A solid state matrix, process of preparation thereof, and process of preparation of theaflavins

Abstract

The present invention relates to a process for the development of a highly efficient solid state matrix by the activation of acrylate based polymer resin having specialized functional groups with 1,1-Carbonyl diimidazole for immobilizing biologically active macromolecules such as oxidases, in particular plant oxidases and the most preferred being tea polyphenol oxidase through indirect covalent bonding / cross linking on such activated polymer resin support, are thermally stable, gives very high number of turnovers in vitro ('n' times); with tea substrate forming exclusive product Theaflavins without any loss of biological activity and leaving the product remaining in vitro without adherence to matrix rendering the matrix safe towards product poisoning and subsequent partial or complete loss of biological activity of the matrix bound enzyme system and thus well adapted to and well suited bioreactors based on such systems. It is unique with respect to its recyclability of otherwise uneconomical tea substrates such as seed and flower substrates into theaflavins both with respect to crude substrate or purified ones.

Description

field of invention [0001] In particular, the invention relates to a solid-phase base material, a preparation method thereof and a preparation method of theaflavin (theaflavin). [0002] More specifically, the present invention relates to the development of efficient solid phase substrates for the immobilization of soluble and bound forms of plant oxidases, in particular tea polyphenol oxidase (EC 1.10.3.1), enabling continuous batch production and Complete conversion of tea-derived substrates into theaflavins from purified tea-derived substrates and crude tea-derived substrates, wherein the base material can completely convert all available substrates into theaflavins, thus providing a A method for preparing theaflavins (w / w) in a medium proportional to the substrate available. Background of the invention [0003] Theaflavins are a group of polyphenolic compounds with the general structure shown in Table 2, which are utilized during black tea production by native tea polyph...

AI Technical Summary

Problems solved by technology

[0008] 1. Long coupling time

[0009] 2. Non-physiological pH leads to inactivation of target molecules

[0010] 3. The product contaminates the activated or deactivated polymer

[0011] 4. Polymer type or by-product toxicity

[0012] 5. Limited application in aqueous solution

[0013] 6. Activated polymer constructs are unstable

[0014] 7. The cyanuric chloride method (refer to Abuchowski et al. (1977a), Journal of Biological Chemistry, 252, 3582-3586) and the carbonyldiimidazole method (refer to Beauchamp et al. (1983), Analytical Biochemistry, 131, 25-33) are common And chloroformic acid phenyl ester (can refer to Veronese et al. (1985), Appl.Biochem.Biotechnol., 11,141-152) and succinimidyl (succinimedyl) active ester method (can refer to Shadle et al.; Katre et al. (1987) , Proc.Natl.Acad.Sci.USA, 84,1487-1491) in the substantial loss of biological activity often seen

The present inventors found the previously described methods to be unsuitable because of the high level of product binding during the reaction of the active enzyme:polymer adduct with the substrate, resulting in substrate poisoning and partial or complete loss of biological activity and the inability to extract from the active substrate The product is recovered so it is not suitable for use in bioreactors or as reusable material

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