Air-filled polymeric microcapsules from emulsions containing different organic phases.

Air-filled polymeric microcapsules for use as a contrast agent in ultrasonography have been prepared by the freeze-drying of different oil-in-water emulsions. The water phases consisted of a block copolymer in water. The organic phases consisted of a biodegradable polyester dissolved in (-)-camphene, cyclooctane, cyclohexane or tricyclene, which were relatively poor solvents for the polyester. A polymeric wall was, therefore, precipitated at the droplet surface early in the process, i.e. during freezing. Removing the solvent during freeze-drying, resulted in air-filled microcapsules. The microcapsules were suspended in saline after freeze-drying. All the suspensions contained echogenic microcapsules with a volume mean diameter of approximately 5-7 microm. Microscopic investigations showed that the microcapsules were spherical and hollow. Tricyclene and, to some degree, (-)-camphene were found unsuitable for industrial production due to melting points above 30 degrees C. Cyclooctane and cyclohexane were investigated as replacements for the initially chosen (-)-camphene, since they are liquids over a wider temperature range. These solvents gave improved yields, measured both as particle volume concentration per amount of polymer in suspension and acoustic attenuation at 3.5 MHz per amount of polymer in suspension, although the freeze-drying cycle was not optimized for these systems.

Preparation of polymeric microcapsules: formulation studies.

Air-filled microcapsules were prepared by freeze-drying different oil-in-water emulsions containing biodegradable polyester as the wall-forming material. The aim of this work was to find an acceptable formulation with respect to the microcapsule suspension and the stability of the emulsion during the production process. The influence of various formulation parameters (concentrations of mannitol, polymer, and surfactant; pH; oil-in-water phase ratio) was investigated in a factorial design. The results were treated by ordinary least-square (OLS) regression and partial least-square regression (PLSR). In a previous work, air-filled microcapsules were successfully made using human serum albumin as the surfactant in the emulsion (1). In the present work, a new block copolymer based on poly(ethylene glycol) (PEG) was implemented as the surfactant to replace human serum albumin. It was found that the new block copolymer is a suitable replacement for human serum albumin. The concentration of the polymer in water and the concentration of the surfactant in the oil phase and the interaction between these variables had a significant influence on the stability of the emulsion at 60 degrees C. A surfactant concentration of approximately 2% (w/v) in water was necessary when the concentration of the wall-forming polymer was below 5% (w/v) in (-)-camphene. The concentration of the polymer in the oil phase influenced the yield, measured as the volume concentration of particles in suspension per milligram of polymer added and as acoustic effect per milligram of polymer. Low levels of polymer concentration in (-)-camphene (< 5% w/v) gave the highest yield. Excess polymer in the oil phase did not form microcapsules, but precipitated in the suspension or was included in the wall of the microcapsules. Addition of mannitol protected the microcapsules from being destroyed during freeze-drying and resulted in freeze-dried products with few cracks, little shrinkage, and higher suspension yield.

Characterization of E. coli cell disintegrates from a bead mill and high pressure homogenizers.

The protein releases, the particle size distribution and the viscosity of disrupted E. coli suspensions from Dyno Mill KDL, Manton Gaulin 15 M-8TA and Microfluidizer M-110 were determined. The effects of these parameters on separation of the cell debris from the protein solution by centrifugation and by filtration were also examined. All three disintegration methods investigated give approximately the same protein and enzyme releases but considerably different physical properties of the cell disintegrates which influences centrifugation and filtration. The separation degree of biomass during centrifugation is only slightly affected by increasing degree of disruption (increasing protein releases) in the bead mill, while an increase in the degree of disruption in the two high pressure homogenizers drastically reduces the centrifugal degree of separation. However, increasing degrees of disruption result in shorter filtration times during filtration for all three disintegration methods. The results show further that the cell concentration only has a minor influence on protein releases in the Microfluidizer high-pressure homogenizer, while an increase in the biomass content reduces the separability of the cell disintegrate both in filtration and in centrifugation.

Selective flocculation with chitosan in Escherichia coli disintegrates: effects of pH and nuclease treatment.

The flocculation of cell debris from a beta-galactosidase constitutive E. coli with chitosan as a flocculant was studied to investigate the possibility of obtaining a selective flocculation in cell disintegrates with high product recoveries. The flocculation removed 98% of the cell debris by 30 min sedimentation under gravity, which should be compared to a separation of the cell debris without flocculation of only 70% by centrifugation at 15,000 g. Optimal flocculation dosages varied between 12 and 43 mg chitosan g-1 dry weight of cells, depending on pH. The yield of the product beta-galactosidase reached 60% at optimal pH. Hydrolysis of the nucleic acids by DNAase and RNAase decreased the optimal flocculation dosages considerably. The study showed that the flocculation is somewhat selective, since chitosan also removed 85% of the nucleic acids and 50% of the proteins, which contributed to the purification of the protein solution.