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SDS-PAGE patterns of the proteins from the grain legume samples. M: Molecular weight marker; lane 1: Partner (Pisum sativum); lane 2: NS-Junior (Pisum sativum); lane 3: Dukat (Pisum sativum); lane 4: NS-Pionir (Pisum sativum); lane 5: Javor (Pisum sativum); lane 6: Jezero (Pisum sativum); lane 7: Kristal (Pisum sativum), lane 8: NS-Sirmijum (Vicia sativa); lane 9: Novi Beograd (Vicia sativa); lane 10: Panonka (Vicia pannonica Crantz); lane 11: NS-Vilosa (Vicia villosa Roth); lane 12: Panorama (Lupinus albus); lane 13: Vesna (Lupinus albus); lane 14: S ˇ arac (Vicia faba); lane 15: Gema (Vicia faba).
Source publication
This study was conducted to analyze the seed storage proteins of grain legumes cultivars using Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Lab-on-a-Chip (LoaC) electrophoresis. Using reference proteins, the relative accuracy and precision in estimating molecular weight was similar when these two methods were compared. H...
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... simulated gel patterns for a wide range of grain legume obtained by LoaC are shown in Fig. 2 and the elution profiles are shown in Fig. 3(a-f). Almost all the protein bands of the grain legume samples from the LoaC gel image are in the same range of MWs as the SDS-PAGE gel (Fig. 1). In contrast to SDS-PAGE, the LoaC uses internal lower and upper markers (lower marker at 4.5 kDa and upper marker at 240 kDa) within each lane to ...
Citations
... 41 These proteins were further fractionated into distinct subunits: β-legumin was observed at approximately 22 kDa, while α-legumin was observed at around 40 kDa. 42 Hence, the bands obtained at ~38 and 46 kDa could be associated with α-legumin for the fenugreek seed protein isolate. Moreover, the visible bands at ~22 and 23 kDa could be associated with the presence of β-legumin. ...
The study aims to optimize the extraction process and characterize the proteins found in fenugreek seeds. The water and oil holding capacities, coagulated protein content, foaming, and emulsification properties of the isolated proteins were investigated under all extraction conditions. Also, solubility, molecular weights, structural and thermal properties were determined. In the extraction processes carried out at different pH (pH 6.0–12.0) and solid:solvent ratios (20–60 g/L), it was determined that the highest extraction yield (94.3 ± 0.3%) was achieved when the pH was 11.47 and the solid-solvent ratio was 34.50 g/L. Three distinct bands (46, 59, and 80 kDa) in the range of 22–175 kDa were determined for the fenugreek seed protein isolate obtained under optimum extraction conditions. Protein secondary structures were determined using Fourier Transform Infrared (FT-IR) spectra and it was determined that β-sheet structures were highly present. In addition, denaturation temperature and denaturation enthalpy were calculated as ~119 °C and 28 mJ/g, respectively.
... The chickpea samples of the Pascià genotype were provided by the Department of Agriculture, Food, Natural Resources and Engineering (DAFNE) of the University of Foggia. The extraction of chickpea grain storage proteins was carried out according to a protocol adapted from the literature [29][30][31], and from De Santis et al. [32]. Briefly, the protein fraction was obtained by suspending 100 mg of chickpea flour with 1 mL of an extraction buffer (50 mM of Tris-HCl, pH 7.8, 5 mM of EDTA, 0.1% 1,4-dithiothreitol) for 1 h at room temperature with constant stirring, and then centrifuging at 10,000× g for 30 min. ...
Chickpea (Cicer arietinum L.) seed proteins show a lot of functional properties leading this legume to be an interesting component for the development of protein-enriched foods. However, both the in-depth proteomic investigation and structural characterization of chickpea seed proteins are still lacking. In this paper a detailed characterization of chickpea seed protein fraction by means of SDS-PAGE, in-gel protein digestion, high-resolution mass spectrometry, and database searching is reported. Through this approach, twenty SDS gel bands were cut and analyzed. While the majority of the bands and the identified peptides were related to vicilin and legumin storage proteins, metabolic functional proteins were also detected. Legumins, as expected, were revealed at 45–65 kDa, as whole subunits with the α- and β-chains linked together by a disulphide bond, but also at lower mass ranges (α- and β-chains migrating alone). Similarly, but not expected, the vicilins were also spread along the mass region between 65 and 23 kDa, with some of them being identified in several bands. An MS structural characterization allowed to determine that, although chickpea vicilins were always described as proteins lacking cysteine residues, they contain this amino acid residue. Moreover, similar to legumins, these storage proteins are firstly synthesized as pre-propolypeptides (Mr 50–80 kDa) that may undergo proteolytic steps that not only cut the signal peptides but also produce different subunits with lower molecular masses. Overall, about 360 different proteins specific of the Cicer arietinum L. species were identified and characterized, a result that, up to the current date, represents the most detailed description of the seed proteome of this legume.
