Enzymes as Biocatalysts for Lipid-based Bioproducts Processing - Food and Industrial Bioproducts and Bioprocessing - page 335

Still more widespread use of bioactive compounds is limited due to issues such as lower
solubility and miscibility in more hydrophobic environments. As such, strategies meant to
alter the physicochemical properties of hydrophilic bioactives have been developed. One
relatively successful approach has been the enzymatic acylation of bioactive compounds
with more hydrophobic groups, whereby the addition of such groups impacts on partitioning
and even on the emulsification properties of the resulting products (Sasaki et al ., 2010 ;
Villeneuve, 2007 ).
Enzymatic acylation to yield bioactive compounds with additional properties and/or
altered functionalities generally takes place via esterification or transesterification reactions.
In simple esterification, the bioactive compound reacts with a fatty acid (or alcohol, based on
its structure) to yield an ester and a molecule of water. In contrast, transesterification reactions
involve acyl exchange between an ester and alcohol to yield structurally different ester and
alcohol species. Both reactions are typically catalyzed by lipase and can, therefore, benefit
from milder reaction conditions, substrate and/or regiospecificity (Chebil et al ., 2006 ).
Based on current research, some important factors to be considered during enzymatic
processing of bioactives include the type of reaction media used, the level and distribution
of water in the system, biocatalyst type and loading, reaction temperature, agitation and
the type and ratio of substrates selected for use in a particular system. Overall, proper
adjustment of the aforementioned parameters can help to push the reaction equilibrium
towards the production of modified products and may also result in a more efficient and cost
effective reaction set-up (Devi et al ., 2008 ).
With regards to enzymatic processing of bioactive compounds, bringing strongly hydrophilic
and hydrophobic substrates together to create an efficient reaction system still poses serious
challenges (Villeneuve, 2007 ; Xanthakis et al ., 2010). Most often, some compromise is
necessary, such as utilizing intermediate polarity solvents or else co-solvent systems to
achieve good contact between the vastly differing substrates and the biocatalyst. Some
success has also been achieved employing ionic liquids as novel media, since many ionic
liquids can dissolve vast quantities of hydrophilic and hydrophobic substrates (Katsoura
et al ., 2006 ; Hu et al ., 2009). Additional advantages of ionic liquids includes their ability
to activate and protect enzymes in certain environments; however, it is also clear that desta-
bilization of enzymes such as lipase by particular ionic liquids can also have a strong impact
on reaction systems (Lau et al ., 2004 ; van Rantwijk and Sheldon, 2007 ; Zhao et al ., 2009 ).
Judicious selection of ionic liquids is therefore of the utmost importance.
Microemulsion systems, also, offer the opportunity for improved miscibility of sub-
strates. For instance, esterification of ferulic acid with pentanol at 40 °C in a water-in-oil
microemulsion resulted in a yield of ferulic ester of up to 60% within the short time frame
of eight hours using a non-immobilized feruloyl esterase from Aspergillus niger (Giuliani et
al ., 2001). While it is advantageous in maximizing bioconversion yields, it should be noted
that microemulsions can pose additional challenges during downstream separation pro-
cesses following enzymatic conversions.
The presence and distribution of water in the system also has a strong influence on the
reaction. Lipases require a small amount of water to maintain their active conformation. Too
little or too much water generally results in suboptimal activity. As a by-product of the
esterification reaction, increasing quantities of water will shift the reaction equilibrium
toward the substrates. Thus, reaction systems containing just sufficient amount of water to
maintain the active lipase conformation without contributing significantly to the competing
hydrolytic reaction are most efficient. Excess free water is often removed from mole-
cules the reaction system through the use of activated molecular sieves (Duan et al ., 2006 ).