Antibacterial polypeptide-bioparticle for oral administration: Powder formulation, palatability and in vivo toxicity approach.

The upsurge of bacterial resistance to conventional antibiotics turned a well-recognized public health threat. The need of developing new biomaterials of effective practical use in order to tackle bacterial resistance became urgent. In this study, a submicrometric bioparticle of known antibacterial activity was produced in powder form with suitable texture and appealing characteristics for effective oral administration. Through complex coacervating a natural-source antimicrobial polypeptide with chitosan-N-arginine and alginate, the bioactive polypeptide was physically incorporated to the bioparticle whose structure positively responds to the pH variations found in gastrointestinal tract. The powder formulation presented high palatability that was evaluated using fish as in vivo animal model. A thorough survey of the fish intestinal tissues, following a systematic oral administration, revealed high penetration potential of the biomaterial through epithelial cells and deeper intestine layers. Despite, no cytotoxic effect was observed in analyzing the tissues through different histology methods. The absence of intestinal damage was corroborated by immune histochemistry, being the integrity of epithelial motor myosin Vb and related traffic proteins preserved. Hematology further endorsed absence of toxicity in blood cells whose morphology was evaluated in detail. The study evidenced the applicability potential of a new biomaterial of appealing and safe oral administration of antibacterial polypeptide.

Promising strategies for improving oral bioavailability of poor water-soluble drugs.

INTRODUCTION: Oral administration of poorly water-soluble drugs (PWSDs) is generally related to low bioavailability, leading to high drug doses, multiple side effects, and low patient compliance. Thus, different strategies have been developed to increase drug solubility and dissolution in the gastrointestinal tract, opening new venues for these drugs. AREAS COVERED: This review outlines the current challenges in PWSD formulation development and the strategies to overcome the oral barriers and increase their solubility and bioavailability. Conventional strategies include altering crystalline and molecular structures and modifying oral solid dosage forms. In contrast, novel strategies comprise micro- and nanostructured systems. Recent representative studies involving how these strategies have improved the oral bioavailability of PWSDs were also reviewed and reported. EXPERT OPINION: New approaches to enhance PWSD bioavailability have sought to improve water solubility and dissolution rates, drug protection by overcoming biological barriers, and increased absorption. Still, only a handful of studies have focused on quantifying the increase in bioavailability. Improving the oral bioavailability of PWSDs remains an exciting unexplored field of research and has become an important issue for successfully developing pharmaceutical products.

Oral Administration as a Potential Alternative for the Delivery of Small Extracellular Vesicles.

Small extracellular vesicles (sEVs) have burst into biomedicine as a natural therapeutic alternative for different diseases. Considered nanocarriers of biological origin, various studies have demonstrated the feasibility of their systemic administration, even with repeated doses. However, despite being the preferred route of physicians and patients, little is known about the clinical use of sEVs in oral administration. Different reports show that sEVs can resist the degradative conditions of the gastrointestinal tract after oral administration, accumulating regionally in the intestine, where they are absorbed for systemic biodistribution. Notably, observations demonstrate the efficacy of using sEVs as a nanocarrier system for a therapeutic payload to obtain a desired biological (therapeutic) effect. From another perspective, the information to date indicates that food-derived vesicles (FDVs) could be considered future nutraceutical agents since they contain or even overexpress different nutritional compounds of the foods from which they are derived, with potential effects on human health. In this review, we present and critically analyze the current information on the pharmacokinetics and safety profile of sEVs when administered orally. We also address the molecular and cellular mechanisms that promote intestinal absorption and that command the therapeutic effects that have been observed. Finally, we analyze the potential nutraceutical impact that FDVs would have on human health and how their oral use could be an emerging strategy to balance nutrition in people.

Arabinoxylans-Based Oral Insulin Delivery System Targeting the Colon: Simulation in a Human Intestinal Microbial Ecosystem and Evaluation in Diabetic Rats.

