Addition Of Chemical Reagents Research Articles

A Review on Some Physico-Chemical Compositions of Bread Wheat Determined by Using Destructive and Non-Destructive Methods

Some physico-chemical compositions of bread wheat (Triticum aestivum) were determined by using destructive and non-destructive methods. Standardized conventional method by Association of Official Analytical Chemists (AOAC) food analysis scheme was used for the destructive approach while near infrared spectrophotometer was used to measure some proximate compositions of wheat as a non-destructive method[1]. Near infrared techniques for obtaining the proximate compositions in the bread wheat was found to be more economical and faster without destroying the actual proximate compositions of the grains. The proximate compositions of bread wheat determined by using non-destructive measurement were moisture content 11.07 ± 0.03%, crude fiber 3.00 ± 0.00% and carbohydrate 71.80 ± 0.35% and the proximate compositions of bread wheat determined by using the destructive measurement were 9.50 ± 0.10%, 2.67 ± 0.07% and 67.28 ± 0.39%, respectively. Statistical analysis showed that there was no significant difference between the values obtained by the destructive and non-destructive methods for the determination of the physico-chemical compositions of bread wheat samples. Therefore, non-destructive method is economical, fast and save time compared to the crushing of the sample and addition of chemical reagents in the samples.

Reagent-free phosphorus precipitation from a denitrified swine effluent in a batch electrochemical system

There is high interest in the recovery of phosphorus (P) from wastewater through crystallization processes. However, the addition of chemical reagents (e.g., sodium hydroxide) to raise the pH may result in high treatment costs and increased concentrations of undesired metal ions (e.g., sodium). As an alternative, in this research we considered electrochemical mediated precipitation at low current densities (0.4–1.2 A m−2) without using chemical reagents. For that purpose, a two-chamber electrochemical system was operated in batch for treating denitrified swine effluent (48 mg P L−1). By applying current at 1.2 A m−2, and targeting pH 11.5, a maximum P removal rate of 33.4 mmol P (L·d−1) was obtained while the P removal efficiency was above 90 %. New solids that formed mostly remained suspended in the catholyte. Before discharge, the catholyte effluent was recirculated to the anodic compartment to neutralize the pH, achieving a final pH of 6.4 ± 0.1. Chlorine (Cl2) production in the anodic compartment was favored by a small anode surface and a high initial pH of the catholyte. Although the production of chlorine achieved was limited (the highest concentration was 8.6 ± 0.1 mg Cl2 L−1) these findings represent a new opportunity for the recovery and onsite use of this side-product. Electrochemical impedance spectroscopy tests confirmed that the deposition of solids inside the cathodic compartment during the experimental period was limited. Membrane analysis revealed significant scaling of carbonate compounds. The electrochemical treatment described above was shown as a promising alternative to sodium hydroxide and sulfuric acid dosage for pH adjustment when crystallizing phosphate salts.

Highly efficient selective extraction of Li from spent LiNixCoyMnzO2 assisted with activated pyrite in a subcritical water system

As the strategic importance of Li in the energy sector increases, selective Li extraction technology from spent lithium-ion batteries (LIBs) is attracting increasing attention. Current Li extraction processes typically suffer from lengthy procedures, high costs, and low efficiency. To improve the efficiency of Li extraction, a novel approach to achieve efficient Li recovery is proposed in this study, namely, reacting pyrite (FeS2) with LiNixCoyMnzO2 (NCM) powder in a subcritical water reduction (SWR) system. The reducing solvent environment created by the enhanced reaction of FeS2 with subcritical water converts the high−valent metals in NCM to a low-valent state, causing the collapse of the stable laminar structure and allowing Li+ to be released smoothly. After dual activation through mechanochemical and roasting processes, more than 99 % of Li is preferentially extracted under optimal conditions. Furthermore, Li+ in solution is converted into highly pure Li2CO3, while other metallic elements remain in the residue. Using inexpensive FeS2 for efficient Li extraction without adding additional chemical reagents is a promising approach for recovering spent LIBs.

