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Fermentative Production and Purification of Fructosyltransferase and Production of Fructooligosaccharides by Transfructosylation.

IR@CSIR-CFTRI

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Relation http://ir.cftri.com/11936/
 
Title Fermentative Production and Purification of Fructosyltransferase and Production of Fructooligosaccharides by Transfructosylation.
 
Creator Narendra Nath, Sanyal
 
Subject 05 Enzymes
 
Description Fructo-oligosaccharides (FOS) are effective prebiotics that can be produced from sucrose using fructosyltransferase (FTase) enzyme. However, the high cost of the enzyme hinders mass production of FOS. In order to reduce the expense, the enzyme can be produced in fungal organisms and purified in simple and convenient steps. In this work Aspergillusoryzae has been cultured in submerged fermenter to produce FTase, with highest titre after 90 hours of incubation. After separation of biomass, by centrifugation and filtration, the extracellular enzyme was concentrated using ultrafiltration with an yield of 38.40% followed by salt precipitation (yield- 14.05%). The protein content in each step was measured by Bradford assay. The salts were removed using Amicon filters (50 KDa) and dialysis (12 KDa), and then loaded into a gel filtration chromatography (GFC) column with a flow-rate of 8mlh-1. The 12th to 15th fractions from the column showed maximum FTase activity (53.628Uml-1) which was determined by GOD-POD method, the yield calculated after GFC was 12.271%. The fractions having highest activity were pooled and concentrated. The concentrated pool was analyzed by Native PAGE, which showed a single band, in both silver staining and activity staining methods. The crude and the purified enzyme were used for FOS production. The yield of FOS was measured by HPLC, and the highest value was found to be 56% for 1:3 enzyme:substrate ratio, after 18 hours of reaction time. It can be inferred from the results that usage of purified enzyme might lead to a decrease in the time of reaction, thus improving rate of production. The results from the study may prove beneficial to industrial FOS production, making the process highly productive and economic, which can be further extrapolated to preparation of FOS-based food products.
 
Contributor Prapulla, S. G.
 
Date 2015
 
Type Student Project Report
NonPeerReviewed
 
Format application/pdf
 
Language en
 
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Identifier http://ir.cftri.com/11936/1/narendra.pdf
Narendra Nath, Sanyal (2015) Fermentative Production and Purification of Fructosyltransferase and Production of Fructooligosaccharides by Transfructosylation. [Student Project Report] (Submitted)