Zein Protein Research Articles

High internal phase pickering emulsions stabilized by zein/whey protein nanofibril complexes: Preparation and lycopene loading

High internal phase Pickering emulsions (HIPPEs) prepared from natural polymers have attracted much attention in the food manufactures. However, single zein-stabilized HIPPEs are poorly stable and prone to flocculation near the isoelectric point. To address this issue, in this study, zein and whey protein nanofibrils (WPN) complex nanoparticles (ZWNPs) were successfully prepared using a pH-driven method, and ZWNPs were further used as HIPPEs stabilizers. The results showed that zein and WPN were combined together through hydrogen bonding and hydrophobic interaction to form ZWNPs, and the HIPPEs stabilized by ZWNPs had excellent stability, which could effectively protect the internally encapsulated lycopene and improve the bioaccessibility of lycopene. In conclusion, this study provides a new strategy for the preparation of stable hydrophobic protein-based HIPPEs, represented by zein.

Rutin-loaded zein gel as a green biocompatible formulation for wound healing application

Wound healing is a challenging clinical problem and efficient wound management is essential to prevent infection. This is best done by utilizing biocompatible materials in order to complete the healing in a rapid manner, with functional and esthetic outcomes. In this context, the zein protein fulfills the criteria of the ideal wound dressing which include non-toxicity and non-inflammatory stimulation. Zein gels containing rutin were prepared without any chemical refinement or addition of gelling agents in order to obtain a natural formulation characterized by antioxidant and anti-inflammatory properties to be proposed for the treatment of burns and sores. In vitro scratch assay showed that the proposed gel formulations promoted cell migration and a rapid gap closure within 24 h (~90 %). In addition, the in vivo activities of rutin-loaded zein gel showed a greater therapeutic efficacy in Wistar rats, with a decrease of the wound area of about 90 % at day 10 with respect to the free form of the bioactive and to DuoDERM®. The evaluation of various markers (TNF-α, IL-1β, IL-6, IL-10) confirmed the anti-inflammatory effect of the proposed formulation. The results illustrate the feasibility of exploiting the peculiar features of rutin-loaded zein gels for wound-healing purposes.

Enhancing plant-based cheese formulation through molecular docking and dynamic simulation of tocopherol and retinol complexes with zein, soy and almond proteins via SVM-machine learning integration

The current study addresses the growing demand for sustainable plant-based cheese alternatives by employing molecular docking and deep learning algorithms to optimize protein-ligand interactions. Focusing on key proteins (zein, soy, and almond protein) along with tocopherol and retinol, the goal was to improve texture, nutritional value, and flavor characteristics via dynamic simulations. The findings demonstrated that the docking analysis presented high accuracy in predicting conformational changes. Flexible docking algorithms provided insights into dynamic interactions, while analysis of energetics revealed variations in binding strengths. Tocopherol exhibited stronger affinity (−5.8Kcal/mol) to zein compared to retinol (−4.1Kcal/mol). Molecular dynamics simulations offered comprehensive insights into stability and behavior over time. The integration of machine learning algorithms improved the classification and the prediction accuracy, achieving a rate of 71.59%. This study underscores the significance of molecular understanding in driving innovation in the plant-based cheese industry, facilitating the development of sustainable alternatives to traditional dairy products.

Tuning the properties of plant-based whipped cream through zein nanoparticles-sodium stearate complex

The fabrication of plant-based whipped creams is proposed to reduce the high saturated fat content of regular whipped creams by using plant proteins and unsaturated oils. The lack of functionality of regular plant proteins and the absence of crystals in vegetable oils make this process challenging. Here, we hypothesize that the functionality of corn protein zein can be improved by the creation of zein nanoparticles-sodium stearate complex (ZNP-SS), and that monoacylglyceride (MAG) based oleogels can increase the solid fat content of vegetable oils. Therefore, plant-based whipped cream can be produced from oil-in-water emulsions stabilized by ZNP-SS and MAG. A serial of ZNP-SS was produced and characterized in terms of molecular interactions, wettability, and surface activity. Moreover, their ability to stabilize oil-in-water emulsions and foams in the presence or absence of MAG was evaluated. Results show that ZNP-SS complex were stabilized through electrostatic and hydrophobic interactions. An increase in the amount of sodium stearate caused an increase in hydrophilicity, which improved the interfacial properties of ZNP-SS. Corn oil-in-water emulsions were successfully stabilized by all the ZNP-SS, while only the ones with the highest amount of sodium stearate (ZNP-0.6) were able to create stable whipped creams because of the partially coalesced oil droplets in the presence of MAG.

