SDS-PAGE Analysis Research Articles

Effect of rosmarinic acid covalent conjugation on the structure and allergenic capacity of ovalbumin

Ovalbumin (OVA), a major allergen in egg-induced food allergy, is widely used in the food industry. This study investigated the structural and antigenic changes of OVA after covalent conjugation with rosmarinic acid (RA). The SDS-PAGE and RA binding equivalent analysis confirmed that both the radical and alkaline methods successfully prepared the OVA-RA conjugates. LC-MS/MS assay identified 20 and 15 modified sites of RA on OVA in the radical and alkaline OVA-RA conjugates, respectively. The structural changes of OVA were indicated by fluorescence spectroscopy, Fourier transform infrared (FTIR) spectroscopy, circular dichroism (CD), and ultraviolet absorption (UV) spectrometry. Following conjugation with RA, the antioxidant capacities of OVA for scavenging DPPH• and ABTS•+ showed significant enhancement. Subsequently, the impact of RA on the antigenicity of OVA was assessed both in vitro and in vivo. The ELISA analysis revealed a notable reduction in the binding capacity of OVA to IgG and IgE after covalent conjugation with RA. Additionally, the passive cutaneous anaphylaxis (PCA) assay demonstrated lower allergenicity in radical and alkaline OVA-RA conjugates compared to native OVA. These findings collectively suggest that OVA-RA conjugates hold promise for the development of hypoallergenic food.

Simultaneous Purification of Human Interferon Alpha-2b and Serum Albumin Using Bioprivileged Fluorinated Ionic Liquid-Based Aqueous Biphasic Systems.

Interferon alpha-2b (IFN-α2b) is an essential cytokine widely used in the treatment of chronic hepatitis C and hairy cell leukemia, and serum albumin is the most abundant plasma protein with numerous physiological functions. Effective single-step aqueous biphasic system (ABS) extraction for the simultaneous purification of IFN-α2b and BSA (serum albumin protein) was developed in this work. Effects of the ionic liquid (IL)-based ABS functionalization, fluorinated ILs (FILs; [C​2C​1Im][C​4F​9SO​3] and [N​1112(OH)][C​4F​9SO​3]) vs. mere fluoro-containing IL ([C​4C​1Im][CF​3SO​3]), in combination with sucrose or [N​1112(OH)][H​2PO​4] (well-known globular protein stabilizers), or high-charge-density salt K​3PO​4 were investigated. The effects of phase pH, phase water content (%wt), phase composition (%wt), and phase volume ratio were investigated. The phase pH was found to have a significant effect on IFN-α2b and BSA partition. Experimental results show that simultaneous single-step purification was achieved with a high yield (extraction efficiency up to 100%) for both proteins and a purification factor of IFN-α2b high in the enriched IFN-α2b phase (up to 23.22) and low in the BSA-enriched phase (down to 0.00). SDS-PAGE analysis confirmed the purity of both recovered proteins. The stability and structure of IFN-α2b and BSA were preserved or even improved (FIL-rich phase) during the purification step, as evaluated by CD spectroscopy and DSC. Binding studies of IFN-α2b and BSA with the ABS phase-forming components were assessed by MST, showing the strong interaction between FILs aggregates and both proteins. In view of their biocompatibility, customizable properties, and selectivity, FIL-based ABSs are suggested as an improved purification step that could facilitate the development of biologics.

The construction and evaluation of secretory expression engineering bacteria for the trans-Cry3Aa-T-HasA fusion protein against the Monochamus alternatus vector.

Pine wood nematode disease is currently the most deadly forest disease in China, and the Monochamus alternatus is its primary vector. Controlling the M. alternatus is crucial for managing pine wood nematode disease. This study, based on the selected HasA (pGHKW4) secretory expression vector, used electroporation to combine the genetically modified high-toxicity toxin Cry3Aa-T with the entomopathogenic bacterium Yersinia entomophaga isolated from the gut of the M. alternatus. The SDS-PAGE and Western blotting techniques were employed to confirm the toxin protein's secretion capability. The engineered bacteria's genetic stability and effectiveness in controlling M. alternatus were assessed for their insecticidal activity. The results of the SDS-PAGE and Western blotting analyses indicate that the HasA system effectively expresses toxin protein secretion, demonstrates certain genetic stability, and exhibits high insecticidal activity against M. alternatus. This study constructed a highly toxic entomopathogenic engineered bacterial strain against M. alternatus larvae, which holds significant implications for controlling M. alternatus, laying the foundation for subsequent research and application of this strain.

