By C. Todd Lopez
WASHINGTON (May 09, 2012) -- At an Army medical lab in San Antonio, the observed stability of a recently concocted "microemulsion" could one day be key to the long-term stockpiling of important vaccines for distribution in case of disaster.
Maj. Jean M. Muderhwa, a biochemist who serves as deputy director of the lab at the Department of Clinical Investigation at Brooke Army Medical Center, formulated the mixture which appears robust for both longevity and temperature variations.
The compound was stored at both room temperature and in the refrigerator with equal results. "They have been stable for eight months; so storing at room temperature does not hurt," he said.
Muderhwa recently presented his findings at the annual meeting of the American Society for Biochemistry and Molecular Biology.
Some vaccines are created in the form of an emulsion, a mixture of an oil and water that requires high energy input for its formulation. While a vaccine is in storage, the ingredients of the emulsion protect and preserve the "protein antigen" portion of the vaccine, its active component.
The emulsion also serves as a medium to deliver the antigen into the body. Additionally, the emulsion can help the antigen stick locally inside the body for a longer time. That gives the immune system of an individual or patient more time to react to the antigen.
But emulsions can separate due to time and temperature changes. When that happens to vaccines in storage they may have to be destroyed, Muderhwa said. Finding an emulsion that is more robust is important to creating vaccines that can be shelf-stable for longer periods of time.
"If you make an emulsion containing the vaccine, the emulsion only lasts a few weeks or few months," Muderhwa said. "When they separate, the components begin to degrade; either by oxidation or other means."
Muderhwa has worked to develop an emulsion that forms readily and sometimes spontaneously and that would last longer by being thermodynamically stable. One way to do that is with a microemulsion. The particles of oil in the microemulsion are smaller than in a regular emulsion.
"When you have an emulsion of the oil-in-water type, and you measure the particle size of the oil, you can make those particles smaller and smaller by adding a co-surfactant, such as a short chained alcohol, for example, isopropanol or pentanol," he said. "By doing that, you ensure the flexibility of the interfacial layer leading to decreased interfacial tension to nearly zero. The result is a transparent solution. And this solution is thermodynamically stable."
In Muderhwa's microemulsion, he didn't use isopropanol or pentanol because of its toxicity if administered internally and because of its effect of destabilizing the system with time. Instead, he used glycerol as the "co-surfactant." A co-surfactant works in addition to two pharmaceutically-acceptable surfactants, Span 80 and Tween 60, in the microemulsion.
"The co-surfactant works with the surfactants to reduce the surface tension to near zero," Muderhwa said. "In an emulsion, the surface tension is not zero. So the molecules repeal themselves with time until they fail because of the tension. So they give up. They fail to stay together for a long time. In a microemulsion, since you are suppressing that interfacial tension, the particles stick closer to one another. "
A vaccine, however, is not simply oil, water, surfactants and in the case of Muderhwa's compound, the co-surfactant glycerol. A vaccine must also contain its active component, the protein antigen, and also sometimes adjuvants such as aluminum hydroxide and aluminum phosphate. While Muderhwa saw that the microemulsion he'd created was stable, he'd also need to add the additional important components to make it a vaccine.
"Microemulsions are very sensitive to change in composition. If you add an extra compound, they also separate quickly like in the case of emulsions," he said. "But what I have found is that, if I add aluminum adjuvant compounds, which are the only ones approved by the Food and Drug Administration for use in human vaccines like influenza, and I mix that microemulsion with the aluminum compounds, it is still stable".
Thus, Muderhwa's concoction is the result of a composite adjuvant formulation consisting of a microemulsion containing aluminum-adsorbed model protein antigens and immunogenic agents, such as microbial cell components, for example monophosphyl lipid A. Muderhwa used both aluminum hydroxide and aluminum phosphate in his compound.
Just how long will the components of the microemulsion stay emulsified? It's been less than a year at this point, but Muderhwa thinks the microemulsion he's put together in his lab could be stable for a long, long time. "I think years. Ten or fifteen or twenty years," he said.
Longer stability for a vaccine means that large amounts of vaccines could be manufactured and stored for emergency use.
"The Army can have this vaccine to be distributed to Soldiers and to the population," he said. "I think the future will be to make, for example, a vaccine against biological weapons. You can make a vaccine containing anthrax or containing neurotoxins, and stockpile them."
Muderhwa's research opens up new possibilities in vaccine research.
"In contrast to emulsions, microemulsions are thermodynamically stable and are easy to formulate," he said. "Also, given their larger interfacial area, i.e., large surface-to-volume ratio, these dispersions will provide the opportunity to study the effect of increasing the surface area increase on the immune response."
Right now, though, the work is still in the experimental stages, he said, and it could be some time before there is benefit to the Army.
"This is just a formulation to be used to test on mice. If it works in mice, I will have to apply for an investigational new drug with the FDA, to allow me to start clinical trials in humans," he said. "It's a long process that will take time."