Recent Progress and Applications of Thermal Lens Spectrometry and Photothermal Beam Deflection Techniques in Environmental Sensing.

This paper presents recent development and applications of thermal lens microscopy (TLM) and beam deflection spectrometry (BDS) for the analysis of water samples and sea ice. Coupling of TLM detection to a microfluidic system for flow injection analysis (µFIA) enables the detection of microcystin-LR in waters with a four samples/min throughput (in triplicate injections) and provides an LOD of 0.08 µg/L which is 12-times lower than the MCL for microcystin-LR in water. µFIA-TLM was also applied for the determination of total Fe and Fe(II) in 3 µL samples of synthetic cloudwater. The LODs were found to be 100 nM for Fe(II) and 70 nM for total Fe. The application of µFIA-TLM for the determination of ammonium in water resulted in an LOD of 2.3 µM for injection of a 5 µL sample and TLM detection in a 100 µm deep microfluidic channel. For the determination of iron species in sea ice, the BDS was coupled to a diffusive gradient in the thin film technique (DGT). The 2D distribution of Fe(II) and total Fe on DGT gels provided by the BDS (LOD of 50 nM) reflected the distribution of Fe species in sea ice put in contact with DGT gels.

Determination of Trace Cr(VI) with Diphenylcarbazide by µFIA-Thermal Lens Microscopy.

The optimum reaction parameters for the interaction of hexavalent chromium [Cr(VI)] with diphenylcarbazide in microfluidic chips (µFIA) with thermal-lens microscopic detection were selected. The characteristic feature of the applied flow scheme is the injection of the reagent into the stream containing the test metal, which enables in-field and real-time monitoring of Cr(VI) simply by flowing the sample continuously through the microchip. The limit of detection of Cr(VI) under the selected conditions (signal generating wavelength, 514.5 nm; excitation power, 100 mW; detection position, 10 cm downstream from the mixing zone of the microchip; flow rate 10 µL min-1; injection volume, 1.4 µL) is 15 ng mL-1 (2.9 × 10-7 mol L-1). The linear range is 40 ng mL-1 - 10 µg mL-1 with a relative standard deviation no higher than 10% in the concentration range 0.1-1 µg mL-1. The online monitoring by this scheme provides the possibility of up to 360 analyses per hour.

Application of complex geometrical optics to determination of thermal, transport, and optical parameters of thin films by the photothermal beam deflection technique.

In this work, complex geometrical optics is, for what we believe is the first time, applied instead of geometrical or wave optics to describe the probe beam interaction with the field of the thermal wave in photothermal beam deflection (photothermal deflection spectroscopy) experiments on thin films. On the basis of this approach the thermal (thermal diffusivity and conductivity), optical (energy band gap), and transport (carrier lifetime) parameters of the semiconductor thin films (pure TiO2, N- and C-doped TiO2, or TiO2/SiO2 composites deposited on a glass or aluminum support) were determined with better accuracy and simultaneously during one measurement. The results are in good agreement with results obtained by the use of other methods and reported in the literature.

Accuracy of Measurements of Thermophysical Parameters by Dual-Beam Thermal-Lens Spectrometry.

Thermal-lens spectrometry is a sensitive technique for determination of physicochemical properties and thermophysical parameters of various materials including heterogeneous systems and nanoparticles. In this paper, we consider the issues of the correctness (trueness) of measurements of the characteristic time of the thermal-lens effect and, thus, of the thermal diffusivity determined by dual-beam mode-mismatching thermal lensing. As sources of systematic errors, major factors-radiation sources, sample-cell and detector parameters, and general measurement parameters-are considered using several configurations of the thermal-lens setups, and their contributions are quantified or estimated. Furthermore, with aqueous ferroin and Sudan I in ethanol as inert colorants, the effects of the intermolecular distance of the absorbing substance on the correctness of finding the thermophysical parameters are considered. The recommendations for checking the operation of the thermal-lens setup to ensure the maximum accuracy are given. The results obtained help reducing the impact of each investigated factor on the value of systematic error and correctly measure the thermophysical parameters using thermal-lens spectrometry.

Non-Contact and Self-Calibrated Photopyroelectric Method for Complete Thermal Characterization of Porous Materials.