The Potential of Pediococcus acidilactici Cell-Free Supernatant as a Preservative in Food Packaging Materials.

This study delves into the production and antimicrobial characteristics of cell-free supernatants from Pediococcus acidilactici (CFSs-Pa). Antimicrobial activity was initially observed in CFS-Pa harvested after 12 h of incubation and increased up to the late stationary phase at 48 h. The increase in antimicrobial activity did not align with total protein content, pointing to other factors linked to the accumulation of organic acids, particularly lactic acid. The SDS-PAGE analysis also indicated that the expected proteinaceous compound (pediocin) was not observed in CFS-Pa. Further investigations suggested that the antimicrobial properties of CFS-Pa were exclusively due to organic acids. The MIC values confirmed potent antimicrobial activity, particularly at a 10% dilution of CFS-Pa in MRS broth. The time-kill assays demonstrated bactericidal activity against EHEC, Listeria monocytogenes, and Staphylococcus aureus by 12 h, 18 h, and 24 h using a 10% dilution of CFS-Pa. Additionally, CFS-Pa exhibited dose-dependent antioxidant activity, requiring a 70% (v/v) concentration to inhibit DPPH scavenging activity by 50%. All the experimental results suggested potential applications of CFS-Pa in food preservation. An attempt to incorporate CFS-Pa into bacterial cellulose (BC) for edible food packaging demonstrated promising antimicrobial results, particularly against L. monocytogenes and S. aureus, with room for optimization.

Expression of chicken epidermal growth factor (cEGF) in Escherichia coli regulates the microflora structure of the duodenum to improve growth performance and intestinal morphogenesis in broilers

ABSTRACT 1. A study used gene synthesis to obtain the functional domains of chicken epidermal growth factor (cEGF) and examined their impact on broiler growth performance, small intestinal morphology, digestive enzyme activities in the intestinal contents and the structure of duodenal microflora. 2. The pET-32a-cEGF recombinant expression vector was constructed. The specific band at 26 KDa was shown by SDS-PAGE analysis and WB results. The purified protein content was shown to be 1687 μg/ml by assay. 3. A total of 180 healthy, one-day-old Arbor Acres male, white-feathered broilers were randomly divided into three dietary treatment groups (six replicate pens, 10 birds per replicate): A control diet (ND); cEGF diet (cEGF), control supplemented with 250 mg/kg cEGF and the control diet (CD) supplemented with 250 mg/kg chlortetracycline. 4. The results showed that feeding the cEGF and CD diet reduced FCR of broilers aged 1–21 d, average daily feed intake (ADFI) at 22–42 d, and the FCR in the whole period (1–42 d; p < 0.05). Compared with the ND group, the cEGF diet increased duodenal α-amylase and alkaline phosphatase activities in the 1–21 d, duodenal lipase, alkaline phosphatase, and ileal alkaline phosphatase activities in the post-period and increased villus height in the duodenum and ileum (p < 0.05). In addition, the ACE and Chao1 index for the birds fed cEGF were higher than the ND group (p < 0.05). At the phyla level, Firmicutes and Proteobacteria were dominant in all groups. At the genus level, the dominant genus was Lactobacillus. The LEfSe analysis showed that the cEGF group was enriched by 11 species including Brevibacillus, Eisenbergiella, Cloacibacterium, Butyricoccus spp. 5. The addition of 250 mg/kg cEGF to the diet can increase growth performance by improving intestinal development and digestive enzyme activity, which may be related to the duodenal intestinal microflora. Therefore, cEGF is an effective alternative to antibiotics in broiler farming.

Efficient Production of a Thermostable Mutant of Transglutaminase by Streptomyces mobaraensis.

