Probing Polarity and pH Sensitivity of Carbon Dots in Escherichia coli through Time-Resolved Fluorescence Analyses.

Intracellular monitoring of pH and polarity is crucial for understanding cellular processes and functions. This study employed pH- and polarity-sensitive nanomaterials such as carbon dots (CDs) for the intracellular sensing of pH, polarity, and viscosity using integrated time-resolved fluorescence anisotropy (FA) imaging (TR-FAIM) and fluorescence lifetime (FLT) imaging microscopy (FLIM), thereby enabling comprehensive characterization. The functional groups on the surface of CDs exhibit sensitivity to changes in the microenvironment, leading to variations in fluorescence intensity (FI) and FLT according to pH and polarity. The FLT of CDs in aqueous solution changed gradually from 6.38 ± 0.05 ns to 8.03 ± 0.21 ns within a pH range of 2-8. Interestingly, a complex relationship of FI and FLT was observed during measurements of CDs with decreasing polarity. However, the FA and rotational correlation time (θ) increased from 0.062 ± 0.019 to 0.112 ± 0.023 and from 0.49 ± 0.03 ns to 2.01 ± 0.27 ns, respectively. This increase in FA and θ was attributed to the higher viscosity accompanying the decrease in polarity. Furthermore, CDs were found to bind to three locations in Escherichia coli: the cell wall, inner membrane, and cytoplasm, enabling intracellular characterization using FI and FA decay imaging. FLT provided insights into cytoplasmic pH (7.67 ± 0.48), which agreed with previous works, as well as the decrease in polarity in the cell wall and inner membrane. The CD aggregation was suspected in certain areas based on FA, and the θ provided information on cytoplasmic heterogeneity due to the aggregation and/or interactions with biomolecules. The combined TR-FAIM/FLIM system allowed for simultaneous monitoring of pH and polarity changes through FLIM and viscosity variations through TR-FAIM.

Noninvasive Nanodiamond Skin Permeation Profiling Using a Phase Analysis Method: Ex Vivo Experiments.

Carbon-based nanoparticles (NPs) are widely used in nanotechnology. Among them, nanodiamonds (NDs) are suitable for biotechnology and are especially interesting for skin delivery and topical treatments. However, noninvasive detection of NDs within the different skin layers or analyzing their penetration ability is complicated due to the turbid nature of the tissue. The iterative multiplane optical properties extraction (IMOPE) technique detects differences in the optical properties of the measured item by a phase-image analysis method. The phase image is reconstructed by the multiplane Gerchberg-Saxton algorithm. This technique, traditionally, detects differences in the reduced scattering coefficients. Here, however, due to the actual size of the NDs, the IMOPE technique's detection relies on absorption analysis rather than relying on scattering events. In this paper, we use the IMOPE technique to detect the presence of the NDs within tissue-like phantoms. In addition, we perform ex vivo pigskin experiments to estimate the penetration of the NDs to the different skin layers and show that their presence reduces at deeper layers. The significance signal of the NDs within the epidermis, dermis, and fat layers gradually reduces, with t test significance values that are smaller than 10-4, 10-3, and 10-2, respectively. The IMOPE results are corroborated by TEM results and Franz-cell experiments. These results confirm that the IMOPE profiled the skin-permeation of the NDs noninvasively.

A Novel Facial Cream Based on Skin-penetrable Fibrillar Collagen Microparticles.

Background: Collagen protein plays a notable role maintaining firm skin. Topical creams containing collagen fibers are widely available, but their usefulness is questionable due to limited skin penetration. When applied in a cream, collagen does not penetrate the skin leaving the skin structure unaffected. Objective: We formulated micronized collagen in a cream base. Using human skin samples, we sought to investigate the ability of the micronized collagen cream to penetrate human skin. Methods: Particle sizes of micronized marine collagen were evaluated using electron microscopy. Optical profilometry was conducted to evaluate skin topography and roughness. The antioxidant activity of the collagen was evaluated using the electron paramagnetic resonance technique by measuring the changes in free radical production. Collagen penetration depth in human skin samples was monitored using a non-invasive optical technique known as iterative multiplane optical property extraction, which works based on the detection of laser light phase changes following the presence of collagen particles in deep skin layers. Results: According to the electron microscopy, collagen particles were found to be of various sizes, the smallest being about 120nm in diameter. Skin topography measurements revealed that the treated collagen cream increased skin smoothness of the samples. Our results derived from the iterative multiplane optical property extraction indicated that micronized collagen in a cream base penetrates both the stratum corneum and the deep epidermal layers toward the dermis. Conclusion: Our investigation suggests that the collagen in the studied cream formulation was able to penetrate the stratum coreum and deep epidermal layers in human skin samples.

