Single Experiment Research Articles

Biogenic fabrication of spinel nickel ferrite imprinted on Bifurcaria bifurcata Macro-Alga activated carbon for the adsorption of ciprofloxacin and metronidazole

The rise in globalization and industrialization has led to an increase in population, increasing improperly treated discharge containing dyes, pharmaceuticals, pesticides etc in the water bodies. This has led to the need to remove these contaminants from water sources. The NiFe@BBAL adsorbent was developed and analyzed using various techniques such as FTIR, XRD, SEM, and XPS. The effectiveness of the NiFe@BBAL in adsorbing Ciprofloxacin (CIP) and metronidazole (MET) in water was tested. The adsorption of CIP and MET, both individually and in mixtures, was studied. The kinetic sorption experiments showed that the adsorption followed a pseudo-second-order model, with a higher R2 of 0.9934 for MET compared to 0.9916 for CIP. This suggests that the rate-limiting step is chemisorption. The primary adsorption mechanisms for both CIP and MET were hydrogen bonding, hydrophobic interactions, and electrostatic interactions, while CIP also involved electrostatic and metal complexation interactions. The NiFe@BBAL effluent showed no toxic effects on bacteria, indicating that no harmful material was leached in the effluent. The NiFe@BBAL demonstrated excellent performance and stability in removing 97.64% CIP and 97.62% MET in single contaminant experiments, and 57.33% CIP and 70.69% MET in cocktail mixture experiments. This can be attributed to the smaller structure of MET compared to CIP, allowing it better access to the active site, leading to higher adsorption. Furthermore, the repeated use of the adsorbent proved to be stable in the removal of CIP and MET over five cycles, demonstrating that the material is sustainable and suitable for large-scale experiments.

A Comprehensive Protocol for Microbead-Induced Ocular Hypertension in Mice.

Glaucoma is a common optic neuropathy characterized by degeneration of retinal ganglion cells (RGCs). Elevated intraocular pressure (IOP), that is, ocular hypertension, is the primary modifiable risk factor for glaucoma and the primary characteristic of most preclinical glaucoma models. Extensive genotype and phenotype diversity at relatively low cost and high accessibility makes laboratory mice an excellent preclinical model for glaucoma. The microbead occlusion model was introduced in 2010 as an inducible model of ocular hypertension in mice and is now one of the most extensively utilized models of rodent glaucoma. Subsequent modifications of the microbead model increased the magnitude and duration of IOP elevation, primarily through modification of injection materials. Despite its popularity, accessibility of the model is hindered by procedural consistency between users. Here we outline an updated and comprehensive protocol for execution of the microbead model that is focused on improving surgical and outcome measure consistency and on enabling single experimenter execution.

Optimization of White Tea Flavanol Extraction Process by the Ultrasonic‐Assisted Deep Eutectic Solvent Method and Determination of the Antioxidant and Antibacterial Activity

ABSTRACTFlavanols have a variety of health benefits and biological activities and become the hotspot of the food development and research. In this research, flavanols were extracted from white tea by ultrasonic‐assisted deep eutectic solvent (DES) method. High‐performance liquid chromatography was used to determine the content of flavanols. The influence of four factors on the flavanol extraction yield was analyzed by single factor experiments, including deep eutectic solvent mole ratio (the mole of hydrogen bond acceptors: the mole of hydrogen bond donors), solid–liquid ratio (white tea sample weight/g: the volume of DES/mL), extraction time and extraction temperature. On the basis of single factor experiments, the factors which had significant effects on the flavanol extraction yield were further optimized by the response surface test. The results showed that the optimized extraction conditions were as follows: the mole ratio of DES 1: 4 (choline chloride: 1,2‐propanediol), the solid–liquid ratio 1:30, the extraction temperature 56°C, the extraction time 53 min, and the ultrasonic power 341 W. Under the above parameters, the extraction yield of white tea flavanols was 9.63 mg/g. After purification with macroporous resin, antioxidation and antibacterial experiments were carried out. The results showed that the antioxidant and antibacterial effects of white tea flavanols were remarkable. The antibacterial effects performed best on Escherichia coli, followed by Bacillus subtilis and Staphylococcus aureus. The maximum diameter of the antibacterial zone on Escherichia coli was 0.95 ± 0.11 mm. The best radical scavenging rates of OH, ABTS+ and DPPH were 97.1%, 97.5%, and 88.4%, respectively, when the flavanol concentration was 9.5 mg/mL. The conclusions of this research can provide theoretical foundation for the further study of white tea flavanols.

