Buffer Concentration Research Articles

Optimized electroporation buffer improves transfection and prime editing efficiency in adult bovine fibroblasts.

For livestock breeding, using somatic cells from adult animals for gene editing and subsequent cloning allows the preservation and enhancement of superior traits from the parent directly in the offspring, while avoiding the loss of genetic gain that can occur through crossbreeding. However, primary cells generally more difficult to transfect and perform gene editing. To date, most related studies have used more vigorous fetal fibroblasts as donor cells, while using somatic cells from adult animals requires more post-editing screening efforts due to the low yield of edited cells. Here, we performed electroporation on adult bovine ear fibroblasts (BEFs) under various conditions such as electrical pulse settings, plasmid dosage, cell density, and concentration of electroporation buffer, and evaluated the transfect efficiency using flow cytometry analysis. We confirmed that the 270 V-10-10 program (270 V, 10 ms, 10 cycles) using 1.5 million cells and 5 µg of EGFP plasmid yielded the highest number of EGFP positive cells. Additionally, we used prime editor (PE) to edit the MSTN locus in BEFs. More importantly, we discovered that lowering the osmolarity of the electroporation buffer improves both electroporation and gene editing efficiency, which relate to the repression of cGAS-STING pathway. Our finding provides valuable references for using electroporation methods in adult bovine primary cell gene editing.

Influence of volume and citrate on perceived pain in patients treated with adalimumab

BackgroundImmune-mediated inflammatory diseases (IMIDs) are a group of disorders characterised by acute or chronic inflammation. Adalimumab and infliximab are the cornerstones of their pharmacotherapy. This study aimed to determine whether...

Effect of the Electrolyte on the Oxygen Reduction Reaction with a MOF Embedded Co-Porphyrin.

Electrocatalysis in metal-organic frameworks is an interplay between the diffusion of charges, the intrinsic catalytic rate, and the mass-transport of reactants through the pores. Here a systematic study is carried out to investigate the role of the electrolyte nature and concentration on the oxygen reduction reaction (ORR) with the PCN-224(Co) MOF in aqueous electrolyte. It was found that the ORR activity is slightly influenced by the nature of the ions in solution, providing that the ionic strength is high enough to minimize the resistivity during the measurement. The ORR activity was found to be 1.3-1.5 times lower in lithium acetate compared to sodium acetate, while the ORR activity in cesium acetate was 1.3-1.6 times higher compared to the activity in sodium acetate. Moreover, there was no dependency found of the ORR catalysis on the size of the anion, buffer concentration, or oxygen concentration. These findings suggest that ORR catalysis in PCN-224(Co) is limited by the intrinsic ORR rate at the active site rather than charge transport through the porous structure or substrate transport in the pores. Therefore, optimization of ORR catalysis with this MOF might be achieved by the optimization of the electronics at the cobalt active site.

Novel methods for the detection of glutathione by surface-enhanced Raman scattering: A perspective review.

Detection of biomolecules, Glutathione (GSH) in particular, is important because it helps assess antioxidant capacity, cellular protection, detoxification processes, and potential disease associations. Monitoring glutathione levels can provide valuable information about overall health and well-being. Many medical disorders have been connected to glutathione levels. Higher glutathione levels have been seen in several cancer cell types, which may increase their resistance to radiation and chemotherapy. Glutathione levels can be measured through various methods, such as colorimetric assays and fluorescent probes. However, surface-enhanced Raman scattering (SERS) has been known as an efficient and selective technique for biomolecule detection. Here in this perspective review, we have reported two distinctive methods based on SERS technique in detection of GSH; heat-induced method and reversed reporting agent method. Several variables that can impact the detection scheme were elaborated in the "heat-induced method," including pretreatment, nanoparticle reduction time, the process temperature, the pH of the colloidal solution, the concentration of citrate buffer, and the concentration of participating nanoparticles. To choose the best reporting agent for a reverse reporting scheme using SERS approaches, several reporting agents were examined in the second method. In order to grasp the situation at hand, biomolecule detection-specifically, GSH detection schemes-was briefly discussed. SERS spectroscopy and its associated terminology were then covered followed by the perspective and outlook of GSH detection at the end. To meet the demands of real-time applications in everyday life and to enhance SERS methods for biomolecule detection-in particular, GSH detection-such a thorough investigation is unavoidable.