Arabinoxylans (AX) microcapsules loaded with insulin were prepared by enzymatic gelation of AX, using a triaxial electrospray method. The microcapsules presented a spherical shape, with an average size of 250 µm. The behavior of AX microcapsules was evaluated using a simulator of the human intestinal microbial ecosystem. AX microcapsules were mainly (70%) degraded in the ascending colon. The fermentation was completed in the descending colon, increasing the production of acetic, propionic, and butyric acids. In the three regions of the colon, the fermentation of AX microcapsules significantly increased populations of Bifidobacterium and Lactobacillus and decreased the population of Enterobacteriaceae. In addition, the results found in this in vitro model showed that the AX microcapsules could resist the simulated conditions of the upper gastrointestinal system and be a carrier for insulin delivery to the colon. The pharmacological activity of insulin-loaded AX microcapsules was evaluated after oral delivery in diabetic rats. AX microcapsules lowered the serum glucose levels in diabetic rats by 75%, with insulin doses of 25 and 50 IU/kg. The hypoglycemic effect and the insulin levels remained for more than 48 h. Oral relative bioavailability was 13 and 8.7% for the 25 and 50 IU/kg doses, respectively. These results indicate that AX microcapsules are a promising microbiota-activated system for oral insulin delivery in the colon.

Microfluidic Synthesis and Purification of Magnetoliposomes for Potential Applications in the Gastrointestinal Delivery of Difficult-to-Transport Drugs.

Magnetite nanoparticles (MNPs) have gained significant attention in several applications for drug delivery. However, there are some issues related to cell penetration, especially in the transport of cargoes that show limited membrane passing. A widely studied strategy to overcome this problem is the encapsulation of the MNPs into liposomes to form magnetoliposomes (MLPs), which are capable of fusing with membranes to achieve high delivery rates. This study presents a low-cost microfluidic approach for the synthesis and purification of MLPs and their biocompatibility and functional testing via hemolysis, platelet aggregation, cytocompatibility, internalization, and endosomal escape assays to determine their potential application in gastrointestinal delivery. The results show MLPs with average hydrodynamic diameters ranging from 137 ± 17 nm to 787 ± 45 nm with acceptable polydispersity index (PDI) values (below 0.5). In addition, we achieved encapsulation efficiencies between 20% and 90% by varying the total flow rates (TFRs), flow rate ratios (FRRs), and MNPs concentration. Moreover, remarkable biocompatibility was attained with the obtained MLPs in terms of hemocompatibility (hemolysis below 1%), platelet aggregation (less than 10% with respect to PBS 1×), and cytocompatibility (cell viability higher than 80% in AGS and Vero cells at concentrations below 0.1 mg/mL). Additionally, promising delivery results were obtained, as evidenced by high internalization, low endosomal entrapment (AGS cells: PCC of 0.28 and covered area of 60% at 0.5 h and PCC of 0.34 and covered area of 99% at 4 h), and negligible nuclear damage and DNA condensation. These results confirm that the developed microfluidic devices allow high-throughput production of MLPs for potential encapsulation and efficient delivery of nanostructured cell-penetrating agents. Nevertheless, further in vitro analysis must be carried out to evaluate the prevalent intracellular trafficking routes as well as to gain a detailed understanding of the existing interactions between nanovehicles and cells.

Encapsulation of Active Pharmaceutical Ingredients in Lipid Micro/Nanoparticles for Oral Administration by Spray-Cooling.

Nanoencapsulation via spray cooling (also known as spray chilling and spray congealing) has been used with the aim to improve the functionality, solubility, and protection of drugs; as well as to reduce hygroscopicity; to modify taste and odor to enable oral administration; and many times to achieve a controlled release profile. It is a relatively simple technology, it does not require the use of low-cost solvents (mostly associated to toxicological risk), and it can be applied for lipid raw materials as excipients of oral pharmaceutical formulations. The objective of this work was to revise and discuss the advances of spray cooling technology, with a greater emphasis on the development of lipid micro/nanoparticles to the load of active pharmaceutical ingredients for oral administration.