Topical Hyaluronic Acid Facial Cream with New Micronized Molecule Technology Effectively Penetrates and Improves Facial Skin Quality: Results from In-vitro, Ex-vivo, and In-vivo (Open-label) Studies.

Background: Topical hyaluronic acid (HA) has shown effectiveness in maintaining skin hydration. Topical creams containing HA are widely available, but their efficacy is limited by their lack of penetration into the skin due to the large molecule size of HA, the result of being formulated into a cream base. Objective: In this three-part study (in vitro, ex vivo, and in vivo), molecule sizes, penetration levels, and antiaging qualities of a topical HA facial cream that was formulated using a new technology that micronizes HA molecules (m-HA) were assessed. Methods and Results: Particle sizes of m-HA were evaluated using electron microscopy, which showed varying sizes, the smallest of which was 100nm in diameter. The antioxidation capabilities of m-HA were measured using electron spin resonance and were found to be higher than original HA. Skin penetration of the m-HA formulation was evaluated via immunohistochemical staining of porcine skin samples, which demonstrated penetration of the formulation into the stratum corneum and the deep epidermal layers toward the dermis. Antiaging qualities of the m-HA formulation were assessed in an open-label clinical study that included 36 healthy adult women. Skin parameters were measured objectively (e.g., Corneometer, Cutometer) and subjectively via patient questionnaire, results of which indicated significant improvements in facial skin hydration, elasticity, and wrinkle depth. Conclusion: The topical HA facial cream with m-HA technology demonstrated penetration into the epidermal skin layer, and, to our knowledge, our formulation is the first HA facial cream to achieve this. Clinical application of the facial cream demonstrated objective and subjective improvements in facial skin quality of healthy adult female subjects. Our results support the use of this new HA facial cream with m-HA technology as an effective antiaging topical therapy. Larger randomized, controlled studies are needed to confirm our findings.

Imparting Pharmaceutical Applications to the Surface of Fabrics for Wound and Skin Care by Ultrasonic Waves.

In this review, we report the functionalization of textiles composed of nanoscale reactive materials in the treatment of wounds and skin diseases such as acne. In view of the growing demand for high-quality textiles, much research is focused on the creation of antimicrobial finishings for fabrics, in order to protect customers from pathogenic or odorgenerating microorganisms. We present coatings from inorganic, organic and biochemical nanoparticles (NPs) on surfaces that impart the ability to kill bacteria, avoid biofilm formation and speed up the recovery of wounds. In all three cases, sonochemistry is used for immobilizing the nanoparticles on the surfaces. The Introduction broadly covers the progress of nanotechnology in the fields of wound and skin care. The first section of this review outlines the mechanism of the ultrasound-assisted deposition of nanoparticles on textiles. The coating can be performed by an in-situ process in which the nanoparticles are formed and subsequently thrown onto the surface of the fabrics at a very high speed. This approach was used in depositing metal-oxide NPs such as ZnO, CuO and Zn-CuO or the organic NPs of tannic acid, chitosan, etc. on textiles. In addition, the sonochemical process can be used as a "throwing stone" technique, namely, previously synthesized or commercially purchased NPs can be placed in the sonication bath and sonicated in the presence of the fabric. The collapse of the acoustic bubble in the solution causes the throwing of the immersed commercial NPs onto the textiles. This section will also outline why sonochemical deposition on textiles is considered the best coating technique. The second section will discuss new applications of the sonochemically- coated textiles in killing bacteria, avoiding biofilm formation and more. Two points should be noted: 1) the review will primarily report results obtained at Bar-Ilan University and 2) since for all textiles tested in our experiments (cotton, polyester, nylon, nonwoven) similar results were obtained, the type of textile used in a specific experiment will not be mentioned - textiles will be discussed in general. It is also worth emphasizing that this review concentrates only on the sonochemical coating of textiles, ignoring other deposition techniques.

