Effect of Excess Nickel on Starch Mobilization in Germinating Rice Grains

The effect of excess nickel (as NiSO₄) on starch mobilization of rice (Oryza sativa L.) grains or dehulled rice grains during germination was investigated. Excess NiSO₄ had no effect on starch content and α-amylase activity in endosperm of germinating rice grains or germinating dehulled rice grains. Evidence is provided to show that the hull is a barrier against influx of Ni²⁺ to endosperm; endosperm per se is less effective in Ni²⁺ uptake; and α-amylase extracted from the endosperm of germinating rice grains is highly resistant to excess Ni²⁺.     

Fructan Content of Rye and Rye Products

The fructan content of Finnish rye grains (13 samples, seven cultivars, harvested in 1998‐2000) varied at 4.6–6.6 g/100 g (db). Commercial whole grain rye flour and rye flakes had fructan content of 4 g/100 g, light refined rye flour had fructan content of 3 g/100 g, and rye bran had fructan content of 7 g/100 g. Fructan content as high as 23 g/100 g was detected in the water‐extractable concentrate of rye bran. Finnish soft rye bread and rye crisp bread contained 2–3 g of fructan/100 g. According to the suggested new definition of dietary fiber, fructans are also classified as dietary fiber. This means that the dietary fiber content of some cereal foods such as rye products may be increased by as much as 20% due to the presence of fructans in the grain.     

Aryl‐Glycosidase Activities in Germinating Maize

Soluble protein extracts of germinating maize seedlings exhibited a limited ability to hydrolyze purified xylans, and specific assays were unable to confirm the presence of endo‐β‐1,4‐xylanase activity. However, extracts contained a variety of aryl‐glycosidase activities, including β‐glucosidase, β‐xylosidase, and α‐l ‐arabinofuranosidase. These activities peaked in three‐ to four‐day seedlings and were particularly concentrated in shoot and root tissues. Maximal levels of β‐glucosidase were two orders of magnitude greater than those of β‐xylosidase or α‐l ‐arabinofuranosidase. Isoelectric focusing gels revealed multiple forms of these enzymes. The principal β‐glucosidase and α‐l ‐arabinofuranosidase protein species were clustered at pI 4.8–4.9 and pI 5.8–6.0, respectively. β‐Xylosidase activity appeared to be associated with both of these enzymes, and no evidence was obtained for a distinct β‐xylosidase.     

Oat Bran Fermentation by Rye Sourdough

Hydration of oat bran including fermentation by rye sourdough was studied. Three types of oat bran suspensions were prepared (a control, one with whole meal rye flour added, and one with rye starter added). The suspensions were incubated for 1, 2, 3 and 4 hr. β‐Glucan content and solubilities of protein and β‐glucan were analyzed. Viscosity of the supernatants of oat bran suspensions was determined. Neither the rye sourdough nor the rye flour alone had a significant effect on the total β‐glucan content of oat bran suspensions. However, the addition of rye, either as whole meal rye flour or as sourdough starter, markedly increased the solubility of β‐glucan and proteins and simultaneously decreased the viscosity of the water‐soluble fraction of oat bran suspension. This suggests that a hydrolysis of β‐glucan had occurred that could change the rheological properties of oat bran in baking and the physiological potential of oat bran in nutrition.     

Degradation of Secalins During Rye Sourdough Fermentation

Rye sourdough (RSD) gives rye bread mildly acidic taste and desired flavor. Flavor precursors (amino acids and small peptides) are generated in the proteolytic breakdown of rye proteins. Our aim was to study the protein degradation during RSD fermentations. Two sourdoughs were prepared of flours derived from two rye cultivars (Amilo and Akusti). RSD samples were collected during fermentations. Three protein fractions were obtained by sequential protein extraction and these were analyzed by SDS‐PAGE. Free amino nitrogen (FAN) was measured with a ninhydrin method. In addition, two rye incubations without starter microorganisms (with antibiotics) were made at pH 3.6 and 6.1, and proteinase profiles of the rye cultivars were analyzed at pH 4.3. SDS‐PAGE analysis showed that during RSD fermentations, rye proteins, especially the alcohol‐soluble secalins, were degraded. Secalins also evidently degraded during the incubation without starter microorganisms at pH 3.6. Aspartic proteinases were in the major proteinase group in both rye cultivars. This study confirms that endogenous proteinases of rye, mainly aspartic proteinases, hydrolyze rye proteins, especially secalins, during RSD fermentation. Protein degradation in rye sourdoughs may thus be enhanced by selecting rye flours with high proteolytic activity toward secalins.     

