Exclusion High Performance Liquid Chromatography Research Articles

The anti-angiogenic, anti-inflammatory and anticoagulant potential of a polysaccharide extracted from the brown alga Cystoseira humilis

The present study aimed to evaluate the properties of a sulfated polysaccharide (PSCH) extracted from the brown alga Cystoseira humilis. We analyzed the chemical composition, anti-angiogenesis activities, anticoagulant, and anti-inflammatory effects of the PSCH, using various techniques encompassing UV–visible spectroscopy, fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (X-RD), scanning electron microscopy (SEM), steric exclusion chromatography (SEC) and high-performance liquid chromatography coupled to refractive index detector (HPLC-RID). The study found that PSCH was a sulfated heteropolysaccharide consisting of glucuronic acid, arabinose, galacturonic acid, xylose and fructose, and has a semi-crystalline nature with a molecular weight of about 379.44 kDa. The PSCH contains 41.77 % carbohydrates, 12.33 % proteins, and 4.59 % sulfates. The PSCH fraction has anti-angiogenic potential highlighted through assays on the chorioallantoic membrane, where vascular branching was inhibited by about 41.65 % and 61.38 % at concentrations of 50 and 100 μg/egg, respectively, compared to the untreated group. PSCH displayed an anti-edematous effect in vivo by reducing malondialdehyde and advanced oxidation protein products in paw tissue after five hours of carrageenan injection. When rats were co-treated with PSCH, there was evidence of a significant increase in antioxidant enzyme activity (superoxide dismutase, glutathione peroxidase, and glutathione) in edematous paw tissues compared to untreated rats. Furthermore, PSCH significantly influenced hematological parameters and the rate of anti-inflammatory serum protein fractions compared to the inflamed group. Interestingly, PSCH may exhibit anticoagulant properties by extending activated partial thrombosis time, thrombin time and prothrombin time. The present study has shown that Cystoseira humilis polysaccharides have antioxidant, anti-angiogenic, anti-inflammatory, and anti-coagulant properties, making them suitable for therapeutic purposes.

A Novel Tetravalent CD95/Fas Fusion Protein With Superior CD95L/FasL Antagonism.

Inhibition of CD95/Fas activation is currently under clinical investigation as a therapy for glioblastoma multiforme and preclinical studies suggest that disruption of the CD95-CD95L interaction could also be a strategy to treat inflammatory and neurodegenerative disorders. Besides neutralizing anti-CD95L/FasL antibodies, mainly CD95ed-Fc, a dimeric Fc fusion protein of the extracellular domain of CD95 (CD95ed), is used to prevent CD95 activation. In view of the fact that full CD95 activation requires CD95L-induced CD95 trimerization and clustering of the resulting liganded CD95 trimers, we investigated whether fusion proteins of the extracellular domain of CD95 with a higher valency than CD95ed-Fc have an improved CD95L-neutralization capacity. We evaluated an IgG1(N297A)-based tetravalent CD95ed fusion protein which was obtained by replacing the variable domains of IgG1(N297A) with CD95ed (CD95ed-IgG1(N297A)) and a hexavalent variant obtained by fusion of CD95ed with a TNC-Fc(DANA) scaffold (CD95ed-TNC-Fc(DANA)) promoting hexamerization. The established N297A and DANA mutations were used to minimize FcγR binding of the constructs under maintenance of neonatal Fc receptor (FcRn) binding. Size exclusion high-performance liquid chromatography indicated effective assembly of CD95ed-IgG1(N297A). More important, CD95ed-IgG1(N297A) was much more efficient than CD95ed-Fc in protecting cells from cell death induction by human and murine CD95L. Surprisingly, despite its hexavalent structure, CD95ed-TNC-Fc(DANA) displayed an at best minor improvement of the capacity to neutralize CD95L suggesting that besides valency, other factors, such as spatial organization and agility of the CD95ed domains, play also a role in neutralization of CD95L trimers by CD95ed fusion proteins. More studies are now required to evaluate the superior CD95L-neutralizing capacity of CD95ed-IgG1(N297A) in vivo.

