M.M. Vidal

Evaluation of poly(2-hydroxyethyl methacrylate) gels as drug delivery systems at different pH values

Studies of dynamic and equilibrium swelling, structural characterisation and solute transport in swollen poly(2-hydroxyethyl methacrylate) gels (pHEMA) cross-linked with tripropyleneglycol diacrylate (TPGDA) were done for a wide range of TPGDA concentrations. The influence of the pH on these pHEMA properties was evaluated. In swelling studies it was found that in changing the pH from 6.5 to 12.0, a large increase in swelling occurred, from approximately 48 to 55%, for the lowest concentration of TPGDA (1 mol%), and from 40 to 80% for the highest concentration (10 mol%). Fourier transform infrared (FTIR) and differential scanning calorimetry (DSC) measurements were made after the equilibrium swelling of the gels at different pH values, to explain these results. The advantage of using these gels as controlled drug delivery systems is illustrated using salicylic acid (SA) as a model drug. The loading and the release of the SA were made at different pH values and the results obtained showed that it is possible to modulate the hydrogel performance by controlling an external factor, the pH at which the drug loading and release were performed.

Antifungal Nanoparticles and Surfaces

Nosocomial fungal infections, an increasing healthcare concern worldwide, are often associated with medical devices. We have developed antifungal nanoparticle conjugates that can act in suspension or attach to a surface, efficiently killing fungi. For that purpose, we immobilized covalently amphotericin B (AmB), a potent antifungal agent approved by the FDA, widely used in clinical practice and effective against a large spectrum of fungi, into silica nanoparticles. These antifungal nanoparticle conjugates are fungicidal against several strains of Candida sp., mainly by contact. In addition, they can be reused up to 5 cycles without losing their activity. Our results show that the antifungal nanoparticle conjugates are more fungistatic and fungicidal than 10 nm colloidal silver. The antifungal activity of the antifungal nanoparticle conjugates is maintained when they are immobilized on a surface using a chemical adhesive formed by polydopamine. The antifungal nanocoatings have no hemolytic or cytotoxic effect against red blood cells and blood mononuclear cells, respectively. Surfaces coated with these antifungal nanoparticle conjugates can be very useful to render medical devices with antifungal properties.

Preparation and characterisation of gels based on sucrose modified with glycidyl methacrylate

Abstract Sugar-based hydrogels were prepared by non-selective modification of sucrose with the introduction of vinyl groups. Sucrose was reacted with glycidyl methacrylate (1:1 molar ratio) in DMSO and in the presence of 4-( N , N -dimethylamino)pyridine (4-DMAP) as catalyst. The structure of the product (SucMA) was established using 1 H NMR, 1 H DQF-COSY NMR and 13 C NMR analysis. Gels were prepared by copolymerisation of hydroxyethyl methacrylate (HEMA) and SucMA in a 70:30 ethylene glycol/water mixed solvent using ammonium persulphate/sodium methabisulphite pair as initiating system. The swelling behaviour of these gels was evaluated as a function of crosslinker content, copolymer composition, pH and ionic strength. In addition, the diffusional behaviour of sodium salycilate (SSA) from these gels and poly(HEMA) gels is also reported.

A solid-phase extraction procedure for the clean-up of thiram from aqueous solutions containing high concentrations of humic substances.

A simple solid-phase extraction (SPE) procedure with an octadecyl bonded phase silica (C(18)) was developed for clean-up of the fungicide thiram from aqueous solutions containing high concentrations of humic substances, for future studies of thiram adsorption onto solid humic substances or soils. Suspensions of humic acids and soil, in aqueous 0.01M CaCl(2) solution, were prepared and used as samples. These extracts were spiked with thiram and immediately applied to a C(18)-SPE cartridge. Thiram was eluted with chloroform and its concentration measured by spectrophotometry at 283nm. Non-spiked aqueous extracts (blanks) and a control sample of thiram in 0.01M CaCl(2) aqueous solution were also prepared and submitted to the same SPE procedure. The results show that humic substances are extensively retained by the C(18) cartridge but are not eluted with CHCl(3). Recoveries of 100-104% were obtained for thiram in the presence of humic substances. The SPE procedure described in this work is an efficient clean-up step to remove the interference of humic substances absorbance and to be coupled to any spectrophotometric or HPLC-UV method, usually used for thiram analysis in food extracts.