Retention and interfacial failure mechanism of single diamond grains in resin-bonded grinding tools

Abrasive grains and the associated bonding agent are the two significant components in the manufacturing of fixed abrasive machining tools. The material properties and interfacial bonding behavior between the grains and the bonding matrix determine machining performance. In precision machining processes with diamond abrasives, the primary failure modes of fixed abrasive tools are grain dislodgement and premature loss, leading to abrupt change in machining load and ultimately causing inaccurate and inefficient machining performance. This study develops a comprehensive model for understanding abrasive grain retention and interfacial failure mechanisms in resin-bonded diamond tools. Finite element analysis of a single diamond grain embedded in a resin matrix was conducted to examine the influence of the grain shape, protruding height, and orientation angle on critical interfacial failure force. A series of single diamond scratching experiments validated the model, revealing that the maximum retention force reached 43.56 N for grains with a 0.9 mm protruding height and a 60° orientation angle. The results also show that, within a specific grain size range, grain shape—quantified by the sphere deviation coefficient proposed in this paper, has the greatest impact on retention and failure behavior. Protruding height plays a secondary role, while the contribution of orientation angle is minimal. These findings provide valuable insights for the design, manufacture, and optimization of precision abrasive machining tools, particularly for applications requiring high precision and reliability.

Diamond Surfaces with Lateral Gradients for Systematic Optimization of Surface Chemistry for Relaxometry - a Low-Pressure Plasma-Based Approach.

Diamond is increasingly popular because of its unique material properties. Diamond defects called nitrogen vacancy (NV) centers allow for measurements with unprecedented sensitivity. However, to achieve ideal sensing performance, NV centers need to be within nanometers from the surface and are thus strongly dependent on the local surface chemistry. Several attempts have been made to compare diamond surfaces. However, due to the high price of diamond crystals with shallow NV centers, a limited number of chemical modifications have been studied. Here, we developed a systematic method to investigate the continuity of different local environments with varying densities and natures of surface groups in a single experiment on a single diamond plate. To achieve this goal, we used diamonds with a shallow ensemble of NV centers and introduced a chemical gradient across the surface. More specifically, we used air and hydrogen plasma. The gradients were formed by a low-pressure plasma treatment after masking with a right-angled triangular prism shield. As a result, the surface contained gradually more oxygen/hydrogen toward the open end of the shield. We then performed wide-field relaxometry to determine the effect of surface chemistry on the sensing performance. As expected, relaxation times and thus sensing performance indeed vary along the gradient.

Image processing tools for petabyte-scale light sheet microscopy data.

Light sheet microscopy is a powerful technique for high-speed three-dimensional imaging of subcellular dynamics and large biological specimens. However, it often generates datasets ranging from hundreds of gigabytes to petabytes in size for a single experiment. Conventional computational tools process such images far slower than the time to acquire them and often fail outright due to memory limitations. To address these challenges, we present PetaKit5D, a scalable software solution for efficient petabyte-scale light sheet image processing. This software incorporates a suite of commonly used processing tools that are optimized for memory and performance. Notable advancements include rapid image readers and writers, fast and memory-efficient geometric transformations, high-performance Richardson-Lucy deconvolution and scalable Zarr-based stitching. These features outperform state-of-the-art methods by over one order of magnitude, enabling the processing of petabyte-scale image data at the full teravoxel rates of modern imaging cameras. The software opens new avenues for biological discoveries through large-scale imaging experiments.

Heteronuclear Polarization Transfer under Steady-State Conditions: The INEPT-SSFP Experiment.

NMR finds a wide range of applications, ranging from fundamental chemistry to medical imaging. The technique, however, has an inherently low signal-to-noise ratio (SNR)─particularly when dealing with nuclei having low natural abundances and/or low γs. In these cases, sensitivity is often enhanced by methods that, similar to INEPT, transfer polarization from neighboring 1Hs via J-couplings. In 1958, Carr proposed an alternative approach to increase NMR sensitivity, which involves generating and continuously detecting a steady-state transverse magnetization, by applying a train of pulses on an ensemble of noninteracting spins. This study broadens Carr's steady-state free precession (SSFP) framework to encompass the possibility of adding onto it coherent polarization transfers, allowing one to combine the SNR-enhancing benefits of both INEPT and SSFP into a single experiment. Herein, the derivation of the ensuing INEPT-SSFP (ISSFP) sequences is reported. Their use in 13C NMR and MRI experiments leads to ca. 300% improvements in SNR/ over conventional J-driven polarization transfer experiments, and sensitivity gains of over 50% over 13C SSFP performed in combination with 1H decoupling and NOE. These enhancements match well with numerical simulations and analytical evaluations. The conditions needed to optimize these new methods in both spectroscopic and imaging studies are discussed; we also examine their limitations, and the valuable vistas that, in both analytical and molecular imaging NMR, could be opened by this development.

