Antioxidant Efficacy and "In Vivo" Safety of a Bentonite/Vitamin C Hybrid.

L-ascorbic acid (LAA), commonly known as vitamin C, is an excellent and recognized antioxidant molecule used in pharmaceutical and cosmetic formulations. Several strategies have been developed in order to preserve its chemical stability, connected with its antioxidant power, but there is little research regarding the employment of natural clays as LAA host. A safe bentonite (Bent)-which was verified by in vivo ophthalmic irritability and acute dermal toxicity assays-was used as carrier of LAA. The supramolecular complex between LAA and clay may constitute an excellent alternative, since the molecule integrity does not seem to be affected, at least from the point of view of its antioxidant capacity. The Bent/LAA hybrid was prepared and characterized through ultraviolet (UV) spectroscopy, X-ray diffraction (XRD), infrared (IR) spectroscopy, thermogravimetric analysis (TG/DTG) and zeta potential measurements. Photostability and antioxidant capacity tests were also performed. The LAA incorporation into Bent clay was demonstrated, as well as the drug stability due to the Bent photoprotective effect onto the LAA molecule. Moreover, the antioxidant capacity of the drug in the Bent/LAA composite was confirmed.

Hybrid Lipid/Clay Carrier Systems Containing Annatto Oil for Topical Formulations.

Nanocomposites formed by clay and lipid carriers (NLCs) show a high potential for providing controlled release and specific delivery of bioactive molecules and have recently gained attention in the pharmaceutical sector due to their ability to transport hydrophilic and hydrophobic drugs. Recent studies have recognized the biological activity of the oil of Bixa orellana L. (AO) with regards to its healing, antioxidant, antibacterial, and anti-leishmanial properties. Therefore, the purpose of this study is the preparation and characterization of hybrid systems based on lipid nanocarriers and laponite for the delivery of AO. NLCs were prepared by the fusion-emulsification method, using cetyl palmitate (CP) or myristyl myristate (MM), AO, and Poloxamer 188. The morphology, hydrodynamic diameters, zeta potential (ZP), polydispersity index (PDI), thermal analysis, X-ray diffraction analysis (XRD), viscosity behavior, and cytotoxicity testing of the hybrid systems were performed. The thermal study and X-ray diffraction analyses (XRD) revealed polymorphic structural changes compatible with the amorphization of the material. Rheological assays highlighted a typical pseudoplastic behavior in all systems (MM and CP with LAP). The hybrid systems' morphology, size diameters, and PDIs were similar, preset spherical and monodisperse structures (≈200 nm; <0.3), without significant change up to sixty days. The ZP values differed from each other, becoming higher with increasing AO concentration. XEDS spectra and elemental X-ray maps show peaks of lipids (organic components, C and O) and inorganic components O, Mg, and Si. All samples showed cell viability above 60%. The results indicated a stable, biocompatible hybrid system that can be an alternative for topical application.

Correlation between Elemental Composition/Mobility and Skin Cell Proliferation of Fibrous Nanoclay/Spring Water Hydrogels.

Inorganic hydrogels formulated with spring waters and clay minerals are used to treat musculoskeletal disorders and skin affections. Their underlying mechanism of action for skin disorders is not clear, although it is usually ascribed to the chemical composition of the formulation. The aim of this study was to assess the composition and in vitro release of elements with potential wound healing effects from hydrogels prepared with two nanoclays and natural spring water. In vitro Franz cell studies were used and the element concentration was measured by inductively coupled plasma techniques. Biocompatibility studies were used to evaluate the potential toxicity of the formulation against fibroblasts. The studied hydrogels released elements with known therapeutic interest in wound healing. The released ratios of some elements, such as Mg:Ca or Zn:Ca, played a significant role in the final therapeutic activity of the formulation. In particular, the proliferative activity of fibroblasts was ascribed to the release of Mn and the Zn:Ca ratio. Moreover, the importance of formulative studies is highlighted, since it is the optimal combination of the correct ingredients that makes a formulation effective.

