Enhanced gentamicin loading and release of PLGA and PLHMGA microspheres by varying the formulation parameters

The purpose of this study was to develop a suitable formulation for gentamicin sulfate (GS) that gives a sustained release of the drug. Therefore this drug was loaded into poly(D,L-lactide-co-glycolide) (PLGA) and poly(lactic-co-hydroxymethyl glycolic acid) (PLHMGA) microspheres. The effects of various formulation parameters (ethanol, surfactant, osmotic value of the external phase, polymer type and concentration) on particle characteristics (size, loading and release) were investigated. The GS loaded microspheres were prepared using a double emulsion evaporation technique. The results demonstrate that neither ethanol nor surfactants had beneficial effects on the drug loading efficiency (around 4-10%). However, an increase in buffer concentration (and thus osmotic pressure) of the external phase resulted in a substantial increase of GS-loading (from 10 to 28%). Further, an increase of concentration of PLGA in DCM from 10% to 15/20% caused a 4-time increase of the drug loading. The best formulation identified in this study had a loading efficiency of around 70% resulting in PLGA microspheres with a 6% (w/w) loading. The particles showed a burst release of the drug depending on their porosity, followed by a phase of 35 days where hardly any release occurred. The drug was then slowly released for around 25 days likely due to degradation of the microspheres. The drug loading efficiency of GS in PLHMGA was not significantly different from PLGA microspheres (64%). The release of GS from PLHMGA microspheres was faster than that of PLGA because the degradation rate of PLHMGA is more rapid than PLGA. This study shows that prolonged release of gentamicin can be obtained by loading this drug into microspheres made of biodegradable aliphatic polyesters.     

Combining Enzymatic Esterification with Conventional Alkaline Transesterification in an Integrated Biodiesel Process

An integrated biodiesel process that combines enzymatic esterification and alkaline transesterification is suggested. With focus on the enzymatic step, the paper provides proof of concept and suggestions for further process development. Hence, palm fatty acid distillate (PFAD) has been enzymatically converted to fatty acid methyl esters in a two-step process using the immobilized lipase Novozym 435 in packed-bed columns. With only a small excess of methanol, the first reaction stage could reduce the free fatty acid (FFA) content from 85% to 5%. After removal of water by simple phase separation, it was possible to lower the FFA content to 2.5% in a second reaction stage. Both reaction stages are relatively fast with suggested reaction times of 15 min in column 1 (productivity 10 kg/kg/h) and 30 min in column 2 (productivity 5 kg/kg/h), resulting in 15% FFA after column 1 and 5% FFA after column 2. A lifetime study indicated that approximately 3,500 kg PFAD/kg Novozym 435 can be treated in the first reaction stage before the enzyme has become fully inactivated. With further optimization, the enzymatic process could be a real alternative to today’s sulfuric acid catalyzed process.     

Classification of protein binders in artist's paints by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry: an evaluation of principal component analysis (PCA) and soft independent modelling of class analogy (SIMCA)

Proteomics techniques are increasingly applied for the identification of protein binders in historical paints. The complex nature of paint samples, with different kinds of pigments mixed into, and degradation by long term exposure to light, humidity and temperature variations, requires solid analysis and interpretation methods. In this study matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectra of tryptic-digested paint replicas are subjected to principal component analysis (PCA) and soft independent modelling of class analogy (SIMCA) in order to distinguish proteinaceous binders based on animal glues, egg white, egg yolk and milk casein from each other. The most meaningful peptide peaks for a given protein class will be determined, and if possible, annotated with their corresponding amino acid sequence. The methodology was subsequently applied on egg temperas, as well as on animal glues from different species. In the latter small differences in the MALDI-TOF mass spectra can allow the determination of a mammal or sturgeon origin of the glue. Finally, paint samples from the 16(th) century altarpiece of St Margaret of Antioch (Mlynica, Slovakia) were analysed. Several expected peaks are either present in lower abundance or completely missing in these natural aged paints, due to degradation of the paints. In spite of this mammalian glue was identified in the St Margaret samples.     

Determination of cholesterol and triglycerides in serum lipoproteins using flow field-flow fractionation coupled to gas chromatography-mass spectrometry

Asymmetric flow field flow fractionation (AsFlFFF) was combined with pyrolysis-gas chromatography mass spectrometry for a sized based fractionation and a detailed compositional study of the triglycerides and cholesterol associated with the various lipoprotein subclasses present in human serum. Serum samples were injected in the AsFlFFF instrument and fractionated with a time-delayed exponential decay cross flow program. The fractions collected after AsFlFFF elution were injected into a programmable temperature vaporizer (PTV) GC-injector, containing a fritted liner. A temperature and split-flow program for the PTV injector was optimized for the thermally assisted hydrolysis and methylation of the compounds of interest. The resulting fatty acid and cholesterol methyl esters were separated by GC and characteristic fragment ions were detected by MS. The system was optimized and calibrated with triglyceride and cholesterol standards for quantitative analysis. The possible interference by phospholipids with the quantitative results was investigated and found to be of minor importance.The concentrations and lipoprotein profiles of triglycerides and cholesterol were determined in a pooled serum sample of healthy volunteers and a serum sample of a sepsis patient. The results obtained with the GC–MS approach were compared with those of a previously developed method based on AsFlFFF with a dual enzymatic reaction detection system. A good agreement of the profiles was found, for cholesterol as well as for the triglycerides, even when the GC–MS method quantifies the fatty acids while with the enzymatic reaction method the glycerol concentrations are determined. Total cholesterol and triglyceride concentration values for the serum samples showed good agreement with the results of the standard enzymatic method as used in practice in the university hospital.     