Proteolytic activity in soil: A review

The aim of this work is to review current knowledge on inputs, sources and regulation of protease activities in soils from different ecosystems, while exploring limitations to proteolysis and N mineralisation. Extracellular proteases enter the soil via microbial production and other sources, including plant root exudates, animal excrements, decomposition processes and leaching from agro-industrial fertilisers. The synthesis and activities of proteases in soil are regulated by many factors, including climate, soil properties and the presence of organic compounds of plant and microbial origin. Two particularly important areas for future research are the regulation of proteolysis by low-molecular-weight organic compounds, including amino acids, sugars, flavonoids, plant hormones and siderophores, as well as the identification and characterisation of proteinaceous protease inhibitors of plant and microbial origin in the soil. Despite all the work that has been performed on soil proteases, our understanding of the roles of extracellular plant root proteases in N nutrition is weak. Furthermore, the regulation of soil proteolytic activities of different ecosystems, especially in terms of pollutant inputs and the impact of climate change, requires investigation. Other areas that pose important questions for the future include assessments of protease inhibitor inputs to the soil, regulation of these inhibitors via naturally occurring soil organic compounds and the interactions between soil organisms.     

Foam‐Forming Capacity of Substances Present in Rye

Aqueous extracts of rye milling products were tested to establish foam‐forming capacity. The foams were characterized according to volume and strength. The extracts were treated chemically, thermally, and enzymatically to demonstrate the influence of the protein and of polymeric nonstarch carbohydrates on foam formation. The foam formation was attributable to a relatively homogeneous water‐soluble protein with a M_r of 12.3 kDa and an isoelectric point at pH 5.97. The other components of the extracts of rye milling products either contribute to a certain extent to stabilizing the foams derived from aqueous extracts (fructosans) or, in some cases, to destabilizing them (pentosans).     

Polymorphism of Aspartic Proteinases in Resting and Germinating Barley Seeds

Both resting and germinated barley seeds (Hordeum vulgare L. ‘Morex’) contain aspartic endopeptidase activities, and the activities increase during germination. We have extracted and partially purified aspartic endopeptidases from both resting seeds and green malt (four-day germinated barley). Six aspartic proteinase activities were found in resting barley seeds while only four activities were detected in green malt. All of the aspartic proteinases had similar pH activity optima (pH 3.5–4.5) and pI values (≈4.5). The purified green malt aspartic proteinases selectively digested a group of barley seed proteins, postulated to serve as defensive proteins, that are coded by the amylase-trypsin inhibitor super gene family. The aspartic proteinases that bound to a pepstatin A affinity column at pH 4.5 cross-reacted with antiserum raised against aspartic proteinases purified from barley seed. However, those that did not bind the affinity column also did not cross-react with the antiserum, indicating that there are two distinct groups of aspartic proteases in germinating barley.     

Incubation of Isolated Wheat Starch with Proteolytic or Lipolytic Enzymes and Different Extraction Media Reveals a Tight Interaction Between Puroindolines and Lipids at Its Granule Surface

Differences in hardness of wheat cultivars have been related to differences in interactions between the starch granule surface and the gluten protein matrix that are mediated by the proteins puroindoline (PIN) A and B. We examined whether or not PINs and (polar) lipids are associated at the starch granule surface, and, if so, how they interact with the starch granule surface itself. Starch was isolated from a soft wheat cultivar containing both wild‐type PINs and incubated with peptidases or lipases, or in extraction media (typically used for defatting). Protein, PIN, and lipid levels revealed that PINs and lipids are tightly associated together at the starch granule surface. Our results imply that PINs need lipids for binding to the granule surface but not vice versa.     

