Polyacrylamide-entrapped cells

Mullen et al. (1989) reported that Bacillus cereus, B. subtilis, E. coli and P. aeruginosa were able to sorb an average of 89% of the total Ag+ and 12-27% of the total Cd2+, Cu2+ and La3+ from a ImM solution. Using polyacrylamide-entrapped cells of Brevibacterium sp strain PBZ, Simine et al. (1998) measured a sorption capacity of 40 mg g-1 and 13 mg g-1 dry biomass for Pb and Cd, respectively. Hall et al. (2001) isolated two bacterial strains of P. syringae that were tolerant to 1000 mg L-1 Cu. Similarly, Amoroso et al. (2001) were able to obtain Streptomyces spp. strains R22 and R25 with a high tolerance to Cr from sediments of the Sail River, Argentina. The cells of R22 and R25 could accumulate 10.0 and 5.6 mg Cr g-1 dry weight, respectively, from a concentration of 50 mg Cr mL 1. Cell fractionation studies with strain R22 showed that most of the chromium... 

Fig. 5. Correlation between heat response and reaction rate of cephalosporin C transformation by immobilized D-amino acid oxidase of Trigonopsis variabilis. Enzyme immobilization techniques entrapment in polyacrylamide gel ( ), cells cross-linked with glutaraldehyde ( ), cells entrapped in polyacrylamide gel (a) [28]...

Enzymes can be immobilized by matrix entrapment, by microencapsulation, by physical or ionic adsorption, by covalent binding to organic or inorganic polymer-carriers, or by whole cell immobilization (5 ). Particularly impressive is the great number of chemical reactions developed for the covalent binding of enzymes to inorganic carriers such as glass, to natural polymers such as cellulose or Sepharose, and to synthetic polymers such as nylon, polyacrylamide, and other vinyl polymers and... 

Aspartase. Tanabe Seiyaku has used aspartase in lysed E. coli cells immobilized by entrapment in polyacrylamide (3) or K-carra-geenan (38) for production of aspartic acid since 1973. Using a substrate stream containing 1 M ammonium fumarate and 1 mM Mg2+ at pH 8.5 and 37°C, a continuous, automated bioreactor with a 120-day half-life will produce L-aspartate at 60% of the cost of a batch fermentation (3). Recently, a process for immobilization of the cells in polyurethane has also been described (37). 

Immobilized cells have either been fixed or bonded to a surface, or entrapped in a gel matrix. Immobilization of vegetative cells for hydroxylations has been achieved in polyacrylamide, polyethylene glycol diacrylate, calcium alginate, and agar gels with varying success30,31. 

This approach was used in the study of the properties of D-amino acid oxidase isolated or fixed in cells of Trigonopsis variabilis and entrapped in calcium pectate or polyacrylamide gel [28]. The approach of a differential reactor (low enzyme activity in the packed bed) was applied. The experimental thermometric data, ATr, were transformed to reaction rates, vobs, according to Eq. (21), whereas parameter a was determined by the calibration shown in Fig. 5. The data were described by the equation... 

Transformations with immobilized enzymes or cells Often the stability of the biocatalyst can be increased by immobilization and many different enzymes and cells have been immobilized by a variety of different methods. The most popular method for the fixation of whole cells is entrapment or encapsulation with calcium alginate. Other natural gels e.g., carrageenan, collagen, chemically-modified natural polymers e.g., cellulose acetate and synthetic gels and polymers e.g., polyacrylamide or polyhydroxyethylmethacrylate can also be used for this type of immobilization. 

We attempted continuous production of L-aspartic acid from fumaric acid and ammonia by immobilized Escherichia coli having high aspartase activity [3, 4, 5]. Various methods were tested for the immobilization of microbial cells, and a stable and active enzyme system was obtained by entrapping whole microbial cells in a polyacrylamide gel lattice. 

Entrapment provides an alternative to covalent immobilization. IVpically, cells are entrapped in a porous matrix and the cells grow throughout the pores/media to result in high cell densities. Cells may be added to the matrix at the time it is formed, or may be added after matrix formation. Many matrices have been used for entrapment including agarose beads, ceramics and silica, collagen microspheres, polyacrylamide, controlled pore glass, and various membranes [68, 69]. Membrane retention of cells, which has been used for biosensors (table 8.7) is, per se, not an immobilized system since the cells are... 

Klebsiella pneumoniae was entrapped on both alginate and polyacrylamide gels and physically adsorbed on vermiculite and particles wood. The concentrations of mercury were measured in both mercurial and oxidizing solutions according times for all immobilized Klebsiella pneumoniae, as well as for free cells. 

As described in the previous section, the addition of a surfactant produced remarkable stimulation during the synthesis of CoA. In the immobilized cell system, some surfactants showed similar stimulative effects. When the gel in which intact cells were trapped was treated with sodium laurylsulfate before the reaction, it was remarkably activated. An addition of sodium laurylsulfate to the reaction mixture with trapped Intact cells caused an increased accumulation of CoA. Similar observations have been reported by other workers. Franks (M) has reported that polyacrylamide gel-entrapped Streptococcus faecalls, which catalyzes the degradative conversion of arginine to putrescine, was activated by treating it with lysozyme. Chlbata l. (15) activated Escherichia coll cells with aspartase activity, which was trapped in polyacrylamide gel, by autolysis. 

This reaction has been carried out by batch procedure, which has disadvantages for Industrial purpose. Thus, we studied the continuous production of L-aspartlc acid using a column packed with immobilized aspartase. As the aspartase is an Intracellular enzyme, it was necessary to extract the enzyme from microbial cells before Immobilization. Extracted Intracellular enzyme is generally unstable, and most of the Immobilization methods we tried resulted in low activity and poor yield. Although entrapment into polyacrylamide gel lattice gave relatively active immobilized aspartase, its operational stability was not sufficient,... 

E. co-Li cells. Among the methods tested, the most active immobilized E. COi-L cells were obtained by entrapping the cells into polyacrylamide gel lattice [3]. 

Using cells entrapped in polyacrylamide, glucose could be determined. However, severe diffusional restrictions of the system resulted in rather low sensitivity of the system. In this method as well as in the one using a microbe thermistor (15), better analytical resolution would be obtained provided tlie immobilization techniques used are satisfactory. 

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