Nonspherical Particle Stabilized Emulsions Formed through Destabilization and Arrested Coalescence.

To form nonspherical emulsion droplets, the interfacial tension driving droplet sphericity must be overcome. This can be achieved through interfacial particle jamming; however, careful control of particle coverage is required. In this work, we present a scalable novel batch process to form nonspherical particle-stabilized emulsions. This is achieved by concurrently forming interfacially active particles and drastically accelerating emulsion destabilization through addition of electrolyte. To achieve this, surfactant-stabilized oil-in-water emulsions in the presence of dopamine were first produced. These emulsions were then treated with tris(hydroxymethyl)aminomethane hydrochloride buffer to both simultaneously initiate polymerization of dopamine in the emulsion continuous phase and reduce the Debye length of the system, thus accelerating droplet coalescence while forming surface-active particles. The concentration of buffer and imposed shear was then systematically varied, and the behavior at the interface was studied using pendent drop tensiometry and interfacial shear rheology. It was found that polydopamine nanoparticles formed in the emulsion continuous phase adsorbed to the reducing interface during coalescence, resulting in anisotropic droplets formed via arrested coalescence. Greater shear rates resulted in accelerated coalescence and formation of secondary droplets, whereas lower shear rates resulted in thicker interfacial films. The efficacy of this method was further demonstrated with a second system consisting of sodium dodecyl sulfate as the surfactant and polypyrrole particles, which also resulted in nonspherical droplets for optimized conditions.

Optimization of Collagen Extraction from Fish Scales Using Tris-Glycine Buffer: A Taguchi Methodological Approach.

Collagen, a critical biomaterial with wide applications in pharmaceuticals, cosmetics, and medical industries, can be sourced sustainably from fish scales. This study optimizes the extraction of collagen using Tris-Glycine buffer from fish scales via the Taguchi method. Various extraction parameters-buffer concentration, temperature, pH, and time-were evaluated to identify optimal conditions. Under optimal conditions (0.5 M of acetic acids, volume of acids of 100 mL, soaking time of 120 min, and Tris-Glycine buffer of 10 mL), the results demonstrate that temperature and buffer concentration significantly influence collagen yield, with a collagen purity of 17.14 ± 0.05 mg/g. R2 value of 73.84% was obtained for the mathematical model). FTIR analysis confirmed the presence of characteristic collagen peaks at 1611 cm-1 (amide I), 1523 cm-1 (amide II), and 1300 cm-1 (amide III), indicating the successful extraction of type I collagen. SDS-PAGE analysis revealed a protein banding pattern consistent with the molecular weight of collagen, and amino acid analysis shows high percentages of glycine (20.98%), proline (15.43%), and hydroxyproline (11.51%), implying fibrous collagen structures. The finding suggests that the Taguchi method offers an efficient and sustainable approach for collagen extraction, reducing waste from fish processing industries. Nevertheless, there is a need for further experimental validation to align with mathematical modeling on the optimized conditions.

A chemoenzymatic cascade for sustainable production of chiral N-arylated aspartic acids from furfural and waste

Both biomass valorization and waste upcycling are important routes to sustain the circular bioeconomy. In this work, we present a chemoenzymatic cascade for selective synthesis of chiral N-arylated aspartic acids from biomass-derived furfural and waste nitrophenols (NPs) by merging robust photo- and electrocatalysis with stereoselective biocatalysis. Concurrent photoelectrocatalytic oxidation of furfural into maleic acid (MA) and fumaric acid (FA) was significantly enhanced by combining catalyst and reaction engineering strategies including identification of a powerful photocatalyst meso-tetra(4-carboxyphenyl)porphyrin, continuous flow technique, enhancing dissolved O2 and paired electrosynthesis. The overall space-time yield (STY) approached 2.8 g L−1 h−1 in a fed-batch process, with the product titer of 28.3 g L−1. Besides, photoelectrosynthesis of MA/FA was effectively fueled by sunlight, with the STY of up to 3.6 g L−1 h−1. Both MA selectivity and yield could be facilely improved to around 89% by reducing the buffer concentrations. Paired electrosynthesis strategy not only resulted in greatly improved MA production at the anode, but also enabled NPs upcycling into value-added aminophenols (APs) at the cathode. The products formed in the two electrode chambers were converted into N-arylated (S)-aspartic acids by a bienzymatic cascade. This work presents a multicatalytic approach for integrating selective biomass valorization and waste upcycling towards sustainable manufacture.

