Method of analysis for deoxynivalenol and zearalenone from cereal grains

A method was developed to determine deoxynivalenol and zearalenone in corn, wheat, oats, rice and barley. The toxins are extracted with methanol/water (50:50, v/v) (2×) and partially purified by partitioning into ethyl acetate and then defatting with acetonitrile-petroleum ether. Toxins are isolated by silica gel column chromatography. Interfering materials are removed from the column with benzene; zearalenone is eluted with benzene/acetone (95:5, v/v), and after a column wash of chloroform/methanol (95:2, v/v), deoxynivalenol is eluted with chloroform/methanol (95:5, v/v). Zearalenone is quantitated by thin-layer chromatography and deoxynivalenol by gas-liquid chromatography of the trimethylsilyl derivative. The detection limit is about 0.02 μg/g for each toxin. Recoveries of added toxins varied with substrate and level of toxins. Recovery of deoxynivalenol ranged from 58% for 1 ppm in rice to 108% for 1 ppm in corn. Average recoveries for all levels (1, 2 and 5 ppm) ranged from 69% for barley to 89% for oats. Recovered zearalenone ranged from 40% for 5 ppm in wheat to 100% for 1 ppm in barley. Average recoveries for zearalenone at 1, 2 and 5 ppm varied from 53% for wheat to 87% for rice.     

Die Aktivität von Lipoxigenasen in Getreide sowie Möglichkeiten der Enzymhemmung

Activity of Lipoxygenase in Cereals and Possibilities of Enzyme Inhibition In different feed components the activity of the enzyme lipoxigenase was determined. In cereals the enzyme activity predominantly decreases in the following order: barley, wheat, corn, oats and rye. During the malt process the activity of barley lipoxigenases increases to a very high level. During storage ungerminated grains are showing increasing (barley) or decreasing (wheat) variations in enzyme activity. The addition of propionic acid decreases the activity of lipoxigenases in cereals. The peroxide value of added fat in mixed feed was not influenced by lipoxigenases. For wheat products positive relations between the content of crude protein as well as crude fat and the enzyme activity were ascertained. Heat treaments with temperatures exceeding 100% C lowered the lipoxigenase activity to values below 10% of the untreated sample. The usual temperatures during feed processing, e.g. after conditioning and pelleting, are sufficient to inhibit the enzyme activity almost completely.     

Tocol levels in milling fractions of some cereal grains and soybean

Tocol levels in the milling fractions of rice, barley, corn, wheat, and soybeans were analyzed by HPLC with a fluorescence detector. Among all milling fractions tested in this study, rice germ had the highest total tocol levels. In the four milling fractions of barley, except pearling flour, all eight tocol isomers were detected, and they were more uniformly distributed than in any other cereal grains measured in this study. The total tocol and α-tocopherol levels of wheat germ were significantly (P<0.05) higher than the other wheat milling fractions. A significantly (P<0.05) higher proportion of γ-tocopherol was obtained from corn germ (71.5%) and endosperm (50.3%) than from corn hulls. Only four tocol isomers (α-, β-, γ-, and δ-tocopherol) were detected in soybean milling fractions; no tocotrienol isomers were detected. The δ-tocopherol level of soybean endosperm, although minor, was significantly higher than those in milling fractions of other cereal grains in this study.     

Lipids of seven cereal grains

Grain samples of representative varieties of barley, corn, oats, rye, sorghum, triticale, and wheat grown commercially in the north central US were analyzed. Chemical constituents of the varieties studied are presented to provide an overview of their characteristics. Lipids of the milled grain samples were solvent extracted, classified by silicic acid column chromatography, and separated by thin layer chromatography. Fatty acid composition of the total lipid was determined by gas liquid chromatography and the fatty acid content was determined by saponification and extraction. Total lipid content of the grains ranged from 2.3% for ‘Polk’ wheat to 6.6% for ‘Chief’ oats. Lipid composition varied considerably. The row crops, corn and sorghum, have a high neutral lipid and low glycolipid content. The small grain varieties have a more balanced distribution among neutral lipids, glycolipids, and phospholipids. Fatty acid composition of the total lipid was similar for all grains. Minor qualitative differences were noted among the lipid classes of the 7 cereals.     