The application of KMnO4 in reverse flotation separation of chalcopyrite and talc and its selective depression mechanism

Talc exhibits excellent floatability and, as a major silicate gangue mineral associated with chalcopyrite, significantly obstructs the purification and smelting of chalcopyrite. This study investigates the reverse flotation separation of chalcopyrite and talc using KMnO4 as a depressant and DDA (dodecylamine) as a collector within a reverse flotation system. Single mineral flotation experiments show that after adding KMnO4, the recovery of chalcopyrite in 38–74 μm is only 6.86 %, whereas the recovery of talc in 45–106 μm and 25–45 μm are 93.47 % and 89.25 %, respectively. Further experiments with artificially mixed ores demonstrate that KMnO4 effectively achieves the separation of chalcopyrite and talc. The depression mechanism of KMnO4 on chalcopyrite and talc was analyzed using Zeta potential, contact angle measurement, AFM (atomic force microscopy), FTIR (Fourier transform infrared spectroscopy), XPS (X-ray photoelectron spectroscopy), and DFT (density functional theory) calculations. Mechanistic analysis indicates that in an alkaline environment, chalcopyrite can undergo a redox reaction with KMnO4, forming hydrophilic hydroxide species (Cu(OH)2, Fe(OH)3) on the chalcopyrite surface, which depresses chalcopyrite flotation. In contrast, the surface properties of talc are relatively stable and difficult to react with KMnO4, which cannot depress the flotation of talc. Therefore, KMnO4 can be used as an efficient and selective depressant for the reverse flotation separation of chalcopyrite and talc.

Influencing factors of bentonite for the directional electric repair of landfill HDPE membrane leakage in calcium ion solution

The timely repair of high-density polyethylene (HDPE) membrane leakage in landfills is an important means of environmental risk control, which is critical for preventing leachate from polluting groundwater and soil. Electric repair technology is expected to become a hot spot in the leakage repair of HDPE films, because it can solve the issues of complex operation, high costs, and security risks during the operation and closure of landfills. To explore the influence law of various technical factors on the electric repair process, we carried out single factor experiments on the main influencing factors such as the operation time, bentonite concentration and dispersant/bentonite dosage ratio, leakage size, and calcium concentration. We then explored the best technological conditions. The results showed that accumulation at the leakage site possessed a good repair effect with a permeability coefficient of 4.22 × 10−6 cm/s. Furthermore, we observed that the Zeta potential played an important role in the electric repair technology, where the higher the absolute value of the Zeta potential, the faster the electrophoretic migration rate of the bentonite particles, and the lower the permeability coefficient of accumulation. This study may serve as a theoretical basis and offer scientific guidance for the directional electric repair of HDPE membrane leakage during the operating period and after the closure of landfills.

Optimisation of fermentation condition for production of hight monacolin k (mk) yield by monascus-strains

Monacolin K (MK), produced by Monascus fermentation, has significant hypolipidemic effects. In this study, under solid-state fermentation (SSF) quinoa was used as substrate to investigate the effect of the MK production by Monascus species quinoa. The results of the single factor experiment showed that the amount of the substrate loaded, addition of water to the substrate, and fermentation temperature change time have a significant effect on the MK production of Monascus-fermented quinoa. It was found that neither M. pilosus-fermented quinoa nor M. ruber-fermented quinoa produced citrinin. Consequently, these three factors were selected for the response surface optimization to determine the optimal fermentation process. The results showed that the optimal fermentation process of M. pilosus-fermented quinoa was as follows: quinoa loading amount of 30 g/300 ml, addition of 20 ml water to substrate, seed liquid inoculation volume of 10%, cultured at 30°C for 2 days, followed by a temperature change to 25°C and continued cultivation until 12 days. Under this fermentation process, the yield of MK reached 2.51 mg/g and without production of any citrinin. The optimal fermentation process of M. ruber-fermented quinoa was as follows: quinoa loading amount of 31 g/300 ml, water addition of substrate of 20 ml, seed liquid inoculation volume of 20%, cultured at 30°C for 2 days, followed by a temperature change to 25°C and continued cultivation until 12 days. Under this fermentation process, the yield of MK reached 3.22 mg/g and without any citrinin production. Bangladesh J. Bot. 53(3): 717-729, 2024 (September) Special