A general theory of a photopyroelectric (PPE) configuration, based on an opaque sample and transparent pyroelectric sensor, backing and coupling fluids is developed. A combined back-front detection investigation, based on a frequency scan of the phase of the PPE signals, followed by a self-normalization of the phases' behavior, leads to the possibility of simultaneously measuring both thermal effusivity and diffusivity of a solid sample. A particular case of this configuration, with no coupling fluid at the sample/backing interface and air instead of coupling fluid at the sample/sensor interface (non-contact method) is suitable for simultaneous measurement ofboth thermal diffusivity and effusivity (in fact complete thermal characterization) of porous solids. Compared with the already proposed configurations for investigations of porous materials, this novel configuration makes use of a fitting procedure with only one fitting parameter, in order to guarantee the uniqueness of the solution. The porous solids belong to a class of materials which are by far not easy to be investigated using PPE. To the best of our knowledge, porous materials represent the only type of compounds, belonging to condensed matter, which were not taken into consideration (until recently) as potential samples for PPE calorimetric investigations. Consequently, the method proposed in this paper complete the area of applications of the PPE method. Applications on some porous building materials and cellulose-based samples validate the theory.

Improved Photopyroelectric (PPE) Configuration for Thermal Effusivity Investigations of Porous Solids.

A new photopyroelectric detection configuration is proposed in order to measure the thermal effusivity of porous solids. Compared with the previously reported detection scheme this configuration makes use of a transparent window in front of the pyroelectric sensor. In such a way, the heat losses by convection at the sensor's irradiated surface are eliminated, and consequently, the conduction remains the only process responsible for the heat propagation in the whole detection cell. In the paper, the mathematical model for this new configuration is developed, with the main conclusion that the sample's thermal effusivity can be finally obtained via a fitting procedure with only two fitting parameters (instead of three as previously reported); in such a way, the possible degeneracy of the results is eliminated. The suitability of the method is demonstrated with application on some porous building materials and cellulose-based pressed powders.

Nanobody-Dependent Detection of Microcystis aeruginosa by ELISA and Thermal Lens Spectrometry.

Nanobodies against cell surface antigens of toxic cyanobacteria Microcystis aeruginosa were recovered by whole-cell biopanning of a naïve phage display library of nanobodies. Six unique sequences were identified and three sub-cloned and purified as fusion immunoreagents together with either green fluorescent protein or AviTag to be used for diagnostics. The yields of nanobody constructs were in the range of 5-10 mg/l and their specificity and sensitivity was initially evaluated by immunofluorescence and by fluorescent enzyme-linked immunosorbent assay (ELISA) using fluorescent nanobodies. The ELISA data confirmed the nanobody specificity but showed that the saturation of the fluorescence signal already in the presence of few hundreds of cells limited the dynamic range of the method. As an alternative, Avi-tagged nanobodies were used in combination with streptavidin-linked horseradish peroxidase for developing a diagnostic colorimetric cell ELISA, the limit-of-detection of which was 3.2 and 4.5 cells/ml for the two tested cyanobacteria strains, whereas the linear range of the assay was expanded from 10 to 10,000 cells. The fluorescent nanobodies were finally exploited for quantifying cyanobacteria by thermal lens spectrometry (TLS) that enabled to reach a limit-of-detection of 1.2 cells/ml and provided a linear range of measurement between 0 and 10,000 cells. No cross-reactivity with unrelated microalgae was detected and both colorimetric ELISA and TLS provided a linear range of detection of few logs. The data indicate that nanobodies are suitable capture reagents and that both TLS and colorimetric ELISA are reliable to monitor variations of cyanobacteria populations.

Photodegradation mechanisms of reactive blue 19 dye under UV and simulated solar light irradiation.

In this work we performed dye photodegradation experiments in presence of TiO2 and Cu/Zr modified TiO2. The changes in the shape of the spectra of RB19 caused by photocatalysts under the simulated solar or UV light were monitored. Since the predominant photocatalytic mechanism can only be observed in very dilute solution of RB19, UV-Vis absorption spectrometry for higher concentrations and thermal lens spectrometry for lower concentrations have been applied to elucidate the mechanistic details of degradation processes. Bleaching of the dye was a characteristic feature, that occurred under both simulated solar and UV lights. It was also evident, that the absorption peak with maximum centered at 592 nm undergoes a slight blue shift during irradiation. The experiments carried out using UV and simulated solar light demonstrated, that two different processes responsible for the RB19 dye degradation occurred. In the initial stage of irradiation one of the processes appears under the UV light and can be recognized by a characteristic blue shift in the absorption spectrum of the solution. The second process is characteristic for irradiation by the simulated solar light which involve a blue shift at longer periods (100 min). These phenomena were attributed to the photocatalytic and photosensitization mechanisms, respectively. However, photocatalytic mechanism was also observed under simulated solar radiation, when the initial dye concentration was decreased to 5 mgL-1, and was recognized by the increase of the thermal lens signal during the initial stages of degradation process. This was possible because the thermal lens spectroscopy technique provides a limit of quantification for RB19 at the concentration level of 0.12 mg L-1, while UV-Vis spectrometry enables quantification of RB19 only down to 4 mg L-1 levels.

