Dissolution of multicomponent microbubbles in the bloodstream: 1. Theory.

The problem of dissolution of a bubble in the bloodstream is examined. The bubble is assumed to be filled with a mixture of a sparingly water-soluble gas (osmotic agent) and air. The dissolution of the bubble has three definite stages. In Stage 1, the bubble quickly swells in air. The swelling ratio depends on the surface tension, blood pressure, level of oxygen metabolism and initial mole fraction of osmotic agent in the bubble. In Stage 2, the osmotic agent slowly diffuses out of the bubble. The squared radius decreases nearly linearly with time, at a rate proportional to the Ostwald coefficient and diffusivity of the osmotic agent. In Stage 3, the partial pressure of the osmotic agent becomes so high that it condenses into a liquid. In order to prolong the lifetime of 5-micron bubbles in the bloodstream from < 1 s (as found with pure air), the osmotic agent must have a low Ostwald coefficient (< or = 10(-4)) and a relatively high saturated vapor pressure at body temperature (> or = 0.3 atm = 3 x 10(4) Pa).

Dissolution of multicomponent microbubbles in the bloodstream: 2. Experiment.

The effect of the nature of the filling gas on the persistence of microbubbles in the bloodstream was studied. All the microbubbles were covered with the same shells. Various perfluorocarbons and perfluoropolyethers alone and as mixtures with nitrogen were used as the filling gases. The persistence time of microbubbles in the bloodstream tau increased with the molecular weight of the filling gas, from approximately 2 min for perfluorethane, to > 40 min for perfluorodiglyme, C6F14O3, and then decreased again to 8 min for C6F14O5. An acceptable ultrasound scattering efficacy was exhibited by the filling gases with intermediate molecular weights that possessed both a high saturated vapor pressure and a comparatively low water solubility (Ostwald coefficient). On the basis of the experimental data, it is concluded that the microbubble persistence tau is controlled primarily by the dissolution of microbubbles and not by the removal of the microbubbles by the reticular endothelial system. Although the qualitative experimental trends are in good agreement with the theoretical model developed previously, there are some quantitative differences. Possible reasons for these differences are discussed.

Influence of perflubron emulsion particle size on blood half-life and febrile response in rats.

Perfluorochemical (PFC) emulsions are particulate in nature and, as such, can cause delayed febrile reactions when injected intravenously. This study investigated the influence of emulsion particle size on intravascular retention and on body temperature changes in unrestrained conscious rats. Concentrated (60% to 90% w/v) emulsions based on perflubron (perfluorooctyl bromide [PFOB]) with mean particle sizes ranging from 0.05 microns to 0.63 microns were tested. Rats were fitted with a chronic jugular catheter and an abdominal body temperature telemetry unit. Fully recovered, conscious rats were monitored for 24 hours after infusion (dose = 2.7 g PFC/kg). Emulsion blood half-life (T1/2) was determined from blood perflubron levels measured by gas chromatography. Emulsions with a particle size of 0.2-0.3 microns caused fevers (6 to 8 hour duration) which peaked at 1-1.5 degrees C above normal (approximately 37.5 degrees C). Fevers could be blocked by i.v. treatment with either cyclooxygenase inhibitors (ibuprofen) or corticosteroids (dexamethasone). Both intensity and duration of the temperature response, quantified by area under the temperature curve, was decreased significantly for emulsions with a particle size < or = 0.12 micron. Blood T1/2 varied inversely with particle size, and was 3 to 4 fold longer for emulsions with a mean particle size < or = 0.2 micron. Thus, smaller emulsion particles more effectively evaded the reticuloendothelial system, which resulted in longer intravascular retention, less macrophage activity, and reduced febrile responses.

Proposed mechanism of pulmonary gas trapping (PGT) following intravenous perfluorocarbon emulsion administration.

PURPOSE: To investigate various hypotheses and identify the most likely mechanism preventing the complete collapse of test animal lungs at sacrifice subsequent to intravenous injection of certain perfluorocarbon emulsions. PROTOCOL: Literature data were reviewed, experimental data were extracted from completed studies and new data were generated in an attempt to delineate reasons why, in certain animals, lungs fail to collapse normally at necropsy if previously injected with certain perfluorocarbon emulsions. The proposed hypothesis involved gas osmosis through endogenous pulmonary surfactant-liquid bridges (micro-bubbles). RESULTS: The observed effect of incomplete lung collapse upon necropsy was found to correlate with perfluorocarbon vapor pressure. Results indicated that failure to collapse could be attributed to the formation of intra-alveolar micro-bubbles induced by the normal pulmonary elimination of perfluorocarbon vapor. These micro-bubbles result in a phenomenon which could be characterized by the term, pulmonary gas trapping. Reduction of the perfluorocarbon concentration gradient across the bubble films by exposure to a perfluorocarbon vapor-containing atmosphere was found to reduce the effect in-vivo and prevent gas osmosis bubble growth in-vitro. CONCLUSION: Experimental observations are consistent with the proposed theory of perfluorocarbon-related gas osmosis through micro-bubbles that prevent complete lung collapse as observed upon opening the thoracic cavity of test animals.

Oxidative assessment of phospholipid-stabilized perfluorocarbon-based blood substitutes.