Adversarial sample attack method based on loss smoothing

Deep neural networks (DNNs) are vulnerable to adversarial examples.Although the existing momentum-based adversarial example generation method can achieve a close 100%white-box attack success rate, it is still not ideal when attacking other models, and the black- box attack success rate is low. To address this, an adversarial example attack method based on loss smoothing is proposed to improve the transferability of adversarial examples. In the iterative process of calculating the gradient at each step, the current gradient is not used directly, but the local average gradient is used to accumulate momentum, so as to suppress the local oscillation phenomenon on the loss function surface, thereby stabilizing the update direction and escaping the local extreme point. A large number of experimental results on the ImageNet dataset show that compared with the existing momentum-based method, the average black-box attack success rate of the proposed method in single model attack experiments is improved by 38.07%and 27.77%, and the average black-box attack success rate in integrated model attack experiments is improved by and.Rising32.50%and28.63%

Modeling photomolecular effect using generalized boundary conditions for Maxwell equations

We recently demonstrated via experiments in hydrogels and at a single air-water interface the photomolecular effect: photons directly cleaving off water molecular clusters in the visible spectrum where bulk water has negligible absorption. To model single interface experiments, here we re-derive generalized boundary conditions for Maxwell equations by assuming a transition region of the electromagnetic fields across the interface, leading naturally to the Feibelman parameters used before to describe surface photoelectric and surface plasmon effects on metals. This generalization leads to modifications of the Fresnel coefficients and an expression for the surface absorptance that can reasonably explain trends in our single-interface experimental data on the angle and polarization dependence of the beam deflection. Our work provides further support for the existence of the photomolecular effect, suggests that surface absorption should exist in many materials, and lays a foundation for assessing the impacts of such surface absorption based on the Maxwell equations.

Glucotoxicity suppresses function of pancreatic beta and duct cells via miR-335-targeted Runx2 and insulin-mediated mechanism.

Pancreatic cell dynamics have important contributions to the development of type 2 diabetes and related diseases such as nonalcoholic fatty pancreas disease. The aim of this study was to investigate the effects of prolonged excessive glucose exposure on the functions of pancreatic beta cells and duct cells in single and co-culture conditions. In this study, we focused on the effects of glucotoxicity on insulin secretion which is the main function of beta cells and on progenitor functions of duct cells. Rat primary INS1 beta cells and ARIP duct cells were exposed to glucose (25mM) for 72h under single or indirect co-culture conditions. Glucotoxicity stimuli increased insulin secretion and decreased insulin expression in single beta cells while stimulating beta-cell differentiation and adipogenesis in single duct cells. On the other hand, glucotoxicity caused functional loss and increased proliferation and apoptosis in beta cells while increasing proliferation but suppressed beta-cell differentiation and adipogenesis in duct cells under co-culture conditions. The expression level of miR-335, a microRNA known to be upregulated by leptin and target Runx2, was measured. As a result, unlike single-cell culture, glucotoxicity upregulated miR-335, downregulated Runx2, and decreased insulin signaling in beta cells while downregulating miR-335 and upregulating Runx2, and decreased insulin signaling in duct cells under co-culture conditions. When the results of single and co-culture experiments are compared, insulin and miR-335 may be seen as important mediators for setting up the relation between beta and duct cells. Our findings are important for preventing the development of type 2 diabetes and nonalcoholic fatty pancreas disease, even developing new diagnosis and treatment strategies.

Exotic mangrove Laguncularia racemosa litter input accelerates nutrient cycling in mangrove ecosystems.

