Development of alginate film filled with halloysite-carbon dots for active food packaging.

The development of functional bionanocomposites for active food packaging is of current interest to replace non-biodegradable plastic coatings. In the present work, we report the synthesis of an alginate-based nanocomposite filled with modified halloysite nanotubes (HNTs) to develop coatings with improved barrier properties for food packaging. Firstly, HNTs were chemically modified by the introduction of carbon dots units (CDs) onto their external surface (HNTs-CDs) obtaining a nanomaterial where CDs are uniformly present onto the tubes as verified by morphological investigations, with good UV absorption and antioxidant properties. Afterwards, these were dispersed in the alginate matrix to obtain the alginate/HNTs-CDs nanocomposite (Alg/HNTs-CDs) whose morphology was imaged by AFM measurements. The UV and water barrier properties (in terms of moisture content and water vapor permeability) were investigated, and the antioxidant properties were evaluated as well. To confer some antimicrobial properties to the final nanocomposite, the synthetized filler was loaded with a natural extract (E) from M. cisplatensis. Finally, the extract kinetic release both from the filler and from the nanocomposite was studied in a medium mimicking a food simulant and preliminary studies on the effect of Alg/HNTs-CDs/E on coated and uncoated fruits, specifically apples and bananas were also carried out.

Kinetic Study and Reaction Mechanism of the Gas-Phase Thermolysis Reaction of Methyl Derivatives of 1,2,4,5-Tetroxane.

Tetroxane derivatives are interesting drugs for antileishmaniasis and antimalaric treatments. The gas-phase thermal decomposition of 3,6,-dimethyl-1,2,4,5-tetroxane (DMT) and 3,3,6,6,-tetramethyl-1,2,4,5-tetroxane (acetone diperoxide (ACDP)) was studied at 493-543 K by direct gas chromatography by means of a flow reactor. The reaction is produced in the injector chamber at different temperatures. The resulting kinetics Arrhenius equations were calculated for both tetroxanes. Including the parent compound of the series 1,2,4,5-tetroxane (formaldehyde diperoxide (FDP)), the activation energy and frequency factors decrease linearly with the number of methyl groups. The reaction mechanisms of ACDP and 3,6,6-trimethyl-1,2,4,5-tetroxane (TMT) decomposition have been studied by means of the DFT method with the BHANDHLYP functional. Our calculations confirm that the concerted mechanism should be discarded and that only the stepwise mechanism occurs. The critical points of the singlet and triplet state potential energy surfaces (S- and T-PES) of the thermolysis reaction of both compounds have been determined. The calculated activation energies of the different steps vary linearly with the number of methyl groups of the methyl-tetroxanes series. The mechanism for the S-PES leads to a diradical O···O open structure, which leads to a C···O dissociation in the second step and the production of the first acetaldehyde/acetone molecule. This last one yields a second C···O dissociation, producing O2 and another acetone/acetaldehyde molecule. The O2 molecule is in the singlet state. A quasi-parallel mechanism for the T-PES from the open diradical to products is also found. Most of the critical points of both PES are linear with the number of methyl groups. Reaction in the triplet state is much more exothermic than the singlet state mechanism. Transitions from the singlet ground state, S0 and low-lying singlet states S1-3, to the low-lying triplet excited states, T1-4, (chemical excitation) in the family of methyl tetroxanes are also studied at the CASSCF/CASPT2 level. Two possible mechanisms are possible here: (i) from S0 to T3 by strong spin orbit coupling (SOC) and subsequent fast internal conversion to the excited T1 state and (ii) from S0 to S2 from internal conversion and subsequent S2 to T1 by SOC. From these experimental and theoretical results, the additivity effect of the methyl groups in the thermolysis reaction of the methyl tetroxane derivatives is clearly highlighted. This information will have a great impact for controlling these processes in the laboratory and chemical industries.

Hectorite/Phenanthroline-Based Nanomaterial as Fluorescent Sensor for Zn Ion Detection: A Theoretical and Experimental Study.

