Ignacio de Ory, Gema Cabrera, Martin Ramirez, and Ana Blandino
In this chapter, protocols and details for the immobilization of a model cell onto polyurethane foam carriers are provided in order to facilitate the use of such systems in laboratory or industrial reactors. Polyurethane foam has recently acquired great relevance as a carrier for its good mechanical properties, high porosity, and large adsorption surface. In addition, it has a very low commercial cost. Two different immobilization protocols have been described, differing in the flow regime or the possibilities for the reactor: immobilization in a stirred tank reactor working in a discontinuous regime (by cycles) and immobilization in a packed column working in continuous operation mode. Protocols for carrier sterilization, analytical methodology, and immobilization are described.
Key Words: Adsorption; packed column; polyurethane foam; stirred tank. 1. Introduction
Adsorption onto surfaces represents a particular form of cellular adhesion based on the ability of certain microorganisms to fix themselves to solid surfaces by means of natural physicochemical bonds (1). Polyurethane foam (PUF) has recently acquired great relevance as a carrier in this immobilization technique, a fact exemplified by the increasing number of applications reported in literature. These applications include removal of organic compounds (2-6), odor waste control (7), acetic acid fermentation (8), hydrocarbon removal (9), and ferrous sulfate oxidation (10). In such studies, several microorganisms were immobilized (or coimmobilized) onto this carrier using different types of reactors such as stirred tanks, packed columns or biofilters.
PUF is an inert material with good mechanical properties (high resistance and elasticity) and very low commercial cost. This material also has a high porosity (near 97%) and therefore has a large adsorption surface. In addition to this, PUF does not suffer from the scale-up drawbacks experienced with other encapsulation matrices because huge volumes of this carrier can be easily prepared. Another
important advantage of this immobilization support is that oxygen diffusion problems can be reduced as a result of its large pore size, which is a particularly relevant factor for aerobic microorganisms. Cubic units of commercial PUF and an electron micrograph of its internal structure are shown in Fig. 1.
One aspect that must be pointed out in order to understand the phenomena involved in the immobilization process on PUF is the adsorption-desorption equilibrium in the biofilm. When the immobilized biofilm is forming (adsorption) a continuous loss of cells is simultaneously occurring (desorption), especially if the reactor is energetically stirred. For this reason, a suitable stirring rate must be established to find a satisfactory balance between the two effects. On the one hand, vigorous stirring improves the general conditions for mass transfer to submerged biomass and, as a consequence, increases the total biomass population and the number of cells adsorbed onto the carrier. However, both erosion effects and collision tensions can be increased by turbulent stirring, seriously hindering surface cell adsorption processes (11,12). At the same time, the amount of interstitial biomass (cells submerged in the liquid remaining within the internal structure of PUF) constitutes a crucial source of potential immobilized biomass for the continuous adsorption-desorption equilibrium and, in its own right, has great relevance for the success of subsequent fermentations (13).
In this chapter, protocols and details for the immobilization of a model cell onto PUF carriers will be provided in order to facilitate the use of such systems in laboratory or industrial reactors. Not all cells have the ability to fix themselves to solid surfaces, but they usually take advantage of the symbiotic action of other microorganisms in a mixed culture. These other microorganisms are adhered but do not contribute to the biological reaction. A number of studies concerning immobilization with mixed cultures or co-immobilization are reported in the literature (14). In general, all bacteria have this single adsorption capacity and so, in order to develop a more generic model, aerobic bacteria will be considered.
Two different immobilization protocols will be discussed and these differ in the flow regime or the possibilities for the reactor: (1) immobilization in a stirred tank reactor working in a discontinuous regime (by cycles) and (2) immobilization in a packed column working in continuous operation mode.
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