Role of the channel geometry on the bubble pinch-off in flow-focusing devices

The formation of bubbles by flow focusing of a gas and a liquid in a rectangular channel is shown to depend strongly on the channel aspect ratio. Bubble breakup consists in a slow linear 2D collapse of the gas thread, ending in a fast 3D pinch-off. The 2D collapse is predicted to be stable against perturbations of the gas-liquid interface, whereas the 3D pinch-off is unstable, causing bubble polydispersity. During 3D pinch-off, a scaling w_(m) approximately tau(1/3) between the neck width w_(m) and the time tau before breakup indicates that breakup is driven by the inertia of both gas and liquid, not by capillarity.     

Adsorptive separation using self-assembly on graphite: from nanoscale to bulk processes

Adsorptive separation is a promising lower-energy alternative for traditional industrial separation processes. While carbon-based materials have a long history in adsorptive removal of organic contaminants from solution or gas mixtures, separation using an adsorption/desorption protocol is rarely considered. The main drawbacks are the limited control in bulk adsorption experiments, as often all organic molecules are adsorbed, and lack of desorption methods to retrieve the adsorbed molecules. Using high-resolution on-surface characterization with scanning tunneling microscopy (STM), an increased understanding of the on-surface adsorption behavior under different conditions was obtained. The insight obtained from the nanoscale experiments was used to develop a highly selective separation method using adsorption and desorption on graphite, which was tested for the separation of quinonoid zwitterions. These experiments on adsorptive separation using self-assembly on graphite show its potential and demonstrate the advantage of combining surface characterization techniques with bulk experiments to exploit different possible applications of carbon-based materials.

Insights from high-resolution on-surface characterization techniques are used to improve the control over adsorption and desorption on graphite in bulk adsorptive separation processes.     

Structure Elucidation of the First Sex-Inducing Pheromone of a Diatom

Diatoms are abundant unicellular microalgae, responsible for ≈20 % of global photosynthetic CO₂ fixation. Nevertheless, we know little about fundamental aspects of their biology, such as their sexual reproduction. Pheromone-mediated chemical communication is crucial for successful mating. An attraction pheromone was identified in the diatom Seminavis robusta, but metabolites priming cells for sex and synchronizing search and mating behavior remained elusive. These sex-inducing pheromones (SIP) induce cell cycle arrest and trigger the production of the attraction pheromone. Here we describe the challenging structure elucidation of an S. robusta SIP. Guided by metabolomics, a candidate metabolite was identified and elucidated by labeling experiments, NMR, ESI MSⁿ analyses, and chemical transformations. The use of negative ion mode MS was essential to decipher the unprecedented hydroxyproline and β-sulfated aspartate-containing cyclic heptapeptide that acts in femtomolar concentrations.     

Dielectric relaxation spectroscopy of amorphous and liquid-crystalline side-chain polycarbonates

The molecular dynamics of amorphous and liquid-crystalline (LC) side-chain polycarbonates was studied by dielectric spectroscopy at frequencies from 10⁻² to 10⁶ Hz and at temperatures from −160 to 180°C. ‘Model’ compounds containing no mesogenic side-groups showed two relaxations, which originate from the carbonate group (α, β_m-relaxation). By contrast, in LC-polycarbonates bearing a mesogenic nitrostilbene side group around and above the glass transition temperature T_g up to three relaxation modes were distinguished (α-, λ₁-, λ₂-process); below T_g four secondary relaxations (γ-, β_m-, β_s-, β_sc-relaxation) were observed. The γ-relaxation was found only in compounds possessing an aliphatic spacer linked to the backbone by an ether bond. Apart from β_m-, two additional β-processes were identified as relaxations associated with the mesogenic unit in the glassy (β_s) or in the crystalline state (β_sc).     

Adsorption kinetics of protein mixtures. A tentative explanation of “the vroman effect”

A model for protein adsorption kinetics is presented. This model includes diffusion limited adsorption, adsorption and desorption rate constants which are dependent on the surface concentration and an interaction term for the mutual influence of the adsorbed protein molecules. It is shown that, in first approximation, the values of the adsorption and desorption rate constants are exponential functions of the surface concentration. Assuming an adequate interaction term it is possible to show with this model for the adsorption kinetics of a mixture of proteins that the ratio of the adsorbed proteins is strongly dependent on the overall surface concentration even if the ratio of the bulk concentrations of these proteins is kept constant. Differences in interaction terms for the different proteins offer a possible explanation for the peculiar behaviour of plasma protein adsorption on a surface at different dilutions of the plasma, the so called “Vroman effect”.