Hydrogel has potential use in the controlled release of drugs, and as 3-D structure for the formation of tissue matrix. A biocompatible gel film that provides a three dimensional support for the formation of a matrix, and also delivers the biological agents is a subject of extensive investigation. Use of natural and synthetic polymers, and hydrogels are known, where the biocompatibility and the biodegradability are important requirements. A gel layer has the potential to hold a significant volume of a biological agent that can diffuse into the host tissue over a period of time. Also the gel layer, loaded with a matrix forming cell can act as a scaffold, over which the tissue regeneration takes place. In these applications, it is important that a substantial porosity is induced in the gel layer. Additionally, the porosity has to be uniformly distributed so that the pore to pore distance remains uniform. This calls for a highly ordered pore structure. Emulsion freeze drying, fiber bonding, solvent casting or particulate leaching, gas foaming, thermal phase separation, electrospinning, and use of supercritical CO2 are some of the methods to induce the voids in a gel layer. Direct introduction of bubbles using a fluidic arrangement is an alternative method that allows better control of the void size and the porosity. Under most circumstances, the bubbles generated by this method are monodisperse. The bubbles rapidly self-assemble, and provide an ordered structure. Also, the gel is not exposed to any chemical or thermal treatment by this method. The method is inexpensive, in comparison with the solid free form fabrication techniques.
In a general co-flow arrangement, the pulled capillaries are arranged one inside the other. The inner gas thread is dragged by the co-flowing liquid until the gas stream snaps to form a bubble. The other variants to this arrangement are flow focusing, where the liquid is injected in a cross-flow manner to impart a direct squeeze on the dispersed phase, or use of a series of such flow focusing joints. In this presentation, a co-flow device is described, where a second constriction beyond the tip of the inner capillary (referred as orifice in throat) ensures further splitting of the bubbles. In traditional co-flow arrangements, the bubbles were found to be an order of magnitude larger than the feature size of the device. Introduction of a second squeeze enables achieving a smaller bubble size, on the order of the feature size of the device. The disengagement of bubbles from such device requires the collapse of the neck that holds the bubble to the tip of the nozzle. Since the collapse is instability driven, a bifurcation from monodisperse cluster to higher levels of dispersion may occur. Other than the velocities, the type and consistency of the fluid may influence such bifurcations.
This presentation reports baseline experiments with glycerol in water, and follow-up experiments with two biopolymers, swelled in water. These biopolymers are alginate and chitosan. Alginate is a naturally occurring polysaccharide, sourced from brown algae that grow in warm areas. Chitosan is the deacetylated form of Chitin, the second most abundantly available polysaccharide in the nature, other than the cellulose. Chitin is commonly sourced from the shells of crabs, prawns, lobsters, shrimps and exoskeleton of insects. Both of these gel systems are known for their biocompatibility, and have potential use in drug delivery and tissue regeneration. The surfactant is added to retain the bubble against coalescence. Alfa olefin sulfonate, pluronic and Lutensol AT 25 are used as surfactant. Nitrogen was used as the gas phase, and was injected into the fluidic device using a mass flow controller from Alicat Scientific U.S.A. The biopolymer, swelled in water was injected using a syringe pump from Harvard Apparatus U.S.A. The two biopolymers were further crosslinked using calcium chloride and formaldehyde respectively.
The sub-millimeter size bubbles, when collected on a petridish in a thin layer of liquid shows self-alignment in a monolayer, without coalescence, or shrinkage. An edge detection algorithm was utilized to compute the bubble size. Davis software from LaVision GmbH was utilized for image processing. The cumulative frequency of less than type as a function of the bubble diameter was fitted to Gompertz function. The derivative of this function provided the bubble size distribution. Multiple functions were considered for a single curve to explore possibilities other than unimodal distributions. The bubble size decreases with increase in liquid to gas flow ratio.
The crosslinked gel film from the petridish was dried in a vacuum oven. The image of the alginate film prior to crosslinking was acquired under digital microscope (Labomed, U.S.A.), and was compared with the images of the dried scaffolds from the scanning electron microscope (JSM 5800, JEOL Limited, Japan). The voids retained their identity at the time of drying, while the diameter got reduced to half. The shrinkage was primarily observed in the thickness of the scaffold. The information on shrinkage and the drying rate shows how the de-saturation of the various parts of the film was phased. The effective diffusivity, based on a mathematical model as function of moisture content shows the de-saturation behavior of void zone, gel matrix and film surface respectively. From the SEM images two levels of porosity are evident. The smaller pores were intrinsically provided by the gel matrix. The larger ones were induced by the fluidic arrangement. The ultimate strength of the gel film due to the introduction of voids is measured using universal testing machine (Tinius Olsen, U.K.).
Subsequently, the dried gel films are soaked in Vitamin B-12 solution. The porosity of the saturated film is measured gravimetrically using the weighing balance with below balance facility from Sartorius. The release of Vitamin B12 in water and phosphate buffer saline respectively on a shaker is studied using UV Vis Spectrophotometer (Perkin Elmer Lambda 35). For comparison, the experiment was repeated with a film that did not have any embedded void. The enhancements in uptake and release of Vitamin B12 due to the presence of voids are estimated. In case of chitosan, the uptake of Vitamin B-12 was compared for two choices of crosslinker. These are formaldehyde and glyoxal. The bubbles, placed in more than one layer enhance the porosity, and the ability to absorb Vitamin B-12 further.
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