The transglutaminase (TGase) from Streptomyces mobaraensis is widely used to improve the texture of protein-based foods. However, wild-type TGase is not heat-resistant, which is unfavorable for its application. In this study, we successfully constructed a S. mobaraensis strain that can efficiently produce TGm2, a thermostable mutant of S. mobaraensis TGase. First, S. mobaraensis DSM40587 was subjected to atmospheric room temperature plasma mutagenesis, generating mutant smY2022 with a 12.2-fold increase in TGase activity. Then, based on the double-crossover recombination, we replaced the coding sequence of the TGase with that of TGm2 in smY2022, obtaining the strain smY2022-TGm2. The extracellular TGase activity of smY2022-TGm2 reached 61.7 U/mL, 147% higher than that of smY2022. Finally, the catalytic properties of TGm2 were characterized. The half-life time at 60 °C and specific activity of TGm2 reached 64 min and 71.15 U/mg, 35.6- and 2.9-fold higher than those of the wild-type TGase, respectively. As indicated by SDS-PAGE analysis, TGm2 exhibited demonstrably better protein cross-linking ability than the wild-type TGase at 70 °C, although both enzymes shared a similar ability at 40 °C. With improved enzyme production and thermal stability, smY2022-TGm2 could be a competitive strain for the industrial production of transglutaminase.

High-Level Secretory Production of Recombinant E2-Spy Antigen Protein via Combined Strategy in Pichia pastoris

E2-Spy (abbreviated as ES) plays a vital role as a component in the Bacterial-Like Particles (BLPs) vaccine against classical swine fever virus (CSFV). This vaccine demonstrates remarkable immunoprotection, highlighting the importance of augmenting ES production in the development of CSFV subunit vaccines. In this study, a Pichia pastoris strain capable of high-yield secretory production of ES was developed through signal peptide engineering, gene dosage optimization and co-expression of molecular chaperones. Initially, a hybrid signal peptide cSP3 was engineered, leading to a 3.38-fold increase in ES production when compared to the control strain 1-α-ES. Subsequently, cSP3 was evaluated for its expression efficiency alongside different commonly used signal peptides under multicopy conditions. SDS-PAGE analysis revealed that 2-αd14-ES exhibited the highest ES production, displaying a 4.38-fold increase in comparison to 1-α-ES. Afterwards, SSA1, YDJ1, BIP, LHS1, and their combinations were integrated into 2-αd14-ES, resulting in a 1.92-fold rise in ES production compared to 2-αd14-ES (equivalent to a 6.18-fold increase compared to 1-α-ES). The final yield of ES was evaluated as 168.3 mg/L through comparison with serially diluted BSA protein bands.

Elemental sulfur concentration can be used as a rapid, reliable, and cost-effective predictor of sulfur amino acid content of soybean seeds

In this study, we have examined the feasibility of using elemental sulfur content of soybean seeds as a proxy for the overall sulfur amino acid content of soybean seeds. Earlier, we have identified by high throughput ionomic phenotyping several high and low sulfur containing soybean lines from the USDA Soybean Germplasm Collection. Here, we measured the cysteine and methionine content of select soybean lines by high-performance liquid chromatography. Our results demonstrate that those soybean lines which had high elemental sulfur content also had a higher cysteine and methionine content when compared to soybean lines with low elemental sulfur. SDS-PAGE and immunoblot analysis revealed that the accumulation of Bowman Birk protease inhibitor and lunasin in soybean seeds may only be marginally correlated with the elemental sulfur levels. However, we found a positive correlation between the levels of trypsin and chymotrypsin inhibitor activities and elemental sulfur and sulfur amino acid content of the seeds. Thus, elemental sulfur content and/or protease inhibitor activity measurement can be utilized as a rapid and cost-effective method to predict the overall sulfur amino acid content of soybean seeds. Our findings will benefit breeders in their endeavors to develop soybean cultivars with enhanced sulfur amino acid content.

Putative Role of Anti-microbial Peptide Recovered from Lactiplantibacillus spp. in Biocontrol Activity.

Antimicrobial peptides (AMPs) stand as a promising alternative to conventional pesticides, leveraging a multifaceted approach to combat plant pathogens. This study focuses on identifying and characterizing the AMP produced by Lactiplantibacillus argentoratensis strain IT, demonstrating potent antibacterial activity against various harmful microorganisms. Evaluation of AMPs' antibacterial activity was conducted through an agar well diffusion assay, a reliable method for assessing secondary metabolite antimicrobial efficacy. The study unveils the antimicrobial potential of the purified extract obtained from Lactiplantibacillus argentoratensis IT, isolated from goat milk. Notably, the AMP exhibited robust antibacterial activity against phytopathogens affecting solanaceous crops, including the Gram-negative Ralstonia solanacearum. Expression conditions and purification methods were optimized to identify the peptide's mass and sequence, utilizing LC-MS and SDS-PAGE. This paper underscores the application potential of Lactiplantibacillus spp. IT as a biocontrol agent for managing bacterial infectious diseases in plants. Results indicate optimal AMP production at 37°C, with a culture broth pH of 5 during fermentation. The obtained peptide sequence corresponded to peaks at 842.5 and 2866.4m/z ratio, with a molecular weight of approximately 5kDa according to tricine SDS-PAGE analysis. In conclusion, this study lays the foundation for utilizing Lactiplantibacillus spp. IT derived AMPs in plant biocontrol strategies, showcasing their efficacy against bacterial phytopathogens. These findings contribute valuable insights for advancing sustainable agricultural practices.