Lipids in wheat from various classes and varieties

Lipids were extracted with petroleum ether and with water-saturated n-butanol from 8 hard red winter, 5 hard red spring, and one each from soft red, durum, and club wheat varieties from 2 harvests. The butanol-extracted lipids were fractionated into nonpolar and polar lipids by silicic acid column chromatography, and the two major fractions were subfractionated by thin-layer chromatography. Durum wheats contained the highest lipid contents, and the highest concentration of nonpolar lipids. The breadmaking wheat varieties had a lipid content which was consistent for the 2 years examined. The total and nonpolar lipid contents of hard red spring wheats were higher than of hard red winter wheats. The polar lipid contents of wheats from the two classes were essentially equal. Total lipid contents were substantially higher in wheats than in flours milled from the wheats. Nonpolar lipids constituted about one-half of the flour lipids and two-thirds of the wheat lipids. Concentrations of triglycerides were higher in wheat than in flour nonpolar lipids. Glycolipids were present in comparable concentrations in wheat and in flour polar lipids; concentration in polar lipids of phosphatidyl choline was higher and of other phospholipids was lower in wheat than in flour polar lipids. No. 547, Kansas Agrieultural Experiment Station, Manhattan. Done in part under contract with the U.S. Department of Agriculture and authorized by the Research and Marketing Act of 1946. Supervised by the Western Utilization Research and Development Div., ARS.     

Nicotinamide Effectively Suppresses Fusarium Head Blight in Wheat Plants

Pyridine nucleotides such as a nicotinamide adenine dinucleotide (NAD) are known as plant defense activators. We previously reported that nicotinamide mononucleotide (NMN) enhanced disease resistance against fungal pathogen Fusarium graminearum in barley and Arabidopsis. In this study, we reveal that the pretreatment of nicotinamide (NIM), which does not contain nucleotides, effectively suppresses disease development of Fusarium Head Blight (FHB) in wheat plants. Correspondingly, deoxynivalenol (DON) mycotoxin accumulation was also significantly decreased by NIM pretreatment. A metabolome analysis showed that several antioxidant and antifungal compounds such as trigonelline were significantly accumulated in the NIM-pretreated spikes after inoculation of F. graminearum. In addition, some metabolites involved in the DNA hypomethylation were accumulated in the NIM-pretreated spikes. On the other hand, fungal metabolites DON and ergosterol peroxide were significantly reduced by the NIM pretreatment. Since NIM is relative stable and inexpensive compared with NMN and NAD, it may be more useful for the control of symptoms of FHB and DON accumulation in wheat and other crops.     

New property of vitamin A and beta-carotene on human iron absorption: effect on phytate and polyphenols as inhibitors of iron absorption

One hundred and seventy four human subjects were studied to find out the interaction of vitamin A or beta-carotene with the inhibitors of iron absorption, from a basal breakfast containing bread from either 100 g of precooked corn flour or 100 g of white wheat flour, 50 g of cheese and 10 g of margarine. Bread was labeled with either 55Fe or 59Fe. This bread was made from commercially flours fortified with iron as ferrous fumarate and vitamins. It was noticed that the percentage of iron absorption from the breakfast prepared with precooked corn flour given alone and with different concentrations of coffee was practically the same, while the iron absorption from the breakfast prepared from wheat flour decreased from 6% when the breakfast was given alone, to less than 2% when it was given with different concentrations of coffee. The only ingredient present in precooked corn flour and not in wheat flour was vitamin A. This difference encouraged the authors to perform further experiments using precooked corn and wheat flours fortified only with ferrous fumarate. These studies demonstrated that vitamin A inhibits the effect of the polyphenol and partially inhibits the effect phytate on iron absorption. HPLC and spectrophotometric studies demonstrated an interaction between vitamin A and iron. Other experiments, which included 100 volunteers, were performed to test the effect of vitamin A and beta-carotene on iron absorption from corn, wheat and rice. The presence of vitamin A increased iron absorption up to 3 times for rice, 2.4 times for wheat and 1.8 times for corn. beta-carotene increased absorption almost 3 times for the three cereals tested, showing that both compounds were capable of preventing the inhibitory effect of phytates on iron absorption. This information suggest that vitamin A and beta-carotene form a complex with iron keeping it soluble in the intestinal lumen and preventing the inhibitory effect of phytates and polyphenols on iron absorption.     

