Plant cells entrapment

Relatively less work has been done on immobilization of plant and animal cells and spores of microbes in silica matrixes. The main drawback is less viability of the cells in sol-gel matrices. Thus more refined methods are required to utilize harness of the whole cells entrapped in sol-gel matrices and biosensing applications. At the same time studies such as interactions between sol-gel matrices and whole cells and metabolic changes during immobilization have to be closely monitored for the exploration of new matrices and methods. 

Plant cell cultures represent a potentially rich source of secondary metabolites of commercial importance and have been shown to produce them in higher concentrations than the related intact plants. However, plant cell cultures often produce metabolites in lower concentrations than desired and commonly store them intracellularly. These limitations can be overcome by product yield enhancement procedures, including immobilization of cultured cells, and permeabilization, or ideally using a combined immobilization/ permeabilization process with retained plant cell viability. Complex coacervate capsules consisting of chitosan and alginate or carrageenan proved to be effective biomaterials for entrapment, controlled permeabilization of cells and to allow control of capsule membrane diffusivity. 

Entrapment methods have been used almost exclusively to immobilize plant cells (17). They can be classified into three general groups (24,25) a. gel formation by ionic crosslinking of a charged polymer (ionotropic) b. gel formation by cooling of a heated polymer (thermal), and c. gel formation by chemical reaction (cross-linking, radical polymerization). 

Immobilization of cultured plant cells by entrapment in a gel matrix appears to provide conditions that are optimum to cell differentiation resulting in higher yields of secondary products. Compact or organized and slow-growing cultures often synthesize higher amounts of secondary products than growing cultures. 

Woerly, S. Plant, G.W. Harvey, A.R. Cultured rat neuronal and glial cells entrapped within hydrogel polymer matrices A potential tool for neural tissue replacement. Neurosci. Lett. 1996, 205, 197-201. 

It is apparent from the data in Table III that the effects of lignin are restricted to the cellulose and hemicellulose carbohydrates of the plant cell wall. One must reject the conclusion of Drapala (23) that availability of cellular contents is lowered by entrapment in lignified cells. 

An improved technique for entrapment of plant cells has been recently developed with Capsiaum frutesaene cell suspensions. Cells were shown to passively invade the pores of a reticulated polyurethane matrix, inside which viability and metabolic activity were better preserved than in other immobilizing matrices (23). This sytem was used in a deteuled study of the synthesis of capsaicin (an ester of vanillylamine), which is the pungent principle of chili pepper fruits. In the absence of specific precursors to capsaicin, the entrapped cells produced 2-3 orders of magnitude higher yields than c ll suspensions over a 5-10 day culture period (4-5 mg capsaicin g dry weight 1 versus 30 yg... 

Fixation of plant cells in a matrix of, for example, polyurethane foam or entrapment of the cells in calcium alginate beads provides an artificial surrounding for the cells, which protects them from hydrodynamic stress, high cell densities inside the matrix also allow cell to cell contact and communication. Inside the immobilized matrix nutrient and product gradients may occur. Furthermore, immobilized biomass is easily separated from the medium, which is useful in production and biotransformation systems. Immobilization of plant cells has been reviewed 103,104). 

Amaranthus tricolor plant cells were entrapped with chitosan gel to determine the polycationic properties of chitosan on plant cell membrane permeability. On the fifth day, maximum tricolor cells were released from chitosan-immobilized cells (Knorr and Teutonico 1986). 

Immobilized enzymes are defined as enzymes physically confined or localized in a certain defined region of space with retention of their catalytic activities, which can be used repeatedly and continuously. This definition is applicable to the enzymes as well as aU types of biocatalysts such as cellular organelles, microbial cells, plant cells, and animal cells. In some cases, these biocatalysts are bound to or within insoluble supporting materials (carriers) by chemical or physical binding. In other cases, biocatalysts are free, but confined to limited domains or spaces of supporting materials (entrapment). 

Entrapment of plant cell Plant cell in hybrid gel (Campostrini, 1996)... 

Techniques of attachment, entrapment, and encapsulation are most widely used for cell immobilization with support materials, which are illustrated in Figure 7.1. These techniques can be applied to essentially all the viable or nonviable wholecell systems of potential interest microorganisms, plant cells, and mammalian and insect cells [2]. Although most of the principles associated with enzyme immobilization are directly applicable to cell immobilization, due to the complete difference in size and biochemical properties between enzymes the cells, the relative importance of these methods is considerably different [10]. 

D-glucose and the three-enzyme system GOx, mutarotase and invertase for sucrose estimation. A common format was adopted to facihtate design and operation, in this case immobilization method, the fact that all enzymes used were oxidases and that a common detection principle, reoxidation of H2O2 generated product, was chosen (except for ascorbic acid which was estimated directly). Pectin, a natural polysaccharide present in plant cells, was used as a novel matrix to enhance enzyme entrapment and stabilization in the sensors. Interferences related to electrochemi-caUy active compounds present in fruits under study were significantly reduced by inclusion of a suitable cellulose acetate membrane diffusional barrier or by enzymatic inactivation with ascorbate oxidase. Enzyme sensors demonstrated expected response with respect to their substrates, on analyte average concentration of 5 mM. 

Physically inaccessible starch (RSi) Type I resistant starch is physically inaccessible and is protected from the action of a-amylase, the enzyme that hydrolyzes the breakdown of starch in the human small intestine. This inaccessibility is due to the presence of plant cell walls that entrap the starch, for example, in legume seeds and partially milled and whole grains. RSi can also be found in highly compact processed food like pasta. The RSj content is affected by disruption of the food structure during processing (e.g., milling) and, to some extent, by chewing. 

Traditional techniques such as physical adsorption and covalent linkage onto solid supports, entrapment in polymer matrices, and microencapsulation have long been used for immobilizing such enzymes as lipases, proteases, hydantoinases, acylases, amidases, oxidases, isomerases, lyases, and transferases [12-18]. Encapsulation and adsorption have also proved their utility in the immobilization of bacterial, fungal, animal, and plant cells [12-21]. However, as biocatalysis applications have grown, so the drawbacks and limitations of traditional approaches have become increasingly evident. The forefront issues now facing bioimmobilization are indicated in Table 1. 

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