Fabrication and characterization of polydopamine-mediated zein-based nanoparticle for delivery of bioactive molecules

In this study, a combination of whey protein (hydrophilic coating) and polydopamine (crosslinking agent) was used to improve the stability and functionality of quercetin-loaded zein nanoparticles. There are two key benefits of the core-shell nanoparticles formed. First, the ability of the polydopamine to bind to both zein and whey protein facilitates the formation of a stable core-shell structure, thereby protecting quercetin from any pro-oxidants in the aqueous surroundings. Second, neutral and hydrophilic whey proteins were used for the surface coating of the nanoparticles to further enhance the sustained and slow release of quercetin, facilitating its sustained release into the body at a slow and steady rate. The results of this study will promote the innovative development of precise nutritional delivery systems for zein and provide a theoretical basis for the design and development of dietary supplements based on hydrophobic food nutrient molecules.

Zein—A plant protein as a promising biopolymer for biomedical applications: A perspective

Recent technological advances in the field of biomaterials and tissue engineering have spurred the interest in biopolymers for various biomedical applications. The advantage of biopolymers is their favorable characteristics for these applications, among which proteins are of particular importance. Proteins are explored widely for 3D bioprinting and tissue engineering applications, wound healing, drug delivery systems, implants etc. and the proteins mainly available include collagen, gelatin, albumin, zein, etc. Zein is a plant protein abundantly present in corn endosperm and it is about 80% of total corn protein. It is a highly renewable source and zein has been reported to be applicable in different industrial applications and of late it is gaining attention in biomedical applications. This research interest on zein is on account of its biocompatibility, non-toxicity and also its certain unique physico-chemical properties. Zein comes under GRAS category and is considered safe for biomedical applications. The hydrophobic nature of this protein imparts it an added advantage and has wider applications in drug delivery. This review focuses on details about zein protein, its properties and potential applications in biomedical sectors.

An in vitro evaluation of the biological activity of biogenic eggshell-derived nanosized bioglass, poly (ε-caprolactone), and zein protein 3D composite scaffolds for bone tissue engineering

An in vitro evaluation of the biological activity of biogenic eggshell-derived nanosized bioglass, poly (ε-caprolactone), and zein protein 3D composite scaffolds for bone tissue engineering

Urine Metabolite Profiles after the Consumption of a Low- and a High-Digestible Protein Meal, and Comparison of Urine Normalization Techniques.

In the context of dietary transition toward plant proteins, it is necessary to ensure protein security in populations. It would thus be of interest to identify biomarkers of altered protein digestibility in populations. We examined the association between urinary metabolites and the acute intake of low- or highly digestible protein in healthy volunteers. The urine samples were collected before and 9 h after the ingestion of a meal containing either no protein, zein (low-digestible) or whey protein isolate (highly digestible). The liquid chromatography-high resolution mass spectrometry metabolomics approach was used for the profiling of the urinary metabolites. For the standardization of metabolomics data sets, osmolality-based, standard normal variates (SNV) and probabilistic quotient normalization (PQN) techniques were used. The ANOVA-based factorial method, AComDim_ICA, was used for chemometrics analysis. The osmolality adjustment has a beneficial effect and the subsequent mathematical normalization improves the chemometric analysis further. Some changes in the urinary metabolomes were observed 9 h after the meal in the three groups. However, there was no difference in the urine metabolome between groups. No biomarker of protein digestibility can be identified after the ingestion of a single meal, even when marked differences in the digestion efficiency of protein have been observed.

