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Synthesis of Feruloyl Ester Using Bacillus subtilis AKL 13 Lipase Immobilized on Celite® 545


Karthikumar Sankarorcid tiny and Anant Achary*orcid tiny

 

Department of Biotechnology, Centre for Research, Kamaraj College of Engineering and Technology,
S.P.G.C. Nagar, 625701 K. Vellakulam, Near Virudhunagar, Madurai District, Tamil Nadu, India


Article history:
Received: May 2, 2017
Accepted: August 1, 2017


Key words:
Bacillus subtilis lipase, Celite® 545, immobilization, esterification, feruloyl ester


Summary:
The lipophilic antioxidants, glyceryl ferulate and feruloyl glyceryl linoleate, were synthesized using lipase from Bacillus subtilis AKL 13. The extracellular lipase was produced by cultivation of the strain in modified minimal medium and the enzyme was recovered by fractionation at 80 % ammonium salt saturation. The concentrated enzyme with the specific activity of (4647±66) U/mg was immobilized on Celite® 545 and crosslinked using glutaraldehyde. The prepared enzyme catalyst was used for esterification of ferulic and linoleic acids with glycerol separately in hexane butane solvent system at 50 °C and 3.144×g agitation. The maximum ester conversion of 94 % of feruloyl glyceryl linoleate was achieved at 48 h, whereas only 35 % of glyceryl ferulate was synthesized. The reaction products were characterized using RP-HPLC, FTIR, 1H NMR, 13C NMR and fluorescence spectrophotometry. The kinetic parameters of esterification reaction were determined according to ping-pong bi-bi model. The Km and υmax were found to be 69.37 and 3.46 mmol, and 0.387 and 1.02 mmol/(min·g) for glyceryl ferulate and feruloyl glyceryl linoleate, respectively. The kinetic parameters were simulated in MATLAB and the experimental data were in good agreement. Furthermore, 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity of the blend of feruloyl ester and palm oil was higher than of the plain palm oil and was closer to α-tocopherol.



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