Oleogel-Based Nanoemulsions for Beverages: Effect of Self-Assembled Fibrillar Networks on Stability and Release Properties of Emulsions.

Reducing the use of stabilizers is one of the main challenges in food emulsions, especially for beverages. This work aimed to produce oleogel-structured nanoemulsions (NEs) without additional surfactants. Lecithin-stearic acid (LSa) and lecithin-sorbitan tristearate (LSt) oleogels formed stable NEs under optimized sonication conditions. Microscopy and rheometry revealed that the presence of self-assembled fibrous networks (SAFiNs) in both dispersed and continuous phases provided steric stabilization to NEs. Lecithin acted as crystal habit modifier of SAFiNs and facilitated their phase partitioning. Notably, the short fibers of LSt showed better emulsifying efficiency than the long fibers of LSa. Curcumin release studies under simulated gastrointestinal conditions demonstrated that SAFiNs affect the release capabilities of NEs. Polydispersity index, zeta potential and oil syneresis data showed that the emulsions are stable for six months. Moreover, NEs showed thermal stability upon curcumin release at 25 and 50 °C. These results suggest that the developed oleogel-based NEs are suitable for the delivery of bioactive agents for beverages and other food applications.

Carboxymethyl chitosan-N-alkylimine derivatives: Synthesis, characterization and use for preservation of symbiotic biofertilizer bacteria on chickpea seeds.

A series of carboxymethyl chitosan-N-alkylimine derivatives with side chain length of 4 to 10 carbons (CMCS-n, n = 4, 6, 8, 10) was prepared in a one-step solvent-free synthesis using Schiff base chemistry. The modified polysaccharides were characterized by their spectral, thermal and physical properties. The prepared polymers demonstrated an ability to spontaneous self-assembly with a clear correlation between critical aggregation concentration and the chain length of the alkyl substituent. N-alkylimine-CMCS derivatives were found to deliver hydrophobic (curcumin) and hydrophilic (ascorbic acid) active agents in unfavorable environments of water and oil, respectively. Then, N-alkylimine-CMCS derivatives were used as a platform for the delivery of symbiotic gram-positive bacteria Bacillus subtilis CJ onto chickpea seeds. These bacteria demonstrated a significantly higher survival rate (106 CFU/mL) in dried CMCS-6 derivative film than in other films tested. The seeds treated with N-alkylimine-CMCS coatings that contained B. subtilis CJ demonstrated up to 100-fold increase of this bacterial population on the seedlings in comparison to the pristine CMCS.

Nanogel Particles Based on Modified Nucleosides and Oligosaccharides as Advanced Delivery System.

This work addresses the challenge of delivering bioactive molecules by designing biocompatible nanogel particles (NGPs) utilizing rationally modified nature-sourced building blocks: capryl-oligochitosan and oxidized inosine. Capryl substituents endowed the resultant NGPs with membrane-penetration capabilities, while purine-containing inosine allowed H-bond/π-π/π-cation interactions. The prepared NGPs were complexed with carboxyfluorescein-labeled single-stranded oligonucleotide (FAM-oligo) and DsRed-encoding plasmid DNA. The successful delivery of FAM-oligo to the cell cytoplasm of the Nicotiana benthamiana plant was observed. Alexa 555-labeled bovine serum albumin (Alexa 555-BSA) was also efficiently encapsulated and delivered to the plant. In addition to delivering FAM-oligo and Alexa 555-BSA separately, NGPs also successfully co-delivered both biomolecules to the plant. Finally, NGPs successfully encapsulated the drug amphotericin B and reduced its toxicity while maintaining its efficacy. The presented findings suggest that NGPs may become a promising platform for the advanced delivery of bioactive molecules in various applications.

Carboxymethyl chitosan-quercetin conjugate: A sustainable one-step synthesis and use for food preservation.