A new azine-based ferrocenophane for azo dyes degradation in dark–ambient conditions

In a novel approach, 1,1′-bis(2,3-diazabuta-l,3‑dien)ferrocenophane (Fc-azine) synthesized by self-condensation of 1,1′-Ferrocenedicarboxaldehyde hydrazone (FcDH) with the aid of Zn(OAc)2·2H2O or acetic acid at 120 °C under high pressure. Fc-azine as an insoluble compound was fully characterized by Fourier transform infrared spectroscopy (FT-IR), diffuse reflectance spectroscopy (DRS), thermogravimetric analysis (TGA), Brunauer-Emmett-Teller analyses (BET), powder x-ray diffraction (XRD) and single crystal x-ray diffraction. The narrow band gap (Eg = 1.93 eV) and the presence of ferrocene and azine functional groups encouraged us to use Fc-azine as an advanced oxidation process (AOP) catalyst. Interestingly, the addition of Fc-azine (1 g L-1) to aqueous solutions of 50 ppm of Congo red (CR) and 10 ppm of methyl orange (MO) led to the fast degradation of CR (over 80 % in 5 min) and the slow degradation of MO (98 % in 48 h) in the dark, without additional energy sources or chemical reagents. It was found that Fc-azine can degrade CR in six cycles with high efficiency (over 80 %). CR degradation by Fc-azine can be performed in acid and basic environments (pH= 3–9), and effective MO degradation happens in slightly acidic and natural environments (pH= 5–7). Possible adsorption, reactive oxygen species production (·OH, O2·−), and the mechanism of CR and MO degradation were also studied.

Green synthesis of ZnO NPs with long-lasting and ultra-high antimicrobial activity

The continual onslaught of pathogenic microorganisms against humans, coupled with the escalating global challenge of antibiotic resistance, has led to a pervasive health crisis. To produce efficient antibacterial agents and combat bacterial resistance, zinc oxide nanoparticles (ZnO NPs) were synthesized, through green synthesis method by using Magnolia officinalis, Goldthread, and Lonicera japonica as extracts, respectively. Notably, there were no additional chemical reagents used in the synthesis route, emphasizing its eco-friendly nature. The antimicrobial efficacy of these ZnO NPs was assessed against Escherichia coli, Staphylococcus aureus, and Botrytis cinerea. The results revealed the ZnO NPs exceptional antimicrobial properties, demonstrating a minimum antimicrobial concentration (MIC) of 10 mg/L. Magnolia officinalis-ZnO NPs (M-ZnO NPs) exhibited 99.99 % antimicrobial activity against both E. coli and Staphylococcus aureus, along with 99.63 % effectiveness against Botrytis cinerea. Additionally, it was observed that the antibacterial characteristics of ZnO NPs exhibited minimal degradation after being stored in ambient air conditions at standard room temperature for a half-year. This study conclusively affirms the potent antimicrobial efficacy of green-synthesized ZnO nanoparticles. Transmission electron microscopy revealed that the particle size of ZnO NPs is 20–40 nm. Plant extracts contribute to the reduction processes of metal ions, and can also control the size and stability of formed nanostructures while avoiding the use of hazardous solvents. This green synthesis results in smaller particle sizes and sustains a long-term high antibacterial effect. Overall, this research expands the prospects for development involving nanomaterials, offering promising avenues for further exploration and innovation in this field.

Constructing sustainable keratin/algae biohydrogels for efficient removal of aniline blue via a pre-extrusion induced self-crosslinking strategy

Two environmentally friendly hierarchical porous biohydrogels, honeycomb keratin hydrogel (HKH) and spongy keratin-algae hydrogel (SKAH), were fabricated using an extrusion approach by using extruded keratin with or without algae. The extruded keratin exhibits exceptional self-crosslinking properties, making the biohydrogels behave with excellent adsorption performance for aniline blue dye removal from wastewater. In addition, the SKAH demonstrated a significantly higher maximum uptake capacity of 442.56 mg/g, surpassing that of the HKH (134.63 mg/g). This enhancement can be attributed to the looser porous structure of the SKAH, which enables better exposure of active groups within the material, which indicates the introduction of algae led to noticeable variations in the microstructure and performance of the adsorbent. Systematic investigation on the adsorption performance of isotherm and kinetics analyses indicated that chemisorption played a predominant role in governing the overall rate of adsorption. Overall, this extrusion approach allows for a simplified and non-toxic method of synthesizing these biohydrogels without the need for additional chemical reagents or biocross-linkers, promoting their eco-friendliness and reducing potential environmental impacts. The unique feature presents significant opportunity to fabricate sustainable and eco-friendly biohydrogel adsorbent for wastewater treatment.