Biomimetic nanocarriers loaded with temozolomide by cloaking brain-targeting peptides for targeting drug delivery system to promote anticancer effects in glioblastoma cells

Glioma is the leading cancer of the central nervous system (CNS). The efficacy of glioma treatment is significantly hindered by the presence of the blood-brain barrier (BBB) and blood-brain tumour barrier (BBTB), which prevent most drugs from entering the brain and tumours. Hence, we established a novel drug delivery nanosystem of brain tumour-targeting that could self-assemble the method using an amphiphilic Zein protein isolated from corn. Zein's amphiphilicity prompted it to self-assembled into NPs, efficiently containing TMZ. This allowed us to investigate temozolomide (TMZ) for glioblastoma (GBM) treatment. To construct TMZ-encapsulated NPs (TMZ@RVG-Zein NPs), the NPs' Zein was clocked to rabies virus glycoprotein 29 (RVG29). To verify that the NPs could penetrate the BBB and precisely target and kill the GBM cancer cell line, in vitro studies were performed. The process of NPs penetrating cancer cell membranes was investigated using enzyme-linked immunosorbent assays (ELISAs) to measure the expressions of nicotinic acetylcholine receptors (nAChRs) on the U87 cell line. Therefore, effective targeted brain cancer treatment is possible by forming NP clocks, a cell-penetrating natural Zein protein with an RVG29. These NPs can penetrate the blood-brain barrier (BBB) and enter the glioblastoma (U87) cell line to release TMZ. These NPs have a distinct cocktail of biocompatibility and brain-targeting abilities, making them ideal for involving brain diseases.

Synthesis, physiochemical characterization, molecular docking study, and anti-breast cancer activity of silymarin loaded zein nanoparticles

Breast cancer is a major cause of death in women worldwide leading to requirement of new therapeutic strategies. Silymarin demonstrated the anti-cancer activity however, due to low bioavailability its use is restricted. This study aimed to improve the solubility of silymarin by developing a silymarin loaded zein nanoparticles (SLNPs) which was stabilized by beta cyclodextrin. Comprehensive physiochemical characterization studies based on DLS, FTIR, UV–Vis Spectroscopy, FE-SEM, TEM, XRD, DSC, NMR and TGA confirmed the successful synthesis of SLNPs via an anti-solvent precipitation method. FE-SEM and TEM images demonstrated the uniform size and spherical shape of nanoparticles with encapsulation and loading efficiencies of 84.32 ± 1.9 % and 15.25 ± 2.4 % respectively. The zein protein interaction with silymarin, and β-cyclodextrin was shown to be beneficial via the use of molecular simulations and binding energy calculations. Cellular studies demonstrated dose and time dependent cytotoxicity of SLNPs on MCF-7 breast cancer cell. FACS, qRT-PCR and Western blotting showed Bax (pro-apoptotic) upregulation while Bcl-2 (anti-apoptotic) downregulation. Our findings suggest that these loaded nanoparticles are more efficient than pure drug, enhancing its bioavailability and paving the path for developing it as a promising nutraceutical to treat breast cancer.

Production of zein nanofibers using an electrospinning technique for the encapsulation of peppermint (Mentha piperita L.) essential oil

Peppermint is a perennial herb with high inhibitory power against bacteria because its chemical profile contains menthol and menthone. In this research, peppermint essential oil (PEO) was encapsulated in various levels (0, 2.5, 5, and 10% w/v) in nanofibers (NFs) obtained from zein using an electrospinning technique. SEM, XRD, TGA, and FTIR tests were performed to investigate the characteristics of NFs after producing them. Also, the resultant NFs were evaluated regarding encapsulation efficiency (EE) and antimicrobial properties via disc-diffusion against Escherichia coli, Staphylococcus aureus, Bacillus cereus, and Salmonella enterica. The results indicated that the morphology was tubular, uniform, and free bead. Fiber diameter increased on average by increasing the amount of PEO (p<0.05). XRD results showed that PEO was amorphous in electrospun zein NFs. TGA and FTIR analyses confirmed the presence of PEO in the electrospun fibers. In addition, the FTIR spectrum showed that the zein protein changed its secondary structure because of the PEO. The results of EE showed that an increase in PEO concentration caused a relevant rise in EE (P<0.05). The electrospun NFs containing PEO showed antimicrobial properties against the target bacteria. According to the current research, NFs obtained from zein via electrospinning can be applied suitably as a substrate to encapsulate PEO as a hydrophobic component.