Reoxidation Behavior of Wheat and Rye Glutelin Subunits

The ability of HMW and LMW subunits of wheat glutelin to form a polymeric gluten network by intermolecular disulfide bonds is responsible for the unique rheological properties and baking quality of wheat dough. Because the mechanism of gluten formation is not fully understood, the reoxidation behavior of HMW and LMW subunits of wheat glutelin and HMW subunits of rye glutelin was studied. The subunits were isolated from wheat flour cv. Rektor (REK) and from rye flour cv. Danko (DAN) with a selective extraction and precipitation method. For reoxidation, different oxidants (KBrO₃ and KIO₃), protein concentrations (0.5, 1.0, and 2.0%), solvent compositions, pH values (2.0 and 8.0), and reaction times (0–360 min) were compared. The characterization of reoxidized products was achieved by the determination of the thiol content with the Ellman's reagent, and of the M_r distribution by gel‐permeation chromatography. The results demonstrated that both HMW and LMW subunits could be slowly reoxidized with KBrO₃ to polymers with M_r up to several millions. Yield and M_r distribution of polymers were dependent both on the protein concentration and on the molar ratio of oxidants to thiol groups. The HMW subunits of wheat glutelin (HMW‐REK) yielded slightly higher quantities of polymeric proteins than did the HMW subunits of rye (HMW‐DAN). Reoxidation with KIO₃ proceeded much faster than with KBrO₃ and led to lower proportions of polymerized proteins for HMW‐REK and HMW‐DAN. Obviously, more intra‐ and fewer intermolecular disulfide bonds were formed by reoxidation with KIO₃ compared with KBrO₃. In contrast, LMW‐REK was reoxidized with KIO₃ to higher amounts of polymeric aggregates, which indicated that LMW subunits formed intermolecular disulfide bonds with both KIO₃ and KBrO₃. Independent of the protein type and the oxidant used for reoxidation, more inter‐ and fewer intramolecular disulfide bonds were formed when the protein concentration was increased. Single subunits 5, 7, and 10 were isolated from HMW‐REK by preparative acid‐PAGE and were reoxidized with KBrO₃ for 360 min. The M_r distribution indicated that x‐type subunit 5 had a greater tendency to form polymers than x‐type subunit 7. The y‐type subunit 10 was characterized by a lower proportion of polymers after reoxidation than x‐type subunits 5 and 7.     

Whole Grains and Digestive Health

Whole grains contain all parts of the grain: the endosperm, germ, and bran. Whole grains are rich in fermentable carbohydrates that reach the gut: dietary fiber, resistant starch, and oligosaccharides. Most research that supports the importance of grains to gut health was conducted with isolated fiber fractions, rather than whole grains. Whole grains are an important source of dietary fiber and grain fibers such as wheat, oats, barley, and rye increase stool weight, speed intestinal transit, get fermented to short chain fatty acids, and modify the gut microflora. Wheat bran is particularly effective in increasing stool weight; wheat bran increases stool weight by a ratio of 5:1. In contrast, many novel fibers that are easily incorporated into beverages and foods increase stool weight only on a ratio of 1:1. In vitro fermentation studies with whole grains have been published. Carbohydrates of oat bran (rich in β‐glucan) were consumed by bacteria faster than those of rye and wheat brans (rich in arabinoxylan). Grain fibers were fermented more slowly than inulin, causing less gas production. Wheat is particularly high in fructo‐oligosaccharides, while wheat germ is high in raffinose oligosaccharides. Some in vivo studies show the prebiotic potential of whole grains. Whole grain breakfast cereal was more effective than wheat bran breakfast cereal as a prebiotic, increasing fecal bifidobacteria and lactobacilli in human subjects. Wheat bran consumption increased stool frequency. Thus, the gut enhancing effects of cereal fibers are well known. Limited data exist that whole grains alter gut health.     

Rye and Triticale as Feedstock for Fuel Ethanol Production

Normal gravity rye and triticale mashes, containing 20–21 g of dissolved solids per 100 mL of mash liquid, were fermented with active dry yeast at 27°C. Fermentations were completed within 48 hr for rye, and within 72 hr for triticale. Supplementation of mashes with urea at a concentration of 8 mM accelerated rates of sugar consumption and fermentation, and reduced fermentation time from 48 to 36 hr for rye, and from 72 to 48 hr for triticale. Rye fermented faster than triticale, due to its higher level of free amino nitrogen. Ethanol yields were 356–363 L/tonne of 14% moisture rye grain, and 362–367 L/tonne of 14% moisture triticale. Fermentation efficiencies, which were 90–91% for triticale, and 91–93% for rye, and ethanol yields were comparable to those obtained from wheat and were not affected significantly by urea supplementation. The replacement of wheats by less expensive crops such as rye and triticale would provide good economic opportunities and alternatives for the fuel alcohol industry.     

Proteolytic activity under nitrogen or sulfur limitation

Sand cultures were used to evaluate the effect of C, N, and S ratio on protein degradation by soil microorganisms. Sand was inoculated with soil and amended with defined nutrient media to produce limitation for C, N, or S. Limitation for N or S resulted in reduced biomass (total protein) and increased proteolytic activity as indicated by measurements of dye released from a commercial protease substrate (azocoll). Carbon limitation had little effect on proteolytic activity. As expected, utilization of carbon (glucose) was dependent upon the availability of N or S. Protein synthesis inhibitors (chloramphenicol and cycloheximide) suppressed proteolytic activity, suggesting a need for new gene expression in the response of organisms to N or S stress. Correlations of proteolytic activity and biomass among treatments revealed distinctly different relationships depending upon the availability of C, N, or S. The results of this experiment support a role of proteolytic activity in response of microorganisms to N or S deprivation and suggest that protease activity in soil is more strongly influenced by regulatory signals than by standing biomass.     

