Formation of protein-oligosaccharide conjugates by laccase and tyrosinase

Proteins and certain carbohydrates contain phenolic moieties, which are potential sites for modification of the function of the biopolymers. In this study, the capability of two different fungal oxidative enzymes, laccase from Trametes hirsuta (ThL) and tyrosinase from Trichoderma reesei (TrT), to catalyze formation of hetero-cross-linking between tyrosine side chains of alpha-casein and phenolic acids of hydrolyzed oat spelt xylan (hOSX) was studied. Formation of reaction products was followed by size exclusion chromatography (SEC), fluorescence spectroscopy, and SDS-PAGE, using specific staining methods for proteins and protein-carbohydrate conjugates. ThL and TrT were observed to differ significantly in their ability to catalyze the formation of protein-carbohydrate conjugates or the linking of the small molecular weight phenolic compounds to alpha-casein. The efficiency of these enzymes to directly cross-link protein also differed notably. TrT was able to cross-link alpha-casein more efficiently than ThL. ThL-catalyzed casein cross-linking was significantly enhanced by ferulic acid, p-coumaric acid, and also hOSX. The main reaction products by ThL appeared to be phenolic acid-bridged alpha-caseins. Indications of hetero-cross-link formation between alpha-casein and hOSX by both oxidative enzymes could be visualized by glycoprotein-specific staining in the SDS-PAGE analysis, although ThL was observed to be more effective in the heteroconjugate formation than TrT.     

Model approach to starch functionality in bread making

We used modified wheat starches in gluten-starch flour models to study the role of starch in bread making. Incorporation of hydroxypropylated starch in the recipe reduced loaf volume and initial crumb firmness and increased crumb gas cell size. Firming rate and firmness after storage increased for loaves containing the least hydroxypropylated starch. Inclusion of cross-linked starch had little effect on loaf volume or crumb structure but increased crumb firmness. The firming rate was mostly similar to that of control samples. Presumably, the moment and extent of starch gelatinization and the concomitant water migration influence the structure formation during baking. Initial bread firmness seems determined by the rigidity of the gelatinized granules and leached amylose. Amylopectin retrogradation and strengthening of a long-range network by intensifying the inter- and intramolecular starch-starch and possibly also starch-gluten interactions (presumably because of water incorporation in retrograded amylopectin crystallites) play an important role in firming.     

Effects of tyrosinase and laccase on oat proteins and quality parameters of gluten-free oat breads

Effects of a Trichoderma reesei tyrosinase (TYR) and a Trametes hirsuta laccase (LAC) on breadmaking performance of gluten-free oat flour were investigated by SDS-PAGE analysis of oat protein fractions, large deformation rheology, and microscopy of the doughs, as well as on the basis of specific volume and firmness of the gluten-free breads. TYR induced the formation of higher molecular weight proteins in the SDS-PAGE assay. Microscopical analysis showed more intensive protein aggregation in the TYR-treated dough than in the dough without TYR. TYR also increased the firmness of the dough, which was assumed to be because of the cross-linking of oat globulins. LAC did not affect the oat globulins. TYR alone, or together with a commercial Thermomyces lanuginosus xylanase (XYL), increased significantly the specific volume of the gluten-free oat bread. A combination of TYR and XYL also increased the softness of the bread, whereas a combination of LAC and XYL improved the specific volume but did not affect the softness of oat bread. The results thus indicate that cross-linking of oat globulins by TYR, especially with the addition of XYL, was beneficial for improving the texture of gluten-free oat bread.     

Reaction kinetics of gliadin-glutenin cross-linking in model systems and in bread making