Bioactive polysaccharide, carboxymethyl chitosan-quercetin (CMCS-q) was prepared by a one-step reaction utilizing Schiff base chemistry. Notably, the presented conjugation method involves neither radical reactions nor auxiliary coupling agents. Physicochemical properties and bioactivity of the modified polymer were studied and compared to those of the pristine carboxymethyl chitosan, CMCS. The modified CMCS-q demonstrated antioxidant activity by TEAC assay and antifungal activity by inhibiting spore germination of plant pathogen Botrytis cynerea. Then, CMCS-q was applied as an active coating on fresh-cut apples. The treatment resulted in enhanced firmness, inhibited browning and improved microbiological quality of the food product. The presented conjugation method allows retaining antimicrobial and antioxidant activity of quercetin moiety in the modified biopolymer. This method can be further used as a platform for binding ketone/aldehyde-containing polyphenols and other natural compounds to form various bioactive polymers.

Oligochitosan and oxidized nucleoside-based bioderived hydrogels for wound healing.

Herein, we report biocompatible hydrogel for wound healing that was prepared using nature-sourced building blocks. For the first time, OCS was employed as a building macromolecule to form bulk hydrogels along with the nature-sourced nucleoside derivative (inosine dialdehyde, IdA) as the cross-linker. A strong correlation was obtained between the mechanical properties and stability of the prepared hydrogels with a cross-linker concentration. The Cryo-SEM images of IdA/OCS hydrogels showed an interconnected spongy-like porous structure. Alexa 555 labeled bovine serum albumin was incorporated into the hydrogels matrix. The release kinetics studies under physiological conditions indicated that cross-linker concentration could also control the release rate. The potential of hydrogels in wound healing applications was tested in vitro and ex vivo on human skin. Topical application of the hydrogel was excellently tolerated by the skin with no impairment of epidermal viability or irritation, determined by MTT and IL-1α assays, respectively. The hydrogels were used to load and deliver epidermal growth factor (EGF), showing an increase in its ameliorating action, effectively enhancing wound closure inflicted by punch biopsy. Furthermore, BrdU incorporation assay performed in both fibroblast and keratinocyte cells revealed an increased proliferation in hydrogel-treated cells and an enhancement of EGF impact in keratinocytes.

A Novel Platform for Root Protection Applies New Root-Coating Technologies to Mitigate Soil-Borne Tomato Brown Rugose Fruit Virus Disease.

Tomato brown rugose fruit virus (ToBRFV) is a soil-borne virus showing a low percentage of ca. 3% soil-mediated infection when the soil contains root debris from a previous 30-50 day growth cycle of ToBRFV-infected tomato plants. We designed stringent conditions of soil-mediated ToBRFV infection by increasing the length of the pre-growth cycle to 90-120 days, adding a ToBRFV inoculum as well as truncating seedling roots, which increased seedling susceptibility to ToBRFV infection. These rigorous conditions were employed to challenge the efficiency of four innovative root-coating technologies in mitigating soil-mediated ToBRFV infection while avoiding any phytotoxic effect. We tested four different formulations, which were prepared with or without the addition of various virus disinfectants. We found that under conditions of 100% soil-mediated ToBRFV infection of uncoated positive control plants, root-coating with formulations based on methylcellulose (MC), polyvinyl alcohol (PVA), silica Pickering emulsion and super-absorbent polymer (SAP) that were prepared with the disinfectant chlorinated-trisodium phosphate (Cl-TSP) showed low percentages of soil-mediated ToBRFV infection of 0%, 4.3%, 5.5% and 0%, respectively. These formulations had no adverse effect on plant growth parameters when compared to negative control plants grown under non ToBRFV inoculation conditions.

Nucleoside-Based Cross-Linkers for Hydrogels with Tunable Properties.