Stability Studies of the Vaccine Adjuvant U-Omp19.

Unlipidated outer membrane protein 19 (U-Omp19) is a novel mucosal adjuvant in preclinical development to be used in vaccine formulations. U-Omp19 holds two main properties, it is capable of inhibiting gastrointestinal and lysosomal peptidases, increasing the amount of co-administered antigen that reaches the immune inductive sites and its half-life inside cells, and it is able to stimulate antigen presenting cells in vivo. These activities enable U-Omp19 to enhance the adaptive immune response to co-administrated antigens. To characterize the stability of U-Omp19 we have performed an extensive analysis of its physicochemical and biological properties in a 3-year long-term stability study, and under potentially damaging freeze-thawing and lyophilization stress processes. Results revealed that U-Omp19 retains its full protease inhibitor activity, its monomeric state and its secondary structure even when stored in solution for 36 months or after multiple freeze-thawing cycles. Non-enzymatic hydrolysis resulted the major degradation pathway for storage in solution at 4 °C or room temperature which can be abrogated by lyophilization yet increasing protein tendency to form aggregates. This information will play a key role in the development of a stable formulation of U-Omp19, allowing an extended shelf-life during manufacturing, storage, and shipping of a future vaccine containing this pioneering adjuvant.

N,N,N-trimethylchitosan-poly (n-butylcyanoacrylate) core-shell nanoparticles as a potential oral delivery system for acyclovir.

This study investigated the feasibility of polysaccharide-coated poly(n-butyl cyanoacrylate) (PBCA) nanoparticles for oral delivery of acyclovir (ACV). PBCA nanoparticles were obtained by the emulsion polymerization method. Chitosan was chemically modified to obtain N,N,N-trimethylchitosan (TMC), which was used to coat the nanoparticles (PBCA-TMC). Nanoparticles were characterized by dynamic light scattering, zeta potential, differential scanning calorimetry (DSC), atomic force microscopy (AFM), cytotoxicity, and the effect on the transepithelial electrical resistance (TEER) of the Caco-2 cells. The size of the coated nanoparticles (296.2 nm) was significantly larger than uncoated (175.0 nm). Furthermore, PBCA nanoparticles had a negative charge (-11.7 mV), which was inverted to highly positive values (+36.5 mV) after coating. DSC analysis suggested the occurrence of the coating, which was confirmed by AFM images. The MTT assay revealed concentration-dependent cytotoxicity for the core-shell nanoparticles. Additionally, PBCA-TMC caused a significant but reversible decrease in the Caco-2 cell monolayer TEER. Entrapped ACV (PBCA-ACV-TMC), a Biopharmaceutical Classification System class III drug substance, increased approximately 3.25 times the Papp of ACV in the Caco-2 permeability assay. The nanoparticles were also able to provide in vitro ACV controlled release using media with different pH values (1.2; 6.8; 7.4). Accordingly, this new core-shell nanoparticle showed the potential to improve the oral delivery of ACV.

Oral Treatment of Spontaneously Hypertensive Rats with Captopril-Surface Functionalized Furosemide-Loaded Multi-Wall Lipid-Core Nanocapsules.

Multi-wall lipid-core nanocapsule (MLNC) functionalized with captopril and nanoencapsulating furosemide within the core was developed as a liquid formulation for oral administration. The nanocapsules had mean particle size below 200 nm, showing unimodal and narrow size distributions with moderate dispersity (laser diffraction and dynamic light scattering). Zeta potential was inverted from -14.3 mV [LNC-Fur(0,5)] to +18.3 mV after chitosan coating. Transmission electron microscopy and atomic force microscopy showed spherical structures corroborating the nanometric diameter of the nanocapsules. Regarding the systolic pressure, on the first day, the formulations showed antihypertensive effect and a longer effect than the respective drug solutions. When both drugs were associated, the anti-hypertensive effect was prolonged. On the fifth day, a time effect reduction was observed for all treatments, except for the nanocapsule formulation containing both drugs [Capt(0.5)-Zn(25)-MLNC-Fur(0.45)]. For diastolic pressure, only Capt(0.5)-Zn(25)-MLNC-Fur(0.45) presented a significant difference (p < 0.05) on the first day. On the fifth day, both Capt(0.5)-MLNC-Fur(0.45) and Capt(0.5)-Zn(25)-MLNC-Fur(0.45) had an effect lasting up to 24 h. The analysis of early kidney damage marker showed a potential protection in renal function by Capt(0.5)-Zn(25)-MLNC-Fur(0.45). In conclusion, the formulation Capt(0.5)-Zn(25)-MLNC-Fur(0.45) proved to be suitable for hypertension treatment envisaging an important innovation.