Molecular Characterization of the Iron-Containing Alcohol Dehydrogenase from the Extremely Thermophilic Bacterium Pseudothermotoga hypogea.

Pseudothermotoga hypogea is an extremely thermophilic bacterium capable of growing at 90 °C and producing ethanol, which is catalyzed by an alcohol dehydrogenase (ADH). The gene encoding P. hypogea ADH (PhADH) was cloned, sequenced and over-expressed. The gene sequence (1164 bp) was obtained by sequencing all fragments of the gene, which were amplified from the genomic DNA. The deduced amino acid sequence showed high identity to iron-containing ADHs from other Thermotoga species and harbored typical iron- and NADP-binding motifs, Asp195His199His268His282 and Gly39Gly40Gly41Ser42, respectively. Structural modeling showed that the N-terminal domain of PhADH contains an α/β-dinucleotide-binding motif and that its C-terminal domain is an α-helix-rich region containing the iron-binding motif. The recombinant PhADH was soluble, active, and thermostable, with a subunit size of 43 ± 1 kDa revealed by SDS-PAGE analyses. The recombinant PhADH (69 ± 2 U/mg) was shown to have similar properties to the native enzyme. The optimal pH values for alcohol oxidation and aldehyde reduction were 11.0 and 8.0, respectively. It was also thermostable, with a half-life of 5 h at 70 °C. The successful expression of the recombinant PhADH in E. coli significantly enhanced the yield of enzyme production and thus will facilitate further investigation of the catalytic mechanisms of iron-containing ADHs.

Heat-induced agglomeration of water-soluble cod proteins toward gelled structures

Cod proteins (CPs) have potential applications in designing desirable gel-based products, and this study aimed to unravel their heat-induced aggregation pattern and further probe the roles in protein gels. SDS-PAGE analysis indicated that high-precipitation-coefficient aggregates (HPCAs) of CPs aggregates were composed of considerable polymers of myosin heavy chains and actin, and their low-precipitation-coefficient aggregates (LPCAs) contained myosin light chains and tropomyosin. Studies from correlation analysis between the structure and aggregation kinetics revealed that the generation of β-sheet and SS bonds were responsible for their spontaneous thermal aggregation induced by heating temperature and protein concentration, respectively. Additionally, as protein denaturation ratio increased, more and larger HPCAs were formed, which was evidenced driving the network formation of protein gels and resulting in higher storage modulus (G') values. These novel findings may be applicable to other animal proteins for better tailoring the manufacturing of muscle gel-based products.

Ptpn23 Controls Cardiac T-Tubule Patterning by Promoting the Assembly of Dystrophin-Glycoprotein Complex.