"The More, the Better?": The Impact of Sugar-to-Protein Molar Ratio in Freeze-Dried Monoclonal Antibody Formulations on Protein Stability.

Lyophilization is widely used to ensure the stability of protein drugs by minimizing chemical and physical degradation in the dry solid state. To this end, proteins are typically formulated with sugars that form an amorphous immobilizing matrix and stabilize hydrogen bonds replacing water molecules. The optimal amount of sugar required and protein stability at low excipient-to-protein molar ratios are not well understood. We investigated this by focusing on the physical stability of formulations, reflecting highly concentrated monoclonal antibody (mAb) lyophilizates at low sucrose to mAb ratios between 25:1 and 360:1. Additionally, the impact of different excipient types, buffer concentrations, and polysorbates was studied. The mAb stability was evaluated over up to three months at 25 and 40 °C. We investigated the "the more, the better" approach regarding excipient usage in protein formulation and the existence of a potential stabilizing threshold. Our findings show efficient monomeric content preservation even at low molar ratios, which could be explained based on the water replacement theory. We identified an exponential correlation between the sucrose to protein molar ratio and aggregate formation and found that there is no molar ratio threshold to achieve minimum stabilization. Sucrose demonstrated the best stabilization effect. Both mannitol, used as a cryoprotectant at low concentrations, and arginine reduced aggregation compared to the pure mAb formulation. The higher ionic strength of 5 mM histidine buffer enhanced protein stability compared to that of 0.1 mM histidine buffer, which was more pronounced at lower molar ratios. The addition of polysorbate 20 contributed an additional interfacial stabilizing effect, complementing the cryoprotective and lyoprotective properties of sucrose. Overall, a model could be developed to optimize the quantity of sugar required for protein stabilization and facilitate a more rational design of protein lyophilizates. The molar ratio of sugar to protein for high-concentration mAb products is limited by the acceptable tonicity, but we showed that sufficient stabilization can be achieved even at low molar ratios.

Wide range luminescence lifetime-based pH sensing with covalently immobilized multi-protonatable Ru(II) complexes

To overcome the limitations of optical pH sensors, the response of which is just ca. one pH unit around the pKa value of the indicator dye and capitalize on the advantages of red-luminescent Ru(II) complexes for optical sensing, multi-pKa members of this family are proposed. Bipyridine ligands bearing protonatable groups (amines and carboxylates) are the basis to design and prepare the indicator dyes. Their photophysical properties and response to pH changes were investigated in the pH 3.5 − 9.0 range. The complex [Ru(DCB)2DEAMB], where DCB and DEAMB stand for 2,2’-bipyridine-4,4’-dicarboxylate and 4,4’-(N,N-diethylaminomethyl)-2,2’-bipyridine, respectively, shows a reversible monotonic change of its emission wavelength maximum (648 – 634 nm), luminescence intensity (1.40x) and lifetime (335 – 429 ns) in solution with increasing pH. This complex and two similar ones were covalently tethered to commercial TentaGel® M Br polymer microbeads for luminescence phase shift fiberoptic pH measurements. The immobilized [Ru(DCB)2DEAMB] displays the widest pH sensitivity over pH 3.5 to pH 8.5. The stability of this sensor was satisfactory when cycling between pH 3.5 and 8.5 for five days as well as under a three-day cycle between each pH unit. Changes of the buffer type, buffer concentration and osmolarity did not significantly influence the sensor response; however, large variations of dissolved oxygen and, naturally, temperature would require correction by the corresponding sensors. The novel, robust luminescent pH sensor has been tested in phase-sensitive mode for cell cultures monitoring in commercial bioreactors, but its response would also be suitable for in situ monitoring of natural waters.

