Drug Delivery through Epidermal Tissue Cells by Functionalized Biosilica from Diatom Microalgae.

Diatom microalgae are a natural source of fossil biosilica shells, namely the diatomaceous earth (DE), abundantly available at low cost. High surface area, mesoporosity and biocompatibility, as well as the availability of a variety of approaches for surface chemical modification, make DE highly profitable as a nanostructured material for drug delivery applications. Despite this, the studies reported so far in the literature are generally limited to the development of biohybrid systems for drug delivery by oral or parenteral administration. Here we demonstrate the suitability of diatomaceous earth properly functionalized on the surface with n-octyl chains as an efficient system for local drug delivery to skin tissues. Naproxen was selected as a non-steroidal anti-inflammatory model drug for experiments performed both in vitro by immersion of the drug-loaded DE in an artificial sweat solution and, for the first time, by trans-epidermal drug permeation through a 3D-organotypic tissue that better mimics the in vivo permeation mechanism of drugs in human skin tissues. Octyl chains were demonstrated to both favour the DE adhesion onto porcine skin tissues and to control the gradual release and the trans-epidermal permeation of Naproxen within 24 h of the beginning of experiments. The evidence of the viability of human epithelial cells after permeation of the drug released from diatomaceous earth, also confirmed the biocompatibility with human skin of both Naproxen and mesoporous biosilica from diatom microalgae, disclosing promising applications of these drug-delivery systems for therapies of skin diseases.

Boronic Acid Moieties Stabilize Adhesion of Microalgal Biofilms on Glassy Substrates: A Chemical Tool for Environmental Applications.

Photosynthetic organisms such as diatoms microalgae provide innovative routes to eco-friendly technologies for environmental pollution bioremediation. Living diatoms are capable to incorporate in vivo a wide variety of chemical species dispersed in seawater, thus being promising candidates for eco-friendly removal of toxic contaminants. However, their exploitation requires immobilization methods that allow to confine microalgae during water treatment. Here we demonstrate that a biofilm of Phaeodactylum tricornutum diatom cells grown on the surface of a glassy substrate bearing boronic acid protruding moieties is stably anchored to the substrate resisting mechanical stress and it is suitable for removal of up to 80 % metal ions (As, Cr, Cu, Zn, Sn, Pb, Sb) in a model polluted water sample. Control experiments also suggest that stabilization of the biofilm adhesion occurs by interaction of boronic acid surface groups of the substrate with the hydroxyl groups of diatoms extracellular polysaccharides.

Supramolecular Biohybrid Construct for Photoconversion Based on a Bacterial Reaction Center Covalently Bound to Cytochrome c by an Organic Light Harvesting Bridge.

A supramolecular construct for solar energy conversion is developed by covalently bridging the reaction center (RC) from the photosynthetic bacterium Rhodobacter sphaeroides and cytochrome c (Cyt c) proteins with a tailored organic light harvesting antenna (hCy2). The RC-hCy2-Cyt c biohybrid mimics the working mechanism of biological assemblies located in the bacterial cell membrane to convert sunlight into metabolic energy. hCy2 collects visible light and transfers energy to the RC, increasing the rate of photocycle between a RC and Cyt c that are linked in such a way that enhances proximity without preventing protein mobility. The biohybrid obtained with average 1 RC/10 hCy2/1.5 Cyt c molar ratio features an almost doubled photoactivity versus the pristine RC upon illumination at 660 nm, and ∼10 times higher photocurrent versus an equimolar mixture of the unbound proteins. Our results represent an interesting insight into photoenzyme chemical manipulation, opening the way to new eco-sustainable systems for biophotovoltaics.

Improving the In Vitro Removal of Indoxyl Sulfate and p-Cresyl Sulfate by Coating Diatomaceous Earth (DE) and Poly-vinyl-pyrrolidone-co-styrene (PVP-co-S) with Polydopamine.

Polydopamine (PDA) is a synthetic eumelanin polymer mimicking the biopolymer secreted by mussels to attach to surfaces with a high binding strength. It exhibits unique adhesive properties and has recently attracted considerable interest as a multifunctional thin film coating. In this study, we demonstrate that a PDA coating on silica- and polymer-based materials improves the entrapment and retention of uremic toxins produced in specific diseases. The low-cost natural nanotextured fossil diatomaceous earth (DE), an abundant source of mesoporous silica, and polyvinylpyrrolidone-co-Styrene (PVP-co-S), a commercial absorbent comprising polymeric particles, were easily coated with a PDA layer by oxidative polymerization of dopamine at mild basic aqueous conditions. An in-depth chemical-physical investigation of both the resulting PDA-coated materials was performed by SEM, AFM, UV-visible, Raman spectroscopy and spectroscopic ellipsometry. Finally, the obtained hybrid systems were successfully tested for the removal of two uremic toxins (indoxyl sulfate and p-cresyl sulfate) directly from patients' sera.