The development of fluorescent materials that can act as sensors for the determination of metal ions in biological fluids is important since they show, among others, high sensitivity and specificity. However, most of the molecules that are used for these purposes possess a very low solubility in aqueous media, and, thus, it is necessary to adopt some derivation strategies. Clay minerals, for example, hectorite, as natural materials, are biocompatible and available in large amounts at a very low cost that have been extensively used as carrier systems for the delivery of different hydrophobic species. In the present work, we report the synthesis and characterization of a hectorite/phenanthroline nanomaterial as a potential fluorescent sensor for Zn ion detection in water. The interaction of phenanthroline with the Ht interlaminar space was thoroughly investigated, via both theoretical and experimental studies (i.e., thermogravimetry, FT-IR, UV-vis and fluorescence spectroscopies and XRD measurements), while its morphology was imaged by scanning electron microscopy. Afterwards, the possibility to use it as sensor for the detection of Zn2+ ions, in comparison to other metal ions, was investigated through fluorescent measurements, and the stability of the solid Ht/Phe/Zn complex was assessed by different experimental and theoretical measurements.

Comparison of Synthetic Pathways for Obtaining Fluorescent Nanomaterials Based on Halloysite and Carbon Dots for Potential Biological Sensing.

Recently, fluorescent sensors have gained considerable attention due to their high sensitivity, low cost and noninvasiveness. Among the different materials that can be used for this purpose, carbon dots (CDs) represent valuable candidates for applications in sensing. These, indeed, are easily synthesized, show high quantum yield and are highly biocompatible. However, it was pointed out that the photoluminescence properties of these nanomaterials are strictly dependent on the synthetic and purification methods adopted. The presence of halloysite nanotubes (HNTs), a natural, low cost and biocompatible clay mineral, has been found to be efficient in obtaining small and highly monodispersed CDs without long and tedious purification techniques. Herein, we report the comparison of synthetic pathways for obtaining halloysite-N-doped CDs (HNTs-NCDs) that could be used in biological sensing. One was based on the synthesis of N-doped CDs by a bottom-up approach on HNTs' surface by a MW pyrolysis process; the other one was based on the post-modification of pristine N-doped CDs with halloysite derivatives. The evaluation of the best synthetic route was performed by different physico-chemical techniques. It was found that the bottom-up approach led to the formation of N-doped CDs with different functional groups onto the HNTs' surface. This evidence was also translated in the different fluorescence quantum yields and the existence of several functional groups in the obtained materials was investigated by potentiometric titrations. Furthermore, the ability of the synthesized nanomaterials as sensors for Fe3+ ions detection was assessed by spectroscopic measurements, and the cellular uptake was verified by confocal/fluorescence microscopies as well.

Carrier capability of halloysite nanotubes for the intracellular delivery of antisense PNA targeting mRNA of neuroglobin gene.

Peptide nucleic acid (PNA) is a DNA mimic that shows good stability against nucleases and proteases, forming strongly recognized complementary strands of DNA and RNA. However, due to its feeble ability to cross the cellular membrane, PNA activity and its targeting gene action is limited. Halloysite nanotubes (HNTs) are a natural and low-cost aluminosilicate clay. Because of their peculiar ability to cross cellular membrane, HNTs represent a valuable candidate for delivering genetic materials into cells. Herein, two differently charged 12-mer PNAs capable of recognizing as molecular target a 12-mer DNA molecule mimicking a purine-rich tract of neuroglobin were synthetized and loaded onto HNTs by electrostatic attraction interactions. After characterization, the kinetic release was also assessed in media mimicking physiological conditions. Resonance light scattering measurements assessed their ability to bind complementary single-stranded DNA. Furthermore, their intracellular delivery was assessed by confocal laser scanning microscopy on living MCF-7 cells incubated with fluorescence isothiocyanate (FITC)-PNA and HNTs labeled with a probe. The nanomaterials were found to cross cellular membrane and cell nuclei efficiently. Finally, it is worth mentioning that the HNTs/PNA can reduce the level of neuroglobin gene expression, as shown by reverse transcription-quantitative polymerase chain reaction and western blotting analysis.

Thixotropic Hydrogels Based on Laponite® and Cucurbituril for Delivery of Lipophilic Drug Molecules.

