SDS-polyacrylamide Gel Research Articles

Comparative study of a liposome and emulsion system with cinnamon essential oil on the quality and proteolysis of refrigerated minced pork.

Essential oils have been recognized for their strong antibacterial property, making them an innovative approach for preserving meat. However, their chemical instability and direct impact on meat proteins limit their application. To overcome these limitations, various loading systems have been explored. This study aimed to compare the effect of cinnamon essential oil (CEO) loaded in a liposome and emulsion system on the proteolysis of minced pork and to evaluate the advantages of each delivery system in preventing microorganism-induced quality deterioration of meat. Minced pork treated with CEO-liposomes exhibited lower pH, total volatile basic nitrogen (TVB-N), and total viable count (TVC) values than CEO-emulsions and provided better protection against microorganisms. SDS-polyacrylamide gel electrophoresis (PAGE) analysis confirmed that CEO-liposome was more effective in protecting proteins from degradation. Moreover, CEO-liposome produced lower amount of bitter amino acids and harmful biogenic amines. Antibacterial mechanisms indicated that CEO-liposome exhibited a stronger inhibitory effect against major spoilage bacteria in meat products by increasing cell membrane permeability. The membrane damage was further supported by an increase in conductivity and the leakage of nucleic acids. Compared to the CEO-emulsion system, CEO-liposome emerged as an effective preservative for minced pork. These results provided important theoretical support for using a bioactive compound delivery system to prevent microorganism-induced quality deterioration in meat.

Unicorn Fish (Naso reticulatus Randall, 2001) Skin Collagens Prepared Using Two Pepsin Sources: An Assessment on Physicochemical Characteristics

Fish collagens have gained considerable attention from numerous researchers due to their attractive traits and are more acceptable from most religious beliefs. This paper aimed to extract and characterize collagens from the skins of unicorn fish (N. reticulatus) influenced by pepsin from porcine (UCP) and pepsin from bovine (UCB). The yield of the UCP sample (15.60%) was significantly higher (P<0.05) compared to the UCB (10.40%). In addition to this, the swelling value of two collagens showed significant differences (P<0.05), with a greater percentage obtained in UCP 9261.23%) rather than UCB (196.75%). Both UCP and UCB were classified as type I collagen owing to the existence of two alpha chains under SDS-polyacrylamide gel electrophoresis. Under infrared and ultraviolet-visible parameters, the triple helical structure of collagens prepared using pepsin from porcine and bovine was preserved, and it was comparable to previous findings in fish collagen literature. All samples showed a different thermostability value, the higher one observed in the UCP (43.63°C) compared to the UCB sample (35.25°C), and their variations of thermostability were in agreement with the hydroxyproline content of UCP (83.57 mg/g) and UCB (81.47 mg/g). The unicorn fish (N. reticulatus) skin may be used as a good source of collagen, mainly utilized for industrial perspective.

In vitro gastrointestinal simulated digestion of three plant proteins: determination of digestion rate, free amino acids and peptide contents

ABSTRACT Cassava protein (CP), barley protein (BP) and yellow pea protein (YPP) are important nutrient and integral constituent of staple in pet foods. It is known that the digestion of proteins directly influences their absorption and utilisation. In the present work, we performed in vitro simulated gastrointestinal digestion of three plant proteins as a staple for dog and cat food. The digestion rate of CP, BP and YPP in dog food was 56.33 ± 0.90%, 48.53 ± 0.91%, and 66.96 ± 0.37%, respectively, whereas the digestion rate of CP, BP, and YPP in cat food was 66.25 ± 0.72%, 43.42 ± 0.83%, and 58.05 ± 0.85%, respectively. Using SDS-polyacrylamide gel electrophoresis to determine the molecular weight (MW) of each protein and the products of their digestion, it was revealed that MW of digestion samples decreased, and MW during the small intestine phase was lower than that during the gastric phase. Peptide sequences of digested products were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and it was found that the total number of peptides in the small intestine digestion samples was higher than that in the gastric phase samples. The MW of peptides obtained from CP was within the range of 1000–1500 Da, while MW of peptides derived from BP and YPP was within the range of 400–2000 Da. In addition, free amino acids were mainly produced in the small intestine phase. Furthermore, the percentage of essential amino acids in the small intestine phase (63 ~ 82%) was higher than that in the gastric phase (37 ~ 63%). Taken together, these findings contribute to the current understanding of the utilisation of plant proteins in dog and cat foods and provide important insights into the selection and application of plant proteins as a staple in dog and cat foods.