Comparison of CIM discs and CPG glass as solid supports for bioanalytical columns used in allergen detection.

This work presents a comparison of convective interaction media (CIM) and controlled pore glass (CPG) as solid supports for immunoglobulin antibodies used in bioanalytical detection of allergens in foodstuffs. A flow-injection manifold with highly sensitive thermal lens spectrometric detection was used for this purpose. Using beta-lactoglobulin, a milk allergen, as a model analyte, CIM disc supports had a higher linear range (0.2-3.5 microg L(-1)), better reproducibility (intra-day RSD = 1%, inter-day RSD = 10%), lower consumption of reagents, and better immunocolumn stability (1 month, over 240 injections of substrate), while providing comparable LODs (0.1 microg L(-1)). Application of CIM discs as solid supports in immunocolumns for allergen detection enables fast and sensitive screening of allergens in foodstuffs with sample throughput of up to eight samples per hour.

A multi-thermal-lens approach to evaluation of multi-pass probe beam configuration in thermal lens spectrometry.

In this work, a recently proposed thermal lens instrument based on multi-pass probe beam concept is investigated and described as a multi-thermal-lens equivalent system. A simulation of the photothermal lens signal formation in a multi-thermal-lens equivalent configuration of the system is performed and validated by comparing the experimental signals of single, dual and ten-pass configurations to theoretically calculated values. The theoretically predicted enhancement of the signal is 9 to 10-fold for a weak thermal lens when comparing the ten-pass configuration with the conventional single-pass thermal lens system. Experimentally achieved signal enhancement in the ten-pass system is 8.3 for pure ethanol sample and between 8 and 9 for solutions with different concentrations of the Fe(II) - 1,10-Phenanthroline complex. Additionally, a value of 9.1 was calculated as the ratio of the slopes of the calibration lines obtained using the ten-pass and single-pass configurations. The achieved limit of detection for determination of Fe(II), in the ten-pass configuration, was 0.4 µgL-1, with a relative standard deviation around 4.5%, which compares favorably with previously reported results for TLS determination of Fe(II) in thin samples using low excitation power. For the multi-pass configuration the linear range of measurement is reduced when compared to the single-pass configuration. This is explained by the theoretical analysis of the photothermal signal under multi-pass condition, which shows the important contribution of nonlinear term in theoretical expression for the photothermal signal. The ten-pass configuration, which is presented and validated experimentally for the first time, offers important signal enhancement needed in recently developed TLS instruments with tunable, low power excitation sources.

Determination of Iron in Environmental Water Samples by FIA-TLS.

The determination of low concentration of iron in natural waters can be difficult due to the complexity of natural water, but primarily because it requires preconcentration of the sample with solvent extraction. In this work we report on results of thermal lens spectrometry (TLS) coupled to flow injection analysis (FIA) as a highly sensitive FIA-TLS method of iron detection. The concentration of iron redox species was determined using 1,10-phenanthroline (PHN), that forms stable complexes with Fe(II) ions which are characterized by an absorption maximum at 508 nm. The TLS system using a 633 nm probe laser and 530 nm pump laser beam was exploited for on-line detection in flow injection analysis, where a PHN solution was used as the carrier solution for FIA. The concentration of the complexing agent affects the quality of the TLS signal, and the optimal concentration was found at 1 mM PHN. The achieved limits of detection (LODs) for Fe(II) and total iron were 33 nM for Fe(II) and 21 nM for total iron concentration. The method was further validated by determining the linear concentration range, specificity in terms of analytical yield and by determining concentration of iron in a water sample from a local water stream.

Determination of Dissolved Iron Redox Species in Freshwater Sediment using DGT Technique Coupled to BDS.