Stretchable zein-coated alginate fiber for aligning muscle cells to artificially produce cultivated meat

Numerous studies have explored the cultivation of muscle cells using non-animal materials for cultivated meat production. Achieving muscle cell proliferation and alignment using 3D scaffolds made from plant-based materials remains challenging. This study introduces a technique to culture and align muscle cells using only plant-based materials, avoiding toxic chemical modifications. Zein-alginate fibers (ZA fibers) were fabricated by coating zein protein onto alginate fibers (A fibers). Zein’s excellent cell compatibility and biodegradability enable high cell adhesion and proliferation rates, and the good ductility of the ZA fibers enable a high strain rate (>75%). We demonstrate mature and aligned myotube formation in ZA fibers, providing a simple way to align muscle cells using plant-based materials. Additionally, cultivated meat was constructed by assembling muscle, fat, and vessel fibers. This method holds promise for the future mass production of cultivated meat.

Cell‐Membrane Inspired Multifunctional Nanocoating for Rescuing the Active‐Material Microenvironment in High‐Capacity Sulfur Cathode

AbstractAchieving healthy active‐material microenvironment (ME@AM) for stable and efficient electron/ion transport around each active‐material particle is crucial for high‐performance battery electrodes. However, this goal has been proved extremely challenging for most high‐capacity AMs such as sulfur, owing to its notable volume change and severe shuttle effect. Here, a multifunctional hybrid material with zein protein reinforced catalytic single Cu atom/carbon composite (Cu─C) (zein/Cu─C) is coated onto sulfur/carbon (SC) particle, to prepare the advanced zein/Cu─C@SC core–shell particle. Similar to the multifunctional cell membrane well‐known in biology, this multifunctional zein/Cu─C coating helps build a healthy and stable ME@AM within sulfur cathode. Specifically, it can simultaneously provide robust protective shell for the AM particle, adsorption and catalyst function to the dissolved polysulfides, and AM surface treatment to improve AM/conductive agent interface. All these functions are the keys to build and maintain a healthy ME@AM in sulfur cathode. This study not only brings effective solutions to the challenges in volume‐change high‐capacity sulfur active materials and beyond, but also helps achieve comprehensive understanding of the key factors controlling the structuring and final quality of ME@AM.

Effect of Recycling on Thermomechanical Properties of Zein and Soy Protein Isolate Bioplastics

Bioplastics are an alternative to reduce the environmental damage caused by petroleum-based plastics. However, the effect of primary recycling (reprocessing) of bioplastics from biomass resources has not yet been well studied. If successful, this would boost the landing of recyclable and biodegradable bio-based materials to the market. In order to meet the challenge of recycling bioplastics, it is necessary to study the reprocessing of bio-based materials that potentially behave as thermoplastics. This study investigated the primary recyclability of Zein- and soy protein isolate (SPI)-based bioplastics by reprocessing. Protein powders were initially mixed with glycerol (Gly), which acts as a plasticizer, and the blends were subjected to injection moulding. Initial specimens were reprocessed by injection moulding up to five times. The effect of reprocessing was evaluated by dynamic mechanical analysis (DMA), tensile test, and water uptake capacity (WUC). Finally, the property–structure relationship was assessed by scanning electron microscopy (SEM). The results showed that the recycled SPI-based bioplastics reduced elongation at break (i.e., ɛMax decreased from 0.8 to 0.3 mm/mm), whereas the parameters from tensile tests did not decrease upon recycling for Zein-based bioplastics (p &lt; 0.05). The results obtained confirm that it is possible to reprocess protein-based bioplastics from two different renewable sources while maintaining the mechanical properties, although the loss of Gly was reflected in tensile tests and WUC. These results highlight the possibility of replacing petroleum-based plastics with bio-based materials that can be recycled, which reduces dependence on natural biopolymers and contributes to sustainable development.

