Phytosterols and lycopene are neutraceuticals that are known antioxidants and cancer inhibitors. Unfortunately, they do not dissolve readily in body fluids and therefore present unique problems concerning their uptake by the blood stream. This project explores the ability to enhance the rate of solubility of lycopene and phytosterols by formulating tham as nano- and micro-particles. OBJECTIVES: Phytosterols and lycopene are known antioxidants and cancer inhibitors but they are lipophilic and therefore present unique problems concerning their uptake by the blood stream. The overarching problem is that there are a limited number of techniques for enhancing the solubility of lipophilic nutrients. This research focuses on the formulation of poorly water soluble compounds as micro- and nano-size particles. Due to their sub-cellular and sub-micron size, nanoparticles can penetrate deep into tissues through fine capillaries and are generally taken up efficiently by the cells. It is only within the last two years that a new technology has brought the production of nano- and micro-particles to an economic basis that offers a unique opportunity to formulate micro-powders for nutraceuticals and food products. Aktiv-Dry's proprietary Carbon Dioxide Nebulization with a Bubble Dryer (CAN-BD) technology uses near-supercritical carbon dioxide as a nebulizing agent but harvests nano- and micro-products at relatively low temperatures and at atmospheric pressure. Our goal for this Phase I SBIR project is to produce nano- and/or micron-size phytosterol and lycopene particles using laboratory scale CAN-BD technology. The experimental program we propose will allow us to evaluate the need for, or the advisability of, adding excipients, surfactants or other compounds that alter the fluid dynamic properties of the solution and/or the emulsion formed when it contacts near supercritical carbon dioxide. Our experiments will also provide insight into the complex mechanisms of CAN-BD nebulization and how they are influenced by process parameters such as drying temperature and liquid flow rates. We will integrate the new knowledge provided by the proposed experiments with results obtained from our previous research with pharmaceutical and biotech compounds to seek optimization of the CAN-BD process for phytosterol and lycopene particle designs. The specific objective of Phase I research is to test the hypothesis that the CAN-BD process can produce phytosterol and lycopene micro-powders. To achieve this objective, our experimental program will determine the effects of processing parameters (e.g., pressure, temperature, flow rates, etc.) and preformulation conditions on the physical properties of powders. The experimental program confirms the hypothesis only if we satisfy all of the following conditions: 1. We can determine preformulation and operating conditions and identify restrictor dimensions that provide a continuous, stable plume of microbubbles that form fine particles of phytosterol and/or lycopene. 2. We can determine preformulation and operating conditions that assure the fine particles do not aggregate significantly during drying. 3. We can collect the particles as a fine powder. 4. Particle size is distributed in the range 100 nm < davg < 5 microns. 5. The rate of dissolution of the compound in water and/or organic solvents after processing significantly exceeds the solubility before processing. APPROACH: We will use CAN-BD laboratory apparatus, similar to that used traditionally by Aktive-Dry and its associates, to produce micron size phytosterol and lycopene particles from solution concentrates. This apparatus will produce a continuous synthesis of aerosol powder through the simultaneous use of two pumps and desolvation of the aerosol, followed by particle collection on a filter. In a typical experiment, phytosterols will be dissolved in ethanol at concentrations varying from 0.1 to 3 % by weight and the solution delivered to a low-dead-volume tee at a rate of approximately 0.3 ml/min. Several techniques may be used to accomplish this. One relatively simple approach is to fit a free floating piston into a high pressure Thar vessel that contains the solution in the downstream volume and water or solvent in the upstream volume. The flow rate is set by either an HPLC or syringe pump containing the barely compressible water or solvent. In the tee, the solution will mix intimately with a stream of near-critical CO2 which has been pressurized to ~ 100 atm at room temperature by a syringe pump. The resulting emulsion will then expand through a restrictor (made of fused silica, PEEK, or stainless steel) to produce a plume consisting of micron-size bubbles. The plume will discharge into a two-liter glass drying chamber maintained at approximately atmospheric pressure. Abrupt decompressive flashing will cause the micro-bubbles to burst and form tiny droplets of phytosterol solution. Warm (30 - 65 C) nitrogen gas flowing through the chamber will dry the droplets to produce fine phytosterol particles that will collect on a filter membrane (likely pore size 0.45 micron) which is attached to the bottom of the drying chamber. Similar experiments will also be carried out with commercial lycopene concentrates. Samples of phytosterol and lycopene powders will be analyzed by a scanning electron microscope (ISI, model SX-30). The aerodynamic particle size distribution will be measured using a Model 3225 TSI Aerosizer, which uses a laser time of flight principle. Alternatively, particle size distributions may be determined from an Andersen Cascade Impactor or dynamic light scattering instrument manufactured by Malvern Instruments. If raw materials (phytosterol and lycopene) are supplied in powder form, these tests will be carried out before and after the CAN-BD process to evaluate the effect of nanonization and micronization on solubility and the particle size distribution and morphology. Nearly twin laboratory-scale CAN-BD apparatus will be assembled at Aktiv-Dry and at Oklahoma State University and dedicated to this project. An initial series of experiments conducted independently at each location will confirm the experimental approach. Thereafter, we anticipate that each laboratory will focus on different specific research objectives.