Cardiac transverse tubules (T-tubules) are anchored to sarcomeric Z-discs by costameres to establish a regular spaced pattern. One of the major components of costameres is the dystrophin-glycoprotein complex (DGC). Nevertheless, how the assembly of the DGC coordinates with the formation and maintenance of T-tubules under physiological and pathological conditions remains unclear. Given the known role of Ptpn23 (protein tyrosine phosphatase, nonreceptor type 23) in regulating membrane deformation, its expression in patients with dilated cardiomyopathy was determined. Taking advantage of Cre/Loxp, CRISPR/Cas9, and adeno-associated virus 9 (AAV9)-mediated in vivo gene editing, we generated cardiomyocyte-specific Ptpn23 and Actn2 (α-actinin-2, a major component of Z-discs) knockout mice. We also perturbed the DGC by using dystrophin global knockout mice (DmdE4*). MM 4-64 and Di-8-ANEPPS staining, Cav3 immunofluorescence, and transmission electron microscopy were performed to determine T-tubule structure in isolated cells and intact hearts. In addition, the assembly of the DGC with Ptpn23 and dystrophin loss of function was determined by glycerol-gradient fractionation and SDS-PAGE analysis. The expression level of Ptpn23 was reduced in failing hearts from dilated cardiomyopathy patients and mice. Genetic deletion of Ptpn23 resulted in disorganized T-tubules with enlarged diameters and progressive dilated cardiomyopathy without affecting sarcomere organization. AAV9-mediated mosaic somatic mutagenesis further indicated a cell-autonomous role of Ptpn23 in regulating T-tubule formation. Genetic and biochemical analyses showed that Ptpn23 was essential for the integrity of costameres, which anchor the T-tubule membrane to Z-discs, through interactions with α-actinin and dystrophin. Deletion of α-actinin altered the subcellular localization of Ptpn23 and DGCs. In addition, genetic inactivation of dystrophin caused similar T-tubule defects to Ptpn23 loss-of-function without affecting Ptpn23 localization at Z-discs. Last, inducible Ptpn23 knockout at 1 month of age showed Ptpn23 is also required for the maintenance of T-tubules in adult cardiomyocytes. Ptpn23 is essential for cardiac T-tubule formation and maintenance along Z-discs. During postnatal heart development, Ptpn23 interacts with sarcomeric α-actinin and coordinates the assembly of the DGC at costameres to sculpt T-tubule spatial patterning and morphology.

Effect of cadmium and lead on the morphology and protein profile of Calligonum comosum seeds

BackgroundSoils contaminated with heavy metals (HMs) pose a risk to human health via the food chain, as many edible plants absorb these metals. In turn, some of these plants could be used as phytoremediators for such soils. Calligonum comosum is an indigenous medicinal shrub that grows naturally in wide swaths of sandy soil in Saudi Arabia and has many advantages that render it a promising candidate for the treatment of HM-contaminated areas. But the impact of HM on this plant remains unknown, especially in the early stages of its development.ObjectiveThis investigation aimed to study the effects of lead (Pb) and cadmium (Cd) on the germination of C. comosum seeds under laboratory conditions, and assess the seeds’ response to these metals.MethodsThe C. comosum seed germination was monitored in Petri dishes containing Pb and Cd at increasing concentrations (25, 50, 75, and 100 µM) for up to 3 weeks. SDS-PAGE was used to examine the protein profile of germinated seeds and the western blot was used to assess the influence of HMs on the activities of the catalase enzyme and the beta subunit of ATP synthase (AtpB).ResultsThe germination rate and speed of C. comosum seeds were delayed by increasing concentrations of either Pb or Cd, but this effect was dose-dependent. SDS-PAGE analysis results revealed that exposure to both metals led to altered protein profiles as indicated by the resulting band intensities and disappearance of some proteins compared with the untreated controls. Further, the western blot analysis detected greater activity of catalase enzyme as well as AtpB in the Cd- and Pb-treated seeds.ConclusionC. comosum seeds treated with Cd or Pb enhance protein degradation and denaturation beside oxidative stress, leading to reduced seed viability. These results suggest oxidoreduction proteins and those involved in ATP synthesis are enhanced in C. comosum seeds in response to Cd and Pb stressors, which is a probable mechanism by which seeds may tolerate heavy metal stress.

Genetic Modification of Aurantiochytrium sp. 18W-13a for Enhancement of Proteolytic Activity by Heterologous Expression of Extracellular Proteases.

A marine thraustochytrid, Aurantiochytrium, is a promising organism to produce docosahexaenoic acid (DHA) and squalene. Utilization of inexpensive substances such as proteins in wastes and by-products from the food industry for cultivation is a considerable option to reduce production cost; however, the proteolytic ability of Aurantiochytrium spp. is low compared to taxonomically close Shizochytrium aggregatum. We previously identified extracellular protease (extracellular protease 1, EP1) in S. aggregatum ATCC 28209 from the supernatant of the culture and found that a similar protease gene (EP2) was located downstream of the EP1 gene. In the present study, we created the transformants expressing SaEP1 and/or SaEP2 to enhance the proteolytic ability of Aurantiochytrium sp. 18W-13a strain and cultivated them in the medium containing casein as a test protein substrate. Through SDS-PAGE analysis, we confirmed that casein in the supernatant was more efficiently degraded by the transformants than the wild type, suggesting that the expressed protease(s) were properly expressed and excreted. After 4-day cultivation in the casein medium, the value of optical density at 660nm and the cell number in the culture of the transformant that expressed both SaEP1 and SaEP2 (designated as EP12 strain) showed 1.48- and 1.38-fold higher than those of wild type, respectively. The DHA and squalene yield of the EP12 strain were respectively 158.3 and 0.23mg L-1, and these values were 1.42- and 2.01-fold higher than those of wild type, respectively, suggesting that the EP12 created in the present study is a favorable strain for the cultivation using protein-containing medium.

