Biochemical aspects of lipase immobilization at polysaccharides for biotechnology.

The design of immobilized enzyme preparations is an important and relevant area of modern sciences and technologies. Immobilization of enzymes from animal sources (component I) on natural carriers (component II) increases the system stability by protecting the active site of the enzyme from deactivation; facilitates the separation and accelerates the recovery of the enzyme. This makes reuse possible and provides a significant reduction in operating costs. Hydrolytic enzymes (such as lipases) and polysaccharides (such as chitosan) are the most promising of such pairs of components. The main attention here is devoted to the discussion on lipase immobilization on polysaccharide (mainly - chitin and chitosan). Based on the analysis of the available literature, the most adequate method is the immobilization of lipase from porcine pancreas (LPP) on polysaccharide particles (such as chitin or chitosan) pre-treated with ultrasound (to increase the particle surface area) and glutaraldehyde (for particle activation) that shows reasonably high LPP activity and stability. In order to increase further the activity of the lipase, some authors proposed to incorporate a spacer in the form of 1,3-diaminopropane (or 1,3-diaminobutane) prior to activation of the surface of the chitosan particles. In particular cases, the use of chitin (instead of chitosan) may be an alternative solution for biotechnological applications. Recently the idea of constructing "supramolecular enzyme systems" realized in the so-called "coimmobilized multienzymatic systems" strategy. The most fascinating example is the combined assay of a mixture of native LPP, glycerol kinase (from Cellulomonas) and glycerol-3-phosphate oxidase (from Aerococcus viridans) linked by glutaraldehyde to chitosan (as shell for inorganic nanoparticle core). This material was placed on a Pt-electrode as biosensor and was successfully applied for amperometric determination of the triglyceride level in the serum of healthy and diseased person. Thus, the whole innovative research-production sequence is described by Aggarwal V. and Pundir C.S.: from simple components to advanced material and further biomedical application. Thus, the following approach of lipase immobilization appears the most promising for future applications: a few types of lipases or the combination of LPP with some other enzymes immobilized simultaneously on multifunctional carriers (as nanohybrids of inorganic core and polysaccharide shell).

Cell Staining by Photo-activated Dye and Its Conjugate with Chitosan.

Photo-activated or "Caged" rhodamine dyes are the most useful for microscopic investigation of biological tissue by various fluorescent techniques. Novel precursor of the fluorescent dye (PFD813) has been studied for photosensitive staining of numerous animal cells. The functional rhodamine dye (Rho813) with intensive fluorescence has been obtained after photoactivation of its precursor PFD813 inside cells. The dye Rho813 has been successfully used for the optical detection of particular features in biological objects (HaCaT cells, HBL-100, MDCK, lymphocytes). Moreover, the chitosan conjugate with PFD molecules ("Chitosan-PFD813″) has been obtained and studied for the first time. The developed procedures and obtained data are important for further applications of novel precursors of fluorescent dyes ("caged" dyes) for microscopic probing of biological objects. As example, the synthesized "Chitosan-PFD813″ has been successfully applied in this study for intracellular transport visualization by fluorescent microscopy.

Multifunctional membranes based on photosensitive crown-ether derivatives with advanced properties.

This review discusses recent works on monolayer, multilayer and polymer films of various crown-ether derivatives. Preparation and investigation of such membrane nanostructures based on photosensitive and surface-active crown-ethers is a rapidly growing field at the "junction" of colloids and polymers, materials sciences and nanotechnology. These membranes can serve as convenient models for studying the self-organization and molecular recognition processes at interfaces that are typical for biomembranes. The results obtained for such structures by absorption and fluorescence spectroscopy, atomic force and Brewster-angle microscopy, surface pressure and surface potential isotherm measurements have been described. The possibility of developing multifunctional materials possessing advanced properties has been demonstrated.

Novel precursors of fluorescent dyes. 1. Interaction of the dyes with model phospholipid in monolayers.

Photoactivated ("caged") fluorescent dyes are modern tools for structure and function studies of cell membranes and subcellular organelles. Recently synthesized precursors of rhodamine fluorescent dyes (abbreviations PFD813 and PFD814) important for microscopic probing of biological objects have been studied in solution. In order to characterize the behavior at interfaces, monolayers of PFD813 and PFD814 on water have been formed and investigated. The interactions of these precursors with the biomembrane component dimyristoylphosphatidylethanolamine in monolayers at the air-water interface and after transfer to glass plates have been studied by measuring monolayer parameters and spectroscopic properties before and after photo-chemical formation of the fluorescent rhodamine dyes Rho813 and Rho814, respectively.

Membrane and Films Based on Novel Crown-Containing Dyes as Promising Chemosensoring Materials.

This paper discusses several works on supramolecular systems such as monolayer and multilayer, polymer films of various crown-containing dyes, surface-active monomers and polymers. Design, production and investigation of the membrane nanostructures based on crown ethers is a rapidly developing field at the "junction" of materials sciences and nanotechnology. These nanostructures can serve as convenient models for studying the self-organization and molecular recognition processes at interfaces that are typical for biomembranes. Based on the results obtained for such structures by absorption and fluorescence spectroscopy, atomic force and Brewster-angle microscopy, surface pressure and surface potential isotherm measurements, the possibility of developing micro- and nanomaterials possessing a set of specified properties (including chemosensor, photochromic and photorefractive materials) is demonstrated.