Invited for this month's cover are the collaborating groups of Prof. Serena Riela at University of Catania, Prof. César Viseras at University of Granada and Dr. Ignacio Sainz-Diaz at Instituto Andaluz de Ciencias de la Tierra. The cover picture shows the possible application of the developed system. In particular, flufenamic acid, anti-inflammatory and anti-pyretic drug, was complexed into cucurbituril cavity and the supramolecular system obtained was used as filler for laponite® hydrogel for its topical delivery. More information can be found in the Research Article by Viseras-Iborra, Riela, and co-workers.

Thixotropic Hydrogels Based on Laponite® and Cucurbituril for Delivery of Lipophilic Drug Molecules.

Nowadays the use of hydrogels for biomedical purposes is increasing because of their interesting features that allow the development of targeted drug delivery systems. Herein, hydrogel based on Laponite® (Lap) clay mineral as gelator and cucurbit[6]uril (CB[6]) molecules were synthetized for the delivery of flufenamic acid (FFA) for potential topical application. Firstly, the interaction between CB[6] and FFA was assessed by UV-vis spectroscopic measurements and molecular modeling calculations. Then, the obtained complex was used as filler for Lap hydrogel (Lap/CB[6]/FFA). The properties of the hydrogel in terms of viscosity and, self-repair abilities were investigated; its morphology was imaged by scanning electron and polarized optical microscopies. Furthermore, the changes in the hydrodynamic radii and in the colloidal stability of CB[6]/Lap mixture were investigated in terms of translational diffusion from dynamic light scattering and ζ-potential measurements. Finally, the kinetic in vitro release of FFA, from Lap/CB[6]/FFA hydrogel, was studied in a medium mimicking the pH of skin and the obtained results were discussed both by an experimental point of view and by molecular modeling calculations.

The Optimization of Pressure-Assisted Microsyringe (PAM) 3D Printing Parameters for the Development of Sustainable Starch-Based Patches.

The aim of this work was to develop sustainable patches for wound application, using the biopolymer starch, created using a low-cost 3D printing PAM device. The composition of a starch gel was optimized for PAM extrusion: corn starch 10% w/w, ß-glucan water suspension (filler, 1% w/w), glycerol (plasticizer, 29% w/w), and water 60% w/w. The most suitable 3D printing parameters were optimized as well (nozzle size 0.8 mm, layer height 0.2 mm, infill 100%, volumetric flow rate 3.02 mm3/s, and print speed 15 mm/s). The suitable conditions for post-printing drying were set at 37 °C for 24 h. The obtained patch was homogenous but with low mechanical resistance. To solve this problem, the starch gel was extruded over an alginate support, which, after drying, becomes an integral part of the product, constituting the backing layer of the final formulation. This approach significantly improved the physicochemical and post-printing properties of the final bilayer patch, showing suitable mechanical properties such as elastic modulus (3.80 ± 0.82 MPa), strength (0.92 ± 0.08 MPa), and deformation at break (50 ± 1%). The obtained results suggest the possibility of low-cost production of patches for wound treatment by additive manufacturing technology.

Nanoformulations based on collagenases loaded into halloysite/Veegum® clay minerals for potential pharmaceutical applications.

The design and development of nanomaterials capable of penetrate cancer cells is fundamental when anticancer therapy is involved. The use of collagenase (Col) is useful since this enzyme can degrade collagen, mainly present in the tumor extracellular matrix. However, its use is often limited since collagenase suffers from inactivation and short half-life. Use of recombinant ultrapure collagenase or carrier systems for their delivery are among the strategies adopted to increase the enzyme stability. Herein, based on the more stability showed by recombinant enzymes and the possibility to use them in anticancer therapy, we propose a novel strategy to further increase their stability by using halloysite nanotubes (HNTs) as carrier. ColG and ColH were supramolecularly loaded onto HNTs and used as fillers for Veegum gels. The systems could be used for potential local administration of collagenases for solid tumor treatment. All techniques adopted for characterization showed that halloysite interacts with collagenases in different ways depending with the Col considered. Furthermore, the hydrogels showed a very slow release of the collagenases within 24 h. Finally, biological assays were performed by studying the digestion of a type-I collagen matrix highlighting that once released the Col still possessed some activity. Thus we developed carrier systems that could further increase the high recombinant collagenases stability, preventing their inactivation in future in vivo applications for potential local tumor treatment.

