Front Matter

which modify the aggregate structure either by remaining in the particles or by being

released. Therefore, in biocatalytic processes changes of the catalytic constants in

course of the reaction must be taken into account. Finally, in the evaluation of acti-

vators or inhibitors of phospholipases, it is difficult to differentiate between effects

caused by molecular interactions of the effector with the phospholipase and those

resulting from modifications of the morphology and charge of the phospholipid in-

terphase.

12.4.4 Catalysis in the presence of organic solvents

In biocatalytic applications of phospholipases (see Section 12.5.), organic solvents

are often involved in the reaction systems. In general, reactions are performed in

shaken emulsion systems, where a nonpolar organic solvent such as diethyl ether

or ethyl acetate containing the dissolved phospholipid is mixed with an aqueous

phase containing the enzyme in buffer solution (see Section 12.5). In such sys-

tems, the peculiarities of the kinetics of phospholipases have scarcely been re-

garded. Indeed, the influence of solvents on the aggregate structures enhance the

complexity of the system. Moreover, an additional reaction partner is often intro-

duced, e.g., an acceptor alcohol in transphosphatidylation by PLD, which may parti-

tion between the aqueous phase, the organic phase and the aggregates. In addition,

due to the heterogeneity of the reaction system, diffusion processes may become

relevant. Finally, denaturation of the enzymes by the organic solvents must be taken

into consideration. As a consequence, most studies on the reactions of phospholi-

pases in the presence of organic solvents barely allow general conclusions.

In a study usingD- andL-serine as substrates in the transphosphatidylation by a

PLD fromStreptomycessp., the following ranking of solvents in emulsion systems

was reported: ethyl acetate>diethyl ether>benzene>chloroform>toluene

(Juneja et al., 1989a). In contrast, in the reaction of 1,2-dipalmitoyl-sn-glycero-3-

phosphocholine with 4-methoxyphenol catalyzed by a PLD from Streptomyces

sp. rates depended on the solvent in the sequence of benzene>dichloromethane

>toluene>diethyl ether>ethyl acetate (Takami et al., 1994a), whereas a ranking

of dichloromethane>diethyl ether>ethyl acetate>benzene was found for the

formation of 1,2-dipalmitoyl-sn-glycero-3-kojic acid by this enzyme (Takami et

al., 1994b). The phosphatidyl glucose production from PC by PLD fromActinoma-

durasp. was high in an emulsion system containing diethyl ether, benzene, toluene,

and dichloromethane, whereas the yield was low in chloroform and no reaction was

observed in hexane (Kokusho et al., 1993).

In the transphosphatidylation reaction between PC and 3-dimethylamino-1-pro-

panol by PLD fromStreptomycesPMF in water – ethyl acetate emulsion sys-

tems, Carrea et al. (1997a) found an interfacial saturation by the enzyme and con-

cluded that only PLD located at the interface of the water – organic phase is active.

By measuring the enzymatic hydrolysis of PC monolayers formed at the polarized

nitrobenzene/water interphase by peanut PLD, Kondo et al. (1992; 1994) showed

that the rate of hydrolysis depends markedly on the potential drop across the inter-

phase. Hirche et al. (1997a) and Hirche and Ulbrich-Hofmann (1999), in analyzing

12.4 Kinetic particularities of phospholipases and their consequences 237