Immobilized cell catalysts

A thermally stable NHase from Comamonas testosteroni 5-MGAM-4D (ATCC 55 744) [22] was recombinantly expressed in Escherichia coli, and the resulting transformant cells immobilized in alginate beads that were subsequently chemically cross-linked with glutaraldehyde and polyethylenimine. This immobilized cell catalyst (at 0.5 % dew per reaction volume) was added to an aqueous reaction mixture containing 32wt% 3-cyanopyridine at 25 °C, and a quantitative conversion to nicotinamide was obtained. The versatility of this catalyst system was further illustrated by a systematic study of substrates, which included... 

The enzymatic production of diverse amino acids has been described by quite a lot of companies over the last 20-30 years and many processes are based on the use of lyases [138-141]. With regard to the use of immobilized biocatalysts only two new processes shall be introduced here the production of L-as-partic acid (169) using an immobilized cell catalyst by Nippon Shokubai Ka-gaku Kogyo, Japan [142, 143] and the use of an immobilized enzyme by BioCatalytics [7] (both Scheme 55). 

From a practical standpoint, the two most important characteristics of an immobilized cell catalyst are its activity and its operational stability. The latter parameter is usually expressed in catalyst half-life. The amount of activity, say in International Units (I.U.), would be a function of cell-to-carrier ratio. As mentioned earlier, a 50% loading ratio has been found to be optimal. 

For Immobilized cell catalysts there are two possibilities to obtain this factor. Firstly, the particles of larger radius can be grinded down to such a small size that pore diffusion becomes negligible. In this case v = Vg g q Due to the size and the simple entrapment of the catalytic species loss from the matrix may be considerable. Therefore, secondly, the free cell activity can be used in the denominator, if the exact concentration of cat-... 

BB-SFG, we have investigated CO adsorption on smooth polycrystaHine and singlecrystal electrodes that could be considered model surfaces to those apphed in fuel cell research and development. Representative data are shown in Fig. 12.16 the Pt nanoparticles were about 7 nm of Pt black, and were immobilized on a smooth Au disk. The electrolyte was CO-saturated 0.1 M H2SO4, and the potential was scanned from 0.19 V up to 0.64 V at 1 mV/s. The BB-SFG spectra (Fig. 12.16a) at about 2085 cm at 0.19 V correspond to atop CO [Arenz et al., 2005], with a Stark tuning slope of about 24 cm / V (Fig. 12.16b). Note that the Stark slope is lower than that obtained with Pt(l 11) (Fig. 12.9), for reasons to be further investigated. The shoulder near 2120 cm is associated with CO adsorbed on the Au sites [Bhzanac et al., 2004], and the broad background (seen clearly at 0.64 V) is from nomesonant SFG. The data shown in Figs. 12.4, 12.1 la, and 12.16 represent a hnk between smooth and nanostructure catalyst surfaces, and will be of use in our further studies of fuel cell catalysts in the BB-SFG IR perspective. 

At the next level of abstraction are measurements performed at a thin film of fuel cell catalyst immobilized on the surface of an inert substrate, such as glassy carbon (GC) or gold (Fig. 15.2c). Essentially, three versions of this approach have been described in the fiterature. In the first case (a porous electrode ), an ink containing catalyst and Nafion ionomer is spread onto an inert nonporous substrate [Gloaguen et al., 1994 Gamez et al., 1996 Kabbabi et al., 1994]. In the second case (a thin-fihn electrode ), the ink does not contain Nafion , but the latter is... 

In recent years, extensive attention has been focused on finding cultured plant cells that can be used as catalysts for organic functional group transformations. A number of transformations employing freely suspended or immobilized plant cell cultures have been reported.24 For example, Akakabe et al.25 report that immobilized cells of Daucus carota from carrot can be used to reduce prochiral carbonyl substrates such as keto esters, aromatic ketones, and heterocyclic ketones to the corresponding secondary alcohols in ( -configuration with enantiomeric excess of 52-99% and chemical yields of 30 63%). 

Immobilized enzymes and immobilized cells can be used in principle as heterogeneous catalysts not only for the enhancement of hydrolytic reactions but also in specific synthetic reactions. Many of the enzymes involved in the synthesis of complex biological compounds -in vivo and in vitro require cofactors which have to be provided in suitable form when immobilized enzymes or cells are used. 

Immobilized enzymes and immobilized cells are being employed as specific heterogeneous catalysts by several of the chemical industries. An evaluation of the mode of action and efficiency of... 

The lure of new physical phenomena and new patterns of chemical reactivity has driven a tremendous surge in the study of nanoscale materials. This activity spans many areas of chemistry. In the specific field of electrochemistry, much of the activity has focused on several areas (a) electrocatalysis with nanoparticles (NPs) of metals supported on various substrates, for example, fuel-cell catalysts comprising Pt or Ag NPs supported on carbon [1,2], (b) the fundamental electrochemical behavior of NPs of noble metals, for example, quantized double-layer charging of thiol-capped Au NPs [3-5], (c) the electrochemical and photoelectrochemical behavior of semiconductor NPs [4, 6-8], and (d) biosensor applications of nanoparticles [9, 10]. These topics have received much attention, and relatively recent reviews of these areas are cited. Considerably less has been reported on the fundamental electrochemical behavior of electroactive NPs that do not fall within these categories. In particular, work is only beginning in the area of the electrochemistry of discrete, electroactive NPs. That is the topic of this review, which discusses the synthesis, interfacial immobilization and electrochemical behavior of electroactive NPs. The review is not intended to be an exhaustive treatment of the area, but rather to give a flavor of the types of systems that have been examined and the types of phenomena that can influence the electrochemical behavior of electroactive NPs. 