Herein, we report bioderived cross-linkers to create biopolymer-based hydrogels with tunable properties. Nucleosides (inosine and uridine) and ribose (pentose sugar lucking the nitrogenous base) were partially oxidized to yield inosine dialdehyde (IdA), uridine dialdehyde (UdA), and ribose dialdehyde (RdA). The dialdehydes were further used as cross-linkers with polysaccharide chitosan to form hydrogels. Depending on the cross-linker type and concentration, the hydrogels showed tunable rheological, mechanical, and liquid holding properties allowing the preparation of injectable, soft, and moldable hydrogels. Computational modeling and molecular dynamics simulations shed light on hydrogel formation and revealed that, in addition to covalent bonding, noncovalent interactions (π-π stacking, cation-π, and H-bonding) also significantly contributed to the cross-linking process. To demonstrate various application possibilities, the prepared hydrogels were used as a growth platform for plant cells, as injectable inks for layer-by-layer 3D printing applications, and as moldable hydrogels for soft lithography to replicate the microstructure of the plant. These findings suggest that the obtained tunable biocompatible hydrogels have the potential to be good candidates for various biotechnological applications.

Rechargeable films for protection of dry foods: A sustainable method for covalent grafting of ß-cyclodextrin-thymol complex on PET/viscose platform.

Rechargeable materials for the effective protection of dry food products were developed. ß-cyclodextrin-thymol inclusion complex was covalently grafted on a solid platform of polyethylene terephthalate-viscose using either traditional cross-linker of epichlorohydrin, or natural cross-linker of citric acid. A correlation between the grafting method and physicochemical properties, loading capacity and release capabilities of the resulted materials was studied. The developed materials demonstrated antimicrobial properties preventing mold propagation in wheat grains. The treated grains showed normal germination abilities verifying that the prepared materials can protect dry food products without using harmful chemicals. The suggested approach can be extended to other applications and active agents. A combination of rechargeable films with natural volatiles can serve as an effective platform for sustainable active materials for food protection and in other fields such as agriculture, cosmetics, and medicine.

Formation of Robust and Adaptive Biopolymers via Non-Covalent Supramolecular Interactions.

Biomass-originated materials are the future's next-tier polymers. This work suggests improving mechanical and barrier properties of nature-sourced polymers using non-covalent supramolecular interactions. Polysaccharide chitosan is modified with amino acids via an esterification pathway using a systematic variation of hydrogen bond and aromatic domains (Degrees of substitution 12-49%). These controlled modifications improve stability due to non-covalent interactions, resulting in biopolymers with tailored thermal (decomposition temperature 232-275 °C), mechanical (Young's modulus 540-2667 MPa), and surface properties (roughness 4-40 nm). Chitosan and natural amino acids that are already manufactured at scale are purposely selected. The facile synthesis, controlled properties, stimuli-responsive potential, and inexhaustible origin of the raw materials provide the presented findings with the potential to become the method for the formation of high-performance biodegradable alternatives to petroleum-based polymers that can be used in packaging, food, agriculture, and medicine.

Synergistic antimicrobial effect of the combination of beta-lactam antibiotics and chitosan derivative on multidrug-resistant bacteria.

Dissemination of multidrug-resistant (MDR) bacteria with CTX-M-type extended-spectrum ß-lactamases (blaCTX-M) has become the greatest challenge in public health care. This study aimed to investigate the synergistic antibacterial potential of N-alkylaminated chitosan nanoparticles (CNPs) combined with conventional ß-lactam antibiotics (BLAs) against multidrug-resistant pathogen with blaCTX-M gene. The results of this study showed that the developed nano-formulation resensitized the studied E. coli MDR strain (E001) to ampicillin (AMP) and piperacillin (PIP) by causing a 1000-10,000-fold decrease in their MIC values (5000-50,000 mg/L to 5 mg/L). The conjugation of CNPs with cefoxitin (FOX) and ceftazidime (CAZ) showed a comparatively lower synergistic inhibitory effect owing to the higher susceptibility (MIC value = 0.5 mg/L-5 mg/L) of E001 to these antibiotics. The results indicate that CNPs could be effectively employed as an additive to augment the antibacterial effect of the BLAs for which MDR strains exhibit higher MIC values.

Hydrophilic Chitosan Derivatives: Synthesis and Applications.