Effect of carboxymethyl chitosan on the storage stability of rice dough during frozen storage

In this study, we aimed to determine the effect of carboxymethyl chitosan (CMCh) and carboxymethyl cellulose sodium (CMCNa) on the quality of frozen rice dough. We used a variety of methods to conduct a thorough investigation of frozen rice dough, including nuclear magnetic resonance (NMR), scanning electron microscopy (SEM), Fourier transform infrared (FT-IR) spectroscopy, size exclusion high-performance liquid chromatography (SE-HPLC), X-ray diffraction (X-RD), differential scanning calorimetry (DSC), and rapid visco analyzer (RVA). Our findings showed that frozen storage caused significant damage to the texture of rice dough, and this damage was reduced by the inclusion of CMCh, which led to a gradual change in the orderly structure of proteins. The degree of cross-linking between CMCh-B (DS:1; 0.5 %, 1 %, and 1.5 %) and the large protein polymer was significantly higher than that between CMCh-A (DS:0.8; 0.5 %, 1 %, and 1.5 %) and CMCNa (DS:1; 1 %), which decreased the ability of bound water to become free water. This resulted in the increase of tan δ, which effectively delayed the structural transformation of frozen rice dough. Furthermore, the introduction of CMCh delayed the immediate order of starch and crystal structure modifications, altering the thermal properties and pasting qualities of the frozen rice dough. Therefore, 1.5 % CMCh-B showed the best protective effect on frozen rice dough.

Purification, and characterization of a new pro-coagulant protein from Iranian Echis carinatus venom

This work aimed to purify the proteins that cause blood coagulation in the venom of the Iranian Echis carinatus snake species in a comprehensive manner. Gel filtration chromatography (GFC), Ion exchange chromatography (IEC), and Size Exclusion High-Performance Liquid Chromatography (SEC-HPLC) were utilized in the purification of the coagulation factors. The prothrombin clotting time (PRCT) and SDS-PAGE electrophoresis were performed to confirm the coagulative fractions. The fraction with the shortest coagulation time was selected. The components of this designated fraction were identified through matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF) following thorough purification. Circular dichroism (CD) was employed to determine the second structure of the coagulation factor. The crude venom (CV) was analyzed and had a total protein concentration of 97%.Furthermore, the PRCT of the crude venom solution at a concentration of 1 mg/ml was determined to be 24.19 ± 1.05 s. The dosage administered was found to be a factor in the venom's capacity to induce hemolysis. According to CD analysis, the protein under investigation had a helical structure of 16.7%, a beta structure of 41%, and a turn structure of 9.8%. CHNS proved that the purified coagulant protein had a Carbon content of 77.82%, 5.66% Hydrogen, 3.19% Nitrogen, and 0.49% Sulphur. In the present investigation, a particular type of snake venom metalloproteinase (SVMP) has undergone the process of purification and characterization and has been designated as EC-124. This purified fraction shows significant efficacy as a procoagulant. Our findings have shown that this compound has a function similar to factor X and most likely it can cause blood coagulation by activating factor II (FII).

Effect of different inactivants and preservatives on the stability of 146S fraction of foot-and-mouth diseases virus.

Foot-and-mouth disease virus (FMDV) cripples livestock by imparting devastating effects to economy. A good vaccine is the key to stopping it, but due to instability of 146S of FMDV, it is becoming difficult. This is bad because only 146S can fight against disease and its dissociation ultimately leads to decreased potency of vaccine. This study aimed to preserve the integrity of 146S in vaccine using different inactivators and preservatives. Foot-and-mouth Disease virus type 'O' was propagated on baby hamster kidney 21 cell lines and inactivated using formalin or binary ethylenimine (BEI). Size exclusion high performance liquid chromatography analysis revealed minimal 146S loss after double inactivation with formalin and BEI. This inactivated virus was further formulated into oil-based vaccine with sodium thiomersal or chloroform as a preservative. Our findings demonstrated that chloroform outperformed thiomersal in maintaining shelf life of vaccine. This claims that the combined approach of double inactivation with formalin and BEI followed by chloroform as preservative offered a promising strategy for developing efficacious FMDV.

