Serge Bérot

Large scale isolation of water-soluble and water-insoluble pentosans from wheat flour

Abstract A procedure has been developed which allows the isolation of enriched pentosan fractions from wheat flour. The procedure is based on mixing flour and water (50 kg/150L). After centrifugation the supernatant is heat-treated and then concentrated by cross-flow ultrafiltration to give water-soluble pentosans, whereas the insoluble residue is treated with Alcalase and then Termamyl to give water-insoluble pentosans. Starting from 50 kg of flour from three different varieties (Soissons, Thesee, Apollo), water-soluble and water-insoluble pentosans containing respectively 100–200 g of water-soluble and 250–350 g of water-insoluble arabinoxylans, were prepared within 2 d. Water-soluble pentosans are constituted of approx. 40% arabinoxylans and are contaminated by proteins, whereas water-insoluble pentosans are composed of arabinoxylans (ca. 30%) and are contaminated by residual starch (ca 40%). Enrichment factors in arabinoxylans ranged from 98 to 66 for water-soluble and 27 to 19 for water-insoluble pentosans as compared with initial flour.

Ultrafiltration to fractionate wheat polypeptides

An ultrafiltration process allowing the fractionation of two kinds of polypeptides issued from limited chymotryptic hydrolysis of wheat gliadins was applied to wheat gluten hydrolysates. Hydrophilic and poorly charged polypeptides were well transmitted through an inorganic ZrO2-based membrane at acidic pH, whereas hydrophobic and positively charged polypeptides were highly retained. By combining reversed-phase and cation-exchange chromatography (CEC), it was proved that the fractionation of the polypeptides was based on electrostatic repulsion of the charged polypeptides by the positively charged membrane. After a continuous diafiltration process, retentates containing 75 to 88% of hydrophobic polypeptide and permeates containing 84 to 90% of hydrophilic polypeptides were recovered, depending on the size of membrane used. Even if the ultrafiltration fractions were less purified than fractions issued from CEC, it was shown that they exhibited very different foaming properties: permeate did not produce nor stabilize foams, whereas retentate was more efficient than the whole hydrolysates and BSA.

Acetylation of pea isolate in a torus microreactor.

Acetylation, which acts on the amino groups of proteins, allows to increase the solubility and the emulsifying properties of pea isolate. Acetylation by acetic anhydride was carried out in a torus microreactor in semibatch and continuous conditions. The mixing characteristics, obtained by a residence time distribution (RTD) method, are the same in batch and continuous processes. The maximum acetylation degree reached by the torus reactor is higher than with the stirred reactor. Torus reactors are more efficient than stirred ones as shown by a conversion efficiency, defined by the quantity of modified lysine groups by consumed acetic anhydride. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 53: 409-414, 1997.

Effects of two wheat cultivars on physico-chemical properties of wheat flours and digesta from two broiler chicken lines (D+ and D-) differing in digestion capacity

1. The current experiment is the second part of a study about the effects of wheat quality on digestibility of pelleted diets for broiler chickens. In the first part, it was shown that a hard cultivar resulted in a negative effect on starch digestibility in two divergent lines of chickens (D+ and D−) selected for digestion capacity. The aim of this second part was to investigate the reasons for this negative effect of a hard cultivar (Baltimor) compared to a soft one (Scipion) in D+ and D− lines. 2. Proventriculus pepsin activity and pancreas proteolytic and amylolytic activities were estimated in 4 pools of birds: ‘D+ line (Baltimor fed)’, ‘D+ line (Scipion fed)’, ‘D− line (Baltimor fed)’ and ‘D− line (Scipion fed)’. Results suggested the greatest amount of pepsin units per g BW for D+ birds and the lowest amount of pancreas proteolytic units per g BW for D+ birds fed Scipion wheat. Pancreas showed very similar α-amylase activities among treatments. 3. In vitro hydrolyses of wheat gluten proteins with proventriculus extracts from pools of D+ and D− birds did not show any differences between hard and soft cultivars, whatever the origin of pools. 4. Pepsin hydrolysis of fine (300 to 425 µm) and coarse (1180 to 1600 µm) fractions from wheat flours (Baltimor or Scipion) showed that the 30 min proteolysis rate was highest for the fine fraction in both cultivars. No difference was observed with extended hydrolysis time. 5. In vitro digestion simulation of whole wheat flours confirmed the results previously obtained in vivo, with a negative effect of hard cultivar on starch digestion rate and no effect on protein digestion. 6. Laser particle size analyses showed that ileum digesta from birds fed with hard wheat cultivar showed the highest proportion of coarse particles. 7. Microscopic analyses of D+ ileum digesta revealed that the concentration of undigested starch granules in the subaleurone area of wheat bran particles was the highest with hard cultivar. 8. The results suggested that physical entrapment of starch granules in coarse particles was a major explanation for decreased starch digestibility values in chickens fed hard wheat diets.