Unmet Quality Needs in Oral Drug Delivery: Contrasts of Drug Content and Uniformity on Distinct Approaches for Achieving Individual Dosing.

Individualized dosing is often required in pharmacotherapy, particularly for pediatric and geriatric patients and adjustment of drugs that demand dose adaptation. This study aimed to evaluate critical quality attributes (CQAs) of doses obtained by distinct approaches for achieving individual dosing. Approaches were evaluated as follows: subdivision of tablets by splitter and hand (haloperidol) and delivery by plastic dropper bottle (haloperidol), glass dropper bottle (clonazepam), dosing cup (sodium valproate), and dosing syringe (carbamazepine), including brand name, generic, and similar marketed products. Measuring devices were packaged with their respective product. Drug content uniformity was assessed to each substance according to pharmacopeial methods. Tablets subdivided by splitter had the poorest performance among all approaches, in which doses ranged around 60% of the labeled amount (Acceptance Value = 58.1 and RSD = 23.2%). The greatest performances were observed for the dosing syringe which fulfilled all the requirements for dose precision and for the glass dropper bottle. There were significant differences in dose delivery between manufacturers of the same medicine when measuring the same volume or number of drops. High drug content variability is extremely harmful to pharmacotherapy and may result in therapeutic failure or toxicity. It is crucial that measuring devices and scoring of tablets be checked for functionality and standardized for different manufacturers of the same medicine. Part of the approaches for achieving individual dosing did not meet the quality needs for drug content and uniformity. Yet, our findings show that more accurate and precise dosing can be accessed when using the dosing syringe and glass dropper bottle.

Enzymatically cross-linked arabinoxylan microspheres as oral insulin delivery system.

Arabinoxylans (AX) microspheres with different insulin/AX mass ratio were prepared by formation of phenoxy radical issued from the ferulic acid by enzymatic oxidation (entrapped in situ of insulin). Phenolic acid content and FT-IR spectrum of unloaded and insulin-loaded AX microspheres revealed that the phenoxy radical issued from the ferulic acid by enzymatic oxidation did not interact covalently with insulin. The microspheres showed a spherical shape, smooth surface and an average diameter of particles of 320 µm. In vitro control release found that AX microspheres minimized the insulin loss in the upper GI tract, retaining high percentage (~75%) of insulin in its matrix. The stability of the secondary structure of insulin was studied by dichroism circular (CD). The CD spectra of insulin released from AX microspheres did not change according to the insulin/AX mass ratio of the microsphere. Significant hypoglycemic effects with improved insulin-relative bioavailability tested on an in vivo murine model revealed the efficacy of these enzymatically cross-linked arabinoxylans microspheres as a new oral insulin carrier.

Preparation and Characterization of Poloxamer 407 Solid Dispersions as an Alternative Strategy to Improve Benznidazole Bioperformance.