Alleviative effects of carboxymethyl chitosan on the quality deterioration of frozen rice dough during freeze thaw cycles

The objective of this study was to examine the impact of carboxymethyl chitosan (CMCh) on rice dough quality during freeze–thaw cycles (FTCs). We conducted a comprehensive analysis of frozen rice dough using various techniques, including scanning electron microscopy (SEM), low–field nuclear magnetic resonance (LF–NMR), confocal laser scanning microscopy (CLSM), Fourier transform infrared spectroscopy (FT–IR), size–exclusion high–performance liquid chromatography (SE–HPLC), small–angle X–ray scattering (SAXS), X–ray diffraction (X–RD), differential scanning calorimetry (DSC), and rapid visco–analysis (RVA). Our results revealed that adding carboxymethyl chitosan (CMCh) to frozen rice dough reduced damage to starch granules and gluten proteins during FTCs compared to dough without CMCh. This resulted in the deceleration of the modification of the ordered protein structure. Furthermore, the inclusion of CMCh resulted in increased cross–linking with the large protein polymer, impairing the capacity of bound water to transition into free water within the frozen rice dough. Consequently, this led to an elevation in the storage modulus (G′), effectively retarding the textural transformation of the frozen rice dough. Introducing CMCh to frozen rice dough subsequently decelerated the alterations in the short-range order and crystal structures of starch, consequently modifying the pasting and thermal properties of the frozen rice dough. Based on the results obtained from our experiments, CMCh has a decelerating effect on the impact of frozen rice dough on FTCs.

Peptide NCTX15 derived from spider toxin gland effectively relieves hyperuricemia in mice

Hyperuricemia is a clinical disease characterized by a continuous increase in uric acid (UA) due to purine metabolism disorder. As current drug treatments are limited, it is imperative to explore new drugs that offer better safety and efficacy. In this study, Nephila clavata toxin gland homogenates were isolated and purified by exclusion chromatography and high-performance liquid chromatography, resulting in the identification and isolation of a short peptide (NCTX15) with the sequence ‘QSGHTFK’. Analysis showed that NCTX15 exhibited no cytotoxicity in mouse macrophages or toxic and hemolytic activity in mice. Notably, NCTX15 inhibited UA production by down-regulating urate transporter 1 and glucose transporter 9 and up-regulating organic anion transporter 1, thus promoting UA excretion. In addition, NCTX15 alleviated the inflammatory response and renal injury by inhibiting the expression of inflammatory factors interleukin-6, interleukin-1β, tumor necrosis factor alpha, NLR family, pyrin domain-containing 3, and pyroptosis-related factor gasdermin D. These results indicate that NCTX15 displayed urate-lowering, anti-inflammatory, and analgesic effects. As the first urate-reducing short peptide isolated from a spider toxin gland homogenate, NCTX15 exhibits considerable potential as a novel drug molecule for anti-gout and hyperuricemia treatment.

Insights into zein on rheological, structural, thermal properties of wheat dough under glutathione and azodicarbonamide, salt, or acidic conditions

Zein exhibited viscoelastic properties similar to those of wheat dough under appropriate conditions. To improve the characteristics of zein in wheat dough, wheat doughs with added zein (3%) were prepared under redox, salt, and acidic conditions. The incorporation of glutathione (GSH) into the zein-added dough (MGFZ) resulted in reduced viscoelasticity and increased creep strain, corresponding to a more relaxed water distribution, similar to what has been reported in wheat dough. In contrast, the salt-added dough exhibited the opposite trend, with a recovery rate reaching 70.11%. Azodicarbonamide (ADA) and acidic conditions showed a tendency to increase the viscoelasticity of MGFZ. Size exclusion high-performance liquid chromatography (SE-HPLC) demonstrated that ADA and salt induced protein aggregation. Moreover, ADA and salt induced a 2.37% and 7.47% decrease in weight loss, respectively, as detected by thermogravimetric analysis (TGA) when compared with MGFZ. The study provided insights into the effects of redox agents, salt, and acidic environments on zein-enriched wheat dough, potentially enhancing the applicability of zein in wheat-based products.