Polymer alternatives sourced from nature have attracted increasing attention for applications in medicine, cosmetics, agriculture, food, water purification, and more. Among them, chitosan is the most versatile due to its full biodegradability, exceptional biocompatibility, multipurpose bioactivity, and low toxicity. Although remarkable progress has been made in its synthetic modification by using C3/C6 secondary/primary hydroxy (-OH) and the C2 amino (-NH2 ) active sites, its solubility under physiological conditions remains limited and has hampered larger-scale adoption. This review summarizes different synthetic methods that increase chitosan's hydrophilicity and water solubility by using covalent modifications, namely amino acid addition, quaternary ammonium formation, phosphorylation, and carboxymethylation. We also review several applications for each type of substitution in fields such as cosmetics, medicine, agriculture, and water purification, and provide an outlook and perspective for future modifications and implementations.

Biocompatible nanocarriers for passive transdermal delivery of insulin based on self-adjusting N-alkylamidated carboxymethyl cellulose polysaccharides.

In this work, we present biocompatible nanocarriers based on modified polysaccharides capable of transporting insulin macromolecules through human skin without any auxiliary techniques. N-Alkylamidated carboxymethyl cellulose (CMC) derivatives CMC-6 and CMC-12 were synthesized and characterized using attenuated total reflectance Fourier transform infrared (ATR-FTIR) and nuclear magnetic resonance (NMR) spectroscopy, gel permeation chromatography and thermogravimetric, calorimetric and microscopic techniques. The prepared modified polysaccharides spontaneously assemble into soft nanoaggregates capable of adjusting to both aqueous and lipid environments. Due to this remarkable self-adjustment ability, CMC-6 and CMC-12 were examined for transdermal delivery of insulin. First, a significant increase in the amount of insulin present in lipid media upon encapsulation in CMC-12 was observed in vitro. Then, ex vivo studies on human skin were conducted. Those studies revealed that the CMC-12 carrier led to an enhancement of transdermal insulin delivery, showing a remarkable 85% insulin permeation. Finally, toxicity studies revealed no alteration in epidermal viability upon treatment and the absence of any skin irritation or amplified cytokine release, verifying the safety of the prepared carriers. Three-dimensional (3D) molecular modeling and conformational dynamics of CMC-6 and CMC-12 polymer chains explained their binding capacities and the ability to transport insulin macromolecules. The presented carriers have the potential to become a biocompatible, safe and feasible platform for the design of effective systems for transdermal delivery of bioactive macromolecules in medicine and cosmetics. In addition, transdermal insulin delivery reduces the pain and infection risk in comparison to injections, which may increase the compliance and glycemic control of diabetic patients.

Oligomers of Carboxymethyl Cellulose for Postharvest Treatment of Fresh Produce: The Effect on Fresh-Cut Strawberry in Combination with Natural Active Agents.

In this study, oligomers of carboxymethyl cellulose (O-CMC) were used as a new postharvest treatment for fresh produce. The oligomers were prepared by green and cost-effective enzymatic hydrolysis and applied to prevent spoilage and improve storability of fresh-cut strawberries. The produce quality was improved by all formulations containing O-CMC in comparison to the control, as indicated by the decrease in decay incidence, weight loss (min ~2-5 times less), higher firmness, microbial load decrease, better appearance, and sensorial quality of the fruits. Natural resources: ascorbic acid, gallic acid, and vanillin were further added to enhance the beneficial effect. O-CMC with vanillin was most efficient in all of the tested parameters, exhibiting the full prevention of fruit decay during all 7 days of refrigerated storage. In addition, fruits coated with O-CMC vanillin have the smallest weight loss (%), minimum browning, and highest antimicrobial effect preventing bacterial (~3 log, 2 log) and yeast/mold contaminations. Based on the obtained positive results, O-CMC may provide a new, safe, and effective tool for the postharvest treatment of fresh produce that can be used alone or in combination with other active agents.

Non-radical synthesis of chitosan-quercetin polysaccharide: Properties, bioactivity and applications.