... Glass's good isolation properties make it perfect for various frequencies [28]. "Lab-on-a-Chip" system with DEP gives better accuracy, shorter analysis time, high resolution, and precision in comparison to sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) of reference protein molecular weight estimation [45]. DEP enhances through the various geometry of the electrodes. ...
... It can be used in blood component fractionation and medical purposes like sorting single objects or aggregates of cells, viruses, organelles, and ribosomes. Lab-on-Chip technology and transient ITP coupled to free zone electrophoresis (t-ITP-MFZE) mode, which helps in the separation and detection of proteins [45,52]. ...
The dielectrophoretic (DEP) chip is used for the separation, focussing, manipulation, and sorting of cells. This review included solutions to cell separation problems like heat generation, single-cell analysis, cell sorting against fluid flow, etc. Cell trapping efficiency increases using optical pressure, DEP, and electroporation at low ionic strength, large multi-layered electrode structure, batch & continuous separation, gel-based sieving method, electrodeless dielectrophoresis (EDEP), DEP impedance measurement (DEPIM), electrodes in a trapezoidal manner. These DEP chips have many applications in biotechnology, biomedicine, cell analysis, etc.
... Glass's good isolation properties make it perfect for various frequencies [28]. "Lab-on-a-Chip" system with DEP gives better accuracy, shorter analysis time, high resolution, and precision in comparison to sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) of reference protein molecular weight estimation [45]. DEP enhances through the various geometry of the electrodes. ...
... It can be used in blood component fractionation and medical purposes like sorting single objects or aggregates of cells, viruses, organelles, and ribosomes. Lab-on-Chip technology and transient ITP coupled to free zone electrophoresis (t-ITP-MFZE) mode, which helps in the separation and detection of proteins [45,52]. ...
The dielectrophoretic (DEP) chip is used for the separation, focussing, manipulation, and sorting of cells. This review included solutions to cell separation problems like heat generation, single-cell analysis, cell sorting against fluid flow, etc. Cell trapping efficiency increases using optical pressure, DEP, and electroporation at low ionic strength, large multi-layered electrode structure, batch & continuous separation, gel-based sieving method, electrodeless dielectrophoresis (EDEP), DEP impedance measurement (DEPIM), electrodes in a trapezoidal manner. These DEP chips have many applications in biotechnology, biomedicine, cell analysis, etc.
... By composition, most legume seeds contain up to 40% of their weight in proteins [15,16], but are low in fat (2-5%). Tissue-specific storage proteins are the most common proteins of legume seeds and their overall quality is influenced by their nutritional and functional properties [5,17]. Compared to animal proteins, plant proteins are low in certain essential amino acids, especially the limiting amino acids. ...
The increased consumption of legume seeds as a strategy for enhancing food security, reducing malnutrition, and improving health outcomes on a global scale remains an ongoing subject of profound research interest. Legume seed proteins are rich in their dietary protein contents. However, coexisting with these proteins in the seed matrix are other components that inhibit protein digestibility. Thus, improving access to legume proteins often depends on the neutralisation of these inhibitors, which are collectively described as antinutrients or antinutritional factors. The determination of protein quality, which typically involves evaluating protein digestibility and essential amino acid content, is assessed using various methods, such as in vitro simulated gastrointestinal digestibility, protein digestibility-corrected amino acid score (IV-PDCAAS), and digestible indispensable amino acid score (DIAAS). Since most edible legumes are mainly available in their processed forms, an interrogation of these processing methods, which could be traditional (e.g., cooking, milling, extrusion, germination, and fermentation) or based on emerging technologies (e.g., high-pressure processing (HPP), ultrasound, irradiation, pulsed electric field (PEF), and microwave), is not only critical but also necessary given the capacity of processing methods to influence protein digestibility. Therefore, this timely and important review discusses how each of these processing methods affects legume seed digestibility, examines the potential for improvements, highlights the challenges posed by antinutritional factors, and suggests areas of focus for future research.