Safety of Nanoclay/Spring Water Hydrogels: Assessment and Mobility of Hazardous Elements.

The presence of impurities in medicinal products have to be controlled within safety limits from a pharmaceutical quality perspective. This matter is of special significance for those countries and regions where the directives, guidelines, or legislations, which prescribe the rules for the application of some products is quite selective or incomplete. Clay-based hydrogels are quite an example of this matter since they are topically administered, but, in some regions, they are not subjected to well-defined legal regulations. Since hydrogels establish an intimate contact with the skin, hazardous elements present in the ingredients could potentially be bioavailable and compromise their safety. The elemental composition and mobility of elements present in two hydrogels have been assessed. Sepiolite, palygorskite, and natural spring water were used as ingredients. The release of a particular element mainly depends on its position in the structure of the hydrogels, not only on its concentration in each ingredient. As a general trend, elements' mobility reduced with time. Among the most dangerous elements, whose presence in cosmetics is strictly forbidden by European legal regulations, As and Cd were mobile, although in very low amounts (0.1 and 0.2 µg/100 g of hydrogel, respectively). That is, assuming 100% bioavailability, the studied hydrogels would be completely safe at normal doses. Although there is no sufficient evidence to confirm that their presence is detrimental to hydrogels safety, legally speaking, their mobility could hinder the authorization of these hydrogels as medicines or cosmetics. In conclusion, the present study demonstrates that hydrogels prepared with sepiolite, palygorskite, and Alicún spring water could be topically applied without major intoxication risks.

Wound Healing Activity of Nanoclay/Spring Water Hydrogels.

BACKGROUND: hydrogels prepared with natural inorganic excipients and spring waters are commonly used in medical hydrology. Design of these clay-based formulations continues to be a field scarcely addressed. Safety and wound healing properties of different fibrous nanoclay/spring water hydrogels were addressed. METHODS: in vitro biocompatibility, by means of MTT assay, and wound healing properties were studied. Confocal Laser Scanning Microscopy was used to study the morphology of fibroblasts during the wound healing process. RESULTS: all the ingredients demonstrated to be biocompatible towards fibroblasts. Particularly, the formulation of nanoclays as hydrogels improved biocompatibility with respect to powder samples at the same concentration. Spring waters and hydrogels were even able to promote in vitro fibroblasts motility and, therefore, accelerate wound healing with respect to the control. CONCLUSION: fibrous nanoclay/spring water hydrogels proved to be skin-biocompatible and to possess a high potential as wound healing formulations. Moreover, these results open new prospects for these ingredients to be used in new therapeutic or cosmetic formulations.

Ground Calcium Carbonate as a Low Cost and Biosafety Excipient for Solubility and Dissolution Improvement of Praziquantel.

Calcium carbonate is an abundant mineral with several advantages to be a successful carrier to improve oral bioavailability of poorly water-soluble drugs, such as praziquantel. Praziquantel is an antiparasitic drug classified in group II of the Biopharmaceutical Classification System hence characterized by high-permeability and low-solubility. Therefore, the dissolution rate is the limiting factor for the gastrointestinal absorption that contributes to the low bioavailability. Consequently, the therapeutic dose of the praziquantel must be high and big tablets and capsules are required, which are difficult to swallow, especially for pediatric and elderly patients. Mixtures of praziquantel and calcium carbonate using solid-solid physical mixtures and solid dispersions were prepared and characterized using several techniques (X-ray diffraction differential scanning calorimetry, thermogravimetric analysis, scanning electron microscopy, laser diffraction, Fourier transform infrared and Raman spectroscopies). Solubility of these formulations evidenced that the solubility of praziquantel-calcium carbonate interaction product increased in physiological media. In vitro dissolution tests showed that the interaction product increased the dissolution rate of the drug in acidic medium. Theoretical models were studied to understand this experimental behavior. Cytotoxicity and cell cycle studies were performed, showing that praziquantel-calcium carbonate physical mixture and interaction product were biocompatible with the HTC116 cells, because it did not produce a decrease in cell viability or alterations in the cell cycle.