Quercetin-chitosan (QCS) polysaccharide was synthesized via non-radical reaction using L-valine-quercetin as the precursor. QCS was systematically characterized and demonstrated amphiphilic properties with self-assembling ability. In-vitro activity studies confirmed that quercetin grafting does not diminish but rather increases antimicrobial activity of the original chitosan (CS) and provided the modified polysaccharide with antioxidative properties. QCS applied as a coating on fresh-cut fruit reduced microbial spoilage and oxidative browning of coated melon and apple, respectively. Notably, QCS-based coatings prevented moisture loss, a major problem with fresh produce (2%, 12% and 18% moisture loss for the QCS-coated, CS-coated and uncoated fruit, respectively). The prepared QCS polysaccharide provides advanced bioactivity and does not involve radical reactions during its synthesis, therefore, it has good potential for use as a nature-sourced biocompatible active material for foods and other safety-sensitive applications.

Quaternized chitosan as a biopolymer sanitizer for leafy vegetables: synthesis, characteristics, and traditional vs. dry nano-aerosol applications.

A series of quaternary dimethyl-(alkyl)-ammonium chitosan derivatives (QACs) was synthesized and studied for physicochemical properties and bioactivity. The QACs tended to spontaneously self-assembly into nanoaggregates. Antimicrobial activity was examined in vitro on Gram-negative Escherichia coli (E. coli) and Gram-positive Listeria innocua (L. innocua) bacteria as well as phytopathogenic fungus Botrytis cinerea. The hexyl chain-substituted QAC-6 demonstrated the highest potency causing 3.0- and 4.5-log CFU mL-1 reduction of E. coli and L. innocua, respectively. QAC-6 was tested for antimicrobial activity on stainless steel coupons and fresh spinach leaves. A traditional 'wet' application (spray) and dry Engineered Water Nanostructure (EWNS) approach were used for spinach decontamination. With both approaches, significant reduction of microbial load on the treated produce was achieved. The wet application showed a greater reduction of microbial load, while the advantages of EWNS were reaching the antimicrobial effect with miniscule dose of active agent leaving treated surface visibly dry.

Double-stranded RNA targeting fungal ergosterol biosynthesis pathway controls Botrytis cinerea and postharvest grey mould.

Pathogenic fungi cause major postharvest losses. During storage and ripening, fruit becomes highly susceptible to fungi that cause postharvest disease. Fungicides are effective treatments to limit disease. However, due to increased public concern for their possible side effects, there is a need to develop new strategies to control postharvest fungal pathogens. Botrytis cinerea, a common postharvest pathogen, was shown to uptake small double-stranded RNA (dsRNA) molecules from the host plant. Such dsRNA can regulate gene expression through the RNA interference system. This work aimed to develop a synthetic dsRNA simultaneously targeting three essential transcripts active in the fungal ergosterol biosynthesis pathway (dsRNA-ERG). Our results show initial uptake of dsRNA in the emergence zone of the germination tube that spreads throughout the fungus and results in down-regulation of all three targeted transcripts. Application of dsRNA-ERG decreased B. cinerea germination and growth in in vitro conditions and various fruits, leading to reduce grey-mould decay. The inhibition of growth or decay was reversed by the addition of ergosterol. While dual treatment with dsRNA-ERG and ergosterol-inhibitor fungicide reduced by 100-fold the required amount of fungicide to achieve the same protection rate. The application of dsRNA-ERG induced systemic protection as shown by decreased decay development at inoculation points distant from the treatment point in tomato and pepper fruits. Overall, this study suggests that dsRNA-ERG can effectively control B. cinerea growth and grey-mould development suggesting its efficacy as a future method for postharvest control of fungal pathogens.

Characterization of the selective binding of modified chitosan nanoparticles to Gram-negative bacteria strains.

