Environmental Engineering Reference
In-Depth Information
Table 4. Comparison of alkali- and enzymatically-catalyzed transesterification for
biodiesel production. Adapted from [1]
Parameter
Alkali-catalyzed process
Lipase-catalyzed process
Reaction temperature
60-70°C
30-40°C
Free fatty acids in raw materials
Formation of undesired saponified
products
Formation of desired methyl
esters
Water in raw materials
Interference with the reaction
No influence
Yield of methyl esters
Normal
Higher
Recovery of glycerol
Difficult
Easy
Purification of methyl esters
Repeated washing required
No washing required
Production cost of catalyst
Low
Relatively high
Studies from the mid-1990s to present showed that, in general, lipase-catalyzed reactions
with longer-chain fatty alcohols proceeded far more readily than those with methanol or
ethanol. The reactions tolerated the presence of up to 20 percent water but were favored by
the presence of organic solvents, which presented problems in that organic solvents were not
suitable for fuel production due to the risk of explosion as well as the difficulty of removing
the solvent. In addition, scientists found that even immobilized lipase preparations could not
be re-used, a problem that would have to be solved to promote commercialization [18]. The
greatest obstacle to this technology is the cost of the lipase enzymes. Two primary approaches
are in progress to address this difficulty: first, genetic engineering of microorganisms to
produce lipases in greater quantities and with greater activities, and second, the investigation
of whole-cell systems to allow in situ regeneration of the catalytic units.
2.3.1. Lipase engineering: production
. The cost of lipase production is the primary
impediment to commercialization of lipase-catalyzed systems. This is a general problem of
enzyme-catalyzed processes, which industrial biotechnology has addressed primarily by the
development of high-enzyme-expression systems and of whole-cell biocatalysts.
Overexpression of lipases requires accurate protein folding and translocation across the
cytoplasmic and, in gram-negative bacteria, outer membranes, processes estimated to involve
up to 30 cytoplasmic proteins in some organisms. Through careful manipulation of lipase
signal sequences and secretory pathways, however, several successes have been achieved:
lipases from various Bacillus species have been overexpressed in E. coli systems, for example
[24]; Rhizopus oryzae lipases have been produced in extracellular, functional form in
Saccharomyces cerevisiae [25, 26]; and the Candida antarctica lipase B has been similarly
secreted in functional form from the yeast Pichia pastoris [27]. Because lipase folding and
secretion are highly specific processes that normally do not function properly in heterologous
hosts, these successes represent breakthroughs that provide wonderful opportunities for
further optimization and increases of lipase overexpression in heterologous hosts.
Work directed toward modifying and accelerating the secretion pathways in native lipase
hosts has also led to great increases in extracellular lipase yield within Pseudomonas
fluorescens and Serratia marcescens, showing that pathway manipulation within native hosts
may be another promising avenue for further improvement [24].