Uncovering the antimicrobial activity of G-type lysozyme 2 derived from Cyprinus carpio mucus against bacterial and fungal pathogens

This study aimed to explore the antimicrobic potential of mucus samples collected from Cyprinus carpio and identify the specific antimicrobial peptides responsible for its activity. The crude extract was tested against various bacterial and fungal pathogens, and its protein content and profile were analyzed. Purification steps, including gel filtration chromatography, were employed to isolate the most active fraction (peak IV), which was further identified via liquid chromatography and mass spectroscopy. The results revealed varying degrees of antimicrobial activity of the crude extract against different bacterial and fungal strains, with Leclercia adecarboxylata, Candida glabrata, and Candida parapsilosis showing the highest susceptibility. SDS-PAGE analysis demonstrated the existence of multiple low molecular weight protein bands in the crude extract, while fraction IV obtained from gel filtration chromatography exhibited the strongest antimicrobial activity. Peak IV displayed a range of minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC) values against the tested pathogens, spanning from 0.038 to 4.960 mg/mL. Further investigation identified the purified peptide derived from peak IV as G-type lysozyme 2, characterized by a molecular weight of 21 kDa. These findings shed light on the existence of a highly effective antimicrobial peptide, G-type lysozyme 2, within the mucus of Cyprinus carpio. This peptide demonstrates notable activity against diverse bacterial and fungal pathogens. The insights from this study enhance our understanding of the fish's antimicrobial defense mechanisms and hold promise for developing novel antimicrobial agents.

Synthesis of New Derivatives of Berberine Canagliflozin and Study of Their Antibacterial Activity and Mechanism.

The isoquinoline alkaloid berberine, derived from Coptidis rhizoma, exhibits antibacterial, hypoglycemic, and anti-inflammatory properties. Canagliflozin is a sodium-glucose cotransporter 2 (SGLT2) inhibitor. We synthesized compounds B9OC and B9OBU by conjugating canagliflozin and n-butane at the C9 position of berberine, aiming to develop antimicrobial agents for combating bacterial infections worldwide. We utilized clinically prevalent pathogenic bacteria, namely Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, to investigate the antibacterial efficacy of B9OC. This was accomplished through the determination of the MIC80 values, analysis of bacterial growth curves, evaluation of biofilm formation using crystal violet staining, assessment of impact on bacterial proteins via SDS-PAGE analysis, and observation of alterations in bacterial morphology utilizing field emission scanning electron microscopy. Meanwhile, the ADMET of compound B9OC was predicted using a computer-aided method. The findings revealed that B9OC exhibited lower minimal inhibitory concentrations against all three bacteria compared to berberine alone or in combination with canagliflozin. The minimal inhibitory concentrations (MICs) of B9OC against the three experimental strains were determined to be 0.035, 0.258, and 0.331 mM. However, B9OBu exhibited a lower level of antimicrobial activity compared to berberine. The compound B9OC exhibits a broad spectrum of antibacterial activity by disrupting the integrity of bacterial cell walls, leading to cellular rupture and the subsequent degradation of intracellular proteins.

Effects of the 1RS.1BL wheat-rye translocationon kernel and bran content of bread wheat (Triticum aestivum L.)

