Ninfa Rangel Pedersen

Synthesis of sucrose laurate using a new alkaline protease

Abstract Sucrose laurate esters were synthesized from sucrose and vinyl laurate in organic solvents using an alkaline protease from a new alkalophilic strain, Bacillus pseudofirmus AL-89. Maximum synthetic activity was observed in the presence of 7.5% v/v water and in the pH range of 7–10. With protease AL-89 esterification occurred predominantly at the 2- O -position while subtilisin A-catalyzed monoester formation predominantly at the 1′- O position. In the absence of enzyme, buffer salts catalyzed non-specific reactions, resulting in the formation of a number of esters. Non-specific catalysis was also observed upon inhibition of the enzyme using a serine protease inhibitor or upon deactivation of the enzyme at pH above 10.

Efficient transesterification of sucrose catalysed by the metalloprotease thermolysin in dimethylsulfoxide.

Thermolysin catalyses the formation of sucrose esters from sucrose and vinyl laurate in dimethylsulfoxide, with a specific activity of 53 nmol/min/mg and 2‐O‐lauroyl‐sucrose as the main product. Such transesterification reactions are normally observed only when the mechanism involves an acyl enzyme intermediate, as with lipases or serine proteases, and not with metalloproteases like thermolysin. A possible reason is the affinity of the active site of thermolysin for sugar moieties, as for the potent inhibitor phosphoramidon. The reaction is not catalysed by other proteins under the same conditions, and is inhibited by removal of the active site zinc.

Visualization of the Anticaging Effect of Ronozyme WX Xylanase on Wheat Substrates

ABSTRACT A fluorescence microscope was used for visualization of the anticaging effect of a commercial xylanase on milled wheat, microtome cuts of wheat grains, and digesta samples obtained from piglets 1 or 4 h after feeding a wheat-based diet (wheat 490 g/kg of diet, barley 100 g/kg of diet, and oats 100 g/kg of diet). Both starchy endosperm and aleurone cell walls were shown to be broken down by Ronozyme WX commercial xylanase. Data obtained by fluorescence microscopy was supplemented with measurements of starch and xylose released as a result of degradation of nonstarch polysaccharides by the xylanase. The results visualize and provide evidence that Ronozyme WX commercial xylanase can overcome the so-called cage effect. This release of nutrients from their encapsulation in cereal cell wall structures has positive impact on nutrient digestibility and partially explains the positive effect of xylanase supplementation on livestock performance.

The degradation of arabinoxylan-rich cell walls in digesta obtained from piglets fed wheat-based diets varies depending on digesta collection site, type of cereal, and source of exogenous xylanase

The objective of the present study was to compare the ability of experimental and commercial xylanases to degrade, in vitro, the arabinoxylan (AX) fraction in digesta from 28-d-old piglets fed a wheat (Triticum aestivum)-based diet (49% wheat). Pigs were euthanized at 1, 2, 3, or 4 h after feeding; stomach and ileum contents were isolated and frozen and later used for the in vitro studies. Xylan solubilization provided information regarding the ability of the enzymes to degrade AX during the harsh in vivo conditions prevailing in the gastrointestinal tract. The hydrolytic capacity of a commercial xylanase was compared with that of an experimental xylanase using stomach digesta (pH 1.8) obtained at 4 h after feeding. Relative to the control, both enzymes increased (P < 0.001) xylan solubilization 3-fold. In the ileal digesta (1 h), xylan solubilization was increased by 36% (P < 0.001). Inclusion of arabinofuranosidases (Ara f) with xylanases increased xylan solubilization in stomach samples (P = 0. 007 and P = 0. 030) but not in ileal samples (P = 0.873 and P = 0.997). Our results illustrate clearly the importance of using different conditions and substrates when enzyme performance is studied in vitro as a prescreening tool for setting up in vivo trials.