Chitosan is a nature-sourced polysaccharide widely used in numerous applications. The antibacterial potential of chitosan has attracted researchers to further develop and utilize this polymer for the formation of biocompatible antibacterial agents for both the food and healthcare industries. The tested hypothesis in this study is that modified N-alkylaminated chitosan nanoparticles (CNPs) have selective binding properties to Gram-negative bacteria strains that result in bacterial aggregation. Various bacterial strains were tested of five Gram-negative bacteria including Erwinia carotovora, Escherichia coli, Pseudomonas aeruginosa, Salmonella, and Serratia marcescens, as well as three Gram-positive bacteria strains including Bacillus licheniformis, Bacillus megaterium, and Bacillus subtilis. The fluorescence microscopy characterization showed that the presence of CNPs caused the aggregation of Escherichia coli bacteria cells, where modified CNPs with a shorter chain length of the substituent caused a higher aggregation effect. Moreover, it was found that the CNPs exhibited a selective binding behavior to Gram-negative as compared to Gram-positive bacteria strains, mainly to Escherichia coli and Salmonella. Also, the scanning electron microscopy characterization showed that CNPs exhibited selective binding to Gram-negative bacteria, which was especially understood when both Gram-negative and Gram-positive bacteria strains were within the same sample. In addition, the bacterial viability assay suggests that CNPs with a lower degree of substitution have a higher inhibitory effect on bacterial growth. CNPs with longer side chains had a less inhibitory effect on the bacterial growth of Gram-negative strains, where a concentration-dependent response pattern was only seen for the cases of Gram-negative strains, and not for the case of Gram-positive strain. To conclude, the further understanding of the selective binding of CNPs to Gram-negative bacteria strains may produce new opportunities for the discovery and characterization of effective antibacterial agents.

Stimuli-Free Transcuticular Delivery of Zn Microelement Using Biopolymeric Nanovehicles: Experimental, Theoretical, and In Planta Studies.

This paper reports one-step synthesis of polysaccharide-based nanovehicles, capable of transporting ionic zinc via plant cuticle without auxiliary stimulation. Delivery of highly hydrophilic nutritive microelements via the hydrophobic cuticle of plant foliage is one of the major challenges in modern agriculture. In traditional nutrition via roots, up to 80% of microelements permeate to soil and get wasted; therefore, foliar treatment is an environmentally and economically preferable alternative. Carboxymethyl cellulose (CMC) was modified to amphiphilic N-octylamide-derivative (CMC-8), which spontaneously self-assemble to nanovehicles. It was found that hydrophobic substituents endow a biopolymer with unexpected affinity toward a hydrophilic payload. CMC-8 nanovehicles effectively encapsulated ionic zinc (ZnSO4) and delivered it upon foliar application to pepper (Capsicum annuum) and tomato (Solanum lycopersicum) plants. Zinc uptake and translocation in plants were monitored by SEM-EDS and fluorescence microscopic methods. In planta monitoring of the carrier was done by labeling nanovehicles with fluorescent carbon dots. Three-dimensional (3-D) structural modeling and conformational dynamics explained the CMC-8 self-assembly mechanism and zinc coordination phenomenon upon introduction of hydrophobic substituents.

The polyaminosaccharide-based buffers as a new type of zwitterionic buffering macromolecules for biochemical applications.

A new type of biocompatible buffers based on zwitterionic polyaminosaccharides is reported. The carboxy- and amino-groups containing carboxymethyl chitosan (CM-CS) was synthesized and reacted with hydrochloric/acetic acid resulting in CM-CS-HCl and CM-CS-HAc buffers with buffering capacity of 20.6 and 15.2 mM/pH. The new buffers were comprehensively characterized for their physicochemical properties and checked on enzymatic reactions of acetylcholinesterase (AChE) and alkaline phosphatase (ALP). Their performance was compared to the phosphate and Tris buffers. The chloride-free, CM-CS-HAc demonstrated excellent buffering activity with Michaelis constants of 0.50 and 1.00 mM and maximum reaction rates of 5.62 and 2.26 µmol/min/mL for AChE and ALP reactions, respectively. Toxicity studies on stress-sensitive bioreporter bacteria verified nontoxicity of CM-CS-HAc. Zwitterionic polyaminosaccharides overcome drawbacks of monomeric buffers, such as interference with enzyme active sites, cell membrane injury and purification difficulties. Therefore, they may become the next generation of effective buffers for biological and biochemical applications.