The gluten proteins gliadin and glutenin are important for wheat flour functionality in bread making, where, during baking, they polymerize through a heat-induced sulfhydryl-disulfide exchange mechanism. A model system was used to study the kinetics of this reaction. Thus, gluten was subjected to hydrothermal treatment with the rapid visco analyzer (RVA) with holding temperatures of 80, 90, and 95 degrees C. At these temperatures, omega-gliadin solubility did not change, but the solubilities of alpha- and gamma-gliadin in 60% ethanol decreased according to first-order reaction kinetics. All reaction rate constants increased with temperature. The activation energies for the heat-induced exchange reaction were 110 and 147 kJ/mol for alpha- and gamma-gliadin, respectively. Starch did not influence the reaction rates of the association of alpha- and gamma-gliadin with glutenin. During gluten-starch model bread baking, glutenin oxidized first, and when the internal crumb temperature reached 100 degrees C, alpha- and gamma-gliadin cross-linked to glutenin, again following first-order reaction kinetics. The experimental findings and similarities in temperature conditions and reaction kinetics suggest that the RVA system can be instrumental in understanding gluten behavior in concentrated food systems, such as bread making.     

Detection of soft wheat in semolina and durum wheat bread by analysis of DNA microsatellites

The aim of this work was to evaluate the analysis of DNA microsatellites for the detection of soft wheat (Triticum aestivum L.) in semolina and durum wheat bread (prepared from Triticum turgidum L. var. durum). The results enabled selection of an efficient D-genome-specific repetitive DNA sequence to detect common wheat in semolina and breads by qualitative PCR with a threshold of 3 and 5%, respectively, lowered to 2.5% by real-time PCR. This is of major importance for checking during production of some typical products recently awarded the European Protected Designation of Origin (PDO) mark such as Altamura bread, which should not contain soft wheat flour. The feasibility of quantification of common wheat adulteration in semolina using real-time PCR was also demonstrated.     

Water uptake mechanism in crispy bread crust

Crispness is an important quality characteristic of dry solid food products such as crispy rolls. Its retention is directly related to the kinetics of water uptake by the crust. In this study, a method for the evaluation of the water sorption kinetics in bread crust is proposed. Two different sorption experiments were used: an oscillatory sorption test and a sorption test in which the air relative humidity (RH) was increased stepwise. These two experiments had different time scales, which made it possible to get a better understanding of the mechanisms involved. Results show that the adsorption and desorption dynamics of the oscillatory sorption test could be described by a single exponential in time. The water uptake rate ( k) was one of the fitting parameters. A maximum in the water uptake rate was found for a RH value between 50 and 70%. The rate parameters of the experiment where RH was increased stepwise were around a factor 10 lower than those derived from oscillatory sorption experiments. This is an important factor when designing experiments for the determination of water uptake rates. In addition, also a parameter describing the time dependence of the rate parameters of the oscillatory sorption experiment was calculated (C), again by fitting a single exponential to the rate parameters. C was in the same range as the rate parameter of the isotherm experiment. This indicates that different (relaxation) processes are acting at the same time in the bread crust during water uptake.     

Molecular cytogenetic characterization of the amphiploid between bread wheat and Psathyrostachys huashanica

A new wheat-Psathyrostachys huashanica amphiploid, PHW-SA, was characterized using molecular cytological tools to evaluate the potential utilization of P. huashanica for wheat improvement. PHW-SA pollen mother cells (PMCs) regularly revealed averagely 0.57 univalents, 24.51 ring bivalents, 3.19 rod bivalents and 0.01 trivalents per cell. Complete homologous chromosome pairing was seen in 81% of the PMCs, indicating a degree of cytological stability. P. huashanica C-banding karyotypes of PHW-SA displayed strong heterochromatin terminal bands at either or both ends. 24 P. huashanica chromosome arms showed telomeric bands. Using P. huashanica DNA as a probe and J-11 DNA as blocker, distinctive P. huashanica chromosomes, which were presented entirely greenish-yellow hybridization signals, were observed in a somatic metaphase of PHW-SA. GISH also revealed the chromosomal breakage and translocation occurring in the amphiploid PHW-SA, which may represent a centromeric fusion between P. huashanica and wheat chromosomes. Seeds storage proteins electrophoresis indicated that PHW-SA expressed some of P. huashanica specific gliadin and glutenin bands, and a few gliadin and glutenin bands of the parents disappeared and new bands appeared. The results indicated that the amphiploid PHW-SA could serve as novel germplasm sources for wheat improvement.     

