scholarly journals Characterization of a Hydrophobic Amylase Inhibitor from Corn (Zea mays) Seeds with Activity Against Amylase from Fusarium verticillioides

2003 ◽  
Vol 93 (8) ◽  
pp. 917-922 ◽  
Author(s):  
Edson L. Z. Figueira ◽  
Elisa Y. Hirooka ◽  
Elizabeth Mendiola-Olaya ◽  
Alejandro Blanco-Labra
Keyword(s):  
Enzyme Inhibitor ◽  
Fusarium Verticillioides ◽  
Human Saliva ◽  
Exchange Chromatography ◽  
Fungal Resistance ◽  
Prostephanus Truncatus ◽  
Porcine Pancreas ◽  
Amylase Inhibitor ◽  
Spectrum Activity ◽  
Corn Kernels

A hydrophobic 19.7-kDa amylase inhibitor (AI) was purified from corn kernels by 95% ethanol extraction and anionic exchange chromatography. The AI has an isoelectric point of 3.6 and was very stable at different pH values and high temperatures, maintaining 47.6% activity after heating to 94°C for 60 min. Amino acid analysis indicated high valine, leucine, glycine, alanine, and glutamic acid/glutamine content, and especially high valine content (41.2 mol%). This inhibitor is not a glycoprotein. It required 30-min preincubation to maximize complex enzyme-inhibitor formation when the amylase from Fusarium verticillioides was tested. The optimal pH of interaction was 6.5. It showed broad-spectrum activity including the following amylases: human saliva, porcine pancreas, F. verticillioides, as well as those from some insects of agricultural importance (Acanthoscelides obtectus, Zabrotes subfasciatus, Sitophilus zeamais, and Prostephanus truncatus). This novel hydrophobic protein not only inhibited the amylase from F. verticillioides but also decreased the conidia germination. Thus, this protein represents an approach to decrease the production of fumonisin in corn, either by using it as a molecular marker to detect fungal resistance or through genetic engineering.

scholarly journals New Amylase Inhibitor Present in Corn Seeds Active In Vitro Against Amylase from Fusarium verticillioides

Plant Disease ◽  
2003 ◽  
Vol 87 (3) ◽  
pp. 233-240 ◽  
Author(s):  
Edson L. Z. Figueira ◽  
Alejandro Blanco-Labra ◽  
Antônio Carlos Gerage ◽  
Elisabete Y. S. Ono ◽  
Elizabeth Mendiola-Olaya ◽  
...  
Keyword(s):  
High Performance ◽  
Fusarium Verticillioides ◽  
Gel Filtration ◽  
Anion Exchange Chromatography ◽  
Human Saliva ◽  
Exchange Chromatography ◽  
Experimental Conditions ◽  
Amylase Inhibitor ◽  
Fusarium Sp

A screening for specific amylase inhibitor levels against amylase from Fusarium verticillioides (Fusarium moniliforme), the most relevant mycotoxigenic fungus in corn, was conducted on 37 corn hybrids. The amylase inhibitor levels in these hybrids ranged from 5.5 to 16.0 amylase inhibitor units per gram of corn (AIU/g) in the MASTER and AG5011 hybrids, respectively. The hybrid with the maximum content of inhibitor was used as the source of this new protein. The inhibitor was partially purified using fractional precipitation, gel filtration on Sephadex G-75 column, high performance liquid chromatography (HPLC) Superose HR 10/30 column, and HPLC anion exchange chromatography, obtaining a 20.7-fold purification. Electrophoresis after denaturing and heating under reductive conditions showed an apparent 23.8 kDa molecular mass and an acidic isoelectric point of 5.4, which differs from previous molecular masses reported for other inhibitors present in corn seeds (14 and 22 kDa). This inhibitor showed activity against amylases from human saliva and pancreas, from the fungi F. verticillioides and Aspergillus flavus, and from the insects Acanthoscelides obtectus, Zabrotes subfasciatus, Tribolium castaneum, and Sitotroga cerealella. The mycoflora found in the corn grain indicated Fusarium sp. as the most prevalent fungi (81.1% of the samples), with a count ranging from 1.5 × 102 to 2.4 × 106 CFU/g of corn. The presence of fumonisins was detected in 21 out of the 37 hybrids studied, ranging from 0.05 to 2.67 μg of FB per gram of corn. No correlation could be established between this amylase inhibitor level in the corn seeds and the presence of Fusarium sp. or with the fumonisin content under the experimental conditions of the test.

scholarly journals Potential Application of Lactic Acid Bacteria to Reduce Aflatoxin B1 and Fumonisin B1 Occurrence on Corn Kernels and Corn Ears