Comparative Protein Profiling of Blood and Milk of Early Pregnant Buffaloes Using SDS-PAGE

Conventional methods for detecting pregnancy in buffalo are inefficient before 30-days after artificial insemination. A proteomic approach for high-resolution analysis of blood plasma and milk proteins by SDS polyacrylamide gel electrophoresis (PAGE) is of great importance. Blood and milk samples were collected on days 0 (before insemination) and 20 and 25 after insemination from retrospective pregnant (n = 6) and non-pregnant (n = 6) animals. An equal volume of samples was loaded in (12%-4%) SDS gel to resolve major plasma and milk proteins. Plasma proteins showed three distinct zones – high, medium, and low molecular weight (MW). In the high MW zone (135 - 250 kDa), ~208 kDa and ~190 kDa proteins were visible. In the moderate MW zone (60 -75 kDa), 5-6 proteins like albumin (~70 kDa), 75 kDa, and 63 kDa consisting of pregnancy-associated glycoproteins, ~73 kDa being dominant. In the low MW zone (25 – 35 kDa), a clear band of 27 kDa was visible. Proteins in milk supernatants showed four distinct zones of proteins – very high, high, medium, and low molecular weight (MW) zones. In the very high MW zone (135 - 250 kDa), ~208 kDa and ~190 kDa proteins were visible. In the high MW zone (60 -75 kDa), proteins like lactoferrin (~78.2 kDa), 73 kDa, albumen (~66 kDa), and a heavy chain of immunoglobulin (IgG) (~54 kDa) were detected. In the medium MW zone (25 – 35 kDa), clear bands of milk caseins, namely thick alpha S2 casein (29 kDa), and other caseins. In the low MW zone (11 – 20 kDa), two thick bands of 18 kDa and 12 kDa were visible. The mean gray values (sum of the gray values of all the pixels in the peak divided by the number of pixels) of the 73 kDa protein peak were analyzed in two-way ANOVA using GraphPad (ver 8.0). Repeated measures of ANOVA were selected where pregnancy status (pregnant and non-pregnant) defined column factor and time (0-, 20- and 25-day) defined row factor. The expression of 73 kDa protein in blood plasma was higher (P = 0.04) in pregnant women than in non-pregnant buffaloes at 20- and 25-days after insemination. However, in milk, there was no difference in the expression of the 73 kDa protein between pregnant and non-pregnant buffaloes across time. Variation in animals within the groups was significantly high. The differential expression of ~73 kDa protein in plasma and precise identification of the protein can be a diagnostic marker for early pregnancy detection in buffaloes.

Preparation and extraction of chorion proteins from Salmo salar embryos at the pigmented eye stage for electrophoresis with SDS-polyacrylamide gel

The chorion fulfills important functions in fish embryos, including protecting the embryo during development. The characterization of the protein profile of this envelope could be used as a bioindicator in the evaluation of the quality of embryonic development. The object of this work was to validate a standardized protocol for protein extraction from chorion of Salmo salar embryos at 280 accumulated thermal units (ATU) by comparing and combining existing methods. The protocol consists of consecutive washing of the chorion samples followed by protein extraction with the solution that was named SDS solution (Tris–HCl 100 mM (pH 8), Urea 8 M, 1% SDS, β-mercaptoethanol 300 mM and EGTA 10 Mm, and 1% protease inhibitor cocktail) and mechanical methods. Protein extraction is enhanced by a working temperature of 75 °C and use of a disperser. The protein concentration was quantified by Bradford Assay. After extraction, the samples were diluted (dilution factor 10) before reading against the calibration curve. After gel electrophoresis with a load of 3 µg of protein, staining showed more than 4 bands, with molecular weights between 25 kDa and 180 kDa.•The protein profile of fish chorion was between 25 kDa and 180 kDa.•Solution containing 1% SDS allows a higher extraction of proteins from the chorion of Atlantic salmon embryos with 280 ATU.•Chorion protein identification is a valuable tool in determining gamete and embryo quality in fish.