Exotic plant litter presents different chemical and physical properties relative to native plant litter and alters ecosystem processes and functions that may facilitate exotic plant dispersal. However, these effects are largely unknown, especially within wetland ecosystems. This study examines whether introducing litter from theexotic mangrove Laguncularia racemosa could result in (1) accelerated community litter decomposition rates and increased nutrient cycling rates and (2)microbial community structure changes in the invaded areas. A single decomposition experiment using litterbags was conducted to examine the short-term effects of L. racemosa litter in the native mangrove forest ecosystem. The soil nutrients and microbial communities of Rhizophora stylosa, L. racemosa, and mixed forests were also compared to explore the long-term cumulative effects of L. racemosa litter in native ecosystems. The results indicated that L. racemosa has lower-quality leaf litter than R. stylosa and a significantly faster decomposition rate. This may result from changes in the soil microbial community structure caused by L. racemosa leaf litter input, which favors the decomposition of its own litter. Both the short-term and cumulative effect experiments demonstrated that L. racemosa leaf litter significantly increased the relative abundance of microbes related to litter decomposition, such as Proteobacteria and Bdellovibrionota, and enhanced the alpha diversity of soil fungi, thus creating a microbial environment conducive to L. racemosa leaf litter decomposition. Moreover, the accumulation of soil nutrients was lower under L. racemosa than under R. stylosa over several years. This may be related to the more rapid growth of L. racemosa, which causes soil nutrient absorption and storage within the plant tissues, thereby reducing the soil nutrient content. Inputting exotic mangrove L. racemosa leaf litter reduced the soil blue carbon content, potentially adversely affecting global climate change. L. racemosa may employ a unique strategy to lower soil nutrient levels in native mangroves based on its low-quality leaf litter, thereby weakening the competitive ability of native plants that are intolerant to low-nutrient conditions and enhancing its own competitive advantage to further spread into these areas. In summary, the input of exotic L. racemosa leaf litter accelerates nutrient cycling in local mangroves.

Quantitative proteomic mass spectrometry of protein kinases to determine dynamic heterogeneity of the human kinome.

The kinome is a dynamic system of kinases regulating signaling networks in cells and dysfunction of protein kinases contributes to many diseases. Regulation of the protein expression of kinases alters cellular responses to environmental changes and perturbations. We configured a library of 672 proteotypic peptides to quantify >300 kinases in a single LC-MS experiment using ten micrograms protein from human tissues including biopsies. This enables absolute quantitation of kinase protein abundance at attomole-femtomole expression levels, requiring no kinase enrichment and less than ten micrograms of starting protein from flash-frozen and formalin fixed paraffin embedded tissues. Breast cancer biopsies, organoids, and cell lines were analyzed using the SureQuant method, demonstrating the heterogeneity of kinase protein expression across and within breast cancer clinical subtypes. Kinome quantitation was coupled with nanoscale phosphoproteomics, providing a feasible method for novel clinical diagnosis and understanding of patient kinome responses to treatment.

Multiplexed chromatin immunoprecipitation sequencing for quantitative study of histone modifications and chromatin factors.

ChIP-seq is a widely used technique for studying histone post-translational modifications and DNA-binding proteins. DNA fragments associated with a specific protein or histone modification epitope are captured by using antibodies, sequenced and mapped to a reference genome. Albeit versatile and popular, performing many parallel ChIP-seq experiments to compare different conditions, replicates and epitopes is laborious, is prone to experimental variation and does not allow quantitative comparisons unless adequate spike-in chromatin is included. We present a detailed protocol for performing and analyzing a multiplexed quantitative chromatin immunoprecipitation-sequencing experiment (MINUTE-ChIP), in which multiple samples are profiled against multiple epitopes in a single workflow. Multiplexing not only dramatically increases the throughput of ChIP-seq experiments (e.g., profiling 12 samples against multiple histone modifications or DNA-binding proteins in a single experiment), but also enables accurate quantitative comparisons. The protocol consists of four parts: sample preparation (i.e., lysis, chromatin fragmentation and barcoding of native or formaldehyde-fixed material), pooling and splitting of the barcoded chromatin into parallel immunoprecipitation reactions, preparation of next-generation sequencing libraries from input and immunoprecipitated DNA and data analysis using our dedicated analysis pipeline. This pipeline autonomously generates quantitatively scaled ChIP-seq tracks for downstream analysis and visualization, alongside necessary quality control indicators. The entire workflow requires basic knowledge in molecular biology and bioinformatics and can be completed in 1 week. MINUTE-ChIP empowers biologists to perform every ChIP-seq experiment with an appropriate number of replicates and control conditions, delivering more statistically robust, exquisitely quantitative and biologically meaningful results.

