adequate package of each derived foodstuff.

Characterization and antimicrobial properties of food packaging methylcellulose films containing stem extract of Ginja cherry.

BACKGROUND: Food contamination and spoilage is a problem causing growing concern. To avoid it, the use of food packaging with appropriate characteristics is essential; ideally, the packaging should protect food from external contamination and exhibit antibacterial properties. With this aim, methylcellulose (MC) films containing natural extracts from the stems of Ginja cherry, an agricultural by-product, were developed and characterized. RESULTS: The antibacterial activity of films was screened by the disc diffusion method and quantified using the viable cell count assay. The films inhibited the growth of both Gram-positive and Gram-negative strains (Listeria innocua, methicillin-sensitive Staphylococcus aureus, methicillin-resistant S. aureus, Salmonella Enteritidis, Escherichia coli). For the films with lower extract content, effectiveness against the microorganisms depended on the inoculum concentration. Scanning electron microscope images of the films showed that those containing the extracts had a smooth and continuous structure. UV-visible spectroscopy showed that these materials do not transmit light in the UV. CONCLUSION: This study shows that MC films containing agricultural by-products, in this case Ginja cherry stem extract, could be used to prevent food contamination by relevant bacterial strains and degradation by UV light. Using such materials in food packaging, the shelf life of food products could be extended while utilizing an otherwise wasted by-product.

Extraction of high added value biological compounds from sardine, sardine-type fish and mackerel canning residues--a review.

Different valuable compounds, which can be employed in medicine or in other industries (i.e. food, agrochemical, pharmaceutical) can be recovered from by-products and waste from the fish canning industries. They include lipids, proteins, bio-polymers, minerals, amino acids and enzymes; they can be extracted from wastewaters and/or from solid residues (head, viscera, skin, tails and flesh) generated along the canning process, through the filleting, cooking, salting or smoking stages. In this review, the opportunities for the extraction and the valorisation of bioactive compounds from sardine, sardine-type fish and mackerel canning residues are examined and discussed. These are amongst the most consumed fishes in the Mediterranean area; moreover, canning is one of the most important and common methods of preservation. The large quantities of by-products generated have great potentials for the extraction of biologically desirable high added value compounds.

Antimicrobial properties of light-activated polyurethane containing indocyanine green.

The aim of this study was to produce novel antimicrobial polymers containing the light-activated antimicrobial agent indocyanine green (ICG). The novel materials were prepared by swelling polyurethane in acetone containing water and ICG, followed by solvent evaporation. The uptake of ICG was dependent upon the ratio of acetone to water. Only at a ratio of 99 parts acetone to 1 part water was there any substantial colouration of the samples. When exposed to laser light from the near infrared spectrum (808 nm), polyurethane-containing ICG exhibited antimicrobial activity against Gram-positive bacteria; a 2 log10 reduction was achieved against methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus epidermidis after 15 min exposure, corresponding to an energy dose of 31.83 J delivered at an energy density of 31.83 J/cm². Under the same conditions, Gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa) appeared to be less susceptible, the viable count being reduced by 0.5 log10. Some of the physical properties of the resulting material were also investigated and it was found that the elasticity (Young's modulus) was reduced by approximately 60%. Furthermore, when the ICG-containing polymer was stretched, the breaking point occurred when the elongation was 6.7 times the initial value, while ICG-free polyurethane samples did not break following a 7-fold elongation. The contact angles for water droplets revealed that the ICG-containing polymer was more hydrophobic than untreated polyurethane. The results of this study show that ICG can be embedded in polyurethane to produce materials which when irradiated with near-infrared light can exert a bactericidal effect particularly against MRSA and S. epidermidis. Such materials may be useful for preparing intravenous catheters, which are often colonized by such organisms.

Frictional properties of light-activated antimicrobial polymers in blood vessels.

The adhesion of microbes to catheter surfaces is a serious problem and the resulting infections frequently lead to longer hospitalisation and higher risk for the patient. Several approaches have been developed to produce materials that are less susceptible to microbial colonisation. One such approach is the incorporation of photoactivated compounds, such as Toluidine Blue O (TBO), in the polymeric matrix resulting in 'light-activated antimicrobial materials'. The insertion and removal of catheters can cause tissue damage and patient discomfort through frictional forces; hence the lubricity of a catheter material is also very important. In this work the tribological performance of silicone and polyurethane containing TBO and gold nanoparticles were evaluated using two different surfaces, the inner part of the aorta and the superior vena cava of sheep. Static and kinetic friction coefficients of these materials were measured using a tribometric device developed for in vitro applications using dry materials and those lubricated with blood. It was found that neither the preparation process nor the presence of TBO or gold nanoparticles, had an effect on the friction factors in comparison to those of untreated materials. In all cases, static and kinetic friction coefficients on aorta tissue were higher than those on vena cava due to higher surface roughness of the aorta. The presence of blood as a lubricant resulted in lower friction coefficients.

The antimicrobial properties of light-activated polymers containing methylene blue and gold nanoparticles.

We report the formation of polysiloxane polymers containing embedded methylene blue and gold nanoparticles incorporated by a swell-encapsulation-shrink method. These polymers show significant antimicrobial activity against methicillin-resistant Staphylococcus aureus and Escherichia coli with up to a 3.5 log(10) reduction in the viable count when exposed for 5 min to light from a low power 660 nm laser. The bacterial kill is due to the light-induced production of singlet oxygen and other reactive oxygen species by the methylene blue. Interestingly, the presence of 2 nm gold nanoparticles significantly enhanced the ability of the methylene blue to kill bacteria.