Soluble-protein-aggregate-assisted improvements in heat-induced gel properties: Effect of genipin-mediated crosslinks on egg white protein

Genipin, a covalent crosslinker, has attracted significant attention from the food industry. Herein, the color, heat-induced gelling properties, and soluble aggregate character of the genipin-treated egg white were investigated to clarify the impact of soluble protein aggregates on the gel properties of egg white protein.After genipin pretreatment, the egg white protein turned dark blue and retained this color after heat-induced gelling. The genipin-pretreated egg white protein gel exhibited up to 2.9-fold and 13% higher breaking strength and strain, respectively, compared to non-treated egg white gel. The gel-breaking strength and strain correlated well with the color, before and after gelling. Gel electrophoresis, size exclusion, and anion exchange high-performance liquid chromatography revealed that a soluble protein aggregate with a negatively charged surface formed during pretreatment. The genipin-treated egg white protein formed a finely structured gel with high breaking strength and strain at all tested NaCl concentrations, indicating that the enhanced negative charge of the surface did not contribute to improving the gel properties.The soluble protein aggregates formed via covalent crosslinking are important for improving gel properties. The proposed method is a novel environmentally friendly method for enhancing the gel properties of egg white protein.

Stability of Opened Durvalumab (IMFINZI) Vials. The Beginning of the End of Costly Product Wastage?

Durvalumab is a monoclonal antibody approved for the treatment of lung, urothelial and biliary tract cancers. Durvalumab is supplied in vials as a solution containing no preservatives. Monographs recommend single use of durvalumab vials, and that any leftovers be discarded within 24 h. Thus, significant portions of unused product from opened vials are wasted on a daily basis, generating considerable financial losses. The objective of the present study was to assess the physicochemical and microbiological stability of durvalumab vials kept at 4 °C or room temperature, at 7 and 14 days after opening. Following pH and osmolality measurements, turbidity and submicronic aggregation of durvalumab solution were evaluated by spectrophotometry and dynamic light scattering, respectively. Moreover, steric exclusion high performance liquid chromatography (SE-HPLC), ion exchange HPLC (IEX-HPLC) and peptide mapping HPLC were used to respectively assess aggregation/fragmentation, charge distribution and primary structure of durvalumab. Microbiological stability of durvalumab was evaluated by incubation of vial leftovers on blood agar. All experiments showed physicochemical and microbiological stability of durvalumab vial leftovers for at least 14 days when aseptically handled and kept at either 4 °C or at room temperature. These results suggest the possible extension of utilization of durvalumab vial leftovers well beyond 24 h.

Behavior of protein aggregates via electrostatic interactions or hydrogen bonds during dough formation

The aggregation behavior of gluten proteins via electrostatic interactions or hydrogen bonds during dough formation was investigated. Proteins were extracted with 50 mM acetic acid or 4 M urea to break electrostatic interactions or hydrogen bonds, respectively. The extracted proteins were analyzed by size exclusion high performance liquid chromatography and two dimensional fluorescence difference gel electrophoresis. Acetic acid- and urea-soluble protein aggregates increased with dough development, indicating that electrostatic interactions and hydrogen bonds weakened during dough mixing. On the other hand, the some monomeric proteins (mainly α-gliadins) in dough aggregated via electrostatic interactions rather than hydrogen bonds. A model is proposed for gluten formation during dough mixing based on non-covalent bonds.

Efficient quenching sheds light on early stages of gold nanoparticle formation.

The formation mechanism of plasmonic gold nanoparticles (Au NPs) by fast NaBH4 induced reduction of the precursors is still under debate. In this work we introduce a simple method to access intermediate species of Au NPs by quenching the solid formation process at desired time periods. In this way, we take advantage of the covalent binding of glutathione on Au NPs to stop their growth. By applying a plethora of precise particle characterization techniques, we shed new light on the early stages of particle formation. The results of in situ UV/vis measurements, ex situ sedimentation coefficient analysis by analytical ultracentrifugation, size exclusion high performance liquid chromatography, electrospray ionization mass spectrometry supported by mobility classification and scanning transmission electron microscopy suggest an initial rapid formation of small non-plasmonic Au clusters with Au10 as the main species followed by their growth to plasmonic Au NPs by agglomeration. The fast reduction of gold salts by NaBH4 depends on mixing which is hard to control during the scale-up of batch processes. Thus, we transferred the Au NP synthesis to a continuous flow process with improved mixing. We observed that the mean volume particle sizes and the width of the particle size distribution decrease with increasing flow rate and thus higher energy input. Mixing- and reaction-controlled regimes are identified.