Influence of pH and salt concentration on the emulsifying properties of native wheat gliadins and of their chymotryptic hydrolysates

Wheat gluten is produced in large amounts in Europe and is used mainly as an improver of the rheological properties of wheat flours in breadmaking. The uses of wheat gluten can be developed and diversified through fractionation and modification. The fractionation of gluten proteins in two fractions enriched respectively in gliadins (monomeric prolamins) and glutenins (aggregated prolamins) was realized at the pilot scale [1]. As expected, the resulting products differed widely in their technological properties [2] and could be used to adapt the rheological properties of wheat flour doughs and of other food systems. Limited enzymatic hydrolysis is another way to enhance the applications of gluten. This treatment increases gluten solubility over a wide range of pHs and makes it possible to express surface activity [3]. Limited hydrolysis of gliadins by chymotrypsin was shown to release essentially two types of peptides. They corresponded to the repetitive and the non-repetitive sequence domains of the proteins, respectively [4 ‐ 6]. When it was solubilized in a good solvent, the nonrepetitive domain isolated from a c-type gliadin exhibited better emulsifying properties than the native proteins, whereas the repetive domain had no stabilizing effect on an alcane/ water interface [7]. In this study we extended our previous work by analysing the emulsifying properties of the hydrolysates from a/b- and c-type gliadins in various conditions of pH and salt concentrations and by identifying the active peptides.

Fractionation of gliadin hydrolysates in water‐ethanol by ultrafiltration with modified or unmodified membranes

Ultrafiltration was applied to the fractionation of neutral vs. charged peptides of similar size. The peptides, produced from gliadins, a major fraction of wheat storage proteins, were obtained by limited hydrolysis with alpha-chymotrypsin in water-ethanol 80/20 (v/v). Peptides, according to their elution by RP-HPLC, were quasineutral (repetitive peptides) irrespective of pH, or positively charged (nonrepetitive peptides) at pH below 5. The transmission through the membranes of the nonrepetitive peptides was less (until sevenfold) than that of the repetitive ones, because of the role of electrostatic repulsion involved in the retention of charged solutes. The difference of transmission was more efficient at acidic pH (3) and low ionic strength with inorganic membranes and in a wider range of pH and ionic strength with membranes modified by coating of positively charged polymers (polyvinylimidazole PVI, polyethyleneimine PEI). A continuous diafiltration process using an inorganic membrane of low molecular cut-off permitted the selective enrichment of the retentate in nonrepetitive peptides (up to 80%) and of the permeate in repetitive peptides (up to 80%) from hydrolysate feed containing about 60/40% of repetitive and nonrepetitive peptides, respectively, with a diafiltration volume of 4.

New methods for chemo-enzymatic galactosidation of 2S rapeseed protein.

Two chemo-enzymatic methodologies to synthesize neoglycoproteins from rapeseed 2S protein (napin) were developed. In the first approach, glycosidases were used to catalyse 1-O-glycosylation of serine residues, whereas in the second one, 6-N-galactosylation was examined using an amino-reduction reaction between the ε-NH2 of lysine residues and 6-oxogalactosides (readily available by means of the oxidation reaction of the corresponding galactosides mediated by galactose oxidase). Our results indicated that glycosidases were unable to glycosylate native proteins. Conversely, this reaction was possible, although in low yields (10%), after the introduction of a hydroxyethylene spacer. The latter modified proteins were obtained via the condensation of ε-NH2 of lysines with ethylene carbonate in basic medium (40% yield). The second approach was much more efficient, as 61% of the lysine residues were shown to be 6-N-galactosylated using sodium cyanoborohydride as a reduction reagent.

Application of a Torus Reactor to Chemical and Enzymatic Modifications of Plant Proteins

The hemodynamic behavior of the torus reactor, a new type of bioreactor suitable for modifying plant proteins, was modeled in batch and continuous conditions. As examples of applications, limited enzymatic hydrolysis of wheat gliadin and acetylation of pea isolate were carried out, and performances were compared with those of classical stirred tank reactors.

Fractionation of Gliadin Hydrolysates by Ultrafiltration

Gliadin was hydrolyzed by α-chymotrypsin in 17 kg/mol peptides differing in their hydrophobicity and electrical charge. These peptides were separated on ultrafiltration membranes at acidic pH. Permeates contained 80 to 96% of neutral peptides, and retentates, which contained 80% of charged peptides, exhibited good emulsifying properties.