Tatiana Andreani

Current State-of-Art and New Trends on Lipid Nanoparticles (SLN and NLC) for Oral Drug Delivery

Lipids and lipid nanoparticles are extensively employed as oral-delivery systems for drugs and other active ingredients. These have been exploited for many features in the field of pharmaceutical technology. Lipids usually enhance drug absorption in the gastrointestinal tract (GIT), and when formulated as nanoparticles, these molecules improve mucosal adhesion due to small particle size and increasing their GIT residence time. In addition, lipid nanoparticles may also protect the loaded drugs from chemical and enzymatic degradation and gradually release drug molecules from the lipid matrix into blood, resulting in improved therapeutic profiles compared to free drug. Therefore, due to their physiological and biodegradable properties, lipid molecules may decrease adverse side effects and chronic toxicity of the drug-delivery systems when compared to other of polymeric nature. This paper highlights the importance of lipid nanoparticles to modify the release profile and the pharmacokinetic parameters of drugs when administrated through oral route.

Design of cationic lipid nanoparticles for ocular delivery: development, characterization and cytotoxicity.

In the present study we have developed lipid nanoparticle (LN) dispersions based on a multiple emulsion technique for encapsulation of hydrophilic drugs or/and proteins by a full factorial design. In order to increase ocular retention time and mucoadhesion by electrostatic attraction, a cationic lipid, namely cetyltrimethylammonium bromide (CTAB), was added in the lipid matrix of the optimal LN dispersion obtained from the factorial design. There are a limited number of studies reporting the ideal concentration of cationic agents in LN for drug delivery. This paper suggests that the choice of the concentration of a cationic agent is critical when formulating a safe and stable LN. CTAB was included in the lipid matrix of LN, testing four different concentrations (0.25%, 0.5%, 0.75%, or 1.0%wt) and how composition affects LN behavior regarding physical and chemical parameters, lipid crystallization and polymorphism, and stability of dispersion during storage. In order to develop a safe and compatible system for ocular delivery, CTAB-LN dispersions were exposed to Human retinoblastoma cell line Y-79. The toxicity testing of the CTAB-LN dispersions was a fundamental tool to find the best CTAB concentration for development of these cationic LN, which was found to be 0.5 wt% of CTAB.

Preparation and characterization of PEG-coated silica nanoparticles for oral insulin delivery

The present study reports the production and characterization of PEG-coated silica nanoparticles (SiNP-PEG) containing insulin for oral administration. High (PEG 20,000) and low (PEG 6000) PEG molecular weights were used in the preparations. SiNP were produced by sol-gel technology followed by PEG adsorption and characterized for in vitro release by Franz diffusion cells. In vitro permeation profile was assessed using everted rat intestine. HPLC method has been validated for the determination of insulin released and permeated. Insulin secondary structure was performed by circular dichroism (CD). Uncoated SiNP allowed slower insulin release in comparison to SiNP-PEG. The coating with high molecular weight PEG did not significantly (p> 0.05) alter insulin release. The slow insulin release is attributed to the affinity of insulin for silanol groups at silica surface. Drug release followed second order kinetics for uncoated and SiNP-PEG at pH 2.0. On the other hand, at pH 6.8, the best fitting was first-order for SiNP-PEG, except for SiNP which showed a Boltzmann behavior. Comparing the values of half-live, SiNP-PEG 20,000 showed a faster diffusion followed by Si-PEG 6000 and SiNP. CD studies showed no conformational changes occurring after protein release from the nanoparticles under gastrointestinal simulated conditions.

Physicochemical characterization of epigallocatechin gallate lipid nanoparticles (EGCG-LNs) for ocular instillation

The encapsulation of epigallocatechin gallate (EGCG) in lipid nanoparticles (LNs) could be a suitable approach to avoid drug oxidation and epimerization, which are common processes that lead to low bioavailability of the drug limiting its therapeutic efficacy. The human health benefits of EGCG gained much interest in the pharmaceutical field, and so far there are no studies reporting its encapsulation in LNs. The purpose of this study has been the development of an innovative system for the ocular delivery of EGCG using LNs as carrier for the future treatment of several diseases, such as dry eye, age-related macular degeneration (AMD), glaucoma, diabetic retinopathy and macular oedema. LNs dispersions have been produced by multiple emulsion technique and previously optimized by a factorial design. In order to increase ocular retention time and mucoadhesion by electrostatic attraction, two distinct cationic lipids were used, namely, cetyltrimethylammonium bromide (CTAB) and dimethyldioctadecylammonium bromide (DDAB). EGCG has been successfully loaded in the LNs dispersions and the nanoparticles analysis over 30 days of storage time predicted a good physicochemical stability. The particles were found to be in the nanometer range (<300 nm) and all the evaluated parameters, namely pH, osmolarity and viscosity, were compatible to the ocular administration. The evaluation of the cationic lipid used was compared regarding physical and chemical parameters, lipid crystallization and polymorphism, and stability of dispersion during storage. The results show that different lipids lead to different characteristics mainly associated with the acyl chain composition, i.e. double lipid shows to have influence in the crystallization and stability. Despite the recorded differences between DTAB and DDAB, both cationic LNs seem to fit the parameters for ocular drug delivery.