Striving for Stability in the Dough Mixing Quality of Spring Wheat under the Influence of Prolonged Heat and Drought.

The effects of prolonged heat and drought stress and cool growing conditions on dough mixing quality traits of spring wheat (Triticum aestivum L.) were studied in fifty-six genotypes grown in 2017 and 2018 in southern Sweden. The mixing parameters evaluated by mixograph and the gluten protein characteristics studied by size exclusion high-performance liquid chromatography (SE-HPLC) in dough were compared between the two growing seasons which were very different in length, temperature and precipitation. The genotypes varying in gluten strength between the growing seasons (≤5%, ≤12%, and ≤17%) from three groups (stable (S), moderately stable (MS), and of varying stability (VS)) were studied. The results indicate that most of the mixing parameters were more strongly impacted by the interaction between the group, genotype, and year than by their individual contribution. The excessive prolonged heat and drought did not impact the buildup and mixing time expressed as peak time and time 1–2. The gluten polymeric proteins (unextractable, %UPP; total unextractable, TOTU) and large unextractable monomeric proteins (%LUMP) were closely associated with buildup and water absorption in dough. Major significant differences were found in the dough mixing parameters between the years within each group. In Groups S and MS, the majority of genotypes showed the smallest variation in the dough mixing parameters responsible for the gluten strength and dough development between the years. The mixing parameters such as time 1–2, buildup, and peak time (which were not affected by prolonged heat and drought stress) together with the selected gluten protein parameters (%UPP, TOTU, and %LUMP) are essential components to be used in future screening of dough mixing quality in wheat in severe growing environments.

Pilot scale production and characterization of next generation high molecular weight and tense quaternary state polymerized human hemoglobin.

Polymerized human hemoglobin (PolyhHb) is being studied as a possible red blood cell (RBC) substitute for use in scenarios where blood is not available. While the oxygen (O2 ) carrying capacity of PolyhHb makes it appealing as an O2 therapeutic, the commercial PolyhHb PolyHeme® (Northfield Laboratories Inc.) was never approved for clinical use due to the presence of large quantities of low molecular weight (LMW) polymerichemoglobin (Hb) species (<500 kDa), which have been shown to elicit vasoconstriction, systemic hypertension, and oxidative tissue injury in vivo. Previous bench-top scale studies in our lab demonstrated the ability to synthesize and purify PolyhHb using a two-stage tangential flow filtrationpurification process to remove almost all undesirable Hb species (>0.2 µm and <500 kDa) in the material, to create a product that should be safer for transfusion. Therefore, to enable future large animal studies and eventual human clinical trials, PolyhHb synthesis and purification processes need to be scaled up to the pilot scale. Hence in this study, we describe the pilot scale synthesis and purification of PolyhHb. Characterization of pilot scale PolyhHb showed that PolyhHb could be successfully produced to yield biophysical properties conducive for its use as an RBC substitute. Size exclusion high performance liquid chromatography showed that pilot scale PolyhHb yielded a high molecular weight Hb polymer containing a small percentage of LMW Hb species (<500 kDa). Additionally, the auto-oxidation rate of pilot scale PolyhHb was even lower than that of previous generations of PolyhHb. Taken together, these results demonstrate that PolyhHb has the ability to be seamlessly manufactured at the pilot scale to enable future large animal studies and clinical trials.

Freeze-Dried Biopharmaceutical Formulations are Surprisingly Less Stable than Liquid Formulations during Dropping.