Electrocatalytic membrane with p-block bismuth atoms for selective oxygen activation to hydroxyl radicals for effective water decontamination

Herein, individual p-block bismuth (Bi) atoms are dispersed onto N-doped Ti3C2Tx MXene (BiN3/MXene) to enhance the 3e− oxygen reduction reaction (ORR), for the electrosynthesis of hydroxyl radicals (HO•) via the activation of molecular oxygen (O2). Theoretical calculations and experiments reveal that the p-orbitals of Bi atoms can readily hybridize with the p-orbitals of O, which enables the transfer of charges, creates sufficient strength of adsorption for oxygen intermediates, lowers the activation energy, and modulates the rate-limiting reaction. The optimized generation of HO• is up to 26.7 μmol/(L·h·cm2) without additional chemical reagents, facilitating efficient removal of micropollutants from complex water matrices. Specifically, the rate constant (kobs) for sulfamethoxazole degradation achieves 1.027 min−1, outperforming the reported processes for micropollutants removal. This work illuminates the atomic-level construction of a p-block BiN3/MXene electrocatalytic membrane to efficiently perform the 3e− ORR and highlights the substantial potential for wastewater decontamination.

Employing electrochemically derived pH gradients for Lab-on-PCB protein preconcentration devices

Protein preconcentration is an essential sample preparation step for analysis in which the targeted proteins exist in low concentrations, such as bodily fluids, water, or wastewater. Nonetheless, very few practical implementations of miniaturized protein preconcentration devices have been demonstrated in practice, and even fewer have been integrated with other microanalytical steps. Existing approaches rely heavily on additional chemicals and reagents and introduce complexity to the overall assay. In this paper, we propose a novel miniaturized isoelectric focusing-based protein preconcentration screening device based on electrochemically derived pH gradients rather than existing chemical reagent approaches. In this way, we reduce the need for additional chemical reagents to zero while enabling device incorporation in a seamlessly integrated full protein analysis microsystem via Lab-on-PCB technology. We apply our previously presented Lab-on-PCB approach to quantitatively control the pH of a solution in the vicinity of planar electrodes using electrochemical acid generation through redox-active self-assembled monolayers. The presented device comprises a printed circuit board with an array of gold electrodes that were functionalized with 4-aminothiophenol; this formed a self-assembled monolayer that was electropolymerized to improve its electrochemical reversibility. Protein preconcentration was performed in two configurations. The first was open and needed the use of a holder to suspend a well of fluid above the electrodes; the second used microfluidic channels to enclose small volumes of fluid. Reported here are the resulting data for protein preconcentration in both these forms, with a quantitative concentration factor shown for the open form and qualitative proof shown for the microfluidic.

Hyaluronic Acid Hydrogels with Phototunable Supramolecular Cross-Linking for Spatially Controlled Lymphatic Tube Formation.

The dynamics of the extracellular matrix (ECM) influences stem cell differentiation and morphogenesis into complex lymphatic networks. While dynamic hydrogels with stress relaxation properties have been developed, many require detailed chemical processing to tune viscoelasticity, offering a limited opportunity for in situ and spatiotemporal control. Here, a hyaluronic acid (HA) hydrogel is reported with viscoelasticity that is controlled and spatially tunable using UV light to direct the extent of supramolecular and covalent cross-linking interactions. This is achieved using UV-mediated photodimerization of a supramolecular ternary complex of pendant trans-Brooker's Merocyanine (BM) guests and a cucurbit[8]uril (CB[8]) macrocycle. The UV-mediated conversion of this supramolecular complex to its covalent photodimerized form is catalyzed by CB[8], offering a user-directed route to spatially control hydrogel dynamics in combination with orthogonal photopatterning by UV irradiation through photomasks. This material thus achieves spatial heterogeneity of substrate dynamics, recreating features of native ECM without the need for additional chemical reagents. Moreover, these dynamic hydrogels afford spatial control of substrate mechanics to direct human lymphatic endothelial cells (LECs) to form lymphatic cord-like structures (CLS). Specifically, cells cultured on viscoelastic supramolecular hydrogels have enhanced formation of CLS, arising from increased expression of key lymphatic markers, such as LYVE-1, Podoplanin, and Prox1, compared to static elastic hydrogels prepared from fully covalent cross-linking. Viscoelastic hydrogels promote lymphatic CLS formation through the expression of Nrp2, VEGFR2, and VEGFR3 to enhance the VEGF-C stimulation. Overall, viscoelastic supramolecular hydrogels offer a facile route to spatially control lymphatic CLS formation, providing a tool for future studies of basic lymphatic biology and tissue engineering applications.

New Insights into Selective Singlet Oxygen Production via the Typical Electroactivation of Oxygen for Water Decontamination.