Interpretations of complications following third molar extraction.

OBJECTIVE: Surgical removal of third molars is often associated with complications. The aim of the present study was to analyze the incidence of complications following extraction of third molars relative to the risk factors. METHOD AND MATERIALS: This retrospective study included 463 patients who had mandibular third molar extraction (performed by a single surgeon, DSA) in the years 2001 to 2011. In total, 665 mandibular third molars were extracted. The average patient's age was 29 ± 11.30 years, median 26 years, and the patient age ranged from 13 to 75 years. Patients' records were obtained for medical/general data. RESULTS: The overall prevalence of postsurgical complications was 17%. Dry sockets showed the highest incidence (11.6%). Partially impacted teeth showed the highest incidence of complications (67.3%). Cigarette smoking correlated with increased complications and dry sockets, and complications were more prevalent on the left side (62.8%). CONCLUSION: Complications after mandibular third molar extraction increase with age, level of impaction, side of extraction, and cigarette smoking.

Sonochemical co-deposition of antibacterial nanoparticles and dyes on textiles.

The sonochemical technique has already been proven as one of the best coating methods for stable functionalization of substrates over a wide range of applications. Here, we report for the first time on the simultaneous sonochemical dyeing and coating of textiles with antibacterial metal oxide (MO) nanoparticles. In this one-step process the antibacterial nanoparticles are synthesized in situ and deposited together with dye nanoparticles on the fabric surface. It was shown that the antibacterial behavior of the metal oxides was not influenced by the presence of the dyes. Higher K/S values were achieved by sonochemical deposition of the dyes in comparison to a dip-coating (exhaustion) process. The stability of the antibacterial properties and the dye fastness was studied for 72 h in saline solution aiming at medical applications.

Enhanced pharmacological activity of vitamin B12 and penicillin as nanoparticles.

Sonochemistry has become a well-known technique for fabricating nanomaterials. Since one of the advantages of nanomaterials is that they have higher chemical activities compared with particles in the bulk form, efforts are being made to produce nano organic compounds with enhanced biological activities that could be exploited in the medical area. This study uses the sonication technique to prepare nano Vitamin B12 and nano Penicillin, and demonstrates their enhanced biological and pharmacological activity. The size and morphology of the nano Penicillin and nano Vitamin B12 were investigated using electron microscopy as well as dynamic light scattering techniques. The sizes of Penicillin and Vitamin B12 nanoparticles (NPs) were found to be 70 and 120-180 nm, respectively. The bactericidal effect of nano Penicillin was studied and found to be higher than that of the bulk form. Reducing the size of Vitamin B12 resulted in their enhanced antioxidative activity as observed using the electron paramagnetic spectroscopy technique. The penetration depth of these organic NPs can be detected by an optical iterative method. It is believed that nano organic drugs fabrication will have a great impact on the medical field.

Toxicity Evaluation of a New Zn-Doped CuO Nanocomposite With Highly Effective Antibacterial Properties.

The increased resistances to conventional antibiotics determine a strong need for new antibacterials, and specific syntheses at the nanoscale promise to be helpful in this field. A novel Zinc-doped Copper oxide nanocomposite (nZn-CuO) has been recently sonochemically synthesized and successfully tested also against multi-drug resistant bacteria. After synthesis and characterization of the physicochemical properties, the new nZn-CuO is here evaluated by the Frog Embyo Teratogenesis Assay-Xenopus test for its toxicological potential and this compared with that of nCuO and nZnO synthesized under the same conditions. No lethal effects are observed, while malformations and growth retardation slightly increase after nZn-CuO exposure. Nevertheless, these effects are smaller than those of nZnO. NP uptake by embryo tissues increase significantly with increasing NP concentrations, while no significant accumulation and adverse effects are seen after exposure to soluble Cu(2+) and Zn(2+) at the concentrations dissolved from the NPs. Key oxidative response genes are upregulated by nZn-CuO, as well as by nCuO and nZnO, suggesting the common mechanism of action. Considering the enhanced biocidal activity shown by the nanocomposite, together with the results presented in this study, we can affirm that the doping of the metal oxide nanoparticles should be considered a useful tool to engineer a safer nano-antibacterial.

New life for an old antibiotic.

