Front Matter

As observed for other lipases (Balcao et al., 1996) its stability may have increased

upon immobilization, though no data are available for this.

It should be noted that all wild-typeGeotrichum candidumstrains are known to

produce at least two different lipases (isozymes) of which only the B-lipase is highly

specific. The other lipase, generally referred to as Lipase-A (Charton and Macrae,

1992) or Lipase-II (Holmqvist et al.,1997) is totally nonspecific. Purification and

isolation of the wild-type B-lipase therefore requires laborious and multi-step separa-

tions (Sidebottom et al., 1991; Diks and Lee, 1999). Alternatively, the Lipase-B can

also be produced inPichia pastoris(Catoni et al., 1997) by applying genetic mod-

ification techniques.

Separating the free fatty acids by distillation, they were re-esterified with glycerol

(molar ratio 3 : 1). This reaction was catalyzed by immobilizedRhizomucor miehei

lipase (Lipozyme) at 60 8 C under continuous removal of water. Under these condi-

tions the product consists of>95 % (w/w) triglycerides (Ergan et al., 1990; McNeill

et al., 1996; Diks and Lee, 1999). Although theRhizomucorlipase is 1,3-regiospe-

cific, the high reaction temperature promotes acyl migration of partial glycerides

(Lortie et al., 1993; Dudal and Lortie, 1995). As a result 1(3)-monoglycerides

and 1,3-diglycerides will be converted in their 2-position isomers, resulting in sub-

sequent conversion into triglycerides. Moreover, acyl migration is promoted by the

ionic carrier material (Duolite) applied (Millqvist Fureby et al., 1996).

Enrichment of DHA and EPA from fish oil

It is now recognized that polyunsaturated fatty acids (PUFA) play an essential role in

human nutrition, as well as having important biomedical properties. Both thex-3 and

x-6 families play important roles in human metabolism, but they cannot be inter-

converted in the body. Despite the fact that there is continuing debate among nutri-

tionists on the ideal dietary ratio ofx-6 tox-3 acids, it is generally accepted that the

intake ofx-6 acids is adequate in developed countries. Consequently, nutritionists

now recommend that the intake ofx-3 acids, particularly the long-chain PUFA

(LCPUFA) docosahexaenoic acid (DHA, C22 : 6) and eicosapentaenoic acid (EPA,

C20 : 5), is increased (Ashwell, 1992).

There are a number of sources of these acids, e.g., fish oils, marine algae and some

micro-organisms. Of these, fish oils are the most commercially significant although

the levels are usually only low to moderate. Oils derived from microbial fermenta-

tion can contain high levels but only at a relatively high cost. This has led to a great

deal of interest in processes for the enrichment of DHA and EPA from commodity

fish oils. Many physical methods have been developed for extracting and enriching

these fatty acids, including crystallization, distillation, and the use of supercritical

carbon dioxide.

The use of lipase biotechnology now offers an alternative method which has the

advantage of mild processing conditions that minimize the degradation/oxidation of

these acids (see also Chapters 8-10). The enrichment processes are based on the

principle that, in general, lipases show low reactivity towards LCPUFA compared

to more common fatty acids (<C 20 ). This can be exploited to enrich LCPUFA from

commodity fish oils. Many lipases also show low reactivity towardsc-linolenic acid

6 1 The Exploitation of Lipase Selectivities for the Production of Acylglycerols