Tau and Alzheimer's disease: Past, present and future.

My journey with tau started when in 1974 for the first time I isolated neurofibrillary tangles of paired helical filaments (PHFs) from autopsied Alzheimer's disease (AD) brains and discovered that they were made up of a ~50-70 KDa protein on SDS-polyacrylamide gels. Subsequently my team discovered that this PHF protein and the microtubule-associated factor called tau were one and the same protein. However, we found that tau in neurofibrillary tangles/PHFs in AD brain was abnormally hyperphosphorylated, and unlike normal tau, which promoted the assembly of tubulin into microtubules, the AD-hyperphosphorylated tau inhibited microtubule assembly. These discoveries of tau pathology in AD opened a new and a major area of research on tau and on the molecular pathology of this major cause of dementia in middle- and old-age individuals. Tau pathology, which without fail is made up of the aggregated hyperphosphorylated state of the protein, is also the hallmark lesion of a family of around 20 related neurodegenerative diseases, called tauopathies. Currently, tau pathology is a major drug target for the treatment of AD and related tauopathies. Both active and passive tau immunization human clinical trials at various stages are underway. Initial results range from negative to partially promising. Future studies will reveal whether tau therapy alone or in combination with drugs targeting Aβ and/or neurodegeneration will be required to achieve the most effective treatment for AD and related disorders.

Improvement of camel milk-clotting: Usefulness of crude extract from green pods of carob (Ceratonia siliqua. L) as a substitute for commercial rennet.

Camel milk transformation into cheese remains an objective to be improved today. This study aimed to improve camel milk clotting using a crude extract from green pods of carob as a substitute for commercial rennet. The composition of the crude carob extract was determined for dry matter and protein content. Milk clotting conditions were studied at different temperatures, pH and CaCl2 concentrations. Milk clotting properties were assessed by milk clotting activity, specific activity and proteolytic activity. Enzymatic hydrolysis of camel milk caseins by crude carob extract and its inhibition were demonstrated by SDS-polyacrylamide gel electrophoresis. Crude carob extract analysis showed a protein and dry matter content of 23.26±0.5 mg/ml and 30.66±0.5 g/l, respectively. Optimal milk clotting activity was observed at 53.6 °C, pH 4.5, and 0.09 M CaCl2. The crude carob extract showed a high milk clotting activity (4.97 U/ml) and a low proteolytic activity (0.04U/ml) with camel milk. The cheese yield of curd produced from camel milk using crude carob extract was the highest (23.95%) compared with that of Camel chymosin (20.5%). The high ratio of milk-clotting to proteolytic activity shows the potential of this extract as a substitute for commercial rennet in the dairy industry.

A novel botybirnavirus with a unique satellite dsRNA causes latent infection in Didymella theifolia isolated from tea plants.

A novel botybirnavirus, infecting the tea plant pathogen Didymella theifolia and tentatively named Didymella theifolia botybirnavirus 1 (DtBRV1), together with an additional double-stranded RNA (dsRNA), was characterized. DtBRV1 comprises two dsRNAs (1 and 2) encapsidated in isometric virions, while dsRNA3 is a satellite. The satellite represents a unique specimen since it contains a duplicated region and has high similarity to the two botybirnavirus dsRNAs, supporting the notion that it most likely originated from a deficient genomic component. The biological characteristics of DtBRV1 were further determined. With their unique molecular traits, DtBRV1 and its related dsRNA expand our understanding of virus diversity, taxonomy, and evolution.

Assessment of intra- and inter-genetic diversity in tetraploid and hexaploid wheat genotypes based on omega, gamma and alpha-gliadin profiles.