... Fermentation with L. helveticus (881315) and Flavourzyme ® causes significant proteolysis. The micro-fluidic Loa-C method provides a unique approach to the real-time separation of major milk proteins as well as giving information on size, concentration and purity of proteins in a single assay [30]. The simulated gel patterns for fermented and un-treated skim milk as control, obtained by Loa-C with the elution profiles are shown ( Figure 1). ...
... The bio-analyser (Agilent 2100; Agilent Technologies, Wald Bonn, Germany), was used to perform the micro-fluidic lab-on-a-chip (Loa-C) using a high sensitivity protein 250 kit and 2100 software. Samples, dye and the preparation of chip were carried out as previously described [30] with minor alterations. Briefly, a reconstituted dye solution (0.5 µL) was mixed with a 5 µL protein molecular weight ladder (5-240 kD) and 5 µL of sample in micro tubes and mixed before being incubated on ice for 30 min. ...
Bioactive peptides are generated during milk fermentation or enzymatic hydrolysis. Lactobacillus (L) helveticus is commonly used to produce some types of fermented milk products. Fermented milk derived bioactive peptides are known to be beneficial in human health. Anti-hypertensive peptides play a dual role in the regulation of hypertension through the production of the vasoconstrictor angiotensin II and its inactivation of the vasodilator bradykinin. MALDI MS/MS, nano-LC/MS/MS and RP-HPLC were used to isolate peptides showing angiotensin converting enzyme inhibition (ACE-I) from 12% fermented skim milk using a combination of L. helveticus and Flavourzyme®. The fermentation procedure facilitated the identification of 133 anti-hypertensive peptides and 75% short chain amino acids, and the three with the highest ACE-I activity reduced blood pressure in a rat model of hypertension. The freeze- dried extract was supplemented in rodent chow. In this study 14-week-old male spontaneously hypertensive rats were fed for 10 weeks with the identified peptides added to chow and compared to controls supplemented with skim milk powder. Blood pressure (BP) decreased significantly (p < 0.05) from 6 to 10 weeks of FS groups (120/65 mmHg) compared with the NFS control groups, where the BP increased significantly (220/150 mmHg) (p < 0.05). The F6 fraction provided bioactive peptides with stronger antihypertensive properties than other fractions. Skim milk fermented by L. helveticus and Flavourzyme® generates several bioactive peptides which have a blood pressure lowering effect in hypertensive disease.
... For the AfLB globulin isolate, the protein profile had four major polypeptides (molecular weights 35, 50, 60 and 69 kDa). This profile was greatly similar to globulins 7S and 11S in legume proteins [21][22][23]. We found a band at 69 KDa, which we named Convicilin in the reference to the major pea globulin. ...
African locust bean (AfLB) protein isolates could be an interesting alternative to the use of soy as an ingredient for the development of new protein-rich products. From AfLB seed flour, the protein extractability yields by aqueous extraction were determined as a function of pH and ionic strength. Then thermally induced gelation of various protein suspensions relating to protein concentration was studied. The most critical factors affecting extractability were the pH and the presence of fat. As a function of the extraction process, the extraction yield of protein from AfLB flour ranged from 30 to 65%. Two major fractions of proteins detected in AfLB seeds were albumins and globulins, comprising four families: legume-like protein, vicilin-like proteins, convicilin, and albumins. The globulin isolate had the lowest solubility at pH 3.5-4 and the highest at pH 8-10. The solubility of albumin isolate was lightly affected by pH and ionic strength. At pH 7, the minimum protein concentration for thermal gel formation ranged from 55 to 120 g/L as function purified state of proteins. The less purified extract with a simpler process made it possible to obtain a gel needing a lower protein concentration. This last way seems promising to the development of new foods based on African locust bean flour.
... The lowest NH 2 concentration of IC well described the role of the cell wall integrity in preventing the enzyme diffusion during gastric digestion, which eventually leads to low digestion of protein in the gastric phase. Moreover, pulse proteins are low in sulfur-containing amino acids and major substrates for stomach proteases [36]. The low protein digestibility is similar to a previous study, which showed that an aspartic protease only degraded 5% of the total protein for red kidney bean in gastric phase [24]. ...