Toxins ◽  
2019 ◽  
Vol 12 (1) ◽  
pp. 21 ◽  
Author(s):  
Tiago de Melo Nazareth ◽  
Carlos Luz ◽  
Raquel Torrijos ◽  
Juan Manuel Quiles ◽  
Fernando Bittencourt Luciano ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Fusarium Verticillioides ◽  
Fumonisin B1 ◽  
Health Concern ◽  
Economic Losses ◽  
Antifungal Effect ◽  
Minimum Fungicidal Concentration ◽  
Sensory Defects ◽  
Corn Kernels

Fungal spoilage is an important issue for the food industry, leading to food sensory defects, food waste, economic losses and public health concern through the production of mycotoxins. Concomitantly, the search for safer natural products has gained importance since consumers began to look for less processed and chemically treated foods. In this context, the aim of this study was to evaluate the antifungal and antimycotoxigenic effect of seven strains of Lactobacillus plantarum. Lactic acid bacteria (LAB) were grown on Man Rogosa Sharpe (MRS) broth at 37 °C in anaerobic conditions. After that, the cell-free supernatant (CFS) were recovered to determine its antifungal activity by halo diffusion agar test. In addition, minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) was determined for each L. plantarum CFS by 96-well microplates method. Additionally, CFS was used as a natural biocontrol agent on corn kernels and corn ears contaminated with Aspergillus flavus and Fusarium verticillioides, respectively. The L. plantarum CECT 749 CFS showed the highest antifungal effect against all essayed strains. Moreover, the employment of this CFS in food reduced the mycotoxin production at a percentage ranging from 73.7 to 99.7%. These results suggest that the L. plantarum CECT 749 CFS could be promising for the biocontrol of corn.

scholarly journals Distinct populations of high-Mr mucins secreted by different human salivary glands discriminated by density-gradient electrophoresis

1995 ◽  
Vol 309 (3) ◽  
pp. 801-806 ◽  
Author(s):  
J Bolscher ◽  
E Veerman ◽  
A Van Nieuw Amerongen ◽  
A Tulp ◽  
D Verwoerd
Keyword(s):  
Monoclonal Antibodies ◽  
Sialic Acid ◽  
Salivary Glands ◽  
Density Gradient ◽  
Human Saliva ◽  
Exchange Chromatography ◽  
Dot Blot ◽  
Cell Organelles ◽  
Sulphate Content ◽  
Gradient Electrophoresis

High-M(r) mucins [mucin glycoprotein 1 (MG1)] isolated from human saliva from the individual salivary glands were chemically characterized. The carbohydrate content of MG1 derived from palatal (PAL), submandibular (SM) and sublingual (SL) saliva was typical of mucins but showed heterogeneity, especially in the amount of sialic acid and sulphated sugar residues. The physicochemical properties of native MG1s make conventional SDS/PAGE and ion-exchange chromatography unsuitable for investigating differences between individual samples. Recently a density-gradient electrophoresis (DGE) device has been developed, primarily for separation based on the charge of entire cells or cell organelles [Tulp, Verwoerd and Pieters (1993) Electrophoresis 14, 1295-1301]. We have used this apparatus to study the high-M(r) salivary mucins. Using DGE, the MG1s of individual glands were seen to have clearly distinct electrophoretic mobilities, as monitored by ELISA using MG1-specific monoclonal antibodies. Even within a particular MG1 preparation, subpopulations could be distinguished. DGE analysis of a chemically and enzymically modified MG1 series, followed by ELISA and dot-blot detection using specific monoclonal antibodies, lectins and high-iron diamine staining, suggests that the high electrophoretic mobility of PAL-MG1 is mainly the result of a high sulphate content, whereas the SL subpopulations differ mainly in binding type and amount of sialic acid. SM-MG1 most resembles the low-mobility subpopulation of SL-MG1, except that it has a lower sulphate content. In conclusion, DGE appears to be a powerful method for analysis of native mucin; it has been used to demonstrate that MG1s from the various salivary glands are biochemically much more diverse than was previously assumed.

scholarly journals The Relationship Analysis on Corn Stalk Rot and Ear Rot According to Fusarium Species and Fumonisin Contamination in Kernels

Toxins ◽  
2019 ◽  
Vol 11 (6) ◽  
pp. 320 ◽  
Author(s):  
Lina Li ◽  
Qing Qu ◽  
Zhiyan Cao ◽  
Zhengyu Guo ◽  
Hui Jia ◽  
...  
Keyword(s):  
Fusarium Species ◽  
Fusarium Verticillioides ◽  
Contamination Rate ◽  
Corn Stalk ◽  
Ear Rot ◽  
Stalk Rot ◽  
Relationship Analysis ◽  
Corn Root Rot ◽  
Corn Kernels ◽  
The Relationship

