Enzymes as Biocatalysts for Lipid-based Bioproducts Processing - Food and Industrial Bioproducts and Bioprocessing - page 339

The main constraint in enzymatic processing of PLs is the high cost of enzymes. For
instance, PLD, which is relatively high cost and available only in free form, is primarily
used for development of pharmaceutical products with high added values. Development of
low cost and robust enzymes through protein engineering and directed evaluation may offer
a solution to the problem (Song et al ., 2005). Besides that, separation and purification
of pure PLs from a complex natural PL mixture is relatively tedious. To date, a simplified
isolation procedure is not yet available. Thus, research for a simplified purification method
of PLs is very much in need.
14.4.4 Enzymatic processing of fatty acid alkyl esters
Fatty acid alkyl esters (FAAE) can be prepared from transesterification of vegetable oils or
animal fats with aliphatic alcohols (Knothe et al ., 2005 ). The transesterification reaction,
also known as alcoholysis, is the exchange of alkoxy group of an ester compound (TAG)
with an aliphatic alcohol (the acyl acceptor) in the presence of a catalyst. The overall reac-
tion is a sequence of three consecutive and reversible reactions in which DAG and MAG are
formed as intermediate products (Ma and Hanna, 1999). The general reaction scheme is
given in Figure 14.5 .
At present, industrial production of FAAE is performed using basic chemical catalysis
with, for example, sodium or potassium hydroxide. This process is energy consuming and
not environmentally friendly (Fjerbæk et al ., 2009). In the early 1990s, lipases were firstly
used in place of chemical catalysts for transesterification reaction (Mittelbach, 1990). Since
then, studies on enzymatic transesterification for FAAE production have increased by leaps
and bounds. Some of the benefits of enzymatic FAAE production worth mentioning include
easier handling operations, milder reaction conditions and the possibility of using alternative oil
sources, such as high FFA containing oils, without problems of soap by-product formation
(Kulkarni and Dalai, 2006 ).
There are many factors affecting enzymatic transesterification (Fjerbæk et al ., 2009 ;
Robles-Medina et al ., 2009). Among the key factors are type, stability and reusability of
lipase, type of the acceptor alcohol, substrate ratios, quality of feedstocks, reaction temperature,
water activity and/or water content.
Lipases used for FAAE production are normally of microbial origin, such as CALB and
TLL lipase. They are often used in immobilized forms, which are more stable and versatile
than their free forms (Shimada et al ., 2002 ; Kojima et al ., 2004 ; Nielsen et al ., 2008 ). The
immobilized form is also more industrially feasible, as they can be easily packed and reused
in industrial reactors. Nevertheless, they are also more costly in terms of enzyme price
(per kg of immobilized enzyme). However, the stable and reasonably high productivity of
the enzyme (kg of biodiesel/kg of immobilized enzyme) during a relatively long lifetime is
more important than the sole price comparison. Whole-cell biocatalysts which are cheaper
and more robust may be appropriate for industrial FAAE production (Antczak et al ., 2009 ).
The activity of whole-cell biocatalysts depends on the fatty acid composition of the cell wall
membrane. As there may be different lipases bound to the cell wall or membrane, the FAAE
yield may vary (Adamczak et al ., 2009 ).
A combination of different enzymes has been shown to increase yield in the enzymatic
production of FAAE. For example, Türkan and Kalay (2006) used three different immobilized
lipases from RML, TLL and Candida . They found lipases from RML and TLL catalyzed the
first step (TAG to DAG) of transesterification faster while lipase from Candida antarctica
catalyzed the second (DAG to MAG) and third (MAG to FAAE and glycerol) steps faster.