Lipase-catalyzed synthesis of caffeic acid bornyl ester
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Date
2018-07-01
Authors
Nurul Nadzirah Mohd Yusof
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Abstract
Caffeic acid bornyl ester (CABE) is a rare caffeic acid derivative and natural
product with significant biological and pharmacological properties. However, the use
of traditional chemical extraction and chemical synthesis method to produce CABE
are uneconomical, inefficient and toxic to human and environment. Thus, the main
objective of this study is to establish a green reaction pathway for the synthesis of
CABE via lipase-catalyzed transesterification reaction. The synthesis of CABE was
conducted by the screening of important parameters such as types of immobilized
enzyme (Novozym 435, Lipozyme TLIM and Lipozyme RMIM), organic solvents
(isooctane, n-hexane, n-heptane, toluene and n-hexane:acetone) and effect of alkyl
group (methyl caffeate and ethyl caffeate) followed by investigating the effect of
reaction parameters such as enzyme loading (25 – 250 U), reaction time (0 – 84 h),
substrate ratio of borneol to ethyl caffeate (1:1, 1.5:1, 2:1, 2.5:1 and 3:1), reaction
temperature (30 – 60°C) and optimum reaction conditions based on traditional one
factor at a time (OFAT) method. The reaction kinetic mechanism was investigated
and kinetic parameters of lipase-catalyzed transesterification reaction were
determined based on the initial reaction rate obtained from the investigation of the
effect of substrates concentration. The optimization of CABE production was also
conducted using response surface methodology (RSM) based on face-centered
central composite design (CCD) to obtain the highest response. Finally, the activation
energy required for the reaction was determined by conducting the transesterification
reaction at various temperatures from 30°C to 55°C. From the screening, it was
found that Novozym 435 gave the highest conversion followed by Lipozyme TLIM
and Lipozyme RMIM. The highest substrate conversion was obtained using mixed
solvents system (n-hexane:acetone, 80:20, %v/v) and ethyl caffeate as substrate. In
the investigation of effect of reaction parameters based on OFAT method, the highest
conversions were obtained by using 125 U of enzyme loading, 48 h of reaction time
and temperature at 55°C. It was observed that the substrate ratio of borneol to EC,
2.5:1 (25 mM borneol: 10 mM EC) resulted in the highest conversion. At the
selected optimum reaction conditions as described above, the highest substrate
conversion of 86.0% was obtained. The reaction kinetics model for CABE synthesis
using lipase Novozym 435 was found to obey the ternary complex mechanism and
the kinetic parameters were as follows; KmEC = 0.091 mM, KmBor = 10.908 mM,
Vmax = 0.187 mM/min and KiBor = 1.943 mM. The optimized condition suggested by
RSM was found to be 245 U of enzyme loading at 36 h and 60°C with 93.64%
conversion. The estimated activation energy value of lipase Novozym 435 for the
synthesis of CABE was observed to be 57.6 kJ/mol.