Durum and bread wheat are well adapted to the Mediterranean Basin. Twenty-three genotypes of each species were grown to evaluate the intra- and inter-genetic diversity based on omega (ω), gamma (γ) and alpha (α)-gliadin profiles. To achieve this purpose, the endosperm storage proteins (both gliadins and glutenins) were extracted from wheat grains and electrophoresed on sodium dodecyl sulfate (SDS)-polyacrylamide gels. The results of SDS-Polyacrylamide Gel Electrophoresis (SDS-PAGE) revealed nine polymorphic loci out of 16 loci with durum wheat genotypes and nine polymorphic loci out of 18 loci with bead wheat genotypes. The polymorphisms revealed by the SDS-PAGE were 56% and 50% in durum and bread wheat genotypes, respectively. Using the cluster analysis, the durum wheat genotypes were clustered into five groups, while the bread wheat genotypes were grouped into six clusters using un-weighed pair group mean analyses based on ω, γ, and α-gliadins profiles. The 46 durum and bread wheat genotypes were grouped into seven clusters based on the combined ω, γ, and α-gliadins profiles revealed by the SDS-PAGE. The in silico analysis determined the intra-genetic diversity between bread and durum wheat based on the sequences of ω, γ, and α-gliadins. The alignment of ω-gliadin revealed the highest polymorphism (52.1%) between bread and durum wheat, meanwhile, the alignment of γ and α-gliadins revealed very low polymorphism 6.6% and 15.4%, respectively. According to computational studies, all gliadins contain a lot of glutamine and proline residues. The analysis revealed that the bread wheat possessed ω and γ -gliadins with a lower content of proline and a higher content of glutamine than durum wheat. In contrast, durum wheat possessed α-gliadin with a lower content of proline and a higher content of glutamine than bread wheat. In conclusion, the SDS-PAGE, in silico and computational analyses are effective tools to determine the intra- and inter-genetic diversity in tetraploid and hexaploid wheat genotypes based on ω, γ, and α-gliadins profiles.

Effect of ultrasonic assisted grafting on the structural and functional properties of mung bean protein isolate conjugated with maltodextrin through maillard reaction

Four different methods of maillard reaction including ultrasound (150 W, 10 min) assisted, classical wet heating (80 °C, 60min), moderate water bath heating (60°C, 12 to 30 h) and dry state method (60 °C, 79 % relative humidity and 48 h) were used to Mung bean protein isolate – Maltodexrtin conjugates (MPI-MD) preparation. The samples prepared under ultrasound and wet heating were chosen for further analysis according to degree of graft and UV-absorbance at 420 nm. Higher glycosylation at short time and lower browning were obtained under ultrasound treatment. Covalent attachment in conjugates confirmed by SDS-polyacrylamide gel electrophoresis. The structural analysis revealed prominent unfolding effect of ultrasound waves on the protein's molecules. The decrease of α-helix content was related to the exposure of buried amino group residues during reaction. Glycation of MPI under ultrasound caused changes in tertiary structure of protein and leads to decrease in the fluorescence intensity compared with native and wet heating treatments. FTIR spectra confirmed the conjugation of the MPI and MD and suggested that protein structure was changed and ultrasound promoted the graft reaction more than wet heating treatment. Conjugated MPI showed higher emulsification and solubility index than MPI, moreover the effect of ultrasonic waves on ameliorated functional properties was impressive than those for wet heating treatment. Overall, this study showed use of ultrasonication in maillard reaction was a suitable method for producing MPI- MD conjugates and improved the efficiency of graft reaction and functional properties of grafts.