Montmorillonite-norfloxacin nanocomposite intended for healing of infected wounds.

Background: Chronic cutaneous wounds represent a major issue in medical care and are often prone to infections. Purpose: The aim of this study was the design of a clay mineral-drug nanocomposite based on montmorillonite and norfloxacin (NF, antimicrobial drug) as a powder for cutaneous application, to enhance wound healing in infected skin lesions. Methods: The nanocomposite has been prepared by means of an intercalation solution procedure. Adsorption isotherm, solid-state characterization of the nanocomposite, drug loading capacity and its release have been performed. Moreover, cytocompatibility, in vitro fibroblast proliferation and antimicrobial activity against Pseudomonas aeruginosa and Staphylococcus aureus were assessed. Results: The clay drug adsorption isotherm demonstrates that the mechanism of NF intercalation into montmorillonite galleries is the adsorption as one single process, due to the charge-charge interaction between protonated NF and negatively charged montmorillonite edges in the interlayer space. Nanocomposite is biocompatible and it is characterized by antimicrobial activity greater than the free drug: this is due to its nanostructure and controlled drug release properties. Conclusion: Considering the results obtained, NF-montmorillonite nanocomposite seems a promising tool to treat infected skin lesions or skin wounds prone to infection, as chronic ulcers (diabetic foot, venous leg ulcers) and burns.

Halloysite nanotubes as tools to improve the actual challenge of fixed doses combinations in tuberculosis treatment.

Halloysite nanotubes (HLNTs) were used as nanocarriers of the tuberculostatic agent isoniazid (INH), a BCS (Biopharmaceutics Classification System) class III drug. Self-assembling nanohybrids (INH-loaded HLNTs) with an average outer diameter of 90 nm and polydispersity index of 0.7 approximately, were obtained by spontaneous adsorption of INH molecules to HLNTs powder in aqueous medium. The nanohybrids were aimed to improve oral drug bioavailability and reduce physicochemical incompatibility of INH with other concomitantly administered tuberculostatic agents. In vitro drug release from INH-loaded HLNTs was successfully fitted to a diffusive kinetic law founded on the adsorption-desorption equilibrium between drug molecules in solution and solid inorganic excipients. INH-loaded HLNTs showed good in vitro biocompatibility toward Caco-2 cells at the concentrations studied (up to 1233 µg/mL), with improved cell proliferation. Permeability tests showed that INH transport across Caco-2 cellular membranes was greatly enhanced and fluorescent microscopy confirmed that the drug encapsulated into nanohybrid was effectively internalized by the cells. INH-loaded HLNTs enhanced stability of the drug in presence of other tuberculostatic agents, both in binary and quaternary combinations. It has been demonstrated that simple interaction between INH with HLNTs leads to drug permeability and stability improvements that could greatly facilitate the design of multiple drug dosage forms, an actual challenge in oral treatment of tuberculosis. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2019.

Conformational polymorphic changes in the crystal structure of the chiral antiparasitic drug praziquantel and interactions with calcium carbonate.

Praziquantel is an antiparasitic drug used for decades. Currently, the praziquantel commercial preparation is a racemic mixture, in which only the levo-enantiomer possesses anthelmintic activity. The knowledge of its properties in the solid state and other chemical-physical properties is necessary for improving its efficacy and applications. Drug solid dispersions were prepared with calcium carbonate at 1:5 drug to excipient weight ratio by solvent evaporation method. Then, the modification of the crystal structure of the racemic polymorph of praziquantel in presence of calcium carbonate has been studied by means of several analytical techniques (DSC, TGA, XRD, SEM, FTIR, Raman spectroscopy and chiral liquid chromatography). This study has been completed with atomistic calculations based on empirical interatomic force fields and quantum mechanics methods applied to the crystal structure of praziquantel and of intermolecular interactions. The results evidenced that calcium carbonate provoked a conformational change in the praziquantel molecule yielding the formation of different polymorphs of praziquantel crystal. These alterations were not observed replacing calcium carbonate with colloidal silica as excipient in the solid dispersion.