Photodegradation of the fungicide thiram in aqueous solutions. Kinetic studies and identification of the photodegradation products by HPLC–MS/MS

In this study, the relevance of photodegradation processes on the persistence of the fungicide thiram in waters was investigated. The photodegradation of thiram in Milli-Q water and in aqueous solutions of humic and fulvic acids, as well as the photodegradation in spiked river water were studied. Both pure thiram and one of its commercial formulations were used to prepare the solutions which were irradiated in a solar light simulator. In general, thiram photodegradation follows pseudo-first order kinetics. The half-life time of thiram 2mgL(-1) in Milli-Q water was 28min. However, the degradation rate of thiram was significantly increased (p=0.02) by the inert components of the thiram commercial formulation as well as by commercial humic acids and by fulvic acids isolated from river water (p<0.004). Thus, the half-life time of thiram decreased to 24min in the presence of the inert formulation components, while, in the presence of both humic and fulvic acids (10mgL(-1)) it decreased to 22min. Furthermore, thiram photodegradation in natural river water showed that there is a significant enhancement of the degradation rate constant of thiram relatively to Milli-Q water, corresponding to a decrease of about 38% in its half-life time. This increase of the degradation rate in river water seems to be higher than that observed in the presence of FA, suggesting that beyond organic matter, other natural river components can increase the thiram photodegradation rate. These results allow us to conclude that photodegradation by solar radiation can be an important degradation pathway of thiram in natural waters. HPLC-MS/MS allowed to identify, for the first time, three products of the photodegradation of thiram in aqueous solution. Three compounds were identified and their structure was corroborated by the MS(n) spectra fragmentation profile. Pathways for the formation of the products from thiram photodegradation are proposed and discussed.

Study of an enzyme coupled system for the development of fibre optical bilirubin sensors.

We have established an analytical procedure for bilirubin (BR) determination by following the variation of its yellow absorption band in the presence of the biocatalytic system haemoglobin/glucose oxidase/glucose. The influence of the pH of the medium, the haemoglobin/hydrogen peroxide ratio, and the concentration of the various components in the mixture on the system activity was evaluated. The solubility of the unconjugated BR was analysed. An anionic detergent in Tris buffer, in which it presents a high critical micellar concentration, was selected for the BR/bovine serum albumin cleavage. The Michaelis-Menten constant of the system (after the optimisation of the various parameters) was determined and compared with Km values reported in the literature for similar catalytic systems. Preliminary results obtained with the haemoglobin and glucose oxidase immobilised either in poly(vinyl alcohol) or by crosslinking with glutaraldehyde, show that this system is promising as a future biosensor.

Effect of long term organic amendments on adsorption-desorption of thiram onto a luvisol soil derived from loess.

The objective of this work was to assess the influence of soil organic amendments on the sorption properties of the fungicide thiram. The organic amendments studied were organic household compost (COM), sewage sludge from municipal water treatment facilities (SLU) and farmyard manure (FYM), which were compared to mineral fertilizer application (MIN). Sorption-desorption experiments were performed using the batch method and the results indicated that the adsorption isotherms were non-linear and were found to conform to the Brunauer-Emmett-Teller (BET) model, suggesting multilayer adsorption and adsorbate-adsorbate interactions after the saturation of the surface layer. In general, distribution coefficient values, K(D), are dependent on, but not proportional to, the initial concentration of thiram. For a fixed thiram initial concentration, a significant correlation (r(2)>0.851; p<0.001) between K(D) values and the soil organic carbon content (OC) was observed. The highest value of K(D) was observed for the soil amended with compost, which is the one with the highest organic carbon content. K(D) values were divided by the soil organic carbon contents in order to obtain organic carbon partition coefficients K(OC). Comparing K(OC) means from 3 (initial concentrations) x 4 (soil organic matter compositions) x 3 (replicates) factorial ANOVA allow us to conclude that there is a significant but not proportional influence of the initial concentration of thiram on those values, but changes in the soil organic matter composition, associated to different soil amendments, have no significant influence on adsorption of thiram. To evaluate the reversibility of thiram adsorption, two consecutive desorption cycles were performed with CaCl(2) 0.01 mol L(-1). The desorption K(D) values were consistently higher (approximately twice) than those for adsorption at the same equilibrium concentrations for all soil samples supporting the existence of hysteresis in the adsorption-desorption behavior of thiram. Despite the fact that the adsorption K(D) values were proportionally increased with increasing total organic carbon content, this was not the case for the desorption K(D) values.