Insights into interspecies protein binding variability using clindamycin as an example.

In the preclinical development of new drugs, animal models are often employed to predict their efficacy in humans, relying on translational pharmacokinetic/pharmacodynamic (PK/PD) studies. We performed in vitro experiments focusing on the comparison of plasma protein binding (PPB) and bacterial growth dynamics of clindamycin, a commonly used antimicrobial agent, across a range of drug concentrations and plasma environments. Human, bovine and rat plasma were used for determining PPB of clindamycin at various antibiotic concentrations in buffer and media containing 20% to 70% plasma or pure plasma using ultrafiltration (UF) and equilibrium dialysis (ED). Also bacterial growth and time-kill assays were performed in Mueller-Hinton broth (MHB) containing various percentages of plasma. Protein binding of clindamycin correlated well between UF and ED. Notably, clindamycin exhibited substantially lower protein binding to rat plasma compared with human and bovine plasma. Staphylococcus aureus growth was significantly reduced in 70% human, bovine, and rat plasma after 4, 8 and 24 h compared with standard MHB. Time-kill data demonstrated that bacterial counts at both 20% and 70% plasmas were less when compared with MHB at drug concentrations lower than MIC after 4 and 8 h of incubation. For rat plasma, the difference was maintained over 24 h of incubation. Furthermore, a complete bacterial killing at 16 mg/L was observed after 24 h in 20% and 70% human and bovine plasma, but not for rat plasma. Recognizing interspecies differences in PB might be essential for optimizing the translational relevance of preclinical studies.

Conjugation of Antibodies and siRNA Duplexes to Polymer Nanoparticles via Maleimide-Thiol Chemistry.

Polymeric nanoparticles (NPs) have shown great promise as highly modifiable platforms that can be applied across many different disease states. They are advantageous because they can encapsulate a range of hydrophobic and hydrophilic cargoes while having customizable surface properties. Depending on the desired biointerfacing capabilities, the surface of polymeric NPs can be modified with moieties, such as antibodies, peptides, nucleic acids, and more. The work presented here is intended to provide mechanistic insight into how different parameters influence the loading of antibodies, small interfering ribonucleic acids (siRNAs), or both on the surface of poly(lactic-co-glycolic acid) (PLGA) NPs via maleimide-thiol chemistry. Some of the conjugation parameters investigated include the buffer concentration, maleimide to protein ratio, and the addition of an excipient such as Tween-20. Through variation in the concentration of FZD7 antibodies added to the reaction mixture, we established tunable conjugation and found the upper limit of their loading density under the conditions tested. We also confirmed antibody conjugation through two different mechanisms: via a thiol-modified antibody or a thiol-modified poly(ethylene glycol) (PEG) linker. Conjugation of thiolated siRNA duplexes targeting β-catenin was also investigated through variations in both Tween-20 concentration and CaCl2 buffer concentration. Finally, the coconjugation of both antibodies and siRNA duplexes was explored. Overall, this work outlines a basis for tunable biomolecule loading on polymer NPs using maleimide-thiol chemistry and reveals the incredible versatility of polymer NP platforms.

The influence of citrate buffer molarity on mRNA-LNPs: Exploring factors beyond general critical quality attributes

Lipid nanoparticles (LNPs) are crucial in delivering mRNA vaccines and therapeutics. The properties of LNPs can be influenced by the choice of lipids and the manufacturing conditions, such as mixing parameters, lipid concentration, and the type and concentration of the aqueous buffer used. In this study, we investigated the impact of the citrate buffer molarity, the buffer commonly used to dissolve mRNA in the preparation of mRNA-LNPs. We prepared SM-102 LNPs containing firefly luciferase mRNA using citrate buffers at molarities of 50 mM, 100 mM, or 300 mM. Our findings revealed that varying the molarity of the citrate buffer did not significantly affect the particle size when considering the average diameter (z-average or Mode). All formulations exhibited low polydispersity index (PDI) and high encapsulation efficiency. Detailed analysis of particle size sub-populations (D10, D50, and D90) and morphology indicated that citrate buffer concentration might influence lipid packing during LNP production, though these differences were subtle. However, using higher citrate molarity (300 mM) to produce LNPs notably reduced cellular internalisation and in vitro transfection efficiency. This trend was also observed in vivo, where similar expression levels were noted in mice receiving the 50 mM and 100 mM LNP formulations, but lower expression was seen for the 300 mM formulation. Our study highlights the importance of buffer molarity in the aqueous phase during mRNA-based LNP preparation and that generally reported critical quality attributes (CQAs) for LNPs may not detect subtle formulation differences.