Polymeric Patches Based on Chitosan/Green Clay Composites and Hazelnut Shell Extract as Bio-Sustainable Medication for Wounds.

Hazelnut shells, the main waste deriving from hazelnut processing, represent an interesting source of active molecules useful in pharmaceutics, although they have not yet been examined in depth. A hydrosoluble extract (hazelnut shell extract, HSE) was prepared by the maceration method using a hydroalcoholic solution and used as the active ingredient of patches (prepared by casting method) consisting of composites of highly deacetylated chitosan and green clay. In vitro studies showed that the formulation containing HSE is able to stimulate keratinocyte growth, which is useful for healing purposes, and to inhibit the growth of S. aureus (Log CFU/mL 0.95 vs. 8.85 of the control after 48 h); this bacterium is often responsible for wound infections and is difficult to treat by conventional antibiotics due to its antibiotic resistance. The produced patches showed suitable tensile properties that are necessary to withstand mechanical stress during both the removal from the packaging and application. The obtained results suggest that the developed patch could be a suitable product to treat wounds.

Exploiting the interaction between halloysite and charged PNAs for their controlled release.

The design and development of nanomaterials that could be used in nanomedicine are of fundamental importance to obtain smart nanosystems for the treatment of several diseases. Halloysite, because of its interesting features, represents a suitable nanomaterial for the delivery of different biologically active species. Among them, peptide nucleic acids (PNAs) have attracted considerable attention in recent decades for their potential applications in both molecular antisense diagnosis and as therapeutic agents, although up to now, the actual clinical applications have been very limited. Herein we report a systematic study on the supramolecular interaction of three differently charged PNAs with halloysite. Understanding the interaction mode of charged molecules with the clay surfaces represents a key factor for the future design and development of halloysite based materials which could be used for the delivery and subsequent intracellular release of PNA molecules. Thus, three different PNA tetramers, chosen as models, were synthesized and loaded onto the clay. The obtained nanomaterials were characterized using spectroscopic studies and thermogravimetric analysis, and their morphologies were studied using high angle annular dark field transmission electron microscopy (HAADF/STEM) coupled with Energy Dispersive X-ray spectroscopy (EDX). The aqueous mobility of the three different nanomaterials was investigated by dynamic light scattering (DLS) and ζ-potential measurements. The release of PNA tetramers from the nanomaterials was investigated at two different pH values, mimicking physiological conditions. Finally, to better understand the stability of the synthesized PNAs and their interactions with HNTs, molecular modelling calculations were also performed. The obtained results showed that PNA tetramers interact in different ways with HNT surfaces according to their charge which influences their kinetic release in media mimicking physiological conditions.

Modification of halloysite lumen with dopamine derivatives as filler for antibiofilm coating.

HYPOTHESIS: Development of nanocomposite coating with antibiofilm properties is of fundamental importance to efficient fight biofilm formation preventing infections in biomedical area. In this context, halloysite nanotubes (HNTs), biocompatible and low-cost clay mineral, have been efficiently used as filler for different polymeric matrices affording several nanocomposites with appealing antimicrobial properties. The modification of HNTs surfaces represents a valuable strategy to improve the utilization of the clay for biological purposes. EXPERIMENTS: Herein, the covalent modification of the HNTs lumen with properly designed dopamine derivatives with different perfluoroalkyl chain length is reported. The obtained nanomaterials are thoroughly characterized by several techniques. As proof of concept the antibiofilm properties on E. coli strain of the nanomaterials are assayed as well. Finally, the HNTs fillers were introduced into a polydopamine matrix allowing for the preparation of functional coatings, resistant to formation of microbial biofilms. FINDINGS: All characterization methods proved the selectivity of the modification and the increased hydrophobicity of the lumen. In particular 27Al solid state nuclear magnetic resonance (NMR) spectra showed a upfield shift of the Al signal. Studies on the antibiofilm properties highlighted different activities according to the length of perfluoroalkyl chains of organic molecules as proved by 19F solid state NMR spectra. The synthetized materials were promising for future application as coatings on medical implants.