Bio-Inspired Redox-Adhesive Polydopamine Matrix for Intact Bacteria Biohybrid Photoanodes.

Interfacing intact and metabolically active photosynthetic bacteria with abiotic electrodes requires both establishing extracellular electron transfer and immobilizing the biocatalyst on electrode surfaces. Artificial approaches for photoinduced electron harvesting through redox polymers reported in literature require the separate synthesis of artificial polymeric matrices and their subsequent combination with bacterial cells, making the development of biophotoanodes complex and less sustainable. Herein, we report a one-pot biocompatible and sustainable approach, inspired by the byssus of mussels, that provides bacterial cells adhesion on multiple surfaces under wet conditions to obtain biohybrid photoanodes with facilitated photoinduced electron harvesting. Purple bacteria were utilized as a model organism, as they are of great interest for the development of photobioelectrochemical systems for H2 and NH3 synthesis, biosensing, and bioremediation purposes. The polydopamine matrix preparation strategy allowed the entrapment of active purple bacteria cells by initial oxygenic polymerization followed by electrochemical polymerization. Our results unveil that the deposition of bacterial cells with simultaneous polymerization of polydopamine on the electrode surface enables a 5-fold enhancement in extracellular electron transfer at the biotic/abiotic interface while maintaining the viability of the cells. The presented approach paves the way for a more sustainable development of biohybrid photoelectrodes.

Polydopamine coating of living diatom microalgae.

Many microorganisms produce specific structures, known as spores or cysts, to increase their resistance to adverse environmental conditions. Scientists have started to produce biomimetic materials inspired by these natural membranes, especially for industrial and biomedical applications. Here, we present biological data on the biocompatibility of a polydopamine-based artificial coating for diatom cells. In this work, living Thalassiosira weissflogii diatom cells are coated on their surface with a polydopamine layer mimicking mussel adhesive protein. Polydopamine does not affect diatoms growth kinetics, it enhances their resistance to degradation by treatment with detergents and acids, and it decreases the uptake of model staining emitters. These outcomes pave the way for the use of living diatom cells bearing polymer coatings for sensors based on living cells, resistant to artificial microenvironments, or acting as living devices for cells interface study.

Nitroxides as Building Blocks for Nanoantioxidants.

Nitroxides are an important class of radical trapping antioxidants whose promising biological activities are connected to their ability to scavenge peroxyl (ROO•) radicals. We have measured the rate constants of the reaction with ROO• (kinh) for a series of 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) derivatives as 5.1 × 106, 1.1 × 106, 5.4 × 105, 3.7 × 105, 1.1 × 105, 1.9 × 105, and 5.6 × 104 M-1 s-1 for -H, -OH, -NH2, -COOH, -NHCOCH3, -CONH(CH2)3CH3, and ═O substituents in the 4 position, with a good Marcus relationship between log (kinh) and E° for the R2NO•/R2NO+ couple. Newly synthesized Pluronic-silica nanoparticles (PluS) having nitroxide moieties covalently bound to the silica surface (PluS-NO) through a TEMPO-CONH-R link and coumarin dyes embedded in the silica core, has kinh = 1.5 × 105 M-1 s-1. Each PluS-bound nitroxide displays an inhibition duration nearly double that of a structurally related "free" nitroxide. As each PluS-NO particle bears an average of 30 nitroxide units, this yields an overall ≈60-fold larger inhibition of the PluS-NO nanoantioxidant compared to the molecular analogue. The implications of these results for the development of novel nanoantioxidants based on nitroxide derivatives are discussed, such as the choice of the best linkage group and the importance of the regeneration cycle in determining the duration of inhibition.

Incorporating a molecular antenna in diatom microalgae cells enhances photosynthesis.

Diatom microalgae have great industrial potential as next-generation sources of biomaterials and biofuels. Effective scale-up of their production can be pursued by enhancing the efficiency of their photosynthetic process in a way that increases the solar-to-biomass conversion yield. A proof-of-concept demonstration is given of the possibility of enhancing the light absorption of algae and of increasing their efficiency in photosynthesis by in vivo incorporation of an organic dye which acts as an antenna and enhances cells' growth and biomass production without resorting to genetic modification. A molecular dye (Cy5) is incorporated in Thalassiosira weissflogii diatom cells by simply adding it to the culture medium and thus filling the orange gap that limits their absorption of sunlight. Cy5 enhances diatoms' photosynthetic oxygen production and cell density by 49% and 40%, respectively. Cy5 incorporation also increases by 12% the algal lipid free fatty acid (FFA) production versus the pristine cell culture, thus representing a suitable way to enhance biofuel generation from algal species. Time-resolved spectroscopy reveals Förster Resonance Energy Transfer (FRET) from Cy5 to algal chlorophyll. The present approach lays the basis for non-genetic tailoring of diatoms' spectral response to light harvesting, opening up new ways for their industrial valorization.