Fusarium diseases, including corn root rot, sheath rot, stalk rot, and ear rot are frequently occurring in maize producing areas of China. Fusarium stalk rot and ear rot are the most serious diseases and often occur at the same time, but it is unclear whether there is a correlation between Fusarium composition and disease occurrence. This study was conducted to clarify the relationship between the two diseases. A total of 49 corn stalk rot samples were collected from 15 regions of eight provinces in China from 2016 to 2018. The pathogens were isolated and identified separately from stalks, ear stems, and kernels. The contents of the fumonisins (FB1 and FB2) were detected in kernels. The results showed that the main Fusarium species were found in corn kernels, ear stems and stalks at the same time. The results showed that 1201 strains of Fusarium verticillioides, 668 strains of Fusarium oxysporum, 574 strains of Fusarium graminearum species complex (FGSC), 318 strains of Fusarium equiseti, 95 strains of Fusarium proliferatum, and 40 strains of Fusarium subglutinans were isolated from 1470 corn kernels, 245 ear stems, and 1225 stalks randomly selected from 49 samples. The contamination rate of fumonisins in the 49 samples was 57.1% with an average content of 1.9 μg/g, of which four samples exhibited higher levels as set by the European Commission (4.0 μg/g). These results provide a certain association between stalk rot and ear rot and lay a foundation to study the relationships among Fusarium maize diseases.

scholarly journals Purification and reaction mechanism of the primary inhibitor of plasmin from human plasma

1978 ◽  
Vol 175 (2) ◽  
pp. 635-641 ◽  
Author(s):  
Ulla Christensen ◽  
Inge Clemmensen
Keyword(s):  
Dissociation Constant ◽  
Human Plasma ◽  
Active Site ◽  
Competitive Inhibition ◽  
Enzyme Inhibitor ◽  
Polyacrylamide Gel Electrophoresis ◽  
Inhibitory Effect ◽  
Exchange Chromatography ◽  
Comparison Of The Results ◽  
Sepharose 4B

The primary inhibitor of plasmin in human plasma was purified by a four-step procedure involving fractional (NH4)2SO4 precipitation, ion-exchange chromatography on a column of DEAE-Sepharose CL-6B and affinity chromatography on both a plasminogen–CH-Sepharose 4B column and a column of 6-aminohexanoic acid covalently coupled through the carboxylate function to AH-Sepharose 4B. No impurities in the final preparation could be detected when tested by immunoelectrophoresis against a range of specific antisera or against rabbit anti-human serum. On polyacrylamide-gel electrophoresis the inhibitor preparation showed a single band. The dissociation constant for the inhibitor–plasminogen complex was determined to be approx. 3μm at pH7.8. The reactions of the inhibitor with human plasmin and with bovine trypsin were studied. Comparison of the results obtained confirms the hypothesis previously presented, namely that the reaction of the inhibitor with plasmin involves at least two steps, the initial rapid formation of an enzyme–inhibitor complex followed by a slow irreversible transition to another complex. The results also indicate that the reaction of the inhibitor with trypsin involves just a single, irreversible step, so that this reaction seems to be less complicated than that of the inhibitor with plasmin. The ways in which 6-aminohexanoic acid influences the reactions were studied. The same value for the dissociation constant (approx. 26μm) for 6-aminohexanoic acid is obtained for both its effect on the reaction of the inhibitor with trypsin and for competitive inhibition of trypsin. The inhibitory effect of 6-aminohexanoic acid thus seems to be due to its blocking of the active site of trypsin. In contrast with this, the inhibitory effects of l-lysine and 6-aminohexanoic acid on the inhibitor–plasmin reaction occur at concentrations much too low to affect the active site of plasmin. The possible dependence of the reaction of the inhibitor with plasmin on a second site(s) on plasmin is discussed.

Growth inhibition of a Fusarium verticillioides GUS strain in corn kernels of aflatoxin-resistant genotypes

2001 ◽  
Vol 57 (5-6) ◽  
pp. 708-711 ◽  
Author(s):  
R. Brown ◽  
T. Cleveland ◽  
C. Woloshuk ◽  
G. Payne ◽  
D. Bhatnagar
Keyword(s):  
Growth Inhibition ◽  
Fusarium Verticillioides ◽  
Corn Kernels ◽  
Resistant Genotypes

scholarly journals Screening and Purification of a Chymotrypsin Inhibitor from Entrolobium Saman Seeds

2010 ◽  
Vol 15 ◽  
pp. 19
Author(s):  
Maher Ali. Maqtari ◽  
A.B. Mohamed. Saad
Keyword(s):  
Molecular Weight ◽  
Enzyme Inhibitor ◽  
Gel Filtration ◽  
Exchange Chromatography ◽  
Molar Ratio ◽  
Apparent Dissociation Constant ◽  
Chymotrypsin Inhibitor ◽  
Ammonium Sulphate Precipitation ◽  
Wide Ph Range ◽  
Ph Range