Use of psychrophilic xylanases provides insight into the xylanase functionality in bread making

The bread-improving potential of three psychrophilic xylanases from Pseudoalteromonas haloplanktis TAH3A (XPH), Flavobacterium sp. MSY-2 (rXFH), and unknown bacterial origin (rXyn8) was compared to that of the mesophilic xylanases from Bacillus subtilis (XBS) and Aspergillus aculeatus (XAA). XPH, rXFH, and rXyn8 increased specific bread volumes up to 28%, 18%, and 18%, respectively, while XBS and XAA gave increases of 23% and 12%, respectively. This could be related to their substrate hydrolysis behavior. Xylanases with a high capacity to solubilize water-unextractable arabinoxylan (WU-AX) during mixing, such as XBS and XPH, increased bread volume more than xylanases that mainly solubilized WU-AX during fermentation, such as rXFH, rXyn8, and XAA. Irrespective of their intrinsic bread-improving potential, the dosages needed to increase bread volume to a similar extent were much lower for psychrophilic than for mesophilic xylanases. The xylanase efficiency mainly depended on the enzyme's temperature activity profile and its inhibition sensitivity.     

Impact of redox agents on the extractability of gluten proteins during bread making

The gluten proteins gliadin and glutenin are important for dough and bread characteristics. In the present work, redox agents were used to impact gluten properties and to study gliadin-glutenin interactions in bread making. In control bread making, mixing increased the extractability of glutenin. The level of SDS-extractable glutenin decreased during fermentation and then further in the oven. The levels of extractable alpha- and gamma-gliadin also decreased during bread baking due to gliadin-glutenin polymerization. Neither oxidizing nor reducing agents had an impact on glutenin extractabilities after mixing. The redox additives did not affect omega-gliadin extractabilities during bread making due to their lack of cysteine residues. Potassium iodate (0.82-2.47 micromol/g of protein) and potassium bromate (1.07-3.17 micromol/g of protein) increased both alpha- and gamma-gliadin extractabilities during baking. Increasing concentrations of glutathione (1.15-3.45 micromol/g of protein) decreased levels of extractable alpha- and gamma-gliadins during baking. The work not only demonstrated that, during baking, glutenin and gliadin polymerize through heat-induced sulfhydryl-disulfide exchange reactions, but also demonstrated for the first time that oxidizing agents, besides their effect on dough rheology and hence bread volume, hinder gliadin-glutenin linking during baking, while glutathione increases the degree of covalent gliadin to glutenin linking.     

Food allergy to wheat products: the effect of bread baking and in vitro digestion on wheat allergenic proteins. A study with bread dough, crumb, and crust.

The effect of baking and digestion on the allergenicity of wheat flour proteins has been studied. Pooled sera of patients suffering from food allergy to wheat products were tested for IgE binding to the proteins of the wheat dough and of the bread crumb and crust, before and after being in vitro digested. During in vitro digestion, the IgE binding protein components of the unheated dough tended to disappear, whereas a permanence of IgE recognition was evident for both the bread crumb and crust. This indicates that the baking process increases the resistance of the potential allergens of the wheat flour to proteolytic digestion, allowing them to reach the gastrointestinal tract, where they can elicit the immunological response. Therefore, the effects of baking must be carefully considered in studying food allergies to wheat products.     

Zinc biofortification of bread wheat,triticale, and durum wheat cultivars by foliar zinc fertilization

Poor zinc (Zn) nutrition of wheat is one of the main causes of poor human health in developing countries. A field experiment with no zinc and foliar zinc application (0.5% ZnSO₄.7H₂O) on bread wheat (8), durum wheat (3), and triticale (4) cultivars was conducted in a randomized block design with three replications in 2 years. The experimental soil texture was loamy sand with slightly alkalinity. The grain yields of bread wheat, triticale, and durum wheat cultivars increased from 43.6 to 56.4, 46.5 to 51.6, and 49.4 to 53.5 t ha^?1, respectively, with foliar application of 0.5% ZnSO₄.7H₂O. The highest grain yield was recorded by PBW 550 (wheat), TL 2942 (triticale), and PDW 291 (durum), which was 5.22, 4.24, and 4.56% and significantly higher over no zinc. Foliar zinc application increased zinc in bread wheat, triticale, and durum wheat cultivars grains varying from 31.0 to 63.0, 29.3 to 61.8, and 30.2 to 62.4?mg kg^?1, respectively. So, agronomic biofortification is the best way which enriching the wheat grains with zinc for human consumption.     