In vitro evaluation of permeation, toxicity and effect of praziquantel-loaded solid lipid nanoparticles against Schistosoma mansoni as a strategy to improve efficacy of the schistosomiasis treatment.

Solid lipid nanoparticles (SLN) are a promising drug delivery system for oral administration of poorly-water soluble drugs because of their capacity to increase the solubility of drug molecules when loaded in their lipid matrices, with the resulting improvement of the drug bioavailability. In the present work, we have developed praziquantel (PZQ)-loaded SLN and explored the biological applications of this system for intestinal permeation of PZQ. The effect in vitro on Schistosoma mansoni culture and the cytotoxicity in HepG2 line cell were also evaluated. The results showed a significant decrease in the intestinal absorption of PZQ loaded in SLN compared to free PZQ, suggesting that the SLN matrix could act as reservoir system. In culture of S. mansoni, we observed that PZQ-loaded SLN were more effective than free PZQ, leading the death of the parasites in less time. The result was proportional to doses of PZQ (25 and 50 μg mL⁻¹) and lipid concentration. Regarding cytotoxicity, the encapsulation of PZQ into SLN decreased the toxicity in HepG2 cells in comparison to the free PZQ. From the obtained results, PZQ-loaded SLN could be a new drug delivery system for the schistosomiasis treatment especially in marginalized communities, improving the therapeutic efficacy and reducing the toxic effects of PZQ.

Biopharmaceutical evaluation of epigallocatechin gallate-loaded cationic lipid nanoparticles (EGCG-LNs): In vivo, in vitro and ex vivo studies.

Cationic lipid nanoparticles (LNs) have been tested for sustained release and site-specific targeting of epigallocatechin gallate (EGCG), a potential polyphenol with improved pharmacological profile for the treatment of ocular pathologies, such as age-related macular edema, diabetic retinopathy, and inflammatory disorders. Cationic EGCG-LNs were produced by double-emulsion technique; the in vitro release study was performed in a dialysis bag, followed by the drug assay using a previously validated RP-HPLC method. In vitro HET-CAM study was carried out using chicken embryos to determine the potential risk of irritation of the developed formulations. Ex vivo permeation profile was assessed using rabbit cornea and sclera isolated and mounted in Franz diffusion cells. The results show that the use of cationic LNs provides a prolonged EGCG release, following a Boltzmann sigmoidal profile. In addition, EGCG was successfully quantified in both tested ocular tissues, demonstrating the ability of these formulations to reach both anterior and posterior segment of the eye. The pharmacokinetic study of the corneal permeation showed a first order kinetics for both cationic formulations, while EGCG-cetyltrimethylammonium bromide (CTAB) LNs followed a Boltzmann sigmoidal profile and EGCG-dimethyldioctadecylammonium bromide (DDAB) LNs a first order profile. Our studies also proved the safety and non-irritant nature of the developed LNs. Thus, loading EGCG in cationic LNs is recognised as a promising strategy for the treatment of ocular diseases related to anti-oxidant and anti-inflammatory pathways.

Cationic solid lipid nanoparticles interfere with the activity of antioxidant enzymes in hepatocellular carcinoma cells.

Solid lipid nanoparticles (SLN) are colloidal drug and/or gene carriers developed from solid lipids and surfactants that are considered safe. Cationic SLN, usually used for formulating poorly water-soluble drugs and for gene delivery purposes, as positively charged particles may attach to cellular surfaces and be internalized more easily than negatively charged SLN, but they can also cause damage. The main aim of this work was to test a set of cationic SLN and investigate its influence on the amount of reactive oxygen species (ROS), on antioxidant enzymes activities and on possible oxidative damage to membrane lipids in HepG2 cells. The Dichlorofluorescein assay revealed great increase in ROS presence after cell exposure to SLN. While the exposure to SLN increased the activities of superoxide dismutase and glutathione peroxidase it decreased glutathione reductase activity. Although no significant increase in thiobarbituric reactive species was found, a decrease in sulfhydryl groups was detected. These results indicate that cationic SLN caused oxidative stress in HepG2 cells, but under reported exposure conditions HepG2 cells could attenuate the stress and thus the damage to cellular components was minimal.