Selective production of singlet oxygen (1O2) as an electrophilic oxidant is crucial for the precise control of chemical targets in environmental fields. Herein, we proposed a strategy to construct a redox interface on electrodes, which can in situ produce inorganic metal hydroperoxides with appropriate oxidative ability during oxygen activation. Benefiting from atomic Cu sites (CuN4) in a copper-carbon aerogel electrode, almost complete production of 1O2 was achieved, thereby refraining the competitive formation of other reactive oxygen species. The fast electron transfer rate between CuN4 and electrogenerated H2O2 promoted the in situ formation of copper hydroperoxide (N4-Cu-OOH), thereby selectively and efficiently oxidizing intermediate O2•- to 1O2. The optimized production of 1O2 was up to 2583 μmol L-1 without additional chemical reagents. We further considered the high production of 1O2 for efficiently removing electron-rich organic pollutants from a complex water matrix. Fast kinetics was achieved and considered for removing various pollutants with electron-donating substituents in a nonradical oxidation pathway. The BPA degradation efficiency is less susceptible to the coexisting natural organic matter (NOM) and inorganic ions. Specifically, the kinetic constant for BPA removal is 34 times higher than that for a nanoparticle of a copper-carbon electrode while producing a hydroxyl radical. Our findings highlight the innovative interfacial surface engineering of an electrocatalytic O2 activation system to selectively generate 1O2 for future potential applications.

Calorimetric analysis of Mg–Li ultra light magnesium alloy

The Thermal-Derivative Analysis, or Computer-Aided Cooling Curve Analysis, is a commonly used method for determining the solidification parameters of metals and alloys. Knowledge of the enthalpy, kinetics and broad characteristics of the transformations taking place during the cooling of an alloy can be obtained with the proper configuration of analytical equipment and arrangement of test techniques. This work practically analyses the changes occurring in the Mg–4.5Li–1.5Al alloy as a result of the modification of the structure by the addition of TiB and Sr. The results indicate that adding grain refinements significantly affects the microstructure and thermal parameters of Mg–4.5Li–1.5Al alloy. The nucleation temperature and solidus temperature decrease with the addition of chemical reagents. Compressive strength improved due to the refinement of grain size. The results confirmed that thermal-derivative analysis is suitable for analysing changes in cast magnesium alloys.

All-natural and high-performance structural material based on lignin-reinforced cellulose

Sustainable materials derived from lignocellulosic resources are attractive alternatives to ubiquitous petrochemical plastics. However, it remains a huge challenge to improve the mechanical properties of lignocellulosic materials through efficient and low-energy processing strategy. Herein, we report a facile lignin-reinforced cellulose strategy to manufacture an all-natural and high-performance structural material from bamboo particles via partial delignification followed by hot-pressing. Partial delignification can regulate the amount of lignin in the bamboo particles and release internal cellulose microfibers. The subsequent hot-pressing drives residual lignin into the cellulose matrix to form a dense structure, which acts as a polyphenolic binder to improve the performance of the structural material. The obtained lignin-reinforced cellulose plate (LRCP) demonstrates high isotropic flexural strength (∼162 MPa), which is higher than that of natural bamboo and typical petrochemical plastics. Moreover, LRCP shows outstanding toughness (∼2.58 MPa m1/2), excellent compressive strength (∼178 MPa), improved wet stability and superior flame resistance. LRCP is all-natural and eco-friendly without the need for additional chemical cross-linking reagents. We believe that this facile and cost-effective strategy provides a promising way to process lignocellulosic resources into all-natural and high-performance structural materials that can serve as powerful plastic alternatives in the future.

Magnetron sputtering onto nonionic surfactant for 1-step preparation of metal nanoparticles without additional chemical reagents

Plasma-based sputtering onto liquids (SoL) is a straightforward approach for synthesizing small metal nanoparticles (NPs) without additional stabilizing reagents. In this work, nonionic surfactant Triton X-100 was used for the first time as a host liquid for the SoL process and the production of colloidal solutions of gold, silver and copper NPs was demonstrated. The average diameter of spherical Au NPs lies in the range from 2.6 to 5.5 nm depending on the conditions. The approach presented here opens the pathway to the production of concentrated dispersions of metal NPs of high purity that can be dispersed in water for future usage, therefore extending further the reach of this synthesis pathway.