Effects of laccase and xylanase on the chemical and rheological properties of oat and wheat doughs

The effects of Trametes hirsuta laccase and Pentopan Mono BG xylanase and their combination on oat, wheat, and mixed oat-wheat doughs and the corresponding breads were investigated. Laccase treatment decreased the content of water-extractable arabinoxylan (WEAX) in oat dough due to oxidative cross-linking of feruloylated arabinoxylans. Laccase treatment also increased the proportion of water-soluble polysaccharides (WSNSP) apparently due to the beta-glucanase side activity present in the laccase preparation. As a result of the laccase treatment, the firmness of fresh oat bread was increased. Xylanase treatment doubled the content of WEAX in oat dough and slightly increased the amount of WSNSP. Increased stiffness of the dough and firmness of the fresh bread were detected, probably because of the increased WEAX content, which decreased the amount of water available for beta-glucan. The combination of laccase and xylanase produced slight hydrolysis of beta-glucan by the beta-glucanase side activity of laccase and enhanced the availability of AX for xylanase with concomitant reduction of the amount and molar mass of WSNSP. Subsequently, the volume of oat bread was increased. Laccase treatment tightened wheat dough, probably due to cross-linking of WEAX to higher molecular weight. In oat-wheat dough, laccase slightly increased the proportion of WSNSP between medium to low molecular weight and increased the specific volume of the bread. Xylanase increased the contents of WEAX and WSNSP between medium to low molecular weight in oat-wheat dough, which increased the softness of the dough, as well as the specific volume and softness of the bread. The results thus indicate that a combination of laccase and xylanase was beneficial for the textures of both oat and oat-wheat breads.     

钾肥用量对面包强筋小麦产量和品质的影响

采用田间试验研究不同钾肥用量对龙 94 - 40 83面包强筋小麦产量与品质的影响 ,结果表明施钾明显增加小麦产量 ,K2 O用量 5 2 .5～ 82 .5 kg/ hm2 比较适宜。施钾能明显改善小麦品质 ,小麦粗蛋白和湿面筋含量、沉降值、稳定时间、最大阻力、延伸性、拉伸面积、面包体积和面包评分均有不同程度的提高 ,尤其是对稳定时间的延长效果更为明显。     

Nonstarch polysaccharides in wheat flour wire-cut cookie making

Nonstarch polysaccharides in wheat flour have significant capacity to affect the processing quality of wheat flour dough and the finished quality of wheat flour products. Most research has focused on the effects of arabinoxylans (AX) in bread making. This study found that water-extractable AX and arabinogalactan peptides can predict variation in pastry wheat quality as captured by the wire-cut cookie model system. The sum of water-extractable AX plus arabinogalactan was highly predictive of cookie spread factor. The combination of cookie spread factor and the ratio of water-extractable arabinose to xylose predicted peak force of the three-point bend test of cookie texture.     

Synthesis and antityrosinase mechanism of benzaldehyde thiosemicarbazones: novel tyrosinase inhibitors

p-Hydroxybenzaldehyde thiosemicarbazone (HBT) and p-methoxybenzaldehyde thiosemicarbazone (MBT) were synthesized and established by (1)H NMR and mass spectra. Both compounds were evaluated for their inhibition activities on mushroom tyrosinase and free-cell tyrosinase and melanoma production from B(16) mouse melanoma cells. Results showed that both compounds exhibited significant inhibitory effects on the enzyme activities. HBT and MBT decreased the steady state of the monophenolase activity sharply, and the IC(50) values were estimated as 0.76 and 7.0 μM, respectively. MBT lengthened the lag time, but HBT could not. HBT and MBT inhibited diphenolase activity dose-dependently, and their IC(50) values were estimated as 3.80 and 2.62 μM, respectively. Kinetic analyses showed that inhibition type by both compounds was reversible and their mechanisms were mixed-type. Their inhibition constants were also determined and compared. The research may supply the basis for the development of new food preservatives and cosmetic additives.     