Evaluation of freshwater heavy metals accumulation effect on oxidative stress, Metallothionein biosynthesis and histopathology of Procambarus clarkii (Girard,1985) collected from three locations in the Delta region, Egypt

BackgroundIn this study, the effect of heavy metals accumulation influence was evaluated on adult crayfish Procambarus clarkii (Decapoda, Astacidea) collected from three different Governmental locations (Kafr El-Shaikh, El-Menofya, and El-Gharbiya) of the Egyptian Delta. The activity of super oxidase dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX) of gills, hepatopancreas, and muscle tissue were measured. SDS Polyacrylamide gel electrophoresis (SDS-PAGE) and West blotting technique were performed to detect MT Protein expression.ResultsThe results revealed that Kafr El-Shaikh reflected the highest Superoxide dismutase (SOD), Catalase, and Glutathione S-transferase (GST) activity levels (97.2 u/100 mg, 28.5 u/100 mg, and 8.3 nmol mg (-1) protein min (-1) respectively. Superior protein polymorphism % (30%) remarked collected Freshwater crayfish P. clarkii from Kafr El-Shaikh location. Varied protein polymorphism % was shown between collected crayfish from El-Menofya, and El-Gharbiya locations (5.5 and 6.2 respectively) Increasing Metallothioneins intensity (15.4%) for collected Freshwater crayfish Procambarus clarkii from Kafr El-Shaikh Location.ConclusionHeavy metal stress influences antioxidant status and also induces increasing Metallothioneins intensity, especially samples that were collected from the Kafr El-Shaikh area.

Using enhanced development tools offered by analytical Quality by Design to support switching of a quality control method.

Quality by Design (QbD) principles play an increasingly important role in the pharmaceutical industry. Here, we used an analytical QbD (AQbD) approach to develop a capillary electrophoresis sodium dodecyl sulfate under reducing conditions (rCE-SDS), with the aim of replacing SDS-polyacrylamide gel electrophoresis (SDS-PAGE) as release and stability test method for a commercialized monoclonal antibody product. Method development started with defining analytical method performance requirements as part of an analytical target profile, followed by a systematic risk assessment of method input parameters and their relation to defined method outputs. Based on this, design of experiments studies were performed to identify a method operable design region (MODR). The MODR could be leveraged to improve method robustness. In a bridging study, it was demonstrated that the rCE-SDS method is more sensitive than the legacy SDS-PAGE method, and a conversion factor could be established to compensate for an off-set due to the higher sensitivity, without losing the correlation to the historical data acquired with the former method. Overall, systematic application of analytical Quality by Design principles for designing and developing a new analytical method helped to elucidate the complex dependency of method outputs on its input parameters. The link of the method to product quality attributes and the definition of method performance requirements were found to be most relevant for derisking the analytical method switch, regarding impact on the control strategy.

The effect of dephosphorylation on the properties of αS1-casein enriched protein

αS1-Casein was dephosphorylated using alkaline phosphatase without the interference from proteolysis. The progressive dephosphorylation was monitored by alkaline urea polyacrylamide gel electrophoresis and mass spectrometry techniques. Little change was observed in the SDS-polyacrylamide gel patterns for the native and dephosphorylated αS1-casein. Dephosphorylation modified the zeta-potential of the αS1-casein, shifting the pH versus zeta potential curves to higher pH as dephosphorylation increased. The isoelectric pH of the dephosphorylated αS1-casein increased by about 0.55 pH units when compared with the native protein. Dephosphorylation increased the surface hydrophobicity of αS1-casein and a negative linear correlation was observed between the surface hydrophobicity and the isoelectric pH of αS1-casein. Dephosphorylation did not affect the secondary structure of the αS1-casein.

Two proteins prepared from defatted Antarctic krill (Euphausia superba) powder: Composition, structure and functional properties

Defatted krill powder, rich in protein, has emerged as a potential, novel food raw material. The present study aimed to prepare Antarctic krill protein from the supernatant (AKP-S) and precipitation (AKP-P) using the ultrasound-assisted alkali dissolution and acid precipitation method, and analyze their composition, structure, and functional properties were analyzed. The protein contents of AKP-P and AKP-S were more than 85% (dry weight), and their total amino acid content was 48.75 and 43.59 g/100 g of protein, respectively. The SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and surface morphology analysis (SEM) results showed that the AKP-S was degraded during acid treatment and had a smaller particle size than AKP-P. The α-helix and random curled structures of AKP-P and AKP-S were few or even disappeared. The solubility of AKP-S was more than 70% in the pH range of 3–13, and the change in AKP-P solubility with pH was consistent with the trend of β-sheets. Except for pH 7.0–9.0, the solubility of AKP-P and AKP-S with pH was positively correlated with surface-hydrophobicity (H0). The emulsifying activity of AKP-P and AKP-S at different pH was approximately positively correlated with the solubility. The minimum gel-forming concentration of AKP-P was 8%, while AKP-S could not form gel at 14%. The gel water holding capacity reached more than 90% when the AKP-P concentration was 12%. The rheological results showed that the storage modulus (G′) predominated over the loss modulus (G″) of AKP-P in the range of 8%–14% concentration.