Efficient oxidation of ammonium to nitrogen gas via accelerated ClO• generation at TiO2/Ru-IrO2 bifacial electrode in a UV-driven photoelectrochemical system

Photoelectrocatalysis is effective for the oxidation of ammonium (NH4+) due to the ability to produce oxidizing species such as •OH and Cl•. Compared to •OH and Cl•, ClO• is a more robust species capable of oxidizing NH4+, which can be produced by the combination of •OH and Cl• with free chlorine. However, the conventional photoelectrochemical (PEC) systems generally prefer to produce Cl• for NH4+ oxidation due to the lack of sufficient HOCl. Herein, a UV-driven PEC system consisting of TiO2/Ru-IrO2 bifacial electrode has been constructed for the enhanced ClO• generation for NH4+ oxidation. Large amounts of HOCl can be generated at the Ru-IrO2 interface, which can be further activated by UV irradiation to generate Cl• and •OH, and then rapidly react with HOCl to generate ClO•. The Cl• and •OH generated at the TiO2 interface could also react with HOCl, further accelerating the generation of ClO•. Under UV irradiation, the oxidation rate of TiO2/Ru-IrO2 was 6.7 times than TiO2, 1.5 times than Ru-IrO2, and even 1.24 times than the sum of individual TiO2 and Ru-IrO2. In addition, the bifacial electrode achieved highly efficient NH4+ oxidation for the treatment of actual wastewater, eliminating 81.0 mg L−1 of NH4+ in 120 min with a TN removal rate of 98.4%. The UV-driven PEC system with TiO2/Ru-IrO2 bifacial electrode is a promising potential technology for NH4+ oxidation due to its ability to rapidly produce ClO• and oxidize NH4+ to N2 without the addition of chemical reagents.

Photo-functionalized TiO2 film for facile immobilization of EpCAM antibodies and efficient enrichment of circulating tumor cells.

The highly efficient capture of circulating tumor cells (CTCs) in the blood is essential for the screening, treatment, and assessment of the risk of metastasis or recurrence of cancer. Immobilizing specific antibodies, such as EpCAM antibodies, on the material's surface is currently the primary method for efficiently capturing CTCs. However, the strategies for immobilizing antibodies usually have the disadvantages of requiring multiple chemical reagents and a complex pre-treatment process. Herein we developed a simple strategy for the immobilization of EpCAM antibodies without additional chemical reagents. By utilizing the positive charge property of the photo-functionalized titanium dioxide (TiO2), the negatively charged carboxyl terminal of EpCAM antibodies was immobilized by electrostatic interaction, allowing the antibodies to expose the antigen binding site fully. The experimental results showed that the photo-functionalized TiO2 surface had a marked positive charge and super-hydrophilic properties that could immobilize large amounts of EpCAM antibodies and keep excellent activity. CTCs capture experiments in vitro showed that the EpCAM antibodies-modified photo-functionalized TiO2 could efficiently capture CTCs. The results of blood circulation experiments in rabbits showed that the EpCAM antibodies-modified photo-functionalized TiO2 could accurately capture CTCs from the whole body's blood. It was foreseen that the strategy of simple immobilization of EpCAM antibodies based on photo-functionalized TiO2 is expected to serve in the efficient capture of CTCs in the future.

Resource-saving technologies for utilization of highly mineralized acidic waste from the ionite part of the combined water treatment plant at the Sterlitamak thermal power plant

PURPOSE. Development of technological solutions to reduce the amount of wastewater and reuse them in the cycle of the modernized water treatment plant at the Sterlitamak thermal power plant.METHODS. To achieve this goal, the methods of system analysis of the chemicaltechnological system were used, the sources and transformation of pollutants were determined.RESULTS. To date, Russia has experience in the disposal of wastewater from water treatment plants of energy enterprises. Basically, this method is based on neutralization with the use of various additional chemical reagents. As a result of the analysis, variants of technological schemes of a modernized water treatment plant were presented, where the concentrate after the reverse osmosis installation undergoes additional purification with the use of H- and Nacationite filters with joint or local use. The variant of the technological scheme will directly depend on the qualitative and quantitative composition of wastewater. A gypsum reactor is provided for the disposal of acidic waste regeneration solution, where gypsum is obtained as a product. These technological schemes make it possible to purify the wastewater of the installation and reuse it. These schemes are low-cost and are promising directions for the development of stations. conclusion. The developed technological solutions do not require the purchase of expensive chemicals and equipment, but only require the re-binding of equipment and the presence of a contact tank. These technologies are resource-saving, as a result, the consumption of raw water is reduced due to the reuse of waste, i.e. the cycle is low-flow. In recent decades, many industries have switched to a low-waste policy, since wastewater carries a large amount of valuable substances that can be recycled and reused.

Green one-step synthesis of mushroom-derived carbon dots as fluorescent sensors for Fe3+ detection.