In this study, the effects of rye translocation on some quality properties and mineral content of whole wheat kernel and bran were investigated.The plant material consisted of 147 recombinant inbred lines (RILs) and five control cultivars, including the parent. The genotypes were grown in Turkey at two different locations for two consecutive years. In the study, lines with and without rye translocation were determined by SDS-PAGE and PCR analysis, and the glutenin subunits of the lines at Glu-A1, Glu-B1, Glu-A3 and Glu-B3 loci were also analyzed. The protein, starch, fat, ash, dry matter, acid detergent insoluble fiber (ADF), neutral detergent insoluble fiber (NDF), potassium (K), phosphorus (P) and magnesium (Mg) contents of kernels and bran of the RILs were determined. Statistically significant differences were found between the genotypes in terms of all the examined characteristics except the amount of dry matter. The RILs carrying translocation 1RS.1BLhad a higher average ADF, NDF, protein content, ash content, K, Mg, and P contents of kernels, as well as a higher average ADF, NDF, ash, and K bran contents than the RILs without thetranslocation. The bran of the RILs without thetranslocationhad higher values in terms of fat, starch, and phosphorus content. The results indicate that wheat- rye translocation1RS.1BL has a significant effect on both whole wheat kernel and bran content, and can be used to enrich the content of wheat bran.

Evaluation of the durum wheat landrace genetic diversity using agro analysis and its benefit for human health

Ancient wheat and landraces (traditional varieties) are fascinating to the consideration of scientists who are re-evaluating the healthy and dietary properties attributed to them by popular tradition. Wheat (Triticum durum) is one of the most significant crops that is grown all over the world. In the present study, 19 landraces were analyzed for genetic diversity through morphological and SDS-PAGE analysis. The highest coefficient of variance (18.91%) was found for spike length, and the lowest for days to maturity (1.51%). The genotype Malakand and Dolram produced a maximum number of seeds (42 and 46 seeds per plant), and the genotypes Dogai and Rasool Banda were found to be the tallest (73 and 71 cm, respectively). The genotype Kamar Kotki was found to mature early (165 days) while Doagi was found to mature late (171 days). Spike length was found to be positively correlated with plant height, leaf width, and leaf length. Leaf width was significant with spike length and the internode distance. The cluster analysis divided all the genotypes into six sub-clusters where the Asban and the Shekhan were placed at the extreme of the dendrogram. Based on the total seed storage proteins analysis, 18 (85.71%) bands were polymorphic while 14.28% bands were found to be monomorphic. Using Ward’s methods, all the genotypes were divided into two main lineages at 25% Euclidian distance and five sub-clusters at 75% distance. Our findings have significance for understanding the variations and relationships between various durum wheat genotypes and could serve as a basis for local elite durum germplasm that has been regionally adapted.

Synthesis and characterization of Restricted Access Magnetic Nanotubes (M-RACNTs) for the extraction of secondary metabolites from Lasiodiplodia sp. fermentation broth

This work proposed the synthesis and the characterization of Restricted Access Magnetic Nanotubes (M-RACNTs) to be used as sorbent in the magnetic dispersive solid-phase extraction (MDSPE) of organic compounds from the fermentation broth of the endophyte Lasiodiplodia sp. Synthesis was performed by functionalizing commercial multiple-walled carbon nanotubes with magnetic nanoparticles (M-CNTs) and by coating the M-CNTs with a layer of bovine serum albumin. The materials were characterized by infrared spectroscopy with Fourier Transform, thermogravimetry, zeta potential, and scanning electron microscopy. Different crude extracts were obtained, depending on the sample pH and the elution solvent tested. Chloroform and ethyl acetate extracts showed activity against a Gram-positive pathogen (Staphylococcus aureus). The crude extract's constitution was investigated. By GC/MS analysis were found hydrocarbons, aldehydes, alcohols, among others, while by LC-MS/MS analysis were found flavonoids, macrocyclic lactones, oleic acids, jasmonic acid derivatives and amino acids. A multivariate optimization, through fractional factorial planning, for four independent variables, was performed for a hydrocarbon (hexadecane) and one possible jasmonic acid derivative, both with antimicrobial activity related in the literature. The results showed that it was possible to obtain crude extracts with high concentrations of these analytes in a single extraction procedure. The increase in the analytical signal was 3.7 times for hexadecane and 1.4 times for the possible jasmonic acid derivative. The M-RACNTs also demonstrated their ability to prevent the adsorption of macromolecules, such as lysozyme, which was confirmed through SDS-PAGE analysis.

Reduction of anti-nutrients factors and study of in-vitro protein digestibility, fractionation, SDS-PAGE analysis and thin layer chromatography of wild apricot kernel press cake protein isolate

Reduction of anti-nutrients factors and study of in-vitro protein digestibility, fractionation, SDS-PAGE analysis and thin layer chromatography of wild apricot kernel press cake protein isolate