In this work we have developed a novel method for determination of iron redox species by the use of diffusive gradients in thin-film (DGT) technique coupled to photothermal beam deflection spectroscopy (BDS). The combination of both methods achieved low limit of detection (LOD) of 0.14 µM for Fe (II) ions. The total Fe concentration determined in the Vrtojbica river sediment (Slovenia, Rozna Dolina, 5000 Nova Gorica) was 49.3 µgL-1. The Fe (II) and Fe (III) concentration amounted to 12.8 µgL-1 and 39.9 µgL-1, respectively. Such an approach opens new opportunities for monitoring the content of iron species in natural waters and sediments and provides highly sensitive chemical analysis and an accurate qualitative and quantitative characteristic of the materials under study.

Inhibition of AChE by malathion and some structurally similar compounds.

Inhibition of bovine erythrocyte acetylcholinesterase (free and immobilized on controlled pore glass) by separate and simultaneous exposure to malathion and malathion transformation products which are generally formed during storage or through natural or photochemical degradation was investigated. Increasing concentrations of malathion, its oxidation product malaoxon, and its isomerisation product isomalathion inhibited free and immobilized AChE in a concentration-dependent manner. KI, the dissociation constant for the initial reversible enzyme inhibitor-complex, and k3, the first order rate constant for the conversion of the reversible complex into the irreversibly inhibited enzyme, were determined from the progressive development of inhibition produced by reaction of native AChE with malathion, malaoxon and isomalathion. KI values of 1.3 x 10(-4) M(-1), 5.6 x 10(-6) M(-1) and 7.2 x 10(-6)M(-1) were obtained for malathion, malaoxon and isomalathion, respectively. The IC50 values for free/immobilized AChE, (3.7 +/- 0.2) x 10(-4) M/(1.6 +/-0.1) x 10(-4), (2.4 +/- 0.3) x 10(-6)/(3.4 +/- 0.1) x 10(-6)M and (3.2 +/- 0.3) x 10(-6) M/(2.7 +/- 0.2) x 10(-6) M, were obtained from the inhibition curves induced by malathion, malaoxon and isomalathion, respectively. However, the products formed due to photoinduced degradation, phosphorodithioic O,O,S-trimethyl ester and O,O-dimethyl thiophosphate, did not noticeably affect enzymatic activity, while diethyl maleate inhibited AChE activity at concentrations > 10mM. Inhibition of acetylcholinesterase increased with the time of exposure to malathion and its inhibiting by-products within the interval from 0 to 5 minutes. Through simultaneous exposure of the enzyme to malaoxon and isomalathion, an additive effect was achieved for lower concentrations of the inhibitors (in the presence of malaoxon/isomalathion at concentrations 2 x 10(-7) M/2 x 10(-7) M, 2 x 10(-7) M/3 x 10(-7)M and 2 x 10(-7) M/4.5 x 109-7) M), while an antagonistic effect was obtained for all higher concentrations of inhibitors. The presence of a non-inhibitory degradation product (phosphorodithioic O,O,S-trimethyl ester) did not affect the inhibition efficiencies of the malathion by-products, malaoxon and isomalathion.

Facile synthesis, structure, biocompatibility and antimicrobial property of gold nanoparticle composites from cellulose and keratin.

A novel, one-pot method was developed to synthesize gold nanoparticle composite from cellulose (CEL), wool keratin (KER) and chloroauric acid. Two ionic liquids, butylmethylimmidazolium chloride and ethylmethylimmidazolium bis(trifluoromethylsulfonyl)imide were used to dissolve CEL, KER and HAuCl4. X-ray diffraction and X-ray photoelectron results show that Au3+ was completely reduced to Au0NPs with size of (5.5±1) nm directly in the composite with NaBH4. Spectroscopy and imaging results indicate that CEL and KER remained chemically intact and were homogeneously distributed in the composites with Au0NPs. Encapsulating Au0NPs into [CEL+KER] composite made the composite fully biocompatible and their bactericidal capabilities were increased by the antibacterial activity of Au0NPs. Specifically, the [CEL+KER+Au0NPs] composite exhibited up to 97% and 98% reduction in growth of antibiotic resistant bacteria such as vancomycin resistant Enterococcus faecalis and methicillin resistant Staphylococcus aureus, and was not cytotoxic to human fibroblasts. While [CEL+KER] composite is known to possess some antibacterial activity, the enhanced antibacterial observed here was due solely to added Au0NPs. These results together with our previous finding that [CEL+KER] composites can be used for controlled delivery of drugs clearly indicate that the [CEL+KER+Au0NPs] composites possess all required properties for successful use as dressing to treat chronic ulcerous infected wounds.