Phytosterols in cereal by-products

Phytosterols are hypocholesterolemic. Like corn fiber oil, the lipid extracts of certain cereal by-products may be rich sources of these health-promoting compounds. The objective of this research was to examine the phytosterol content and composition of various cereal by-products. Total lipids in rice bran, wheat bran, wheat germ, durum wheat (bran and germ mixture), oat bran, oat hull, and corn fine fiber were extracted, and the sterol profiles of the extracted lipids were analyzed by GC. Rice bran contained the most lipids (22.2%), followed by wheat germ, durum wheat, oat bran, wheat bran, and oat hull; corn fine fiber contained the least amount of lipids (1.7%). Sitosterol, campesterol, and stigmasterol were the major phytosterols in these lipid extracts, whereas brassicasterol was detected only in wheat samples. Rice bran oil contained considerable amounts of cycloartenol and 24-methylenecycloartanol, which were unique to these samples. Total sterol concentrations in extracted lipids were similar for rice bran, wheat bran, wheat germ, and durum wheat (21.3–15.1 mg/g), but they were very low in oat bran lipids and oat hull lipids (3.4 and 8.2 mg/g, respectively). Corn fine fiber lipids contained the highest amount of sterols (48.3 mg/g). Rice bran appears to be the best source of phytosterols, with the highest oil content and high concentration of sterols.     

Pea–barley intercropping and short-term subsequent crop effects across European organic cropping conditions

Grain legumes are known to increase the soil mineral nitrogen (N) content, reduce the infection pressure of soil borne pathogens, and hence enhance subsequent cereals yields. Replicated field experiments were performed throughout W. Europe (Denmark, United Kingdom, France, Germany and Italy) to asses the effect of intercropping pea and barley on the N supply to subsequent wheat in organic cropping systems. Pea and barley were grown either as sole crops at the recommended plant density (P100 and B100, respectively) or in replacement (P50B50) or additive (P100B50) intercropping designs. In the replacement design the total relative plant density is kept constant, while the additive design uses the optimal sole crop density for pea supplementing with ‘extra’ barley plants. The pea and barley crops were followed by winter wheat with and without N application. Additional experiments in Denmark and the United Kingdom included subsequent spring wheat with grass-clover as catch crops. The experiment was repeated over the three cropping seasons of 2003, 2004 and 2005. Irrespective of sites and intercrop design pea–barley intercropping improved the plant resource utilization (water, light, nutrients) to grain N yield with 25–30% using the Land Equivalent ratio. In terms of absolute quantities, sole cropped pea accumulated more N in the grains as compared to the additive design followed by the replacement design and then sole cropped barley. The post harvest soil mineral N content was unaffected by the preceding crops. Under the following winter wheat, the lowest mineral N content was generally found in early spring. Variation in soil mineral N content under the winter wheat between sites and seasons indicated a greater influence of regional climatic conditions and long-term cropping history than annual preceding crop and residue quality. Just as with the soil mineral N, the subsequent crop response to preceding crop was negligible. Soil N balances showed general negative values in the 2-year period, indicating depletion of N independent of preceding crop and cropping strategy. It is recommended to develop more rotational approaches to determine subsequent crop effects in organic cropping systems, since preceding crop effects, especially when including legumes, can occur over several years of cropping.     

Metabolomics to Decipher the Chemical Defense of Cereals against Fusarium graminearum and Deoxynivalenol Accumulation

Fusarium graminearum is the causal agent of Fusarium head blight (FHB) and Gibberella ear rot (GER), two devastating diseases of wheat, barley, and maize. Furthermore, F. graminearum species can produce type B trichothecene mycotoxins that accumulate in grains. Use of FHB and GER resistant cultivars is one of the most promising strategies to reduce damage induced by F. graminearum. Combined with genetic approaches, metabolomic ones can provide powerful opportunities for plant breeding through the identification of resistant biomarker metabolites which have the advantage of integrating the genetic background and the influence of the environment. In the past decade, several metabolomics attempts have been made to decipher the chemical defense that cereals employ to counteract F. graminearum. By covering the major classes of metabolites that have been highlighted and addressing their potential role, this review demonstrates the complex and integrated network of events that cereals can orchestrate to resist to F. graminearum.     