Cellulose Functionalization Using N-Heterocyclic-Based Leaving Group Chemistry.

There has been continuous interest in developing novel activators that facilitate the functionalization of cellulosic materials. In this paper, we developed a strategy in which trisubstituted triazinium salts act as cellulose preactivators. As leaving groups, these triazinium salts utilize N-heterocycles (pyridine, imidazole, and nicotinic acid). Initially, we optimized the synthetic route for developing these novel cellulose preactivators (triazinium salts), whose structures were confirmed using NMR spectroscopy. The surface zeta potential of cellulose changed from a negative value to a positive one after preactivation due to the cationic nature of these preactivators. To enhance the scope of the study, we functionalized the cellulose-preactivated materials with a series of amine- or hydroxy-containing aliphatic and aromatic hydrocarbons, nucleophilic amino acids (cysteine), colorants (2-aminoanthraquinone and 2-amino-3-methyl-anthraquinone), and biopolymer (zein protein). The treated samples were analyzed using FTIR, time-gated Raman spectroscopy, and reflection spectroscopy, and the success of the functionalization process was validated. To widen the scope of such chemistries, we synthesized four reactive agents containing N-heterocyclic-based leaving groups (pyridine and nicotinic acid) and successfully functionalized cellulose with them in one step. The proposed single- and two-step functionalization approaches will provide opportunities for chemically linking various chemical compounds to cellulose for different applications.

Breeding strategies for quality protein maize: A Review

Zein proteins in maize seeds regulate food and feed quality and belong to several subfamilies. Most maize prolamin is α-zeins, which are rich in glutamine, leucine, and proline but lacking in critical amino acids (lysine and tryptophan). RNAi experiments show that 22kD zeins downregulate the quality protein maize (QPM) phenotype, not 19kD. The opaque-2 mutation caused a chalky, soft endosperm and many defects. QPM genotypes with opaque-2 alleles and firm kernels were improved. As opaque-2 is recessive and biochemical lysine and tryptophan analysis is costly, conventional backcrossing alone is not very useful. Markerassisted selection (MAS) allowed top inbreds to be homozygous o2o2 utilizing opaque-2 gene-specific markers. Vivek QPM-9, a mixture of two QPM introgression lines, benefits various clients and customers. This review highlights the breeding strategies for QPM.

Plant-Derived Zein as an Alternative to Animal-Derived Gelatin for Use as a Tissue Engineering Scaffold.

Natural biomaterials are commonly used as tissue engineering scaffolds due to their biocompatibility and biodegradability. Plant-derived materials have also gained significant interest due to their abundance and as a sustainable resource. This study evaluates the corn-derived protein zein as a plant-derived substitute for animal-derived gelatin, which is widely used for its favorable cell adhesion properties. Limited studies exist evaluating pure zein for tissue engineering. Herein, fibrous zein scaffolds are evaluated in vitro for cell adhesion, growth, and infiltration into the scaffold in comparison to gelatin scaffolds and are further studied in a subcutaneous model in vivo. Human mesenchymal stem cells (MSCs) on zein scaffolds express focal adhesion kinase and integrins such as αvβ3, α4, and β1 similar to gelatin scaffolds. MSCs also infiltrate zein scaffolds with a greater penetration depth than cells on gelatin scaffolds. Cells loaded onto zein scaffolds in vivo show higher cell proliferation and CD31 expression, as an indicator of blood vessel formation. Findings also demonstrate the capability of zein scaffolds to maintain the multipotent capability of MSCs. Overall, findings demonstrate plant-derived zein may be a suitable alternative to the animalderived gelatin and demonstrates zein's potential as a scaffold for tissue engineering.