Influence of Fulvic Acids and Copper Ions on Thiram Determination in Water

The literature concerning the application of solid-phase extraction (SPE) to the concentration of thiram (bis(dimethyldithiocarbamoyl) disulfide) from natural waters is scarce, the available results being contradictory or with no analytical significance. To clarify these contradictory results, a C18-SPE procedure combined with HPLC-UV was applied to thiram analysis in river water, and the influence of several factors on recoveries was studied. This procedure gave thiram recoveries of about 100% when applied to thiram standard solutions. However, when the same procedure was applied to river water samples spiked with thiram, the recoveries depended on the equilibration time after spiking. The influence of river fulvic acids (FAs) and Cu(II) on thiram recoveries from standard solutions was studied as a possible interference for such a result. In the presence of FA, thiram recoveries were always higher than 85%. In the presence of Cu(II), thiram recoveries decreased significantly, due to complexation, but the addition of an excess of EDTA before C18-SPE eliminated that interference, and thiram was completely recovered. However, in river water samples the addition of EDTA had to be done before thiram spiking to obtain a recovery >90%. Thiram standard solutions containing both river FA and Cu(II) showed a behavior similar to the one observed in river water samples. On the basis of these results, the catalytic effect of Cu(II) on the degradation of thiram by FA, with formation of a Cu(II)-dimethyldithiocarbamate complex, was hypothesized.

Swelling and Thermal Properties of Poly(Vinyl Alcohol) Containing Hemoglobin Membranes

Poly(vinyl alcohol) (PVA), a hydrophilic polymer bearing hydroxyl functional groups, readily forms gels on the addition of a bifunctional agent such as glutaraldehyde. This gelation is caused by the formation of acetal bonds between the aldehyde groups and the hydroxyl groups, in the presence of HCL. The crosslink density is easily controlled by changing the concentration of glutaraldehyde, offering a way to control the total water content and, consequently, the permeability. Strong transparent films were obtained by drying these gels. These were suitable for supporting biologically active molecules, providing a viable, better alternative to the usual PVA gels which are physically crosslinked. To establish the relationship between their thermal properties and the occurrence of the physical or chemical gelling, their water vapor sorption and thermal behavior were investigated and compared. PVA/glutaraldehyde membranes were prepared with immobilized hemoglobin for bilirubin analysis. Assisted by the respective equilibrium swelling ratio and thermal stability data, it was found that, for these particular PVA/hemoglobin membranes, the occurrence of either chemical or physical crosslinking is mainly determined by the Hb/HCl ratio.

Adsorption-Desorption Behavior of Thiram onto Humic Acid

The adsorption/desorption behavior of pure thiram (Thi-P) and formulated thiram (Thi-F) onto commercial humic acids (HA) was studied using a batch equilibration procedure. Results of adsorption kinetic experiments showed that thiram adsorption is a fast process since 85% of the equilibrium concentration is reached within two hours. Experimental K(D) values between 0.110 to 0.210 L g(-1) were obtained for the adsorption of both Thi-P and Thi-F onto HA, suggesting that thiram is strongly sorbed by humic acids. In general, for both Thi-P and Thi-F, the lower the initial thiram concentration, the stronger is its adsorption (higher K(D) and percentage adsorption values). The adsorption isotherms were found to match the BET model. The results show that thiram adsorption onto condensed humic acids cannot be explained only in terms of specific interactions, such as those identified in studies of adsorption of thiram with humic acids in solution. The comparison of sorption and desorption results allowed the observation of hysteresis phenomena. Desorption K(D) values were consistently higher than those for adsorption at the same equilibrium concentration. Hysteresis was lower for the formulated thiram suggesting that adsorption is more reversible in the presence of the formulation components turning the pesticide more susceptible to be leached.