In vivo functionalization of diatom biosilica with sodium alendronate as osteoactive material.

Bisphosphonates are a class of drugs widely used in the clinical treatment of disorders of bone metabolism, such as osteoporosis, fibrous dysplasia, myeloma and bone metastases. Because of the negative side effects caused by oral administration of bisphosphonates, various silica mesoporous materials have been investigated for a confined and controlled release of these drugs. Here, we propose biosilica from diatoms as suitable substrate for alendronate local activation of bone cells. Following a novel strategy, sodium alendronate can be in vivo incorporated into biosilica shells of cultured Thalassiosira weissflogii diatoms, by feeding the algae with an aqueous solution of the drug. After acid/oxidative treatments for removing organic matter, the resulting bisphosphonate-functionalized mesoporous biosilica was characterized and tested as osteoinductive support. Effects on osteoblast growth and anti-osteoclast activity have been examined by evaluating SaOS-2, BMSC, J774 cell viability on the alendronate-"doped" biosilica. The loading percentage of sodium alendronate into biosilica, estimated as 1.45% w/w via TGA, was able to decrease metabolic activity of J774 osteoclasts-like cells till 5% over glass control. We demonstrated a good osteoconductive ability and activation of a tissue regeneration model together with osteoclasts inhibition of the functionalized biosilica, opening the way to interesting applications for diatom microalgae as a bioinspired mesoporous material for tissue engineering.

Data from in vivo functionalization of diatom mesoporous biosilica with bisphosphonates.

Diatoms are unicellular photosynthetic microalgae that produce a sophisticated mesoporous biosilica shell called frustule. Easy to achieve and extract, diatom frustules represent a low-cost source of mesoporous biocompatible biosilica. In this paper, the possibility to in vivo functionalize the diatom biosilica with bisphosphonates (BPs) was investigated. In particular, two BPs were tested: the amino-containing sodium alendronate (ALE) and the amino-lacking sodium etidronate (ETI). According to first SEM-EDX analysis, the presence of the amino-moiety in ALE structure allowed a better incorporation of this BP into living diatom biosilica, compared to ETI. Then, diatom growth was deeply investigated in presence of ALE. After extraction of functionalized frustules, ALE-biosilica was further characterized by XPS and microscopy, and ALE release was evaluated by ferrochelation assay. Moreover, the bone regeneration performances of ALE-functionalized frustules were preliminarily investigated on bone osteoblast-like cells, via Comassie staining. Data are related to the research article "In vivo functionalization of diatom biosilica with sodium alendronate as osteoactive material".

Highly Stable and Red-Emitting Nanovesicles Incorporating Lipophilic Diketopyrrolopyrroles for Cell Imaging.

Diketopyrrolopyrroles (DPPs) have recently attracted much interest as very bright and photostable red-emitting molecules. However, their tendency to form nonfluorescent aggregates in water through the aggregation-caused quenching (ACQ) effect is a major issue that limits their application under the microscope. Herein, two DPP molecules have been incorporated into the membrane of highly stable and water-soluble quatsomes (QS; nanovesicles composed of surfactants and sterols), which allow their nanostructuration in water and, at the same time, limits the ACQ effect. The obtained fluorescent organic nanoparticles showed superior structural homogeneity, along with long-term colloidal and optical stability. A thorough one- (1P) and two-photon (2P) fluorescence characterization revealed the promising photophysical features of these fluorescent nanovesicles, which showed a high 1P and 2P brightness. Finally, the fluorescent QSs were used for the in vitro bioimaging of Saos-2 osteosarcoma cell lines; this demonstrates their potential as nanomaterials for bioimaging applications.

Multiple Routes to Smart Nanostructured Materials from Diatom Microalgae: A Chemical Perspective.

Diatoms are unicellular photosynthetic microalgae, ubiquitously diffused in both marine and freshwater environments, which exist worldwide with more than 100 000 species, each with different morphologies and dimensions, but typically ranging from 10 to 200 µm. A special feature of diatoms is their production of siliceous micro- to nanoporous cell walls, the frustules, whose hierarchical organization of silica layers produces extraordinarily intricate pore patterns. Due to the high surface area, mechanical resistance, unique optical features, and biocompatibility, a number of applications of diatom frustules have been investigated in photonics, sensing, optoelectronics, biomedicine, and energy conversion and storage. Current progress in diatom-based nanotechnology relies primarily on the availability of various strategies to isolate frustules, retaining their morphological features, and modify their chemical composition for applications that are not restricted to those of the bare biosilica produced by diatoms. Chemical or biological methods that decorate, integrate, convert, or mimic diatoms' biosilica shells while preserving their structural features represent powerful tools in developing scalable, low-cost routes to a wide variety of nanostructured smart materials. Here, the different approaches to chemical modification as the basis for the description of applications relating to the different materials thus obtained are presented.