Viscosity Concerns with Rye Mashes Used for Ethanol Production

Rye cultivars leading to high-viscosity extracts were easily mashed— even under very high gravity (VHG) conditions—when enzymes were added to reduce viscosity caused by pentosans. The enzymes were so effective that for the fuel alcohol industry, the need for special genetic selections of rye with reduced extract viscosity was not indicated. High-starch cultivars of rye, however, were still most beneficial when alcohol was the end product of value. Both normal and VHG mashes fermented out within 48 hr under the conditions described. Yields (L/t of starch) were, on average, 5.3% higher under VHG conditions than under normal gravity.     

Compaction of Corn Distillers Dried Grains

Corn distillers dried grains (DDGS) were compacted into cylindrical pellets (3.5 cm in length, 1.5 cm in diameter) utilizing a closed‐end die under axial stress from a vertical piston applied by an Instron universal testing machine. The effects of independent variables, including the raw material moisture content (25–35% db), processing temperature (100–120°C), pressure (12.5–37.5 MPa), and dwell time (5–15 sec) on pellet density, durability, and stability were determined using response surface methodology. Moisture content, temperature, and pressure significantly affected (P < 0.05) the properties of DDGS pellets, while the influence of dwell time was negligible (P > 0.05). Increasing temperature initially increased and then decreased unit density. High moisture and pressure had favorable effects on unit density and durability rating. The density ratio increased with increasing pressure and moisture content. The results suggested technical feasibility of compacting DDGS. For the range of variables tested, optimum levels were identified as 34.6% moisture content, 107°C press temperature, and 36.8 MPa pressure to obtain maximum durability and density and acceptable dimensional stability.     

Comparison of Endoproteinases of Various Grains

Two‐dimensional isoelectric focusing (IEF) × PAGE gels were used to compare the endoproteolytic (gelatinase) activities of germinated barley with those of bread and durum wheat, rye, triticale, oat, rice, buckwheat, and sorghum. Barley was used as the standard of comparison because its endoproteinase complement has been studied previously in the greatest detail. The characteristics of the grain proteases were appraised from their migration patterns and by how they were affected by pH levels. All of the germinated grains contained multiple enzyme activities and their separation patterns and pH levels were at least similar to those of barley. The proteinases of the bread and durum wheats, rye, oat, and sorghum were most similar to those of barley, whereas the other grains provided more varied patterns. The rice and buckwheat proteinases developed much more slowly than those of the other grains. The activity patterns of the triticale resembled those of the parents, wheat and rye, but the triticale contained many more activities and higher overall proteolytic activities than any of the other species. These results should be applied to scientific or commercial procedures with caution because grains contain potent endogenous proteinase inhibitors that could inactivate some of these enzymes in various tissues or germination stages.     

Protein Extraction from Triticale Distillers Grains

Triticale is being actively explored as a feedstock for bioethanol production in Western Canada. Triticale distillers grains, an important coproduct of the bioethanol industry, are used mainly as animal feed. This study aims to develop methods of protein extraction from triticale distillers wet grains and distillers dried grains with solubles. Osborne fractionation showed low protein extractability because excessive protein denaturation occurred during sample preparation. Five methods were used to extract proteins: pH shifting, 60% ethanol, alkaline‐ethanol solution, glacial acetic acid, and enzyme‐aided extraction. Extracts obtained with the alkaline‐ethanol and glacial acetic acid methods showed comparatively higher protein contents (≈61–65%) when compared with the other extraction methods (≈35–57%). Enzyme‐aided extraction with Protex 6L yielded 75–82% protein at a content of 43–57%, depending on the types of raw materials. Establishing methods of protein extraction from triticale distillers grains would facilitate further studies on new uses of triticale proteins.     

Soil Inorganic Nitrogen with Incubation of Rye Cover Crop Biomass

A soil incubation study was conducted to evaluate the effect of winter cereal rye (Secale cereale L.) cover crop (CC) biomass and fertilizer nitrogen (N) addition on soil inorganic-N. Rye aboveground biomass was collected following corn (Zea mays L.) and soybean [Glycine max (L.) Merr.], and incubated at equivalent field temperatures for 105 d at rates of 1120, 2240, and 3360 kg dry matter (DM) ha^?1. Despite N addition from the rye biomass at any rate, there was no real effect on ammonium (NH₄)-N, and only from 63 d to 105 d a limited net increase in nitrate (NO₃)-N and inorganic-N was observed compared to no-rye. Nitrate-N and inorganic-N concentrations change per heat unit (HU) accumulation was negative with rye addition through 7 d, but was positive consistently across the remaining incubation period with or without rye. Overall, the rye CC biomass had only a neutral to small positive effect on soil inorganic-N.