A combination of interdisciplinary analytical tools for evaluation of multi-layered coatings on medical grade stainless steel for biomedical applications.

In this comprehensive study several analytical techniques were used in order to evaluate multi-layered biomedical surface coatings composed of a drug (diclofenac) and a polymer (chitosan). Such a thorough examination is of paramount importance in order to assure safety and prove efficiency of potential biomedical materials already at the in vitro level, hence leading to their potentially faster introduction to clinical trials. For the first time a novel technique based on thermal diffusivity and conductivity measurements (photothermal beam deflection spectroscopy - BDS) was employed in order to analyse in a non-destructive way the thickness of respective layers, together with their thermal diffusivity and conductivity. In addition to attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR), BDS confirmed successive surface layers of the prepared coatings. Scanning electron microscopy and atomic force microscopy were used to examine structural information on the macro- and micro/nano-scale, respectively. Surface hydrophobicity was measured with the contact angle analysis, which clearly showed differences in hydrophobicity between coated and non-coated samples. Considering the targeted application of the prepared coatings (as implant in orthopaedic treatments), the in vitro drug release was analysed spectrophotometrically to examine the coatings potential for a controlled drug release. Furthermore, the material was also tested by electrochemical impedance spectroscopy and cyclic polarisation techniques, which were able to detect even minor differences between the performance of the coated and non-coated materials. As the final test, the biocompatibility of the coatings with human osteoblasts was determined.

Trace detection and photothermal spectral characterization by a tuneable thermal lens spectrometer with white-light excitation.

In the thermal lens experimental set-up we replaced the commonly employed pump laser by a halogen lamp, combined with an interference filter, providing a tuneable, nearly monochromatic pump source over the range of wavelengths 430-710 nm. Counter-propagating pump and probe beams are used and a 1 mm path-length sample cell together with the interference filter makes an optical cavity, providing amplification of the thermal lens signal, which leads to enhancement of the measurement sensitivity, and enables detection of absorbances on the order of 5 × 10-6. Amplified thermal lens signal allows us to replace the typical lock-in amplifier and digital oscilloscope with a silicon photodetector, Arduino, and a personal computer, offering the possibility for a compact, robust and portable device, useful for in-field absorption measurements in low concentration or weakly absorbing species. The use of a white light source for optical pumping, an interference filter for wavelength selection and direct diagnostic of the thermal lens signal increase the versatility of the instrument and simplifies substantially the experimental setup. Determination of Fe(II) concentrations at parts per billion levels was performed by the described white-light thermal lens spectrophotometer and the absorption spectrum for 50 µgL-1 Fe(II)-1,10-phenanthroline was well reproduced with an average measurement precision of 4%. The obtained limits of detection and quantitation of Fe(II) determination at 510 nm are 3 µgL-1 and 11 µgL-1, respectively. The calibration curve was linear in the concentration range of LOQ-500 µgL-1 with reproducibility between 2% and 6%, confirming that this instrument provides good spectrometric capabilities such as high sensitivity, tuneability and good reproducibility. In addition, the versatility of the instrument was demonstrated by recording the photothermal spectrum of gold nanostructured material and determination of excitation wavelength with most efficient optical to thermal energy conversion, which differs considerably (cca 100 nm) from the absorption maximum of the investigated sample.

Application of chromogenic reagents in surface plasmon resonance (SPR).

In this paper, a new simple approach for sensitivity optimization in surface plasmon resonance (SPR) chemosensors based on colorimetric ligands is presented. A new design of SPR sensor with tunable analytical wavelength (lambda(SPR)) was constructed for this purpose, to perform studies on the ligand absorbance spectra related sensitivity enhancement. Unlike commercial SPR sensors which operate at one lambda(SPR), the new device can be used for sensitivity analysis at selected lambda(SPR) in the range 550-750 nm, offering the possibility to identify the highest sensitivity lambda(SPR) in regard to the spectral changes of the selected ligand. Measurements can be easily done in ligand bulk solutions without immobilization steps. Sensitivity enhancement analysis and optimization of lambda(SPR) on chromogenic reagents with hypsochromic shift in their absorption spectra are demonstrated in this contribution. Optimal selection of analytical wavelength, set at the absorbance peak of chromogenic reagent Eriochrome Black T (EBT) was observed to result in up to two times increased SPR sensitivity to Cd(2+) compared to wavelengths selected in other parts of the ligand absorbance spectra, with a limit of detection (LOD) 0.2 ppm. The sensitivity enhancement at optimal lambda(SPR) was observed to be related to increased refractive index (n), drop in extinction coefficient (alpha) and simultaneous hypsochromic shift of the EBT absorbance spectra causing the lambda(SPR) to match the absorbance peak shoulder.