Effect of mucoadhesive polymers on the in vitro performance of insulin-loaded silica nanoparticles: Interactions with mucin and biomembrane models

The present paper focuses on the development and characterization of silica nanoparticles (SiNP) coated with hydrophilic polymers as mucoadhesive carriers for oral administration of insulin. SiNP were prepared by sol-gel technology under mild conditions and coated with different hydrophilic polymers, namely, chitosan, sodium alginate or poly(ethylene glycol) (PEG) with low and high molecular weight (PEG 6000 and PEG 20000) to increase the residence time at intestinal mucosa. The mean size and size distribution, association efficiency, insulin structure and insulin thermal denaturation have been determined. The mean nanoparticle diameter ranged from 289 nm to 625 nm with a PI between 0.251 and 0.580. The insulin association efficiency in SiNP was recorded above 70%. After coating, the association efficiency of insulin increased up to 90%, showing the high affinity of the protein to the hydrophilic polymer chains. Circular dichroism (CD) indicated that no conformation changes of insulin structure occurred after loading the peptide into SiNP. Nano-differential scanning calorimetry (nDSC) showed that SiNP shifted the insulin endothermic peak to higher temperatures. The influence of coating on the interaction of nanoparticles with dipalmitoylphosphatidylcholine (DPPC) biomembrane models was also evaluated by nDSC. The increase of ΔH values suggested a strong association of non-coated SiNP and those PEGylated nanoparticles coated with DPPC polar heads by forming hydrogen bonds and/or by electrostatic interaction. The mucoadhesive properties of nanoparticles were examined by studying the interaction with mucin in aqueous solution. SiNP coated with alginate or chitosan showed high contact with mucin. On the other hand, non-coated SiNP and PEGylated SiNP showed lower interaction with mucin, indicating that these nanoparticles can interdiffuse across mucus network. The results of the present work provide valuable data in assessing the in vitro performance of insulin-loaded SiNP coated with mucoadhesive polymers.

d-α-tocopherol nanoemulsions: Size properties, rheological behavior, surface tension, osmolarity and cytotoxicity

The aim of this study was the assessment of the physicochemical stability of d-α-tocopherol formulated in medium chain triglyceride nanoemulsions, stabilized with Tween®80 and Lipoid®S75 as surfactant and co-surfactant, respectively. d-α-tocopherol was selected as active ingredient because of its well-recognized interesting anti-oxidant properties (such as radical scavenger) for food and pharmaceutical industries. A series of nanoemulsions of mean droplet size below 90 nm (polydispersity index < 0.15) have been produced by high-pressure homogenization, and their surface electrical charge (zeta potential), pH, surface tension, osmolarity, and rheological behavior, were characterized as a function of the d-α-tocopherol loading. In vitro studies in Caco-2 cell lines confirmed the safety profile of the developed nanoemulsions with percentage of cell viability above 90% for all formulations.

Toxicological impact of cadmium-based quantum dots towards aquatic biota: Effect of natural sunlight exposure

Cadmium-based quantum dots (QDs) are increasingly applied in existent and emerging technologies, especially in biological applications due to their exceptional photophysical and functionalization properties. However, they are very toxic compounds due to the high reactive and toxic cadmium core. The present study aimed to determine the toxicity of three different QDs (CdS 380, CdS 480 and CdSeS/ZnS) before and after the exposure of suspensions to sunlight, in order to assess the effect of environmentally relevant irradiation levels in their toxicity, which will act after their release to the environment. Therefore, a battery of ecotoxicological tests was performed with organisms that cover different functional and trophic levels, such as Vibrio fischeri, Raphidocelis subcapitata, Chlorella vulgaris and Daphnia magna. The results showed that core-shell type QDs showed lower toxic effects to V. fischeri in comparison to core type QDs before sunlight exposure. However, after sunlight exposure, there was a decrease of CdS 380 and CdS 480 QD toxicity to bacterium. Also, after sunlight exposure, an effective decrease of CdSeS/ZnS and CdS 480 toxicity for D. magna and R. subcapitata, and an evident increase in CdS 380 QD toxicity, at least for D. magna, were observed. The results of this study suggest that sunlight exposure has an effect in the aggregation and precipitation reactions of larger QDs, causing the degradation of functional groups and formation of larger bulks which may be less prone to photo-oxidation due to their diminished surface area. The same aggregation behaviour after sunlight exposure was observed for bare QDs. These results further emphasize that the shell of QDs seems to make them less harmful to aquatic biota, both under standard environmental conditions and after the exposure to a relevant abiotic factor like sunlight.