Shift of grain protein composition in bread wheat under summer drought events

Climate change bears the risk of more frequent drought stress in the northern hemisphere with more frequent early summer drought events affecting main grain crops. Winter wheat (Triticum aestivum L.) is susceptible for such drought events at the flowering and grain filling stages. After drought, the grain yield decrease of three hybrids was about 20% lower compared to three wheat lines analyzed. Wheat grain proteins are classified into four main components such as albumin and globulin, gliadin, and glutenin. The latter two are closely related to the baking quality of flour and might be affected by drought. However, detailed knowledge about the influence of drought on the synthesis of specific storage protein fractions is scarce. By analyzing the grain protein fractions by means of SDS‐PAGE technique, we detected an increase in grain protein content as well as in HMW and some LMW glutenin sub‐fractions. The glutenin fraction seems to be most variable in gene expression under different environmental scenarios such as drought. However, the protein yield as well as the grain yield may be strongly decreased, which might be not acceptable in practice.     

氮钾营养对面包强筋小麦产量和品质的影响

采用田间试验研究N、K不同用量及其配比对龙94-4083强筋小麦产量及品质的影响。结果表明,不同N、K营养对小麦产量、养分吸收及其构成和品质有明显的影响。适宜的N、K配比及用量可提高小麦干重、有效穗数、穗长、穗粒数、千粒重,提高小麦植株对N、P、K养分的吸收。小麦产量及其构成均表现出随N、K用量的增加而增加的趋势。所有处理中以N112.5K75处理获得了最高的产量。品质分析结果显示,小麦子粒粗蛋白、湿面筋、沉降值和稳定时间均表现出随N、K用量的提高而增加的趋势。全部处理中以N112.5K75处理获得了较高的沉降值、粗蛋白和湿面筋含量及较长的稳定时间。N、K用量及其配比适宜既能提高小麦产量,又能改善小麦品质。     

Effect of spelt wheat flour and kernel on bread composition and nutritional characteristics

Spelt wheat seeds (Triticum aestivum subsp. spelta cv. Ostro) were used to obtain white spelt flour (64.5% yield), wholemeal spelt flour (100% yield), and scalded spelt wheat kernels. From these materials, white spelt wheat bread (WSB), wholemeal spelt wheat bread (WMSB), and spelt wheat bread with scalded spelt wheat kernels (SSKB) were made and were compared to the reference white wheat bread (WWB). The spelt wheat flours and breads contained more proteins in comparison to wheat flour and bread. Among the samples the highest rate of starch hydrolysis was noticed in WSB. During the first 30 min of incubation this particular bread was shown to have significantly more (P < 0.05) rapidly digestible starch than the WMSB and later on also more starch than in WWB and SSKB, respectively. The WMSB had the lowest hydrolysis index (HI = 95.7). However, the result did not differ significantly from that in the reference common wheat bread. On the other hand, the most refined spelt wheat flour resulted in a bread product (WSB) that was statistically withdrawn (P < 0.05) as one with the highest HI (112.6).     

Assessment of iron bioavailability in whole wheat bread by addition of phytase-producing bifidobacteria

In this study, the influence of phytase-producing Bifidobacterium strains during the breadmaking process (direct or indirect) on final bread Fe dialyzability and ferritin formation in Caco-2 cell as a measure of cell Fe uptake was assessed. The addition of bifidobacteria significantly reduced the InsP(6) + InsP(5) concentrations compared to control samples. Fe-dialyzable contents for samples with bifidobacteria were increased 2.3-5.6-fold, and dialyzability was improved by 2.6-8.6% compared to controls. However, this was not reflected in an increase of Fe uptake by Caco-2 cells as was predicted by the phytate/Fe molar ratios. The results demonstrated the usefulness of phytase-producing bifidobacteria to reduce phytate during the breadmaking process and to increase Fe accessibility, although the effects appeared to be still insufficient to improve Fe bioavailability in Caco-2 cells. Further refinement of the use of phytase-producing bifidobacterial strains and/or breadmaking technological processes is deserved for improving Fe uptake.