3. Carriers

Cell immobilization in fixed-bed reactors with macroporous carriers is fairly simple compared to other methods such as entrapping in gels (e.g., alginate). Cell loading is often carried out by simply pumping a cell suspension through the bed of carriers, and cells are kept under same physiological conditions for the immobilization. As only the natural properties of the surface and cells interact, there are no toxic effects arising from activating reagents compared to cell entrapment within polymers. Additionally, high load of cells can be avoided by desorption of cells from

Process parameters that have to be optimized during process development comprise selection of carriers, medium selection, appropriate flow velocity, and longterm performance, among others. All these information are required to evaluate the overall performance, e.g., productivity, and to layout the scale-up strategy. In the following, a platform for development of processes for immobilized cells is introduced (Figure 2). As a start, suitability of different carriers can be compared in a small-scale multi-well system. After this, bioreactor systems of different sizes can be used to work out the required process parameters. The first, very small scale of 10 mL working volume is the multi-fixed-bed bioreactor "Multiferm" [9]. The next step is an axial-flow 100 mL fixed-bed system, which can be operated continuously with reasonable effort to investigate the performance and long-term stability of the culture [6]. As a first approach for scale-up, a radial-flow 1 L fixed-bed reactor is applied [6, 10]. Even if this is probably not the final industrial scale, the reactor system has already been the main characteristics of a large-scale system, mainly the radius. For further increase of the volume, just the height has to be increased [11]. For all three systems, a "proof of concept" has been shown before [10]. In Chapter 4 the performance of these three fixed-bed systems is compared for fixed-bed cul-

Platform for development of processes for immobilized cells. From left to right: multi-well plates with special inserts for the first evaluation of appropriate carriers under high-throughput, static conditions; multi-well flow chamber for evaluation of carriers under flow conditions; multi-fixed-bed reactor "Multiferm" (max. 12 small fixed-bed units containing approx. 10 mL carriers for evaluation of carriers under different process parameters (type of carrier, flow rate, medium, oxygen concentration, pH, etc.)); 100 mL axial-flow fixed-bed for longterm continuous culture under steady-state conditions; and 1 L radial-flow fixed-bed representative for a pilot

scale for long-term continuous cultures under steady-state conditions.

the solid surface to the cell suspension.

Growing and Handling of Bacterial Cultures

2.2 Concept for design and operation

tures of LABs.

Figure 2.

84

In cell immobilization, properties of the carrier materials play an important role. This type of immobilization on solid synthetic materials firstly has the advantage that the microorganisms attach independently to the carrier (interaction with the surface) and thus no additional process steps and reagents are required for immobilization. At this point, carrier materials have to demonstrate several certain characteristics. Atkinson et al. [12] and Pörtner and Märkl [5] summarized these properties for cell immobilization such as simple and nontoxic material, high cell loading capacity, mechanical stability, stable at appropriate operational pH values, autoclavable, resistant to microbial degradation, cost appropriate to the application, density appropriate to reactor type used, as well as reusable, if possible. Examples are given in [4, 6]. In our own studies, carriers made of glass [Siran (QVF, Mainz, Germany), VitraPOR® (ROBU® Glasfilter-Geräte GmbH)] or ceramics [(CERAMTEC EO 19/30 (CeramTec, Marktredwitz, Germany) (Figure 3) or Sponceram (Zellwerk, Oberkrämer, Germany)] were applied. All carriers showed similar results with respect to immobilized cell density and lactic acid productivity for immobilized LAB strains [6].

#### Figure 3.

Examples for carriers applied in fixed-bed cultures: (A) CERAMTEC EO 19/30 (α-aluminum, ring, diameter 3–8 mm, height 8 mm, porosity 65%; manufacturer, CeramTec; up-right, carrier; up-left, carrier in fixed-bed cultivation of Lactococcus lactis; down, SEM of carrier); (B) VitraPOR® (glass, sphere, diameter 4 mm; manufacturer, ROBU; up-left, carrier; up-right, carrier in fixed-bed cultivation of L. lactis; down, SEM of carrier).
