Cinnamate esterase

Cinnamate esterase hydrolyzes cinnamic acid esters, such as chlorogenic acid, ethyl cinnamate, and tartaric acid esters. 

Fig. 8.7. Vinyl-phenol formation mechanism in must clarified using a pectinase preparation with cinnamate esterase...

The cinnamate esterase in commercial pectinases has been purified (Barbe, 1995). It is a 240000 Da glycoprotein, consisting of two subunits of 120000 Da. It is strongly inhibited in the presence of ethanol, so it is only active during the... 

These findings have enconraged some pecti-nase prodncers to offer preparations without any contaminant cinnamate esterase, so that there is no longer a risk of increasing the volatile phenol content of white wines when pectinases are used as adjuvants in clarifying white wine must (Table 8.3). 

The pure enzyme was tested for activity against several methylated phenolic and cinnamic acids (Table 2). The enzyme was active on methyl esters of cinnamic acids caffeio p-coumaric> ferulic, and is therefore termed a cinnamoyl esterase (CinnAE). 

Dugelay, I., Giinata, Z., Bitteur, S., Sapis, J.C., Baumes, R., Bayonove, C. (1992a). Formation of volatile phenols from cinnamic precursors during wine making the role of cinnamoyl esterase from commercial enzymic preparations. In P. Schreier P. Winterhalter (Eds.), Progress in flavour studies (pp. 189-193). Carol Stream Ils. Allured Publishing Co. 

Activity of the purified en2ymes was assessed on methyl ferulate (methyl 4-hydroxy 3-methoxy cinnamate, MF). Assays were run for 30 min at 50 °C in 50 mM eitrate buffer at pH 4.8 with initial MF concentrations of 50 to 750 pM and an enzyme concentration of 50 nM. The reactions were terminated after 30 min by boiling for 10 min and analyzed for MF and ferulic acid content via C18 high performance liquid chromatography (HPLC) over a 0 to 100% acetonitrile gradient with 0.1% formic acid in all solutions. Preliminary isothermal titration calorimetry (ITC) on both ferulic acid esterase proteins was conducted in SEC buffer on a Microcal VP-ITC system. All reactions were carried out at 37 °C using MF as a substrate. 

The use of certain pectolytic enzyme preparations to facilitate the extraction or clarification of white must may lead to an increase in the vinyl-phenol content of white wines and a deterioration of their aromatic qualities (Chatonnet et al 1992a Dugelay etal., 1993 Barbe, 1995). Indeed, certain industrial pectinases, made from Aspergillus niger cultures, have a cinnamyl esterase (CE) activity. This enzyme catalyzes the hydrolysis of tartrate esters of hydroxycinnamic acids in must during the pre-fermentation phase (Figure 8.7). Feruhc and j9-coumaric acids are then converted into vinyl-phenols during alcohohc fermentation due to the cinnamate decarboxylase activity of Saccharomyces cerevisiae. 

Ferulic acid is the major cinnamic acid found in a variety of plant cell walls. Com fiber contains about 3% femlic acid. Wheat bran is another source of ferulic acid (0.5-1%). Faulds et al. 226) developed a laboratory scale procedure to produce free femlic acid (5.7 g) from wheat bran (1 kg) by using a Trichoderma xylanase preparation and A, niger femlic acid esterase. Using filamentous fungi, a two-stage process for vanillin formation was developed in which a strain of A. niger was first used to convert femlic acid to vanillic acid, which was then reduced to vanillin by a laccase-deficient strain of Pycnoporus cinnabarinus 227). 

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Feruloyl Esterases. Psychrophilic feruloyl esterases are employed in industry for biomass degradation, biotransformation, and isolation of ferulic acid derivatives useful as antioxidant, antimicrobial, and photoprotectant properties. For instance, one studied feruloyl esterase from the psychrophilic bacterium, P. haloplanktis TAG 125, is a family 1 carbohydrate esterase and displays significant activity toward pNP-acetate, a- and yS-naphthyl acetate, and 4-methylumbelliferyl p-trimethylammonio cinnamate chloride (a model substrate for determining ferulolyl esterase activity) (23). 

Methanol (8-11) is a member of the homologous series of saturated aliphatic alcohols found in plants in the form of various esters, most often in pectins and esters of aromatic carboxylic acids (benzoic, salicylic, cinnamic and others). Free methanol results mainly by hydrolysis of pectin catalysed by plant pectin esterases therefore, it occurs regularly in small amounts as a component of aU natural fruit juices and is present in larger amounts in wines and fruit distillates. 

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