Citation
Mat Radzi, Salina
(2006)
Enzymatic Synthesis of Oleyl Oleate, a Liquid Wax Ester, in a Stirred Tank Reactor.
Doctoral thesis, Universiti Putra Malaysia.
Abstract
High performance enzymatic synthesis of oleyl oleate, a liquid wax ester
was successfully synthesized via enzymatic esterification reaction of oleic
acid and oley alcohol. Immobilised Candida antartica lipase B (Novozym
435) was used as biocatalyst. The study was divided into four parts which
are the optimisation of reaction synthesis at different scales, the reactor
study in term of mixing efficiency, the stability of immobilised enzyme and
the analysis and characterization of the product of the reaction.
Preliminary synthesis of oleyl oleate was carried out in a small scale
reaction with a total volume of 3.5 mL using screw-capped vials.
Optimisation reaction study via conventional method of varying one
parameter at-a-time approach was carried out. A high percentage conversion yield of >90% was achieved at optimum reaction time of 5
min, reaction temperature of 40-6O0C1 molar ratio of substrates (oleyl
alcoholloleic acid) of 2:1, amount of enzyme of 0.4 g and organic
solvents of Log P23.5 at fixed agitation speed of 150 rpm.
Investigation in larger scale production of oleyl oleate was performed
using 2 L stirred tank reactor (STR). The reaction was scaled-up to 300X
with a total volume of 1.05 L. A high percentage conversion of oleyl
oleate was achieved of >95% by conventional experiment method at
reaction time of 30 min, agitation speed of 400 rpm, reaction temperature
of 45-50°C, molar ratio of substrate (oleyl alcoholloleic acid) of 2:l and
amount of enzyme of 90 g.
The reaction synthesis was further optimised by response surface
method (RSM) based on five-level, three-variable central composite
rotatable design (CCRD) to evaluate the interactive effects of important
parameters in larger scale processing. Generally, simultaneously
increasing amount of enzyme, agitation speed and reaction temperature
would improved the yields. A high percentage conversion of 97.4% was
achieved under the optimum condition, which compared well with the
maximum predicted value of 97.7%.
In order to improve the production and productivity of oleyl oleate to the
highest amount that can be produced in a 2 L STR, the reaction was synthesized in a solvent-free system. Maximum scaling-up of substrate
concentration that can be achieved in the reactor vessel was 900X as
compared to 300X previously. The production and productivity of oleyl
oleate were successfully improved from 295.39 g/L/h to 705.76 g1Uh and
31 0.16 glh to 952.78 glh, respectively.
Reactor study on the performance of 2 L STR as a mixing device was
evaluated to improve the mixing efficiency. The rheological property of
the reaction mixture exhibited Newtonian behaviour. Rushton turbine
impeller showed better performance in degree of mixing, as compared to
AL Hydrofoil impeller whereby a high Reynolds number of >lo4 was
achieved at 400 rpm, which exhibit a turbulent flow pattern. There was
significant effect to the mixing improvement on the enzyme particles
distribution by using a 2 impellers system with spacing of 30 mm.
The enzyme showed high stability against heat as shown by the high
percentage conversion of wax ester. Novozym 435 retained its synthetic
activity up to 9 uses and 4 uses in screw-capped vials and STR,
respectively. The effect of shear forces due to the mechanical agitation
speed on the enzyme morphology was determined by scanning electron
microscope (SEM). Although small rupture on the surface of enzyme was
observed when increasing the agitation speed, the enzyme activity was
very high even at high agitation speed.Analysis of product was evaluated by spectroscopy method of Fourier
transform-infrared spectroscopy (FT-IR) and gas chromatography-mass
spectroscopy (GC-MS) to identify the product obtained. Characteristics of
oleyl oleate were also examined, which include iodine value,
saponification value, acid value and ester value. Solubility of oleyl oleate
in methanol and ethanol was comparatively lower as compared to the
solubility at higher chain length of alcohols. This compound seems
compatible in most of oils and stable even after heating up to 90°C and
overnight storage at room temperature.
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