Front Matter

PUFA-HPODs are potentially cytotoxic compounds as they can induce radical

chain reactions that are deleterious to living organisms. In eukaryotes, PUFA-

HPODs are catabolized by various pathways, leading to more stable products.

Furthermore, these metabolites generally possess extremely important physiologi-

cal roles such as jasmonic acid (Hamberg and Gardner, 1992) in plants or leuko-

trienes (Maycock et al., 1989) and lipoxins (Serhan, 1994) in mammals.

16.2 How to use lipoxygenases

16.2.1 The reaction catalyzed by lipoxygenases

As described in Section 2.1, the reaction catalyzed by LOXs is generally highly

specific, leading to near-optically pure PUFA-HPODs. As the total chemical synth-

esis of such HPODs is an extremely difficult task, lipoxygenation of PUFAs is the

most obvious and simple way to obtain access to such compounds. Although this is

an example of where enzymatic catalysis has overshadowed chemical synthesis, the

catalytic properties of LOXs must nevertheless be taken into account when perform-

ing enzymatic synthesis of HPODs. In the following, examples of how to use LOXs

on a synthetic scale based on their catalytic properties will be discussed. As only one

LOX is available commercially (i.e., SBLOX-1) our knowledge of LOXs is derived

mainly from studies dealing with that isoenzyme. It is the consideration of the

authors that the conclusions drawn from SBLOX-1 are broadly applicable to other

LOXs.

Perhaps one of the most serious drawbacks in the use of LOXs on a preparative

scale has been the low solubility of PUFAs in an aqueous environment. Indeed, it has

been shown that LA has a critical micellar concentration of 1.5
10 –4M in borate

buffer (Lagocki et al., 1976). Above this concentration a strong substrate inhibition

has been noted, showing that free fatty acids are the true substrates of LOXs (Galpin

and Allen, 1977). The addition of a co-solvent (ethanol) or surfactants (Tween, Tri-

ton) has long been used to improve the solubility of the substrate to 5
10 –3M. At this

point of the discussion it should also be stressed that the presence of the product of

the reaction (i.e., PUFA-HPODs) is necessary in order for the enzyme to be fully

active (Jones et al., 1996). Descriptions of how the above-mentioned problems

have been solved to allow the enzymatic synthesis of PUFA-HPODs to be con-

ducted on a multigram scale, and at high substrate concentration (up to 0.1 M),

will be discussed in the following text.

16.2.2 First examples of synthetic uses of soybean lipoxygenase-1

The enzymatic production of 15(S)-HPETE from AA and SBLOX-1 was first studied

in 1975, using a buffered system and a substrate concentration of 1.6
10 –3M(1gin

2 L), together with gentle bubbling of oxygen (Baldwin et al., 1975). After reduction

(with NaBH 4 ), 15(S)-HETE was obtained in 45–50 % yield. This was the first report

16.2 How to use lipoxygenases 339