Evaluation of photochemical degradation of digoxin by Na,K-ATPase assay.

A simple Na,K-ATPase assay is described as a suitable method for testing of digoxin photodegradation. The exposure of Na,K-ATPase to the photodegraded samples exhibited reduced inhibition of the enzyme, compared to the unirradiated samples containing equal initial concentrations of drug. The degree of inhibition was dependent on the irradiation time. The concentrations of digoxin in irradiated samples were evaluated by HPLC analysis. Excellent agreement of the results obtained by both methods was observed. The investigation of the influence of irradiated samples on Na,K-ATPase inhibition revealed no side products acting as Na,K-ATPase inhibitors. The cytokinesis block micronucleus test (CBMN) was applied in order to investigate the cytotoxicity of the possible degradation products after exposure to UV irradiation. The results confirmed that the photochemical treatment did not induce the cytotoxic side products. Zero order kinetics, which was observed for digoxin photodegradation and the associated reaction mechanism are also discussed.

One-Pot Synthesis of Biocompatible Silver Nanoparticle Composites from Cellulose and Keratin: Characterization and Antimicrobial Activity.

A novel, simple method was developed to synthesize biocompatible composites containing 50% cellulose (CEL) and 50% keratin (KER) and silver in the form of either ionic (Ag+) or Ag0 nanoparticles (Ag+NPs or Ag0NPs). In this method, butylmethylimmidazolium chloride ([BMIm+Cl-]), a simple ionic liquid, was used as the sole solvent and silver chloride was added to the [BMIm+Cl-] solution of [CEL+KER] during the dissolution process. The silver in the composites can be maintained as ionic silver (Ag+) or completely converted to metallic silver (Ag0) by reducing it with NaBH4. The results of spectroscopy [Fourier transform infrared and X-ray diffraction (XRD)] and imaging [scanning electron microscopy (SEM)] measurements confirm that CEL and KER remain chemically intact and homogeneously distributed in the composites. Powder XRD and SEM results show that the silver in the [CEL+KER+Ag+] and [CEL+KER+Ag0] composites is homogeneously distributed throughout the composites in either Ag+ (in the form of AgClNPs) or Ag0NPs form with sizes of 27 ± 2 or 9 ± 1 nm, respectively. Both composites were found to exhibit excellent antibacterial activity against many bacteria including Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, methicillin-resistant S. aureus (MRSA), and vancomycin-resistant Enterococus faecalis (VRE). The antibacterial activity of both composites increases with the Ag+ or Ag0 content in the composites. More importantly, for the same bacteria and the same silver content, the [CEL+KER+AgClNPs] composite is relatively more toxic than [CEL+KER+Ag0NPs] composite. Experimental results confirm that there was hardly any Ag0NPs release from the [CEL+KER+Ag0NPs] composite, and hence its antimicrobial activity and biocompatibility is due not to any released Ag0NPs but rather entirely to the Ag0NPs embedded in the composite. Both AgClNPs and Ag0NPs were found to be toxic to human fibroblasts at higher concentration (>0.72 mmol), and for the same silver content, the [CEL+KER+AgClNPs] composite is relatively more toxic than the [CEL+KER+Ag0NPs] composite. As expected, by lowering the Ag0NPs concentration to 0.48 mmol or less, the [CEL+KER+Ag0NPs] composite can be made biocompatible while still retaining its antimicrobial activity against bacteria such as E. coli, S. aureus, P. aeruginosa, MRSA, and VRE. These results, together with our previous finding that [CEL+KER] composites can be used for the controlled delivery of drugs such as ciprofloxacin, clearly indicate that the [CEL+KER+Ag0NPs] composite possesses all of the required properties for it to be successfully used as a high-performance dressing to treat chronic ulcerous infected wounds.