Restoring the antibacterial properties of existing antibiotics is of great concern. Herein, we present, for the first time, the formation and deposition of stable antibiotic nanoparticles (NPs) on graphene oxide (GO) sheets by a facile one-step sonochemical technique. Sonochemically synthesized graphene oxide/tetracycline (GO/TET) composite shows enhanced activity against both sensitive and resistant Staphylococcus aureus (S. aureus). The size and deposition of tetracycline (TET) nanoparticles on GO can be controlled by varying the sonication time. The synthesized NPs ranged from 21 to 180 nm. Moreover, ultrasonic irradiation does not cause any structural and chemical changes to the TET molecule as confirmed by Fourier transform infrared spectroscopy (FTIR). The virtue of π-π stacking between GO and TET additionally facilitate the coating of TET NPs upon GO. A time dependent release kinetics of TET NPs from the GO surface is also monitored providing important insights regarding the mechanism of antibacterial activity of GO/TET composites. Our results show that the GO/TET composite is bactericidal in nature, resulting in similar values of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). This composite is found to be active against TET resistant S. aureus at a concentration four times lower than the pristine TET. The sensitive S. aureus follows the same trend showing six times lower MIC values compared to pristine TET. GO shows no activity against both sensitive and resistant S. aureus even at a concentration as high as 1 mg/mL but influences the biocidal activity of the GO/TET composite. We propose that the unique structure and composition manifested by GO/TET composites may be further utilized for different formulations of antibiotics with GO. The sonochemical method used in this work can be precisely tailored for the stable deposition of a variety of antibiotics on the GO surface to reduce health risks and increase the spectrum of applications.

Making the hospital a safer place by sonochemical coating of all its textiles with antibacterial nanoparticles.

The ability to scale-up the sonochemical coating of medical textiles with antibacterial nanoparticles is demonstrated in the current paper. A roll-to-roll pilot installation to coat textiles was built taking into consideration the requirements of the sonochemical process. A long-run experiment was conducted in which 2500 m of fabric were coated with antibacterial ZnO nanoparticles (NPs). The metal oxide NPs were deposited from an ethanol:water solution. In this continuous process a uniform concentration of coated NPs over the length/width of the fabric was achieved. The antibacterial efficiency of the sonochemically-coated textiles was validated in a hospital environment by a reduction in the occurrence of nosocomial infections. NP-coated bed sheets, patient gowns, pillow cover, and bed covers were used by 21 patients. For comparison 16 patients used regular textiles. The clinical data indicated the reduced occurrence of hospital-acquired infections when using the metal oxide NP-coated textiles. In order to reduce the cost of the coating process and considering safety issues during manufacturing, the solvent (ethanol:water) (9:1 v:v) used for the long-run experiment, was replaced by water. Although lesser amounts of ZnO NPs were deposited on the fabric in the water-based process the antibacterial activity of the textiles was preserved due to the smaller size of the particles.

Antibiotic nanoparticles embedded into the Parylene C layer as a new method to prevent medical device-associated infections.

Tetracycline nanoparticles (NPs) were synthesized and simultaneously deposited on Parylene-C coated glass slides using ultrasound irradiation. The optimization of the process conditions, the specific reagent ratio and the precursor concentration resulted in the formation of uniform NPs with an average size of ∼50 nm. These novel tetracycline NP coated-surfaces were tested against two common bacterial pathogens, Escherichia coli and Staphylococcus aureus, and were found to be extremely potent against both bacteria, suggesting that these antibiotic NPs provide the Parylene surface with self-sterilizing properties. Finally, the mechanism describing the formation of tetracycline NPs and their subsequent deposition on the Parylene C surface is presented.

Ultrasound coating of polydimethylsiloxanes with antimicrobial enzymes.