Highly Sensitive and Practical Detection of Plant Viruses via Electrical Impedance of Droplets on Textured Silicon-Based Devices.

Early diagnosis of plant virus infections before the disease symptoms appearance may represent a significant benefit in limiting disease spread by a prompt application of appropriate containment steps. We propose a label-free procedure applied on a device structure where the electrical signal transduction is evaluated via impedance spectroscopy techniques. The device consists of a droplet suspension embedding two representative purified plant viruses i.e., Tomato mosaic virus and Turnip yellow mosaic virus, put in contact with a highly hydrophobic plasma textured silicon surface. Results show a high sensitivity of the system towards the virus particles with an interestingly low detection limit, from tens to hundreds of attomolar corresponding to pg/mL of sap, which refers, in the infection time-scale, to a concentration of virus particles in still-symptomless plants. Such a threshold limit, together with an envisaged engineering of an easily manageable device, compared to more sophisticated apparatuses, may contribute in simplifying the in-field plant virus diagnostics.

Data from two different culture conditions of Thalassiosira weissflogii diatom and from cleaning procedures for obtaining monodisperse nanostructured biosilica.

Diatoms microalgae produce biosilica nanoporous rigid outershells called frustules that exhibit an intricate nanostructured pore pattern. In this paper two specific Thalassiosira weissflogii culture conditions and size control procedures during the diatoms growth are described. Data from white field and fluorescence microscopy, evaluation of cell densities and cell parameters (k value and R value) according to cell culture conditions are listed. Different cleaning procedures for obtaining bare frustules are described. In addition, FTIR and spectrofluorimetric analyses of cleaned biosilica are shown. The data are related to the research article "Chemically Modified Diatoms Biosilica for Bone Cell Growth with Combined Drug-Delivery and Antioxidant Properties" [1].

Biosilica from Living Diatoms: Investigations on Biocompatibility of Bare and Chemically Modified Thalassiosira weissflogii Silica Shells.

In the past decade, mesoporous silica nanoparticles (MSNs) with a large surface area and pore volume have attracted considerable attention for their application in drug delivery and biomedicine. Here we propose biosilica from diatoms as an alternative source of mesoporous materials in the field of multifunctional supports for cell growth: the biosilica surfaces were chemically modified by traditional silanization methods resulting in diatom silica microparticles functionalized with 3-mercaptopropyl-trimethoxysilane (MPTMS) and 3-aminopropyl-triethoxysilane (APTES). Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy analyses revealed that the -SH or -NH2 were successfully grafted onto the biosilica surface. The relationship among the type of functional groups and the cell viability was established as well as the interaction of the cells with the nanoporosity of frustules. These results show that diatom microparticles are promising natural biomaterials suitable for cell growth, and that the surfaces, owing to the mercapto groups, exhibit good biocompatibility.

Chemically Modified Diatoms Biosilica for Bone Cell Growth with Combined Drug-Delivery and Antioxidant Properties.

This month's cover is dedicated to the joint project coordinated by Prof. Gianluca M. Farinola involving the Università degli Studi di Bari Aldo Moro, CNR ICCOM and IMIP in Bari, Università Politecnica delle Marche, and Jaber Innovation. The cover picture shows an SEM image of nanostructured biosilica produced by- Thalassiosira weissflogii- diatoms covalently functionalized with the radical scavenger TEMPO (green disks) and loaded with the antibiotic Ciprofloxacin (red/white capsules), which is used to combat infections related to orthopedic implants. Read the full text of the article at 10.1002/cplu.201402398.

Chemically Modified Diatoms Biosilica for Bone Cell Growth with Combined Drug-Delivery and Antioxidant Properties.

Nanostructured biosilica produced by Thalassiosira weissflogii diatoms is covalently functionalized with the cyclic nitroxide 2,6,6-tetramethylpiperidine-N-oxyl (TEMPO), an efficient scavenger of reactive oxygen species (ROS) in biological systems. Drug delivery properties of the TEMPO-functionalized biosilica are studied for Ciprofloxacin, an antimicrobial thoroughly employed in orthopedic or dental implant related infections. The resulting TEMPO-biosilica, combining Ciprofloxacin drug delivery with anti-oxidant properties, is demonstrated to be a suitable material for fibroblasts and osteoblast-like cells growth.