Utility of green chemistry for spectrofluorometric determination of fingolimod via metal complexation; Application to dosage and spiked biological fluid

The goal of the current research was to establisha quick and practical fluorimetric technique for fingolimod analysis. The approach relied on the drug's complex formation with the zinc ion to produce a high-fluorescence product. The fluorescence was further enhanced by adding sodium dodecyl sulfate, and it was observed at 325 nm following excitation at 475 nm. With a correlation coefficient of 0.9999, the association between emission intensity and fingolimod concentration was linear between 5 and 150 ng mL-1. The quantitation limit was 2.059 ng mL-1,while the detection limit was 0.679 ng mL-1. The buffer type, pH and concentration, type of surfactant and concentration, and finally the diluting solvent were among the reaction conditions that were closely examined. With great precision and reliability, the drug in question was quantified using this method in both spiked human plasma and capsule formulations. The proposed method's level of greenness was assessed using two methodologies: the analytical greenness metric (AGREE) and the Green Analytical Procedure Index (GAPI).

Sensitive HPLC Method to Support Pre-formulation Studies and to Determine Critical Quality Attributes of Therapeutic Contact Lenses Containing Dorzolamide Hydrochloride

Background: Dorzolamide hydrochloride (DRZ) is a carbonic anhydrase inhibitor used to treat glaucoma and ocular hypertension. Drug-eluting contact lenses, such as Acuvue Theravision ™ with Ketotifen, offer improved drug delivery and reduced side effects compared to eye drops. Drug-loaded nanoparticle-loaded contact lenses can sustain drug release and enhance comfort for extended wear. Objective: High buffer concentration and low pH increase the risk of damage to silica-bonded columns. Therapeutic contact lenses face challenges related to critical lens parameters, including the estimation of drug incorporation and release due to interference of lens matrix leaching. There is currently no analytical method available for estimating DRZ in contact lenses. Method: The HPLC method, which was developed and validated using ICH Q2 (R1) criteria, used a C18 column (250 mm × 4.6 mm, 5 μm) as a stationary phase and methanol:water (70:30 v/v) as the mobile phase. The detecting wavelength was 253 nm. Moreover, to support the efficiency of the developed method, the marketed formulation of DRZ eye drops, drug purity, and loading in contact lenses were analysed. The method was also employed to determine the Critical Quality Attributes (CQAs) of therapeutic contact lenses and drug release and drug leaching during the sterilization process. Result: The developed HPLC method shows Rt for DRZ at 2.881 minutes with good linearity (r2 > 0.998) between 2-32μg/mL, precision (RSD < 2%), accuracy (Recovery > 99.5%), sensitivity, and specificity for quantifying DRZ in marketed formulations and therapeutic contact lenses. The developed method is devoid of any buffer or modifier in the mobile phase, making it safer for the stationary phase. This method mitigates the interference of lens matrix leaching, which induces an overestimation of DRZ. All the result for therapeutic contact lenses was found to be closely aligned with theoretically expected results, confirming the reliability of the developed HPLC method for therapeutic contact lenses. Conclusion: This method is specific, accurate, and precise for quantifying DRZ in commercial formulations and newly developed therapeutic contact lenses. It effectively evaluates the critical quality attributes of these lenses, demonstrating their reliability for assessing their performance and ensuring quality in therapeutic applications.

Investigation on the Retention and Separation of Glucosinolates With a Mixed-Mode Reversed-Phase/Weak Anion-Exchange Column.