There is an urgent need for antimicrobial functionalization of urinary catheters to prevent microbial colonization and biofilm formation on them. Here, the antimicrobial hydrogen peroxide (H2O2) producing enzyme cellobiose dehydrogenase (CDH) was for the first time grafted onto polydimethylsiloxanes (PDMS) using an ultrasound assisted coating method. This resulted in the development of an effective in situ continous H2O2 producing system able to continuously prevent microbial colonization and biofilm formation on catheters. This enzyme has an added advantage that it uses various oligosaccharides including expolysaccharides (an important part of the bioflim produced by the microbes while colonizing biomaterials) as electron donors to produce H2O2. Successful immobilization of active CDH nanoparticles on PDMS was confirmed by ESEM and AFM analysis as well as quantification of H2O2. Depending on the initial enzyme concentration, CDH-nanoparticles of varying sizes from 65 ± 17 nm to 93 ± 17 nm were created by the ultrasonic waves and subsequently deposited on the PDMS surface. PDMS sheets treated for 3 min produced 18 µM of H2O2 within 2 hours which was sufficient to significantly reduce the amount of viable S. aureus cells as well as the total amount of biomass deposited on the surface. The ultrasound assisted coating of antimicrobial enzymes therefore provides an easy approach to immobilize enzymes and create a surface with antimicrobial properties.

Tetracycline nanoparticles as antibacterial and gene-silencing agents.

The spread of antibiotic-resistant bacteria and parasites calls for the development of new therapeutic strategies with could potentially reverse this trend. Here, a proposal is presented to exploit a sonochemical method to restore the antibiotic activity of tetracycline (TTCL) against resistant bacteria by converting the antibiotic into a nanoparticulate form. The demonstrated sonochemical method allows nanoscale TTCL assembly to be driven by supramolecular hydrogen bond formation, with no further modification to the antibiotic's chemical structure. It is shown that tetracycline nanoparticles (TTCL NPs) can act as antibacterial agents, both against TTCL sensitive and against resistant bacterial strains. Moreover, the synthesized antibiotic nanoparticles (NPs) can act as effective gene-silencing agents through the use of a TTCL repressor in Trypanosome brucei parasites. It is demonstrated that the NPs are nontoxic to human cells and T. brucei parasites and are able to release their monomer components in an active form in a manner that results in enhanced antimicrobial activity relative to a homogeneous solution of the precursor monomer. As the TTCL NPs are biocompatible and biodegradable, sonochemical formation of TTCL NPs represents a new promising approach for generation of pharmaceutically active nanomaterials.

Detecting nanoparticles in tissue using an optical iterative technique.

Determining the physical penetration depth of nanoparticles (NPs) into tissues is a challenge that many researchers have been facing in recent years. This paper presents a new noninvasive method for detecting NPs in tissue using an optical iterative technique based on the Gerchberg-Saxton (G-S) algorithm. At the end of this algorithm the reduced scattering coefficient (µs'), of a given substance, can be estimated from the standard deviation (STD) of the retrieved phase of the remitted light. Presented in this paper are the results of a tissue simulation which indicate a linear ratio between the STD and the scattering components. A linear ratio was also observed in the tissue-like phantoms and in ex vivo experiments with and without NPs (Gold nanorods and nano Methylene Blue). The proposed technique is the first step towards determining the physical penetration depth of NPs.

Visible light-induced OH radicals in Ga2O3: an EPR study.

Reactive oxygen species (ROS) were found to exist in water suspensions of several metal oxide nanoparticles (NPs), such as CuO, TiO2 and ZnO. Visible light irradiation enhanced the capability of TiO2 and ZnO NPs to generate ROS, thus increasing their antibacterial effects. Because of the possible toxic effects on the host tissue it is desired to find nano-metal oxides which do not produce ROS under room light, but only upon a strong external stimulus. Using the technique of electron-spin resonance (ESR) coupled with spin trapping, we examined the ability of Ga2O3 submicron-particle suspensions in water to produce reactive oxygen species with and without visible light irradiation. We found that in contrast to ZnO and TiO2 NPs, no ROS are produced by Ga2O3 under room light. Nevertheless blue light induced hydroxyl radical formation in Ga2O3. This finding might suggest that NPs of Ga2O3 could be used safely for infected skin sterilization.

Eradication of multi-drug resistant bacteria by a novel Zn-doped CuO nanocomposite.

