Aline Boussard

Effect of Mixing Conditions on the Behavior of Lipoxygenase, Peroxidase, and Catalase in Wheat Flour Doughs

ABSTRACT The effect of mixing has been tested on the extractable activities of lipoxygenase, peroxidase, and catalase from dough after 2, 5, and 20 min of mixing, and 30 min of rest period after 20 min of mixing. Different mixing conditions have been studied including temperature, atmosphere, speed, amount of water added to the dough, buffer solutions between pH 3.6 and 7.5 added to the dough, and different additives (linoleic acid, guaiacol, hydrogen peroxide, ascorbic acid, cysteine, yeast, and sodium chloride). In all the mixing conditions tested, the dough peroxidase activity remains equivalent to the initial flour activity, whereas losses in lipoxygenase and catalase activities largely varied according to mixing conditions. The results show that a self-destruction mechanism as well as physicochemical denaturation are responsible for these losses. Lipoxygenase losses seem mainly associated with the former mechanism, whereas catalase losses are highly increased in acidic conditions (physicochemical den...

Effect of Adding Exogenous Oxidative Enzymes on the Activity of Three Endogenous Oxidoreductases During Mixing of Wheat Flour Dough

ABSTRACT The behavior of different exogenous enzymes (soybean lipoxygenase [SLOX], horseradish peroxidase [HPOD], catalase from bovine liver [BCAT], and glucose oxidase [GOX] from Aspergillus niger) added to dough was studied during mixing. The effect of adding these exogenous oxidoreductases on the activity of three oxidative enzymes present in wheat flour (lipoxygenase [WLOX], peroxidase [WPOD], and catalase [WCAT]) was examined. Proper assay conditions were established to differentiate between added WLOX, WPOD, and WCAT and the corresponding activities present in wheat flour. For doughs with added SLOX, an immediate loss of extractable SLOX (≈40%) was observed which remained constant during further mixing. When compared with the control dough, addition of SLOX decreased the losses in WLOX and WCAT activities, whereas WPOD activity was unaffected. With doughs supplemented by HPOD, an immediate loss of 20% in the HPOD activity was observed which did not change after 20 min of mixing. Compared with contro...

Effect of Storage Temperature and Flour Water Content on Lipids, Lipoxygenase Activity, and Oxygen Uptake During Dough Mixing

ABSTRACT Flours differing in water content of 10% (F10), 12% (F12), and 14% (F14) were stored for 16 weeks at 22, 32, and 45°C. The major changes in lipids concerned the free fatty acids (increase) and the triglycerides (decrease). In all cases, the changes increased with increasing storage temperature and water content. After 16 weeks of storage, the losses in lipoxygenase (LOX) activity increased with increasing flour moisture and storage temperature from 10% for F10 at 22°C to 100% for F14 at 45°C. At the end of storage at 22 and 32°C, the bread volumes decreased by 10 and 25%, respectively, with no statistical differences (P < 0.05) between the samples. At 45°C, the volume losses were equal to 35, 46, and 61% for the F10, F12, and F14 samples, respectively. In the same time, the flour oxidative ability (oxygen uptake during dough mixing) increased for the F10 and F12 samples with increasing storage temperature, whereas it decreased for the F14 samples stored at 45°C. Therefore, provided the residual L...

3D-front-face fluorescence spectroscopy and independent components analysis: a new way to monitor bread dough development

Following bread dough development can be a hard task as no reliable method exists to give the optimal mixing time. Dough development is linked to the evolution of gluten proteins, carbohydrates and lipids which can result in modifications in the spectral properties of the various fluorophores naturally present in the system. In this paper, we propose to use 3-D-front-face-fluorescence (3D-FFF) spectroscopy in the 250-550nm domain to follow the dough development as influenced by formulation (addition or not of glucose, glucose oxidase and ferulic acid in the dough recipe) and mixing time (2, 4, 6 and 8min). In all the 32 dough samples as well as in flour, three regions of maximum fluorescence intensities have been observed at 320nm after excitation at 295nm (Region 1), at 420nm after excitation at 360nm (Region 2) and 450nm after excitation at 390nm (Region 3). The principal components analysis (PCA) of the evolution of these maxima shows that the formulations with and without ferulic acid are clearly separated since the presence of ferulic acid induces a decrease of fluorescence in Region 1 and an increase in Regions 2 and 3. In addition, a kinetic effect of the mixing time can be observed (decrease of fluorescence in the Regions 1 and 2) mainly in the absence of ferulic acid. The analysis of variance (ANOVA) on these maximum values statistically confirms these observations. Independent components analysis (ICA) is also applied to the complete 3-D-FFF spectra in order to extract interpretable signals from spectral data which reflect the complex contribution of several fluorophores as influenced by their environment. In all cases, 3 signals can be clearly separated matching the 3 regions of maximal fluorescence. The signals corresponding to regions 1 and 2 can be ascribed to proteins and ferulic acid respectively, whereas the fluorophores associated with the 3rd signal (corresponding to region 3) remain unidentified. Good correlations are obtained between the IC score values of the 3 signals and the fluorescence intensities in Region 1, Region 2 and Region 3. Ferulic acid addition increases fluorescence in Region 2 and decreases fluorescence in Region 1, probably via a reabsorption of the protein fluorescence by ferulic acid. These phenomena are less pronounced when glucose oxidase is present. The enzymatic oxidation of ferulic acid by the glucose oxidase-peroxidase association could explain some of these effects.

Use of ESR and HPLC to follow the anaerobic reaction catalysed by lipoxygenases

The measurement of the 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPOL) consumption by using ESR allows to follow the anaerobic reaction between linoleic acid (LH) and its 13-hydroperoxide (LOOH) catalysed by lipoxygenase. During this reaction, two types of radicals are initially obtained, alkyl (L) and alkoxyl (LO) radicals which formed two types of adducts (LT and OLT) with TEMPOL as characterised by HPLC. The stoichiometry of the adduct formation is two mole of TEMPOL consumed for one mole of LH and one mole of LOOH. Using ESR, the kinetic parameters and the mechanism of the anaerobic reaction have been determined at pH 6.5 for three different lipoxygenases, soybean, horse bean and wheat and compared to the values obtained at pH 9 for soybean lipoxygenase. Wheat lipoxygenase is very weakly active compared to the other enzymes. An uncompetitive inhibition of the anaerobic reaction catalysed by soybean and horse bean lipoxygenases was observed with 2,6-di-tert-butyl-4-methylphenol (BHT).