Glucosinolates, a crucial group of secondary metabolites in Brassica vegetables, present significant chromatographic separation challenges due to their anionic form, structure diversities, and co-existence of other phenolic compounds. This study comparatively investigated the retention and separation of seven glucosinolates using a mixed-mode reversed-phase/weak anion-exchange column and a conventional reversed-phase C18 column. Separation factors for each glucosinolate with its adjacent peaks were over 1.0 on the mixed-mode column, while co-eluting was observed on the C18 column. The effects of mobile phase additives and pH on the separation and retention of glucosinolates were also investigated. Results showed that glucosinolate retention was inversely related to both buffer concentration and pH. The optimized method for the mix-mode column was applied to the complex Brassica vegetable samples. In addition to the 17 well-resolved glucosinolate peaks, 34 peaks for phenolic compounds were identified in broccoli microgreen, suggesting the successful application scenarios for qualitative analysis in comparison with the single mode reverse phase C18 column. This study demonstrates that the mixed-mode reversed-phase/weak anion-exchange column can be used as a promising separation tool for organic anions in a complex sample matrix.

Rapid Determination of Galantamine in Human Plasma by Microchip Electrophoresis With a Highly Integrated Contactless Conductivity Detector.

A novel and rapid method was developed for the determination of galantamine in human plasma by microchip electrophoresis with a highly integrated contactless conductivity detector (CCD). The instrumental parameters affecting the response of the detector, such as excitation frequency and excitation voltage, were examined and optimized. The electrophoresis conditions that influenced the separation and detection of galantamine, including the composition of buffer solution, buffer pH, buffer concentration, additives, injection time, and separation voltage were systematically investigated. Under the optimal conditions, the peak height had a good linear relationship with the concentration of galantamine in human plasma from 10 to 160 µg/L, and the correlation coefficient was 0.9992, the limit of detection reached 1.1 µg/L. The recoveries were between 98.6% and 102.1%. This sensitive, rapid, and convenient method is a good alternative to existing methods for galantamine determination. Also, this highly integrated CCD holds great promise in clinical biochemical analysis.

Enhancement effect of urea toward electroporation-mediated plasmid transfection efficiency in the HEK-293 cell line.

Intracellular delivery is crucial in biological and medical studies. Although many molecular tools have been created for cell-based gene therapies, it remains challenging to introduce external molecules into cells. As one of the most popular non-viral transfection methods, electroporation induces transient pores in the cell membrane by applying an external electric field. Unsatisfactory transfection efficiency and low cell viability are the major drawbacks of electroporation. To overcome these issues, the current study investigated the effect of urea on electroporation-mediated transfection efficiency. Three voltages of electroporation, including 100, 120, and 140 V, and 3 concentrations of urea buffer, including 0.25%, 0.5%, and 1% W/V, were considered as variables in this study. The HEK-293 cell line was used for transfection, and green fluorescent protein (GFP) expression was evaluated using flow cytometry and fluorescence microscopy. The results showed that the combination of electroporation and urea increased electroporation efficacy, but the effect depended on voltage and urea concentration. When different concentrations of urea were added to HEK-293 cells at a voltage of 100 V, the number of cells transfected by pEGFP-N1 increased (from 12.3 ± 0.2% in untreated cells to 17.35 ± 0.55%, 23.3 ± 0.3%, and 14 ± 0.1% at urea concentrations of 0.25%, 0.5%, and 1% W/V, respectively). The electroporation buffer containing 0.5% W/V urea showed the highest EGFP expression (23.3 ± 0.3%) and high cell viability (over 90%). This research offers a new perspective for improving gene transfection efficiency once electroporation is utilized.