Zinc-doped copper oxide nanoparticles are synthesized and simultaneously deposited on cotton fabric using ultrasound irradiation. The optimization of the processing conditions, the specific reagent ratio, and the precursor concentration results in the formation of uniform nanoparticles with an average size of ≈30 nm. The antibacterial activity of the Zn-doped CuO Cu0.88Zn0.12O in a colloidal suspension or deposited on the fabric is tested against Escherichia coli (Gram negative) and Staphylococcus aureus (Gram positive) bacteria. A substantial enhancement of 10,000 times in the antimicrobial activity of the Zn-CuO nanocomposite compared to the pure CuO and ZnO nanoparticles (NPs) is observed after 10 min exposure to the bacteria. Similar activities are observed against multidrug-resistant bacteria (MDR), (i.e., Methicillin-resistant S. aureus and MDR E. coli) further emphasizing the efficacy of this composite. Finally, the mechanism for this enhanced antibacterial activity is presented.

The synergistic effect of visible light and gentamycin on Pseudomona aeruginosa microorganisms.

Recently there were several publications on the bactericidal effect of visible light, most of them claiming that blue part of the spectrum (400 nm-500 nm) is responsible for killing various pathogens(1-5). The phototoxic effect of blue light was suggested to be a result of light-induced reactive oxygen species (ROS) formation by endogenous bacterial photosensitizers which mostly absorb light in the blue region(4,6,7). There are also reports of biocidal effect of red and near infra red(8) as well as green light(9). In the present study, we developed a method that allowed us to characterize the effect of high power green (wavelength of 532 nm) continuous (CW) and pulsed Q-switched (Q-S) light on Pseudomonas aeruginosa. Using this method we also studied the effect of green light combined with antibiotic treatment (gentamycin) on the bacteria viability. P. aeruginosa is a common noscomial opportunistic pathogen causing various diseases. The strain is fairly resistant to various antibiotics and contains many predicted AcrB/Mex-type RND multidrug efflux systems(10). The method utilized free-living stationary phase Gram-negative bacteria (P. aeruginosa strain PAO1), grown in Luria Broth (LB) medium exposed to Q-switched and/or CW lasers with and without the addition of the antibiotic gentamycin. Cell viability was determined at different time points. The obtained results showed that laser treatment alone did not reduce cell viability compared to untreated control and that gentamycin treatment alone only resulted in a 0.5 log reduction in the viable count for P. aeruginosa. The combined laser and gentamycin treatment, however, resulted in a synergistic effect and the viability of P. aeruginosa was reduced by 8 log's. The proposed method can further be implemented via the development of catheter like device capable of injecting an antibiotic solution into the infected organ while simultaneously illuminating the area with light.

Visible light-induced antibacterial activity of metaloxide nanoparticles.

OBJECTIVE: The purpose of this article was to review studies that use visible light instead of dangerous ultraviolet (UV) radiation, for inducing antibacterial properties in metal oxide nanoparticles (NPs). BACKGROUND DATA: Metal oxide NPs such as ZnO, CuO, and TiO2 are frequently studied for their antibacterial effects, based on their capability to generate reactive oxygen species (ROS) in their water suspensions, following UV light absorption. METHODS: Research articles on shifting metal oxide NPs absorption into the visible light region, published up to 2011, were retrieved from library sources, as well as PubMed and MEDLINE(®) databases. RESULTS: The studies indicated that doping metaloxide NPs with transition metals ions, or attaching the metal oxide nanoparticles to an organic molecule, enhanced their activity in the visible and near infrared (NIR) range. Moreover, ZnO and TiO2 nanoparticles were found to have an absorption peak in UV-A, with a marked absorption in the blue region. CONCLUSIONS: It is possible to extend the absorption region of metal oxide NPs to the red/NIR, increasing their antibacterial activity without inducing damage to tissues and cells.

Low-level visible light (LLVL) irradiation promotes proliferation of mesenchymal stem cells.

Low-level visible light irradiation was found to stimulate proliferation potential of various types of cells in vitro. Stem cells in general are of significance for implantation in regenerative medicine. The aim of the present study was to investigate the effect of low-level light irradiation on the proliferation of mesenchymal stem cells (MSCs). MSCs were isolated from the bone marrow, and light irradiation was applied at energy densities of 2.4, 4.8, and 7.2 J/cm(2). Illumination of the MSCs resulted in almost twofold increase in cell number as compared to controls. Elevated reactive oxygen species and nitric oxide production was also observed in MSCs cultures following illumination with broadband visible light. The present study clearly demonstrates the ability of broadband visible light illumination to promote proliferation of MSCs in vitro. These results may have an important impact on wound healing.