This article describes an interesting phenomenon in which optimized freeze-dried (FD) biopharmaceutical formulations are generally more prone to degradation than their liquid counterparts during dropping and proposes an underlying cause for this surprising phenomenon. Two monoclonal antibodies (mAbs) and a fusion protein (FP) were used as model biopharmaceuticals. The stability after dropping stress was determined by ultraviolet-visible (UV-Vis), size exclusion high-performance liquid chromatography (SE-HPLC), micro-flow imaging (MFI), and dynamic light scattering (DLS). Contrary to what we would normally assume, the FD formulations of the three biopharmaceuticals studied here generally showed much higher amounts of protein sub-visible particles (SbVPs) than liquid formulations after applying the same dropping stress as determined by MFI and DLS. Traditional techniques, such as UV-Vis and SE-HPLC, could hardly detect such degradation. We propose that the higher temperature caused by dropping for the FD powders than the liquid formulations was probably one of the root causes for the higher amount of particles formed for the FD powders. We also recommend that dropping stress should be included for early-stage screening and choosing liquid versus FD biopharmaceutical formulations.

Structural Characterization of RC28-E, a Recombinant Fusion Protein With Dual Targets on VEGF and FGF2

Vascular endothelial growth factor (VEGF) and fibroblast growthfactor (FGF) play important roles in angiogenesis-related diseases. RC28-E is a soluble fusion protein composed of the human VEGF receptor 1 (VEGFR1) extracellular domain 2 (ECD 2), VEGFR2 ECD 3, FGFR1 ECDs 2 and 3, and the Fc regions of human immunoglobulin G1. By targeting both VEGF and FGF2, RC28-E may represent a useful antiangiogenetic agent, but structural and functional characterizations of this fusion protein are needed. Liquid chromatography–tandem mass spectrometry, size exclusion high-performance liquid chromatography, capillary electrophoresis-sodium dodecyl sulfate, imaged capillary isoelectric focusing, and bio-layer interferometry were used to characterize the properties of RC28-E. The purity of RC28-E was confirmed to be 98% or greater. The glycosylation modification of RC28-E was found to be very complicated, with 11 potential N-linked glycosylation points and 23 types of N-glycans, causing high heterogeneity of the protein. The primary modifications of the amino acid sequence of RC28-E protein included C-terminal K truncation, N-deamidation, and M-oxidation modification. Notably, RC28-E demonstrated a higher affinity for both VEGF and FGF2 than VEGF trap or FGF trap for their respective targets.

Partial depolymerization of capsular polysaccharides isolated from Streptococcus pneumoniae serotype 2 by various methods

Partial depolymerization of bacterial capsular polysaccharides (CPS) is an essential process carried out before its use as an antigenic preparation in a vaccine industry. Choice of CPS depolymerization methods depends on the process robustness, reproducibility, yield, retention of CPS bioactivity, etc. Partial depolymerization methods based on chemicals, enzymes, mechanical, thermal, etc. have been subject of many investigations before. Partial depolymerization of Streptococcus pneumoniae serotype 2 purified CPS was conducted by methods such as acid hydrolysis, microfluidization, ultrasonication, thermal and microwave. Partial depolymerization of the CPS was evaluated by size exclusion high performance liquid chromatography, whereas structural identity and conformity of CPS was ensured by 1H NMR spectroscopy. The antigenicity of CPS was assessed by bead based competitive inhibition assay. Microwave and thermal methods effectively depolymerized CPS and reduced the concentration of cell wall polysaccharide (CWPS) impurity, but both methods have a negative impact on the antigenicity of CPS. Whereas the trifluoroacetic acid treatment not only depolymerized the CPS but completely removed the CWPS while retaining the antigenicity of 92 ± 4% and this method is advantageous over other methods.