Benznidazole (BZL), the first line drug for Chagas disease treatment, presents a low solubility, limiting the possibilities for its formulation. In this work, solid dispersions' (SDs) technology was exploited to increase BZL kinetic solubility and dissolution rate, seeking for an improvement in its bioperformance. A physical mixture (PM) and an SD using Poloxamer 407 as carrier were prepared and characterized. Dissolution tests were performed, and data were analyzed with the lumped model, which allowed to calculate different parameters of pharmaceutical relevance. A bioactivity assay was also carried out to probe the SD anti-trypanocidal activity. Among the most relevant results, the initial dissolution rate of the BZL SD was near 3, 4 and about 400-fold faster than the PM, a commercial formulation (CF) and an extracted BZL, respectivley. The times needed for an 80% of drug dissolution were 3.6 (SD), 46.4 (PM), and 238.7 min (CF); while the dissolution efficiency values at 30 min were 85.2 (SD), 71.2 (PM), and 65.0% (CF). Survival curves suggested that using Poloxamer 407 as carrier did not alter the anti-trypanocidal activity of BZL. These results allow to conclude that SDs can be an effective platform for immediate release of BZL in an oral administration.

Delivering Resveratrol on the Buccal Mucosa Using Mucoadhesive Tablets: A Potential Treatment Strategy for Inflammatory Oral Lesions.

BACKGROUND: Resveratrol is a polyphenol that has gained momentum in therapeutics in the last few years. OBJECTIVE: In this study, we hypothesised that resveratrol could act prophylactically and/or treat inflammatory lesions of the oral cavity after being delivered by two different formulations of buccal mucoadhesive tablets (F1 and F2, which differed in terms of viscosity agents used). METHODS: This hypothesis was assessed through permeation studies, to verify diffusion of the drug through the buccal mucosa using a porcine model to predict human in vivo behaviour. RESULTS: F2 (tablet with less viscosity agent) presented better permeation than F1, but the amount of drug that crossed the mucosa was still low compared to the amount retained within it (35.90 µg found in the receptor medium and 15.63 mg quantified within the mucosa). CONCLUSION: This accounted for a local effect rather than a systemic one, which is desirable for local processes, such as oral mucositis, lichen planus, erythema multiforme, nicotinic stomatitis, recurrent aphthous stomatitis, among others. In this sense, resveratrol-loaded mucoadhesive tablets appear to be a prominent alternative to prevent and/or cure inflammatory lesions of the oral cavity.

Self-aggregates of 3,6-O,O'-dimyristoylchitosan derivative are effective in enhancing the solubility and intestinal permeability of camptothecin.

The aim of this work was to investigate the potential of a new 3,6-O,O'-dimyristoyl derivative amphiphilic chitosan (DMCh), in improving the solubility of camptothecin (CPT), a hydrophobic anticancer drug, and its potential oral delivery. FTIR, 1H NMR and solid-state 13C NMR spectroscopy were used to characterize DMCh and to determine its average degree of substitution (DS¯=6.8%). DMCh/CPT micelles size ranged from (281-357nm), zeta potential (+32-50mV) of encapsulation efficiency of 42-100%. The in vitro cell viability showed that DMCh/CPT micelles were able to reduce the toxicity of CPT. The in vitro permeability of CPT through Caco-2 and Caco-2/HT29-MTX intestinal models was increased up to ten fold when formulated into DMCh micelles, underlining the mucoadhesive properties of the nanocarrier. DMCh/CPT micelles are able to enhance CPT solubility and bioavailability while reduce its cytotoxicity, showing the great potential for intestinal delivery of hydrophobic drugs.

Mucoadhesive Properties of Thiolated Pectin-Based Pellets Prepared by Extrusion-Spheronization Technique.

The aim of this study was to develop mucoadhesive pellets on a thiolated pectin base using the extrusion-spheronization technique. Thiolation of pectin was performed by esterification with thioglycolic acid. The molecular weight and thiol group content of the pectins were determined. Pellets containing pectin, microcrystalline cellulose, and ketoprofen were prepared and their mucoadhesive properties were evaluated through a wash-off test using porcine intestinal mucosa. The in vitro ketoprofen release was also evaluated. Thiolated pectin presented a thiol group content of 0.69 mmol/g. Thiolation caused a 13% increase in polymer molecular weight. Pellets containing thiolated pectin were still adhering to the intestinal mucosa after 480 min and showed a more gradual release of ketoprofen. Conversely, pellets prepared with nonthiolated pectin showed rapid disintegration and detached after only 15 min. It can be concluded that thiolated pectin-based pellets can be considered a potential platform for the development of mucoadhesive drug delivery systems for the oral route.