A chymotrypsin inhibitor was isolated and purified from the seeds of Enterolobium saman  (Leguminaceae family) by extraction with 100 mM phosphate buffer, heat treatment, ammonium sulphate precipitation, ion-exchange chromatography on DEAEcellulose and filtration through Sephadex G-75. The final preparation appeared to be homogeneous by both chromatographic and electrophoretic analyses. ESCI had a molecular weight of about 17,890 and an isoelectric point of 5.8. ESCI inhibited bovine chymotrypsin at an inhibitor-enzyme molar ratio of 1:2. The inhibition mode of chymotrypsin inhibitor was competitive on bovine chymotrypsin. Investigation has been carried out on the complex formed between chymotrypsin and chymotrypsin inhibitor by physico-chemical methods. An apparent dissociation constant (Ki) of 9.05 X 10-8 M has been calculated for the complex. This enzyme-  inhibitor complex was isolated by gel filtration on Sephadex G-75 and a molecular weight of 43.000 was estimated for the complex. The inhibitor did not have any effect on other proteinases, such as papain, bromelin, elastase, α -amylase, trypsin and pepsin. The chemical modification of lysine residues indicated that –NH2  groups are not essential for the activity of ESCI toward chymotrypsin. The inhibitor was an acidic protein and was stable over a wide pH range of 2-12 and temperature range of 10o C-97o C. 

scholarly journals Molecular diversity of the Fusarium fujikuroi species complex from maize

2020 ◽  
Vol 158 (4) ◽  
pp. 859-877
Author(s):  
Emilia Jabłońska ◽  
Krzysztof Piątek ◽  
Marcin Wit ◽  
Ewa Mirzwa-Mróz ◽  
Wojciech Wakuliński
Keyword(s):  
Genetic Diversity ◽  
Random Amplified Polymorphic Dna ◽  
Fusarium Verticillioides ◽  
Elongation Factor ◽  
Sensu Stricto ◽  
Fusarium Fujikuroi ◽  
Ear Rot ◽  
Polymorphic Loci ◽  
Polymorphic Bands ◽  
Corn Kernels

Abstract Random amplified polymorphic DNA (RAPD) markers were used to estimate genetic relationship and diversity of Fusarium species associated with pink ear rot of maize. Three Fusarium species – F. verticillioides, F. temperatum and F. subglutinans sensu stricto were collected from contaminated corn kernels. A collection of 41 randomly selected Fusarium isolates (16 F. verticillioides, 16 F. temperatum and nine F. subglutinans sensu stricto) was screened for polymorphism with thirty decamer primers. Seventeen primers that had generated a high number of polymorphic loci were used for the analysis of genetic diversity. The primers yielded a total of 146 polymorphic loci, an average of 8.59 polymorphic bands per primer. The percentage of polymorphic bands (%P) was 46.1%. The RAPD-based UPGMA analysis demonstrated that although F. temperatum and F. subglutinans s.s. form two distinct groups they are more similar to each other than to F. verticillioides. Fusarium verticillioides was the most heterogenous species (Nei’s genetic diversity 0.180), while the least heterogenous one was F. subglutinans s.s. whose genetic diversity was 0.098. The analysis of the translation elongation factor 1-α and β-tubulin gene sequences as well as the mating type ratio were employed to support the RAPD data.

scholarly journals Natural Occurrence of Mycotoxin-Producing Fusaria in Market-Bought Peruvian Cereals: A Food Safety Threat for Andean Populations

Toxins ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 172
Author(s):  
Christine Ducos ◽  
Laetitia Pinson-Gadais ◽  
Sylvain Chereau ◽  
Florence Richard-Forget ◽  
Pedro Vásquez-Ocmín ◽  
...  
Keyword(s):  
Food Safety ◽  
Fusarium Verticillioides ◽  
Daily Basis ◽  
Natural Occurrence ◽  
Safety Standards ◽  
Traditional Markets ◽  
Food Grains ◽  
Toxic Risk ◽  
Corn Kernels ◽  
Fungal Contaminants

Consumption of cereals contaminated by mycotoxins poses health risks. For instance, Fumonisins B, mainly produced by Fusarium verticillioides and Fusariumproliferatum, and the type B trichothecene deoxynivalenol, typically produced by Fusarium graminearum, are highly prevalent on cereal grains that are staples of many cultural diets and known to represent a toxic risk hazard. In Peru, corn and other cereals are frequently consumed on a daily basis under various forms, the majority of food grains being sold through traditional markets for direct consumption. Here, we surveyed mycotoxin contents of market-bought grain samples in order to assess the threat these mycotoxins might represent to Peruvian population, with a focus on corn. We found that nearly one sample of Peruvian corn out of six was contaminated with very high levels of Fumonisins, levels mostly ascribed to the presence of F. verticillioides. Extensive profiling of Peruvian corn kernels for fungal contaminants could provide elements to refine the potential risk associated with Fusarium toxins and help define adapted food safety standards.

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