Recent Approaches for Wound Treatment.

Wounds are a serious global health problem [...].

Exploring the cellular uptake of hectorite clay mineral and its drug carrier capabilities.

In the last years, the use of clay minerals for pharmaceutical purposes has increased due to their interesting properties. Hectorite (Ht) is a clay belonging to the smectite group which has attracted attention for applications in biology, tissue engineering and as drug carrier and delivery system. However, the mechanisms involved in Ht cellular uptake and transport into cells, are still unclear. Herein, we used a labeled Ht (Ht/1Cl) to study both the cellular uptake, by confocal laser scanning microscopy, and internalization pathways involved in the cellular uptake, by various endocytosis-inhibiting studies and fluorescence microscopy. These studies highlighted that Ht can penetrate the cellular membrane, localizing mainly in the cytoplasm. The main intracellular transport mechanisms are the ATP-dependent ones and those where filaments and microtubules are involved. Finally, as proof of concept for the potential of Ht as carrier system, we envisaged the covalent grafting of the anticancer molecule methotrexate (MTX), chosen as model, to obtain the Ht-MTX nanomaterial. The covalent linkage was confirmed by several techniques and the morphology of the obtained nanomaterial was imaged by SEM and TEM investigations. The kinetic release of the drug from the Ht-MTX nanomaterial in physiological conditions was studied as well. Furthermore, cytotoxic studies on different cell lines, namely, HL-60, HL-60R, MCF-7, 5637, UMUC3 and RT112 showed that Ht could be a promising material for anticancer therapy.

Nanocarrier based on halloysite and fluorescent probe for intracellular delivery of peptide nucleic acids.

The development of systems able to deliver genetic material into a target site is a challenge for modern medicine. Single-stranded peptide nucleic acids have attracted attention as promising therapeutic molecules for diagnostic and gene therapy. However, their poor cell membrane permeability represents a drawback for biomedical applications. Halloysite nanotubes (HNTs) are emerging materials in drug delivery applications both for their ability to penetrate cell membranes and for enhancing the solubility of drugs in biological media. Herein, we report the first example of the use of a nanocarrier based on halloysite labelled with fluorescent switchable halochromic oxazine molecules, to deliver a single-stranded peptide nucleic acids tetramer (PNAts) into living cells. The PNAts is covalently attached to halloysite (HNTs-PNA), whereas the fluorescent probe supramolecularly interacts with HNTs. The ability of the nanomaterial to bind complementary single-stranded DNA was assessed by resonance light scattering measurements. Finally, studies of cellular uptake were carried out by confocal laser scanning microscopy on normal and tumoral cell lines. This work highlights the usefulness of the covalent approach to generate HNTs-PNA nanomaterials for the potential targeting of future specific nucleic acids in living cells, which could open the doorway to novel possibilities for theranostic and gene therapy applications.

Prodrug based on halloysite delivery systems to improve the antitumor ability of methotrexate in leukemia cell lines.

The prodrug approach, as well as the development of specific systems able to deliver a chemotherapeutic agent in the target site, decreasing the side effects often associated with its administration, are still a challenging. In this context, both methotrexate drug molecules (MTX) and biotin ligand moieties, whose receptors are overexpressed on the surface of several cancer cells, were loaded on halloysite nanotubes (HNTs) to develop nanomaterial based on multifunctional and "smart" delivery systems. To highlight the crucial role played by biotin, carrier systems based on HNTs and MTX were also synthetized. In detail, several approaches were envisaged: i) a supramolecular interaction between the clay and the drug; ii) a covalent grafting of the drug onto the HNTs external surface and, iii) a combination of both approaches. The nanomaterials obtained were characterized by thermogravimetric analysis, FT-IR, and UV-vis spectroscopies, DLS and ζ-potential measurements and the morphologies were imaged by HAADF/STEM investigations. Kinetic release experiments at different pH conditions were also performed. Finally, as a proof-of-concept application of our pro-drug delivery systems based on HNTs in cancer therapy, the cytotoxic effects were evaluated on acute myeloid leukemia cell lines, HL60 and its multidrug resistance variant, HL60R. The obtained results showed that both the MTX prodrug system and the biotinylated ones played a crucial role in the biological activity and, they are promising agents for the cancer treatments.