Blue photoluminescent carbon dots were synthesized from Lentinus polychrous Lèv. via a simple hydrothermal process without additional chemical reagents or functionalization. The carbon dots (hereafter referred to as LCDs) were quasi-spherical with an average diameter of 6.0 nm. The strong fluorescence emissions of LCDs were utilized as the basis of efficient turn-off probes for Fe3+. The quenching phenomenon could be used to rapidly determine Fe3+ concentrations in the range of 0.0-2.0 mM in aqueous solution, with a limit of detection (LOD) of 16 μM. In the presence of interference, LCDs demonstrated good sensitivity and selectivity towards Fe3+ in both solution-based and paper-based systems. The LCDs also exhibited excellent photostability and an eco-friendly nature, making them an ideal choice for environmental monitoring with significant potential for diagnostic applications.

Natural magnetite/coke composite: A novel promising microwave absorption material

Fe3O4 and carbon materials are the two most common materials in research on microwave-absorbing materials. However, in previous studies, the material sources of Fe3O4 and carbon materials mostly came from the addition of chemical reagents, which meant that expensive raw materials and complex preparation processes restrict the large-scale production and application of microwave-absorbing materials from the raw material level. The preparation of highly efficient electromagnetic wave-absorbing materials from natural mineral resources may be the key way to solve this problem. In this work, pure magnetite minerals and original coking coal were used as the sources of Fe3O4 and carbon materials in the composite material. Through the combination of mixed ball milling and high-temperature roasting, the two natural mineral materials were successfully combined to prepare a magnetite/coke composite material with a porous network structure (Ma/Coke). After carbonization, a continuous network was formed to effectively coat or load magnetic magnetite particles, which formed the electromagnetic cooperative loss mechanism. By optimizing the material ratio, the microstructure of the composite was effectively regulated, and the impedance matching of the material was improved, showing excellent electromagnetic wave-absorbing performance. Ma/Coke-0.5 had a minimum reflection loss (RLmin) of − 57.86 dB and an effective absorption bandwidth of up to 5.47 GHz. This work provides a low-cost, high-efficiency, environment-protecting, and large-scale preparation method for electromagnetic wave-absorbing materials based on natural minerals and provides a new feasible suggestion for the functional application of natural mineral materials.

Study on the reaction mechanism of the potassium bicarbonate alkali activation process in black liquor

The alkaline hydrothermal separation of cellulose is a crucial step in biofuel generation, wherein black liquor containing lignin and alkali is produced as a by-product. This study explored the reaction mechanism of the residual alkali during the activation process. In this reaction, the impregnated K reagent exhibited unusual behavior owing to its bonding with the lignin structure, which was confirmed by the K 2p peak shift in x-ray photoelectron spectra. This behavior resulted in the formation of a microporous and mesoporous substance with a high surface area of 2146 m2 g−1 without requiring additional chemical reagents. Furthermore, the activated carbon derived from black liquor possessed outstanding properties, allowing its use in supercapacitors. This study demonstrates that giant miscanthus-derived black liquor can be used as an activated carbon precursor for application in electric double-layer capacitors owing to its superior electrochemical capabilities.

Manipulation of mRNA translation elongation influences the fragmentation of a biotherapeutic Fc-fusion protein produced in CHO cells.

Mammalian cells, particularly Chinese hamster ovary cells, are the dominant system for the production of protein-based biotherapeutics, however, product degradation, particularly of Fc-fusion proteins, is sometimes observed that impacts the quality of the protein generated. Here, we identify the site of fragmentation of a model immunoglobulin G1 Fc-fusion protein, show that the observed clipping and aggregation are decreased by reduced temperature culturing, that the fragmentation/clipping is intracellular, and that reduced clipping at a lower temperature (<37°C) relates to mesenger RNA (mRNA) translation elongation. We subsequently show that reduced fragmentation can be achieved at 37°C by addition of chemical reagents that slow translation elongation. We then modified mRNA translation elongation speeds by designing different transcript sequences for the Fc-fusion protein based on alternative codon usage and improved the product yield at 37°C, and the ratio of intact to a fragmented product. Our data suggest that rapid elongation results in misfolding that decreases product fidelity, generating a region susceptible to degradation/proteolysis, whilst the slowing of mRNA translation improves the folding, reducing susceptibility to fragmentation. Manipulation of mRNA translation and/or the target Fc-fusion transcript is, therefore, an approach that can be applied to potentially reduce fragmentation of clipping-prone Fc-fusion proteins.