Physicochemical properties of plant-based cheese alternatives fortified with calcium

Fortification with calcium is proposed as an effective strategy to improve the nutritional profile of plant-based cheese alternatives (PBCAs). In this study, the effects of two calcium salts, individually or as a blend, on the physicochemical properties of PBCAs were investigated. Three formulations were obtained based on the calcium (Ca) salts employed, CaChloride, CaCitrate, and CaChlo:CaCitr, with the latter having a 50:50 contribution of calcium from calcium chloride and calcium citrate; a control with no added calcium salt was also analysed. Zein protein isolate and chickpea protein concentrate were used in combination with other ingredients to formulate the PBCAs. Different additions of lactic acid were necessary to adjust the pH to a target of ∼4.5 for all samples, due to the differing buffering capacity of calcium salts. Fortification with calcium led to decreased meltability, while stretching properties of calcium fortified samples were enhanced compared with that of the control. Texture profile analysis demonstrated that calcium fortification significantly affected the texture of samples. The results of this work will help in understanding the role of different calcium salts in formulating PBCAs with improved nutritional profile and the impact thereof on relevant physicochemical properties.

Nanofibers based on zein protein loaded with tungsten oxide for cancer therapy: fabrication, characterization and in vitro evaluation

Plant proteins have become attractive for biomedical applications such as wound dressing and drug delivery. In this research, nanofibers from pristine zein (plant protein) and zein loaded with tungsten oxide (WO3) were prepared (WO3@zein) using less toxic solvents (ethanol and acetic acid). Morphological and biological properties of the zein nanofiber were determined. Prepared nanofibers were defined by thermogravimetric analysis (TGA), X-ray diffraction (X-RD), Fourier-transform infrared spectroscopy (FT-IR), and scanning electron microscopy. The average fiber diameter was unchanged with an increase in WO3 concentration from 0.001 to 0.008%. FT-IR spectroscopy and X-RD indicated the presence of WO3 in WO3@zein nanofibers. In comparison to WO3-free, WO3@zein nanofibers showed higher safety and preserved the anticancer effect of WO3 against human melanoma cell line (A375) melanoma cells compared to WO3-free. Moreover, both WO3-free and WO3@zein caused a fourfold increase in the cellular proliferation of reactive oxygen species (ROS) in the treated A375 cells compared to untreated cells. ROS elevation led to apoptosis-dependent cell death of A375 cells as evidenced by up-regulating the expression of p53-downstream genes (p21 and Bax) (tumor-suppressor gene) while down-regulating the expression of key oncogenes (BCL2 and cyclin D). In conclusion, the prepared nanofiber represents a promising and safe candidate for anticancer applications.

Electrolyte-induced aggregation of zein protein nanoparticles in aqueous dispersions

Ion specific effects on the charging and aggregation features of zein nanoparticles (ZNP) were studied in aqueous suspensions by electrophoretic and time-resolved dynamic light scattering techniques. The influence of mono- and multivalent counterions on the colloidal stability was investigated for positively and negatively charged particles at pH values below and above the isoelectric point, respectively. The sequence of the destabilization power of monovalent salts followed the prediction of the indirect Hofmeister series for positively charged particles, while the direct Hofmeister series for negatively charged ones assumed a hydrophobic character for their surface. The multivalent ions destabilized the oppositely charged ZNPs more effectively and the aggregation process followed the Schulze-Hardy rule. For some multivalent ions, strong adsorption led to charge reversal resulting in restabilization of the suspensions. The experimental critical coagulation concentrations (CCCs) could be well-predicted with the theory developed by Derjaguin, Landau, Verwey and Overbeek indicating that the aggregation processes were mainly driven by electrical double layer repulsion and van der Waals attraction. The ion specific dependence of the CCCs is owing to the modification of the surface charge through ion adsorption at different extents. These results are crucial for drug delivery applications, where inorganic electrolytes are present in ZNP samples.