Effects of Semicontinuous and Batch System Ozonation on Wheat and Corn Starches

Wheat starch-water and corn starch-water suspensions (10%; w/w) were exposed to ozone gas by using two different contact systems; semicontinuous and batch systems. In semicontinuous contact mode, fresh ozone was applied to starch suspensions at a concentration of 4.2 mg dissolved ozone L^?1 water for 30 min, while in closed batch contact mode, starch suspensions were stirred with ozone gas at a rate of 60 g/h for 30 min. Gelatinization temperatures and enthalpies of wheat and corn starch were increased in a semicontinuous system. However, no change was observed for the gelatinization characteristics of starch samples ozonated- in batch system. Structures and birefringences of wheat and corn starch ozonated in both semicontinuous and batch operations were not differentiated clearly. Higher lightness and whiteness values were observed for both wheat and corn starch ozonated in semicontinuous system than the controls and ozonated samples in batch system. Corn starch ozonated in semicontinuous system has lower chroma and yellowness values than the others. Total color difference for the samples ozonated in semicontinuous system is significantly different from the samples ozonated in batch system. Semicontinuous ozonation system is more effective on the thermal and color properties of wheat and corn starches than the batch ozonation system.     

Evaluation of the antimicrobial action of honey against Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, Escherichia coli, Salmonella enteritidis, Listeria monocytogenes and Aspergillus niger. Evaluation of its microbiological charge

The evaluation of the microbiological charge present in Costa Rican samples as the evaluation of its antimicrobial activity over different microorganisms, including those associated to wound infections, will allow to emit criteria referred to its use in therapeutic treatments, specially as alternative therapy for cases involving antibiotic resistant bacteria. The microbiological charge of 25 honey samples, acquired in Costa Rican markets was evaluated through several indicators including total plate aerobic count, total plate anaerobic count, total aerobic spore count, total anaerobic spore count and molds and yeast count. Also, samples were inoculated in tubes with chopped meat media and plated in egg yolk agar in order to determine the presence of Clostridium botulinum. For the antimicrobial activity evaluation, the diffusion method in Muller Hinton agar was performed, testing different honey concentrations (100, 75, 50, 25, 12,5 and 6,25 % v/v) against Staphylococcus aureus (ATCC 25923), Staphylococcus epidermidis (UCR 2902), Pseudomonas aeruginosa (ATCC 9027), Escherichia coli (ATCC25922), Salmonella enteritidis (ATCC 13076), Listeria monocytogenes (ATCC 19116) and Aspergillus niger. The results obtained for the microbiological characterization of honey show that 91% of samples had counts equal or lower than 1,0 x 10(1) CFU/g. No positive result was obtained for the isolation of C. botulinum. 24 of the samples analyzed inhibited the growth of S. aureus even in a 25% v/v concentration, nevertheless, A. niger was no inhibited by any of the samples tested.     

Plant Sterol and Policosanol Characterization of Hexane Extracts from Grain Sorghum, Corn and their DDGS

Plant sterols (PS) and policosanols (PC) have been attributed with plasma cholesterol-lowering properties in humans. Hexane extracts from grain sorghum, corn and their distillers dried grain with solubles (DDGS), an important co-product of ethanol production, contain these health promoting compounds, which could be used to develop health promoting dietary products. However, limited information is currently available regarding optimal methods of extraction and their influence on plant sterol or PC levels and compositions. Therefore, the objective of this study was to quantify lipids, particularly PS and PC extracted from grain sorghum and its DDGS using reflux and Soxtec methods. Corn and its DDGS were also extracted to compare lipid yields and PS/PC compositional profiles in these two related cereals. Intact grains were extracted either as whole or ground kernels. Lipid yields from all the corn sources were typically greater than those obtained from grain sorghum. Lipid yields from DDGS were the highest among all the forms of the grains used whereas corn/sorghum DDGS contained the highest levels of PS and PC, respectively. Additional study demonstrated that hydrolysis (acidic and alkaline) of ground sorghum or sorghum DDGS increased the levels of total extractable plant sterols. Overall, the Soxtec method extracted higher amounts of lipids than the reflux method.     