Synthesis and Characterization of Nanomaterial Based on Halloysite and Hectorite Clay Minerals Covalently Bridged.

Halloysite is an aluminosilicate clay with a predominantly hollow tubular structure (HNTs) able to act as a nanocontainer for the encapsulation of several chemicals. However, HNTs possess low affinity for metal ions in their pristine form and they need to be modified for improving their adsorption capabilities. Therefore, to overcome this issue herein we report a straightforward approach for the covalent modification of the external surface of halloysite nanotubes with hectorite clay. Compared to halloysite, hectorite possesses a lamellar structure with higher cation exchange capacity. The covalent linkage between the two clays was verified by several techniques (FTIR spectroscopy, 13C CP-MAS NMR, TGA, ζ-potential, DLS, and XRD measurements) and the morphology was imaged by TEM investigations. As proof of concept the adsorption ability of the obtained nanomaterial in comparison to pristine clays was proved using ciprofloxacin and silver ions chosen as models for their different chemical characteristics.

Pyrazole[3,4-d]pyrimidine derivatives loaded into halloysite as potential CDK inhibitors.

Uncontrolled cell proliferation is a hallmark of cancer as a result of rapid and deregulated progression through the cell cycle. The inhibition of cyclin-dependent kinases (CDKs) activities is a promising therapeutic strategy to block cell cycle of tumor cells. In this work we reported a new example of nanocomposites based on halloysite nanotubes (HNTs)/pyrazolo[3,4-d]pyrimidine derivatives (Si306 and Si113) as anticancer agents and CDK inhibitors. HNTs/Si306 and HNTs/Si113 nanocomposites were synthesized and characterized. The release kinetics were also investigated. Antitumoral activity was evaluated on three cancer cell lines (HeLa, MDA-MB-231 and HCT116) and the effects on cell cycle arrest in HCT116 cells were evaluated. Finally, molecular dynamics simulations were performed of the complexes between Si113 or Si306 and the active site of both CDK 1 and 2.

Polymeric Bioadhesive Patch Based on Ketoprofen-Hydrotalcite Hybrid for Local Treatments.

Ketoprofen (KET) represents one of the most common drugs used in the topical treatment of pain and inflammations. However, its potential is rather limited due to the very low solubility and photochemical instability. The local administration of KET by conventional products, such as gels, emulgels, creams, and foams, does not guarantee an efficacious and safe treatment because of its low absorption (due to low solubility) and its sensitivity to UV rays. The photodegradation of KET makes many photoproducts responsible for different adverse effects. In the present work, KET was intercalated into the lamellar anionic clay ZnAl-hydrotalcite (ZnAl-HTlc), obtaining the hybrid ZnAl-KET with improved stability to UV rays and water solubility in comparison to the crystalline form (not intercalated KET). The hybrid was then formulated in autoadhesive patches for local pain treatment. The patches were prepared by casting method starting from a hydrogel based on the biocompatible and bioadhesive polymer NaCMC (Sodium carboxymethycellulose) and glycerol as a plasticizing agent. The introduction of ZnAl-KET in the patch composition demonstrated the improvement in the mechanical properties of the formulation. Moreover, a sustained and complete KET release was obtained within 8 h. This allowed reducing the frequency of anti-inflammatory administration, compared to the conventional formulations.

New Mussel Inspired Polydopamine-Like Silica-Based Material for Dye Adsorption.

A straightforward and economic procedure has been developed for the synthesis of a new polydopamine-like silica-based material that has been obtained by oxidation of catechol with KIO4 followed by reaction with 3-aminopropyltrimethoxysilane. All techniques adopted for characterization showed that the obtained material is rich in different functional groups and the morphological analyses revealed dimensions in the nanometric range. The hybrid material has been characterized by several techniques showing its polydopamine-like nature, and preliminary observations for dye adsorption have been reported.