Glycosylphosphatidylinositol-anchoring is required for the proper transport and extensive glycosylation of a classical arabinogalactan protein precursor in tobacco BY-2 cells.

Many precursors of plant arabinogalactan proteins (AGPs) contain a C-terminal glycosylphosphatidylinositol (GPI)-anchoring signal. Using NtAGP1, a classical tobacco AGP, as a model, and green fluorescent protein (GFP) and sweet potato sporamin (SPO) as tags, we analyzed the localization and modification of AGP and its mutant without GPI-anchoring signal (AGPΔC) in tobacco BY-2 cells. The NtAGP1 fusion proteins migrated as large smear on SDS-polyacrylamide gel, and these proteins also localized preferentially to the plasma membrane. In contrast, fusions of AGPΔC with GFP and SPO yielded several forms: The largest were secreted, whereas others were recovered in the endomembrane organelles, including vacuoles. Comparison of the glycan structures of the microsomal SPO-AGP and the secreted SPO-AGPΔC using antibodies against the glycan epitopes of AGP indicated that the glycan structures of these proteins are different. These observations indicate that GPI-anchoring is required for the proper transport and glycosylation of the AGP precursor.

Characterization of Milk Proteins from Vechur and Kasargod Dwarf Breeds of Cattle using SDS-Polyacrylamide Gel Electrophoresis

Characterization of Milk Proteins from Vechur and Kasargod Dwarf Breeds of Cattle using SDS-Polyacrylamide Gel Electrophoresis

A Novel Molecular Approach for Enhancing the Safety of Ozone in Autohemotherapy and Insights into Heme Pocket Autoxidation of Hemoglobin.

Major ozone autohemotherapy (MAH) is a popular clinical practice for treating a variety of pathological conditions due to the mild and controlled oxidative stress produced by the reaction of ozone gas with other biological components. Previous studies have shown that blood ozonation leads to structural changes in hemoglobin (Hb); therefore, in the present study, the molecular effects of ozonation on Hb of a healthy individual were assessed by ozonating whole blood samples with single doses of ozone at 40, 60, and 80 μg/mL or double doses of ozone at 20 + 20, 30 + 30, and 40 + 40 μg/mL ozone to investigate whether ozonating once versus twice (but with the same final ozone concentration) would have varying effects on Hb. Additionally, our study aimed to verify whether using a very high ozone concentration (80 + 80 μg/mL), despite mixing it with blood in two steps, would result in Hb autoxidation. The pH, oxygen partial pressure, and saturation percentage of the whole blood samples were measured through a venous blood gas test, and the purified Hb samples were analyzed using several techniques including intrinsic fluorescence, circular dichroism and UV-vis absorption spectroscopies, SDS-polyacrylamide gel electrophoresis, dynamic light scattering, and a zeta potential analyzer. Structural and sequence analyses were also used to study the Hb heme pocket autoxidation sites and the residues involved. The results showed that the oligomerization and instability of Hb can be reduced if the ozone concentration to be used in MAH is divided into two doses. Indeed, our study demonstrated that two-step ozonation with 20, 30, and 40 μg/mL of ozone instead of single-dose ozonation with 40, 60, and 80 μg/mL of ozone reduced the potential adverse effects of ozone on Hb including protein instability and oligomerization. Moreover, it was found that for certain residues, their orientation or displacement leads to the entry of excess water molecules into the heme moiety, which can contribute to Hb autoxidation. Additionally, the autoxidation rate was found to be higher in alpha globins compared to beta globins.