Soluble and insoluble dietary fiber in cereals and legumes cultivated in Chile

Insoluble, soluble and total dietary fiber (DF) were determined in 35 varieties of certified whole seeds (without processing) of cereals (rice, oat, rye, and wheat) and legumes (pea, cowpea, beans, chikpea, lentil and lupine). The enzymatic method of Asp, Johansson and Siljestrom was used, with modifications in relation to time of incubation with alpha amylase, filtration system and volumes of the filtrates. Results were expressed as g/100 g dry weight. Total DF for cereals showed a range from 10.1 (wheat var. Chasqui) to 22.2 (rice var Quella). Rye, var. Tetra Baer and oats var. Pony Baer presented the highest soluble fiber content (3.3 and 3.9, respectively). In legumes, total DF fluctuated between 12.7 (pea, var. yellow) and 36.6 (lupine, var. Multolupa). Bean, var. Pinto INIA and lupine var. Multolupa presented the highest soluble fiber values (5.8 for both). Based on the results of this research work, it might be concluded that great variation exists in regard to the amount of total soluble and insoluble DF in cereals and legumes, a fact which impedes generalization as to its content in each food item.     

Trichothecenes: Natural occurrence and potential hazard

The trichothecene toxins are a chemical group of fungal metabolites characterized by a tetracyclic 12, 13-epoxy-trichothec-9-ene skeleton. There are over 40 naturally occurring derivatives produced predominantly by species ofFusarium but also produced by species ofCephalosporium, Myrothecium, Trichoderma, andStachybotrys. The trichothecene derivatives most commonly encountered in feedstuff are: T-2 toxin, diacetoxyscirpenol (DAS), deoxynivalenol (vomitoxin) and nivalenol. Deoxynivalenol has been isolated from corn and mixed feed more frequently than the other derivatives reported, and moreover, it is frequently found together with the fungal estrogen zearalenone. Deoxynivalenol has frequently been associated with corn refused by swine, vomiting in dogs, and corn used by humans in South Africa. Deoxynivalenol is produced byFusarium roseum, an organism that is commonly found infecting corn in the field and corn stored on-the-cob in cribs in the Midwest. The trichothecenes have been implicated in diseases of man called alimentary toxic aleukia in the USSR and have been found in corn in the Transkei where the incidence of esophageal cancer is high. Among domestic animals, trichothecenes such as T-2 toxin, DAS and deoxynivalenol have been implicated in diseases characterized by a hemorrhagic syndrome in cows and swine, vomiting in swine and dogs, decrease in avian egg production and body weight, refusal of feed by swine and infertility in swine. The data on the residue of the administered T-2 toxin in domestic animals suggests that trichothecenes can be transmitted into humans.     

Mycoflora and Co-Occurrence of Fumonisins and Aflatoxins in Freshly Harvested Corn in Different Regions of Brazil

Natural mycoflora and co-occurrence of fumonisins (FB₁, FB₂) and aflatoxins (AFB₁, AFB₂, AFG₁ and AFG₂) in freshly harvested corn grain samples from four regions of Brazil were investigated. Fusarium verticillioides was predominant in all samples. Analysis of fumonisins showed that 98% of the samples were contaminated with FB₁ and 74.5% with FB₁ + FB₂, with toxin levels ranging from 0.015 to 9.67 μg/g for FB₁ and from 0.015 to 3.16 μg/g for FB₂. Twenty-one (10.5%) samples were contaminated with AFB₁, seven (3.5%) with AFB₂ and only one (0.5%) with AFG₁ and AFG₂ Co-contamination with aflatoxins and fumonisins was observed in 7% of the samples. The highest contamination of fumonisins and aflatoxins was observed in Nova Odessa (SP) and Várzea Grande (MT), respectively. The lowest contamination of these mycotoxins was found in Várzea Grande and Nova Odessa, respectively.     