... Fig. 5 represents the SDS-PAGE profiles of supernatants (a) and pellets (b) obtained from isolated protein, BC, and IC samples at the nondigestion (N), gastric digestion (G), and at the end of small intestine digestion (I) stages. At the non-digestion stage, the supernatant of the pea isolated protein, BC, and IC samples showed a large amount of protein molecular weight profile in the molecular range of 10 to 70 kDa [36]. Smaller polypeptides at 22 and ≤14 kDa indicated the degradation of vicilin by digestive enzymes [24,42]. ...
The cell wall microstructure has been recognized to modulate the digestibility and bioaccessibility of nutrients in whole pulse foods, while the role of cell wall integrity is unclarified in the hydrolysis of intracellular nutrients during human gastrointestinal transit. Intact pea cells were isolated to prepare a series of cell wall integrity subjected to cooking and followed by the in vitro hydrolysis of starch and protein properties using the INFOGEST 2.0 in vitro simulation. Thermal properties showed that cell samples either in raw or cooked form with different wall integrity exhibited similar and higher starch gelatinization temperatures compared to the isolated starch counterpart. It was found that intact pea cells showed the limited hydrolysis extent of the maltose (16.2%) and NH2 (6.7%) compared to the damaged cells. In addition, intact cells also withheld the cell wall integrity throughout gastrointestinal digestion with minor rupture, and presented the higher protein molecular weight (70 kDa) in the SDS-PAGE profiles. Results suggested that the in vitro starch and protein digestion properties are modulated by the cell wall integrity, which may lead to lower glycemic response and open up the possibilities of designing health food products.
... Soluble proteins were extracted from a protocol adapted from [17]. Briefly, 100 mg of grounded flour was suspended with 1 mL of extraction buffer (50 mM Tris-HCl, pH 7.8, 5 mM EDTA, 0.1% 1,4-dithiothreitol) for 1 h at room temperature with constant mixing and centrifuged at 10,000 g for 30 min. ...
... Despite a moderate genetic effect on protein concentration, a large variability in terms of protein composition was observed in the current study. In recent years, the evidence of an existing genetic diversity in terms of seed protein composition was exploited in cultivated pulses, including chickpeas [8,17,27,28]. Most of the information on pulse protein composition was obtained by electrophoretic separation and is relative to genetic diversity [8,29,30], while less data are available on the effect of environmental [31] or agronomic factors [32]. ...
Chickpea is a key crop in sustainable cropping systems and for its nutritional value. Studies on agronomic and genetic influences on chickpea protein composition are missing. In order to obtain a deep insight into the genetic response of chickpeas to management in relation to agronomic and quality traits, a two-year field trial was carried out with eight chickpea genotypes under an organic and conventional cropping system. Protein composition was assessed by SDS-PAGE in relation to the main fractions (vicilin, convicilin, legumin, lectin, 2s-albumin). Crop response was highly influenced by year and presumably also by management, with a −50% decrease in grain yield under organic farming, mainly due to a reduction in seed number per m2. No effect of crop management was observed on protein content, despite significant differences in terms of protein composition. The ratio between the major globulins, 7s vicilin and 11s legumin, showed a negative relationship with grain yield and was found to be higher under organic farming. Among genotypes, black-seed Nero Senise was characterized by the highest productivity and water-holding capacity, associated with low lectin content. These findings highlight the importance of the choice of chickpea genotypes for cultivation under organic farming in relation to both agronomic performance and technological and health quality.
... Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was used to investigate the seed storage protein patterns in many legumes including Vicia in the same study [59], who analysed the seed protein patterns of 47 accessions belonging to 11 species and four tribes of legumes including Vicia. Another study [60] isolated the protein samples from 15 legume species and cultivars (vetches, pea, white lupin, and field bean) and analysed them by SDS-PAGE electrophoresis of the seed storage proteins and estimated their molecular weight and quantify their relative quantities. ...
Background: Genus Vicia is a member of family Fabaceae and comprises 180 to 210 species. The most important species is faba bean (Vicia faba) which is still one of the most favourable grain legumes over all the world. The genus contains some additional food crops and a number of forage plants and some other weedy strains such as Vicia angustifolia and Vicia cordata. The aim of the present investigation is to elucidate the phylogenetic relationships among four Vicia species, two species (Vicia angustifolia L. ssp. Angustifolia (2n = 12) and Vicia cordata wulfen ex Hoppe (2n = 10)) belong to section Vicia, Vicia dalmatica A. Kern (2n = 12, section Cracca), and Vicia johannis tamamsch (2n = 14, section Faba).