Folk pharmaceutical formulations in western Mediterranean: identification and safety of clays used in pelotherapy.

ETHNOPHARMACOLOGICAL RELEVANCE: Clays are naturally occurring ingredients of many natural health products, being included in most of ancient Mediterranean/European medical texts and currently used to prepare therapeutic hot-muds (peloids) in several thermal stations of the Mediterranean region. Clays are included in the formulation of peloids as vehicles of the mineral-medicinal water, to obtain inorganic gels with rheological and thermal properties suitable to be topically applied. Knowledge about formulations and preparation procedures of these traditional medicines has been orally transmitted since ancient times. Increasing recognition of the therapeutic utility of these traditional and natural health care substances make necessary a full ethnopharmaceutic research to ascertain those compositional characters that allow to establish quality attributes and corresponding requirements for these materials and products, including identity, purity, richness and safety. MATERIALS AND METHODS: Five clay samples (A, B, C, D and E) currently used in various spa centers of southern European/Mediterranean countries were studied. X-Ray diffraction (XRD) and X-ray fluorescence (XRF) data were used to asses sample identity and richness. Elemental impurities and microbiological contaminants were also determined and compared to normative limits. Particle size distribution was related to their safety as powder materials. RESULTS: Samples A, C, D and E were identified as "high purity clay", while sample B was identified as a mix of clay minerals and carbonates. The presence of carbonates in this sample could compromise its suitability for pelotherapy. The studied clays meet the main normative limits for metals impurities, with the exception of arsenic in sample A and nickel in sample B. The samples comply with the microbiological limits proposed by European legislation for medicinal products. According to the particle size of the studied samples, prevention and control of dust exposure must be considered. CONCLUSIONS: Despite their demonstrated longevity, the use of clays in traditional medicine formulations as peloids greatly requires comprehension of their identity and safety attributes. Continuity of these mineral substances as recognized health care ingredients oblige to conduct interdisciplinary research to know the features that sustain their traditional use in the preparation of medicines (ethnopharmaceutics).

Montmorillonite-chitosan-silver sulfadiazine nanocomposites for topical treatment of chronic skin lesions: in vitro biocompatibility, antibacterial efficacy and gap closure cell motility properties.

Silver compounds and especially silver sulfadiazine (AgSD) are reported as effective antimicrobial agents against almost all known bacteria, fungi and some viruses. However, AgSD has been shown to be cytotoxic toward fibroblasts and keratinocytes in vitro and consequently to retard wound healing in vivo. The aim of the present work was to evaluate the in vitro biocompatibility (cytotoxicity and proliferation), antimicrobial efficacy and cell motility gap closure (assay of wound closure) of MT/CS nanocomposites loaded with silver sulfadiazine (AgSD). It is envisioned to be administered as a powder or a dressing for cutaneous application in the treatment of skin ulcers. The loading of AgSD in MT/CS nanocomposites aimed at preventing the delay in wound healing, by decreasing the cytotoxicity of AgSD and maintaining its antimicrobial properties. Nanocomposites were prepared by using different amounts of MT (100-2000 mg) and 40 ml of a 1% (w/w) chitosan glutamate aqueous solution. The relative amounts of AgSD and chitosan in the systems were assessed by suitable analytic methods. The nanocomposite prepared using 100mg of MT was characterized for in vitro biocompatibility and proliferation and for wound healing using normal human dermal fibroblasts (NHDF). Antimicrobial properties were evaluated against four reference bacterial strains: Staphylococcus aureus, Streptococcus pyogenes, Escherichia coli, and Pseudomonas aeruginosa. AgSD loaded in the 100 MT/CS nanocomposite showed good in vitro biocompatibility and gap closure properties (fibroblasts) and maintained AgSD antimicrobial properties, especially against P. aeruginosa, that often complicates skin lesions.