The impact of pH and acid-activated proteases on the decrease of immunological properties of parvalbumin from freshwater fish

Fermentation has been found as an effective method to decrease the hypersensitive reactions of consumers by freshwater fish, but the reason is not clear. Therefore, the aim of this study was to investigate the impact of pH and acid-activated proteases on the decrease of immunological properties of parvalbumin from freshwater fish, by the indicators of SDS-polyacrylamide gel electrophoresis, western blotting, enzyme-linked immunosorbent assay, surface hydrophobicity and circular dichroism. The results showed that black carp (Mylopharyngodon piceus) had the highest level of immunological properties reflected by quantitation, the content of linear epitopes and conformational epitopes of parvalbumin, and IgE-binding rate. Afterwards, purified parvalbumin from black carp was obtained with purity higher than 95%. Then, for acidification and enzyme models, it indicated that acid-activated proteases were the reason for the decrease of immunological properties. Ulteriorly, the mechanism on the level of protein structure was provided, which showed the decrease of α-helix and the increase of β-turn and random coil, as well as the decrease of surface hydrophobicity had joint contribution for the breaking of conformational epitopes. This study could provide a novel guidance for the decrease of immunological properties of fish and promote fish processing industry further.

Comparison of Synthetic Methods and Identification of Several Artificial Antigens of Deoxynivalenol.

The purpose of this experiment was to study the design and modification of hapten molecules and artificial antigen molecules of deoxynivalenol (DON), and to compare the preparation and identification methods of four artificial antigens. According to the characteristics of the molecular structure of DON, four artificial antigen coupling methods were designed-namely, N,N'-carbonyldiimidazole (CDI), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), isobutyl chloroformate (IBCF), and N-hydroxysuccinimide (NHS)-to prepare artificial antigens and detection antigens. Through ultraviolet (UV), infrared (IR), and SDS-polyacrylamide gel electrophoresis (SDS-PAGE), along with other physical and chemical identification methods and animal immunisation, the best artificial antigen coupling method was screened. The results showed that the CDI method achieved the best effect among the synthesis methods. The titre of anti-DON polyclonal antibody (pAb) produced by animal immunisation reached 1: (6.4 × 103). The half inhibitory concentration (IC50) was 47.75 ng/mL, the cross-reaction rate with 3-acetyldeoxynivalenol (3-AcDON) was slightly higher at 35.3%, and there was no cross-reaction with other compounds; therefore, four artificial antigens were successfully prepared by using the molecular structure of DON. Through identification, the CDI method was screened as the best artificial antigen synthesis method, with the highest DON pAb titre, the best sensitivity, and the strongest specificity. This will lay a solid antigenic foundation for the preparation of better anti-DON monoclonal antibodies (mAbs) in the future.

Study on the Structure, Function, and Interface Characteristics of Soybean Protein Isolate by Industrial Phosphorylation

The impacts of industrial phosphorylation on the structural changes, microstructure, functional, and rheological features of soybean protein isolate (SPI) were spotlighted. The findings implied that the spatial structure and functional features of the SPI changed significantly after treatment with the two phosphates. Sodium hexametaphosphate (SHMP) promoted aggregation of SPI with a larger particle size; sodium tripolyphosphate (STP) modified SPI with smaller particle size. SDS-polyacrylamide gel electrophoresis (SDS-PAGE) results showed insignificant alterations in the structure of SPI subunits. Fourier transform infrared (FTIR) and endogenous fluorescence noted a decline in α-helix quantity, an amplification in β-fold quantity, and an increase in protein stretching and disorder, indicating that phosphorylation treatment fluctuated the spatial structure of the SPI. Functional characterization studies showed that the solubility and emulsion properties of the SPI increased to varying degrees after phosphorylation, with a maximum solubility of 94.64% for SHMP-SPI and 97.09% for STP-SPI. Emulsifying activity index (EAI) and emulsifying steadiness index (ESI) results for STP-SPI were better than those for SHMP-SPI. Rheological results showed that the modulus of G' and G″ increased and the emulsion exhibited significant elastic behavior. This affords a theoretical core for expanding the industrial production applications of soybean isolates in the food and various industries.