Reaction selectivity of Burkholderia cepacia (PS‐30) lipase as influenced by monoacylation of sn‐glycerol

Abstract

Reaction selectivities were determined in multicompetitive reactions mediated by Burkholderia cepacia lipase (Amano PS-30) at a water activity of 0.19 in hexane. Saturated FA (C4-C18 even chain) and oleic acid (C18:1) were reacted with a single alcohol, glycerol, or α-or β-MAG, containing C4, C10, C16, or C18:1 individually as alcohol cosubstrate. Similar ordinal patterns of FA selectivity, with C8, C10, and C16 preferred over others, were generally observed for incorporation of FA into specific acylglycerol (AG) pools of the 24 specific cases evaluated. The three exceptions were enrichment of C14 and C18 in the MAG pool with α-C16-MAG substrate, and a general suppression of >C8 incorporation into the TAG pool for reactions with α-C10-and α-C16-MAG. PS-30 lipase selectivity toward MAG was in descending order: α/β-C4-MAG > β-C10-MAG > β-C16-MAG > α/β-C18:1-MAG > α-C10-MAG > α-C16-MAG. Selectivity in channeling CX of the original CX-MAG substrates into higher AG species was in descending order: α-Generally, MAG were better acyl donors than FA for esterification reactions leading to DAG formation. These observations are relevant to the design of biocatalytic processes intended to yield specifically structured TAG.A focus of over a decade of recent research on exploiting the synthetic power of lipases in organic media is the prospect of preparing value-added or functional "structured glycerides" for food and pharmaceutical industries (1). Examples of lipids modified by lipase to confer additional value and/or functionality include low-calorie fats (2), cocoa butter substitutes (3), and nutritional lipids (4,5). The applications-driven research in this area has focused largely on using specific lipid mixtures or even pure reaction components to prepare specific targets of synthesis. Much of this body of literature has been rather empirical and sparingly reliant on features of lipase selectivity, such as regioselectivity. Indeed, the commercial and economic justifications for using a lipase over any chemical modification process are embedded in the multifaceted patterns of selectivity offered by an enzymatic reaction.To facilitate the broadest possible application of lipases to modify lipids for added value, it is imperative to develop an understanding of lipase selectivity patterns in a manner that allows the application of such knowledge to a host of possible reactions, where the identity and levels of lipid substrates may be quite varied. One approach that fulfills this need is the determination of relative kinetic constants that define the substrate selectivity patterns of various lipases. Selectivity constants may be used to predict the pattern of lipase reactivity under a broad range of conditions where substrate identity and levels are variable, since kinetic constants are not concentration-dependent. Many groups have used this approach to quantify the discriminatory power of various lipases among substrates for esterification reactions between FA and alcohols (6-10).Our approach has focused on kinetically chara...

Reaction selectivity of rhizomucor miehei lipase as influenced by monoacylation of sn‐glycerol

“…Enhanced selectivity for C18 FA incorporation into the MAG pool was observed (Fig. 6A), as was also noted for reactions mediated by PS-30 lipase (16). A likely explanation is the hydrolysis of the corresponding C16,CX-DAG species, despite rather low levels of DAG accumulated in this reaction system (Table 1; Figs.…”

“…TAG accumulated readily in reactions employing β-CX-MAG, but only for reactions with α-C4-MAG and α-C18:1-MAG among the α-CX-MAG series. The lack of TAG accumulation in cases of α-C10-MAG and α-C16-MAG represents a lack of reactivity, which is a direct reflection of enzyme selectivity and which may be founded on steric constraints as advanced previously (16).…”

“…General discussion. The overall objective of this work was to establish the patterns of reaction selectivity of RM lipase in terms of successive steps in assembling TAG from MAG or glycerol and FA to provide a comparison and contrast with the patterns established for PS-30 lipase (16). The same caveats offered in the preceding paper also apply to the present study.…”

“…Sources of sn-glycerol-1(3)-monobutyrin (α-C4-MAG), sn-glycerol-1(3)-monocaprin (α-C10-MAG), snglycerol-1(3)-monopalmitin (α-C16-MAG), sn-glycerol-1(3)-monoolein (α-C18:1 MAG), sn-glycerol-2-monobutyrin (β-C4-MAG), sn-glycerol-2-monocaprin (β-C10-MAG), snglycerol-2-monopalmitin (β-C16-MAG), sn-glycerol-2-monoolein (β-C18:1-MAG), and other reagents were as identified in the companion paper (16).…”

“…Glycerol, when used, was immobilized on silica gel prior to introduction into the reaction mixture. Reactions were carried out in n-hexane (20 mL), and water activity (a w ) was poised at 0.19 by the salt hydrate pair (1.2 g each) of anhydrous Na 2 HPO 4 and Na 2 HPO 4 ·(H 2 O) 2 (16).…”