Colloidal Pt/RuO c- (C5 0.4nm) stabilized by a surfactant was prepared by co-hydrolysis of PtCU and RuCls under basic conditions. The Pt Ru ratio in the colloids can be between 1 4 and 4 1 by variation of the stoichiometry of the transition metal salts. The corresponding zerovalent metal colloids are obtained by the subsequent application of H2 to the colloidal Pt/Ru oxides (optionally in the immobilized form). Additional metals have been included in the metal oxide concept [Eq. (10)] in order to prepare binary and ternary mixed metal oxides in the colloidal form. Pt/Ru/WO c is regarded as a good precatalyst especially for the application in DMECs. Main-group elements such as A1 have been included in multimetallic alloy systems in order to improve the durability of fuel-cell catalysts. PtsAlCo.s alloyed with Cr, Mo, or W particles of 4—7-nm size has been prepared by sequential precipitation on conductant carbon supports such as highly disperse Vulcan XC72 [70]. Alternatively, colloidal precursors composed of Pt/Ru/Al allow... 

Phase transfer catalysts have been grafted onto the surface of porous capsules to facilitate product purification after reaction, and many types of immobilized cells, mycelia, enzymes, and catalysts have been encapsulated in polymers such as PDMS, PVA, or cellulose. In the specific case of PVA, they are named Lenti-kats, as commercialized by Genialab and used for nitrate and nitrite reduction and in the synthesis of fine chemicals. These beads show minimized diffusion limitations caused by the swelling of the polymeric environment under the reaction conditions. To avoid catalyst leaching, enlargement can be realized by linking them to, e.g., chitosan. 

Unfortunately, most enzymes do not obey simple Michaelis-Menten kinetics. Substrate and product inhibition, presence of more than one substrate and product, or coupled enzyme reactions in multi-enzyme systems require much more complicated rate equations. Gaseous or solid substrates or enzymes bound in immobilized cells need additional transport barriers to be taken into consideration. Instead of porous spherical particles, other geometries of catalyst particles can be apphed in stirred tanks, plug-flow reactors and others which need some modified treatment of diffusional restrictions and reaction technology. 

A biocatalytic enantioselective addition of ammonia to a C=C bond of an afl-unsaturated compound, namely fumaric acid, makes the manufacture of L-aspar-tic acid, l-27, possible [30], This L-amino acid represents an important intermediate for the production of the artificial sweetener aspartame. The biocatalytic production process, which is applied on an industrial scale by, e.g., Kyowa Hakko Ko-gyo and Tanabe Seiyaku, is based on the use of an aspartate ammonia lyase [E.C.4.3.1.1] [31]. As a biocatalyst, an immobilized L-aspartate ammonia lyase from Escherichia coli [32, 33] as well as Brevibacterium flavum whole-cell catalysts [32 a, 34] have been applied successfully. 

Beshay et al. reported conditions for the short-term continuous cultivation of D. discoideum cells on porous supports (SIRAN beads) in HL-5C medium ]108]. D. discoideum cells actively colonized the porous carrier (Fig. 5.5), after which the colonized beads can be freely suspended in medium. Gell densities of free amoebae remained at about 10 per mL for most of the cultivation time, whereas the cell density on the SIRAN beads reached up to 10 per mL and remained constant for at least 16 days of fermentation [108, 109]. By using broken pumice or CeramTec (a ceramic catalyst support), the immobilized cells reach... 

A thermophilic bacterium (growth temperature 50°C), Thermomonospora curvata JTS 321 was identified as a catalyst for the same reaction96-97. Three different reactor systems have been compared, namely packed-bed and fluidized-bed reactors with immobilized cells (polyacrylamide- hydrazide gels) as well as a hollow-fiber reactor96. The highest productivity (up to 1400mgh 1 L-197 was observed with the latter. 

Much effort has gone, in recent years, in setting up alcoholic fermentations based on immobilized cell technology (J)). Some of the systems have proved to be highly productive, but are faced with drawbacks of leakage of cells, and sterical hindrances. Fermentation in two-phase system, on the other hand, has been successfully carried out with macromolecular substrates such as starch and cellulose (7 10 ). It is also easier to control a reaction system involving a number of enzymes, in a two-phase system as compared to the immobilized systems for example, there is a possibility to add more of the labile catalyst during the continuous operations. 

The industrial use of bio catalysts has been reviewed in many excellent papers from industrial and academic experts in recent years [6-20]. These publications clearly show that immobilized systems find only limited use in present bioprocesses. Straathof et al. recently investigated 134 industrial biotransformations and came to the conclusion that only 20 confirmed processes rely on immobilized bio catalysts [15]. This is due to the fact that immobilization can be a considerable cost factor and is frequently used in combination with less common continuous reactors. In addition, many transformations belong to the class of redox reactions and require a cofactor for the reaction to occur. Such processes can in many cases be realized perfectly under fermentative conditions by the use of living or resting cells [17,21-25]. 

The calculation of r requires axact knowledge of the concentration of catalyticaTly active immobilized cells per unit volume of catalyst particles. The immobilization technique guarantees the complete entrapment of the cells suspended in the polymer solution before crosslinking. If this cell concentration is denoted... 

This new system had been operating industrially since 1973 in Tanabe Seiyaku Co., Ltd. The overall production cost by this system was reduced to above 60% of the conventional batch process using intact cells because of the marked reduction in cost for the preparation of catalysts and of the reduction of labor cost by automation. Furthermore, the procedure employing immobilized cells is advantageous from the standpoint of waste treatment. 

---