First insight in trace element distribution in the intestinal cytosol of two freshwater fish species challenged with moderate environmental contamination

Cytosolic distribution of six essential elements and nonessential Cd among biomolecules of different molecular masses was investigated in the intestine of brown trout (Salmo trutta) from the karst Krka River and Prussian carp (Carassius gibelio) from the lowland Ilova River. Fish were sampled at two locations (reference and contaminated) and in two seasons (autumn and spring). Analyses were conducted by size exclusion high performance liquid chromatography and high resolution inductively coupled plasma mass spectrometry. Although studied salmonid and cyprinid fish have different biological characteristics, obtained profiles often showed mostly similar patterns in both species. Specifically, Cd and Cu were dominantly bound to metallothioneins in both species, but the same association was not observed for Zn, whereas Mo distribution was similar in the intestine of both fish species with two well shaped and clear peaks in HMM (100–400 kDa) and VLMM (2–8 kDa) range. In brown trout, Se was mostly associated with biomolecules of very low molecular masses (VLMM, <10 kDa), whereas significant additional elution in HMM region (30–303 kDa) was observed only in Prussian carp. Iron binding to VLMM biomolecules (1.8–14 kDa) was observed only in brown trouts, and of Zn in Prussian carps. Cobalt was mostly bound to HMM biomolecules (85–235 kDa) in brown trout and to VLMM biomolecules (0.7–18 kDa) in Prussian carp. Comparison of intestinal profiles with previously published data on liver and gills revealed some similarities in distribution, but also organ-specific differences due to the different function and composition of each organ. As so far there is no published data on intestinal trace metal distribution, the obtained results represent the novel findings, and the key point for the exact identification of specific metal-binding biomolecules which could eventually be used as biomarkers of metal exposure or effects.

Solvent Retention Capacity and Gluten Protein Composition of Durum Wheat Flour as Influenced by Drought and Heat Stress.

Drought and temperature stress can cause considerable gluten protein accumulation changes during grain-filling, resulting in variations in wheat quality. The contribution of functional polymeric components of flour to its overall functionality and quality can be measured using solvent retention capacity (SRC). The aim of this study was to determine the effect of moderate and severe drought and heat stress on SRC and swelling index of glutenin (SIG) in six durum wheat cultivars with the same glutenin subunit composition and its relation with gluten protein fractions from size exclusion high performance liquid chromatography. Distilled water, sodium carbonate and sucrose SRC reacted similarly to stress conditions, with moderate heat causing the lowest values. Lactic acid SRC and SIG reacted similarly, where severe heat stress highly significantly increased the values. SIG was significantly correlated with sodium dodecyl sulphate sedimentation (SDSS) and flour protein content (FPC) under all conditions. Lactic acid SRC was highly correlated with FPC under optimal and moderate heat stress and with SDSS under moderate drought and severe heat. SIG was negatively correlated with low molecular weight glutenins under optimal and drought conditions, and combined for all treatments. The relationship between SRC and gluten proteins was inconsistent under different stress conditions.

Innovatively processed quinoa (Chenopodium quinoa Willd.) food: chemistry, structure and end-use characteristics.

Quinoa (Chenopodium quinoa Willd.) flour and processed traditional Peruvian quinoa breakfast foods were studied to evaluate the effect of extrusion and post-processing on protein properties, morphology and nutritional characteristics (amino acids and dietary fibers). The extrusion increased quinoa protein crosslinking and aggregation observed by size exclusion high-performance liquid chromatography and the amount of soluble fibers, as well as decreasing the amounts of insoluble fibers in the processed foods. The post-processing drying resulted in additional crosslinking of large protein fractions in the quinoa products. The microstructure of the extruded quinoa breakfast flakes and heat-post-processed samples studied by scanning electron microscopy and X-ray tomography differed greatly; post-drying induced formation of aerated protein microstructures in the heat-treated samples. Nanostructures revealed by small-angle and wide-angle X-ray scattering indicated that extrusion imparted morphological changes in the quinoa protein and starch (dominance of V-type). Overall, extrusion processing only reduced the content of most of the essential amino acids to a minor extent; the content of valine and methionine was reduced to a slightly greater extent, but the final products met the requirements of the Food and Drug Organization. This study presents innovative examples on how extrusion processing and post-processing heat treatment can be used to produce attractive future food alternatives, such as breakfast cereal flakes and porridge powder, from quinoa grains. Extrusion of quinoa flour into Peruvian foods was shown to be mostly impacted by the processing temperature and processing conditions used. Protein crosslinking increased due to extrusion and post-processing heating. Starch crystallinity decreased most when the product was dried after processing. © 2021 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.