Solid state characterisation of silver sulfadiazine loaded on montmorillonite/chitosan nanocomposite for wound healing.

Biopolymer chitosan/montmorillonite nanocomposites loaded with silver sulfadiazine for wound healing purposes were prepared via intercalation solution technique. Structure and morphology of loaded nanocomposites were studied and compared with pure components and unloaded nanocomposites. X-ray diffraction, Fourier transformed infrared spectroscopy, high resolution transmission electron microscopy coupled with energy-dispersion X-ray analysis, thermal and elemental analysis were employed for the characterisation. The results confirmed that the drug was effectively loaded in the three-dimensional nanocomposite structures, in which chitosan chains were adsorbed in monolayers into the clay mineral interlayer spaces.

Intestinal permeability of oxytetracycline from chitosan-montmorillonite nanocomposites.

A nanocomposite based on chitosan and montmorillonite was developed as carrier to improve oral bioavailability of oxytetracycline. The nanocomposite was prepared by simple solid-liquid interaction and loaded with the drug. The loaded nanocomposite was characterized by X-ray powder diffraction, thermal analysis, FTIR spectroscopy and zeta potential. Caco-2 cell cultures were used to evaluate in vitro cytotoxicity and drug permeation. Confocal laser scanning microscopy was also performed to evaluate the eventual entrapment of drug into the Caco-2 cells. Results showed that the nanocomposite was internalized into the cells and effectively enhanced drug permeation, being also biocompatible towards Caco-2 cells.

Networking and rheology of concentrated clay suspensions "matured" in mineral medicinal water.

This work studied the influence of "maturation" conditions (time and agitation) on aggregation states, gel structure and rheological behaviour of a special kind of pharmaceutical semisolid products made of concentrated clay suspensions in mineral medicinal water. Maturation of the samples was carried out in distilled and sulphated mineral medicinal water, both in static conditions (without agitation) and with manual stirring once a week, during a maximum period of three months. At the measured pH interval (7.5-8.0), three-dimensional band-type networks resulting from face/face contacts were predominant in the laminar (disc-like) clay suspensions, whereas the fibrous (rod-like) particles formed micro-aggregates by van der Waals attractions. The high concentration of solids in the studied systems greatly determined their behaviour. Rod-like sepiolite particles tend to align the major axis in aggregates promoted by low shearing maturation, whereas aggregates of disc-like smectite particles did not have a preferential orientation and their complete swelling required long maturation time, being independent of stirring. Maturation of both kinds of suspensions resulted in improved rheological properties. Laminar clay suspensions became more structured with time, independently from static or dynamic maturation conditions, whereas for fibrous clay periodic agitation was also required. Rheological properties of the studied systems have been related to aggregation states and networking mechanisms, depending on the type of clay minerals constituents. Physical stability of the suspensions was not impaired by the specific composition of the Graena medicinal water.

Supramolecular structure of 5-aminosalycilic acid/halloysite composites.

This paper assesses the supramolecular structure of nanocomposites prepared by including the anti-inflammatory drug 5-aminosalycilic acid in halloysite nanotubes. Halloysite tubes have sub-micron individual lengths with outer diameters ∼0.1 µm, as observed by FESEM. The mercury intrusion plots showed bimodal profiles with pore dimensions ∼10 and 0.06 µm. X-ray diffraction and thermogravimetric results revealed changes in the hydration form of the clay after the interaction. The groups associated to the interaction were studied by FTIR. The location of the drug in the composites was determined after uranium staining of its amino groups by X-EDS microanalysis coupled with HREM. The drug was located both inside and on the surface of the halloysite nanotubes. These results confirm the occurrence of two concomitant interaction mechanisms: rapid adsorption of 5-ASA at the external halloysite surface followed by slow adsorption of the drug inside the tubes.