Methods based on the HPLC separation with subsequent UV detection and spectrofluorimetry have been developed to monitor the formation of oxidative decomposition products of phospholipids in perfluorocarbon emulsions. Catalytic, as well as emulsion oxidative stability studies have been conducted utilizing egg yolk phospholipid (EYP) and perfluorocarbons of varied compositions/purity in order to assess their effect on susceptibility to oxidation. Our studies indicate that phospholipid composition, degree of unsaturation, perfluorocarbon purity and the presence of oxygen and trace metals have a significant effect on the formation of oxidative decomposition products. A combination of methods has proven useful in monitoring the levels of oxidative decomposition products of phospholipids in phospholipid-stabilized perfluorocarbon-based blood substitutes. Such an approach has proven beneficial in the development of pharmaceutical agents of potentially higher quality and storage stability.

Effects of buffer pH and phosphate concentration on the droplet size and EYP hydrolysis of perflubron/EYP emulsions.

Oil-in-water emulsions containing perflubron (perfluorooctyl bromide; PFOB) and stabilized with egg yolk phospholipid (EYP) have potential applications as contrast agents and oxygen carriers. In this study, the effects of buffer pH and total phosphate concentration on the emulsion droplet size and EYP hydrolysis were evaluated. 90% w/v perflubron emulsions with NaH2PO4-Na2HPO4 buffers of different pH (4.7-8.7) and phosphate concentrations (30 and 60 mM) were prepared with a high-pressure homogenizer. Emulsions were stored at 40 degrees C and tested at 0, 1, 2 and 3 months. The pH dropped quickly in emulsions with pH 8.7 buffer whereas acidic and neutral buffered emulsions exhibited minor pH drops. The concentration of free fatty acids (FFA) vs emulsion pH can be fitted to a parabolic curve with a minimum at about pH 6.0. The droplet growth rates in emulsions with the pH 4.7 buffer were about 2.5 times of those in emulsions with the pH 8.7 buffer. Total phosphate concentration had only a minor effect. This study emphasizes the importance of the careful selection of buffer pH and capacity to control EYP hydrolysis and possibly emulsion droplet size.

Effects of formulation, processing and storage parameters on the characteristics and stability of perflubron emulsion.

In this study, the effects of formulation, processing and storage parameters on perflubron (perfluorooctyl bromide; PFOB) emulsions were investigated. Emulsions with varying concentrations of perflubron and egg yolk phospholipid (EYP) were prepared with different processing parameters and placed at different storage temperatures. Their characteristics and stability were compared. The emulsion droplet growth rate was nearly proportional to the perflubron percentage in the range of 15-110% w/v. The initial droplet size of perflubron emulsions was inversely proportional to the concentration of EYP until a certain lower limit of droplet size was reached. The initial droplet size and droplet growth rate of perflubron emulsion were strongly dependent upon the processing parameters. The logarithmic value of the droplet growth rate decreased linearly with l/T in the range of 5-40 degrees C. The formulation and processing parameters are the key variables to be optimized to achieve better emulsion characteristics and stability.

Development of highly fluid, concentrated and stable fluorocarbon emulsions for diagnosis and therapy.

A challenging aim in developing injectable fluorocarbon emulsions is to combine good flow characteristics (especially at low shear rates) with the high fluorocarbon concentration required for high oxygen delivery or effective contrast in imaging, long shelf life, and biological acceptability. A good balance of these sometimes conflicting objectives has been achieved with 90% w/v concentrated emulsions of various fluorocarbons, including the radiopaque oxygen carrier perfluorooctylbromide (PFOB, perflubron). The sterile emulsions have viscosities of about 20 cPs at a shear rate of 1 sec-1; the viscosity decreases rapidly with fluorocarbon concentration, and at 60% w/v the viscosity is less than that of human blood. The emulsions are suitable for injection as prepared, and are stable unfrozen for over a year.

Stabilization of perflubron emulsions with egg yolk phospholipid.

Egg Yolk Phospholipid(EYP) has been used extensively as the primary surfactant in parenteral fat emulsions for many years. The simplicity, functionality and physiologic tolerance of EYP has contributed greatly to its success in the intravenous emulsion arena. The mechanism of stabilization in triglyceride emulsions is well understood; however, this is not the case with perfluorocarbon emulsions. Interfacial models, as well as emulsion stability studies, have been conducted utilizing EYP of varied composition in order to derive a structure/function relationship. Our studies indicate that minor components, total unsaturation, acyl chain length and presence of charged species have significant impact on the functional properties of EYP and the subsequent stability of the emulsion product. These findings contribute to our ability to design and manipulate natural surfactants with superior properties for use in medical applications of perfluorocarbon emulsions.

Drop size stability assessment of fluorocarbon emulsions.

The aging of fluorocarbon emulsions prepared with natural egg yolk phospholipids (EYP) has been studied and a linear variation (r2 greater than 0.95) of the mean average volume of the droplets with time has been observed. The slope of the experimental lines, called "Stability Parameter, S" can thus be taken as a representation of the rate of aging of the emulsions. Examples are given of use of parameter S to assess the effect of formulation and processing parameters on the stability of diverse fluorocarbon emulsions. S is a useful tool to compare emulsions and ascertain any factors of stabilization/destabilization.