Fumonisins,Trichothecenes and Zearalenone in Cereals

Fumonisins are phytotoxic mycotoxins which are synthesized by various species of the fungal genus Fusarium such as Fusarium verticillioides (Sacc.) Nirenberg (ex F.moniliforme Sheldon) and Fusarium proliferatum. The trichothecene (TC) mycotoxins are secondary metabolites produce by species that belong to several fungal genera, especially Fusarium, Stachybotrys, Trichothecium, Trichoderma, Memnoniella and Myrothecium. Fusarium mycotoxins are widely dispersed in cereals and their products. Zearalenone (ZEA) is an estrogenic compound produced by Fusarium spp. such as F. graminearum and F. culmorum. Fumonisins, the TCs and ZEA are hazardous for human and animal health. Contamination with TCs causes a number of illnesses in human and animal such as decrease in food consumption (anorexia), depression or inhibition on immune system function and haematoxicity. The purpose of this paper is to give a review of the papers published on the field of fumonisin, TC and ZEA mycotoxins in cereals consumed in the world.     

Phytotoxicity of water-soluble substances from alfalfa and barley soil extracts on four crop species

Problems associated with continuously planting alfalfa (Medicago saliva L.) or seeding to thicken depleted alfalfa stands may be due to autotoxicity, an intraspecific form of allelopathy. A bioassay approach was utilized to characterize the specificity and chemical nature of phytotoxins in extracts of alfalfa soils as compared to fallow soil or soil where a cereal was the previous crop. In germination chamber experiments, water-soluble substances present in methanol extracts of soil cropped to alfalfa or barley (Hordeum vulgare L.) decreased seedling root length of alfalfa L-720, winter wheat (Triticum aestivum L. Nugaines) and radish (Raphanus sativa L. Crimson Giant). Five days after germination, seedling dry weights of alfalfa and radish in alfalfa soil extracts were lower compared to wheat or red clover (Trifolium pralense L. Kenland). Growth of red clover was not significantly reduced by soil extracts from cropped soil. Extracts of crop residue screened from soil cropped to alfalfa or barley significantly reduced seedling root length; extracts of alfalfa residue caused a greater inhibition of seedling dry weight than extracts of barely residue. A phytotoxic, unidentified substance present in extracts of crop residue screened from alfalfa soil, which inhibited seedling root length of alfalfa, was isolated by thin-layer chromatography (TLC). Residues from a soil cropped continuously to alfalfa for 10 years had the greatest phytotoxic activity.     

New PCR Assays for the Identification of Fusarium verticillioides, Fusarium subglutinans, and Other Species of the Gibberella fujikuroi Complex

Fusarium verticillioides and Fusarium subglutinans are important fungal pathogens of maize and other cereals worldwide. In this study, we developed PCR-based protocols for the identification of these pathogens targeting the gaoB gene, which codes for galactose oxidase. The designed primers recognized isolates of F. verticillioides and F. subglutinans that were obtained from maize seeds from several producing regions of Brazil but did not recognize other Fusarium spp. or other fungal genera that were either obtained from fungal collections or isolated from maize seeds. A multiplex PCR protocol was established to simultaneously detect the genomic DNA from F. verticillioides and F. subglutinans. This protocol could detect the DNA from these fungi growing in artificially or naturally infected maize seeds. Another multiplex reaction with a pair of primers developed in this work combined with a pre-existing pair of primers has allowed identifying F. subglutinans, F. konzum, and F. thapsinum. In addition, the identification of F. nygamai was also possible using a combination of two PCR reactions described in this work, and another described in the literature.     

A national survey of mycotoxins in canada

Mycotoxin contamination of animal feedstuffs continues to be an area of concern in various regions of Canada. A survey of research institutions across Canada revealed that the mycotoxins, zearalenone, ochratoxin A, tricothecenes and aflatoxin, have been detected in suspected feed samples. These mycotoxins have produced mycotoxicoses in poultry, hogs and cattle. Estrogenic syndromes in hogs may be caused by the ingestion of zearalenone produced by Fusarium molds on grain com grown in southern Ontario. Overwintered barley contaminated with T-2 toxin was suspected in an outbreak of fusariotoxicoses in swine in northern Alberta. Central and eastern Canada, which seem to be the major focus of mycotoxin occurrence, will be the object of further field level examination to discover patterns of occurrence and economic impact.