Acoustic Droplet Vaporization Efficiency and Oxygen Scavenging in Whole Blood

ObjectiveAcoustic droplet vaporization (ADV) is the liquid-to-gas phase transition of perfluorocarbon (PFC) droplets to microbubbles upon ultrasound insonation. After ADV, gases dissolved in the surrounding fluid diffuse into microbubbles, enabling oxygen scavenging. Characterization of oxygen scavenging and transition efficiency in whole blood has so far been limited. In this work, oxygen scavenging and PFC droplet transition efficiency in a saline buffer and whole bovine blood were evaluated using blood-gas analysis and flow cytometry. MethodsOxygen scavenging from whole blood via ADV was determined using an in vitro flow phantom with droplets comprising a phospholipid shell and either a decafluorobutane (DFB) or a perfluoropentane (PFP) core. Fluorescent droplets were used to determine ADV transition efficiency in whole blood via flow cytometry. Finally, a mathematical model predicting oxygen scavenging from whole blood was developed based on the experimental transition efficiency values. ResultsDFB droplets enabled greater oxygen scavenging and higher transition efficiency when compared to PFP droplets in both buffer and whole blood. Increasing droplet concentration resulted in a greater amount of hemoglobin-bound and dissolved oxygen scavenging from whole blood. ADV of DFB droplets at a concentration of 5 × 10-4 mL/mL yielded a total oxygen reduction of 913 μM. The transition efficiency decreased with increasing droplet concentration in both buffer and whole blood. Experimental oxygen scavenging data in whole blood aligned with the predicted values from the mathematical model. ConclusionIncreased oxygen scavenging and transition efficiency were achieved with DFB droplets relative to PFP droplets.

Utilization of water chestnut waste for biohydrogen production and enhanced power generation by stacked microbial fuel cell

A novel two-stage approach combining dark fermentation with microbial fuel cell (MFC) technology is proposed which enable biohydrogen production and bioelectricity generation from residual substrate energy. In the present study, batch fermentation of water chestnut waste using Enterobacter aerogenes (MTCC 2822) resulted in the production of biohydrogen. In the batch process, the highest production was 3.2 L/L. Further, single-parameter optimization and multi-parameter optimization were conducted via Response Surface Methodology (RSM) using the Central Composite Design (CCD) model. The maximum biohydrogen reached 3.44 g/L with 55% COD removal. The biohydrogen yield was 7.163 g H2/kg CODreduced with a maximum production rate of 712 mL/L/h. Further, the waste fermentation medium or spent media was used as a substrate in a Microbial Fuel Cell (MFC) to produce power using Pseudomonas aeruginosa PA1_NCHU as inoculum. MFCs were operated with various concentrations of phosphate buffer in the anolyte. As the output is limited in a single MFC, MFCs were operated in parallel stacks to increase power output. A maximum of 28% increase in the power density was observed in stacked MFCs. The energy recovered from the dark fermentation process was around 10.39% and a single MFC was around 10.67%. Hence, this study highlights the innovative use of agricultural waste and the effective combination of dark fermentation with stacked MFC, presenting a sustainable method of maximizing biohydrogen production and bioelectricity from spent media. By leveraging stacked MFC configuration, the potential of higher power output is demonstrated, underscoring the significance of this integrated system for energy recovery.

Pioneering simultaneous determination of breast cancer medications and metformin through sustainable micellar electrokinetic chromatographic method

The current study is the first report for the capillary electrophoretic simultaneous determination of five breast cancer medications, including febuxostat, quercetin, letrozole and doxorubicin in addition to metformin which is frequently co-administered with these drugs. The studied drugs are frequently purchased in different co-formulations used for breast cancer patients. A green micellar electrokinetic chromatographic (MEKC) method is proposed and optimized assisted by factorial design. The factors optimized include pH and concentration of buffer, surfactant concentration, as well as the applied voltage and injection time. The experimental results prove the optimum separation of the five drugs at pH 9.7 and buffer concentration of 10 mM with sodium dodecyl sulfate (SDS) concentration of 40 mM with +25 kV applied voltage. The developed method is validated considering linearity, precision, accuracy, and robustness adopting official guidelines. Good linearity is observed for all drugs, with correlation coefficients greater than 0.99. The range of percentage recoveries was from 98% to 102% with less than 2% relative standard deviation demonstrating the precision and accuracy of the established method. The study includes in-vitro assay of febuxostat, metformin, and doxorubicin in spiked human plasma. In addition, Accurate determination of the analytes under study in a variety of pharmaceutical dosage forms is accomplished by application of the proposed MEKC method. The greenness profile of the investigated method is confirmed using analytical eco-scale in addition to green analytical procedure index (GAPI) approaches.