Development of Nutraceutical Emulsions as Risperidone Delivery Systems: Characterization and Toxicological Studies.

Emulsions are gaining increasing interest to be applied as drug delivery systems. The main goal of this work was the formulation of an oil/water nutraceutical emulsion (NE) for oral administration, enriched in omega 3 (ω3) and omega 6 (ω6), and able to encapsulate risperidone (RISP), an antipsychotic drug widely used in the treatment of autism spectrum disorders (ASD). RISP has low solubility in aqueous medium and poor bioavailability because of its metabolism and high protein binding. Coadministration of ω3, ω3, and vitamin E complexed with RISP might increase its bioavailability and induce a synergistic effect on the treatment of ASD. Here, we developed an easy and quick method to obtain NEs and then optimized them. The best formulation was chosen after characterization by particle size, defects of the oil-in-water interface, zeta potential (ZP), and in vitro drug release. The formulation selected was stable over time, with a particle size of around 3 µm, a ZP lower than -20 mV and controlled drug release. To better understand the biochemical properties of the formulation obtained, we studied in vitro toxicity in the Caco-2 cell line. After 4 h of treatment, an increase in cellular metabolism was observed for all RISP concentrations, but emulsions did not change their metabolic rate, except at the highest concentration without drug (25 µg/mL), which showed a significant reduction in metabolism respect to the control. Additionally, locomotor activity and heart rate in zebrafish were measured as parameters of in vivo toxicity. Only the highest concentration (0.625 µg/mL) showed a cardiotoxic effect, which corresponds to the decrease in spontaneous movement observed previously. As all the materials contained in the formulations were US FDA approved, the NE selected would be good candidate for clinical trials.

Orally disintegrating films containing propolis: properties and release profile.

The objective of this work was the production and characterization of orally disintegrating films of gelatin and hydrolyzed collagen containing the ethanol extract of propolis. The films were produced by casting with different concentrations of hydrolyzed collagen with and without the extract. The mechanical properties, mucoadhesive properties, swelling degree, in vitro release kinetics, stability of active compounds, Fourier transform infrared spectroscopy (FTIR), and antimicrobial activity of the films were evaluated. The films with the highest concentration of hydrolyzed collagen were less resistant and more elastic, and films containing the extract were more resistant than the control. In addition, the films with the extract showed higher mucoadhesion, which is important for ensuring the release of active compounds in the oral cavity. Generally, all formulations showed a high swelling capacity, which may have contributed to the quick release also demonstrated by the release kinetics model. Interactions between the extract compounds and the polymeric matrix were observed by FTIR spectroscopy, which may have contributed to an improvement in the mechanical properties. Films containing the extract had good stability and effective antimicrobial properties against Staphylococcus aureus, which shows that these films can potentially be used to release active compounds in the oral mucosa.

Starch-based microspheres for sustained-release of curcumin: preparation and cytotoxic effect on tumor cells.

Curcumin (CUR) has been proved to be highly cytotoxic against different tumor cell lines. However, its poor solubility in aqueous medium and fast degradation in physiological pH are the common drawbacks preventing its efficient practical use. Herein, we report the development of original microspheres based on the biopolymer starch crosslinked with N,N-methylenebisacrylamide (MBA) to be applied as an efficient delivering system for CUR. The starch-based microspheres showed high loading efficiency even in loading solution with different CUR concentrations. In vitro release assays data showed that the CUR release is governed by anomalous transport (n=0.73) and it is pH-dependent. Cytotoxicity assays showed that starch microspheres could improve the cytotoxicity of CUR toward Caco-2 and HCT-116 tumor cell lines up to 40 times than that found for pure CUR. This behavior was attributed to the slowly and sustained release of CUR from the microspheres.