Screening the characteristic hyperspectral wavelength variables of peanut leaves based on coupled algorithms to predict SPAD value in peanut

ABSTRACT The accurate acquisition and prediction of the chlorophyll content in peanuts can provide reference for scientifically sound planting and management of peanut crops. In this study, peanut canopy leaves analysed during the flowering and pegging period were taken as the research object, and the spectral data collected by a portable hyperspectral spectrometer were used as the data source to study the inversion model of peanut leaf soil and plant analyser development (SPAD). To address the issues of randomness, the need for multiple iterations, and strong subjectivity in threshold selection in the initial variable set of the random frog (RF) algorithm, an improved RF algorithm was developed based on the sparse group lasso tool. By analysing and comparing the results of variable extraction of the improved RF algorithm and six characteristic wavelength screening algorithms, three algorithms were selected for pairwise coupling. These six algorithms were the correlation coefficient method (CC), successive projections algorithm, iteratively retains informative variables (IRIV), uninformative variable elimination (UVE), competitive adaptive reweighed sampling, and a genetic algorithm. The results show that: (1) the results of CC, UVE, and IRIV algorithms are better in a single algorithm experiment; and (2) in the coupled algorithms experiment, CC-UVE, CC-IRIV, and UVE-IRIV methods can effectively reduce dimensions, and the model has a certain level of stability. Among them, the UVE-IRIV-Extreme gradient boosting model had the highest accuracy, R 2 = 0.652, RMSE = 1.867, and the variable compression rate was as high as 97.84%. The research results can provide a theoretical basis for rapid and accurate monitoring of the SPAD values of peanut leaves.

B-286 Single Experiment, Single Plate Solid Phase Extraction (SPE) Optimization using 47 Drug Analytes as an Example

Abstract Background SPE of biological matrices isolates analytes of interest from interfering matrix components that can affect LC-MS/MS quantitation. Optimal SPE conditions are dependent upon the sample matrix and analyte properties and typically require multiple experiments to determine optimal conditions for SPE extraction. Here, a 96-well method development plate was utilized to streamline method development for a panel of 47 clinical research drug analytes: 14 antipsychotics, 22 antidepressants and 11 anticonvulsants. Methods An SPE plate with four polymeric sorbents: reversed phase (X), strong and weak cation-exchange and weak anion-exchange mixed-mode phases (X-C, X-CW and X-AW) was used. The 47 drugs analytes were extracted on all four sorbents using three sets of conditions: acidic pretreatment/wash (aq) with basic elution (AB), neutral pretreatment/wash (aq) with neutral elution (NN), and basic pretreatment/wash (aq) with acidic elution (BA). A 50/50 methanol/water wash was also included. Elution used neutral or pH adjusted ethyl acetate/isopropanol or methanol/acetonitrile. A reagent blank, matrix blank, extracted serum samples spiked post-elution (n=2), and spiked serum (n=4) were extracted on each sorbent using each set of conditions. Optimal conditions were determined by calculating absolute percent recovery for each analyte. Results Recovery for antipsychotics was 85-97% using X-CW-AB, except Chlorpromazine (59%). It had better recovery (80%) using X-AB. Recovery for the antidepressants was 75-112% using X-C-AB, except Bupropion (35%) and Selegiline (10%). Bupropion had better recovery (83% and 95%) using X or X-CW and BA. Selegiline had better recovery using X, X-CW and X-AW with BA (83-92%). Anticonvulsants comprise zwitterionic, sulfonamides, neutral, acidic, and basic compounds with widely varying pKa and polarity. Targeting a single method for these analytes is difficult. The best recovery for anticonvulsants used X-CW-AB with several exceptions. Lacosamide, Felbamate, and Levetiracetam are neutral and among the most polar analytes and had 7-31% recovery under all SPE conditions tested. Zwitterionic Gabapentin and Pregabalin had 50-53% recovery from X-CW and X-C sorbents. The stronger protic elution (ammoniated methanol/acetonitrile vs ethyl acetate/isopropanol ) improved solubility for complete elution. Oxcarbazepine had 37% recovery from X-CW-AB, but recovery was 71% under BA conditions. Neutral, polar Zonisamide and Topiramate (Sulfonamide compounds) recovery was 25-26% using X-CW-AB. However, recovery was better (47% and 83%) under NN conditions, suggesting the analytes didn’t undergo complete elution from the X-CW sorbent. A stronger elution solvent would improve recovery from X-CW. Conclusions Most antipsychotics were recovered with X-CW-AB. Most antidepressants could be extracted using X-C-AB. Some analytes had higher recovery using hydrophobic interaction (X) SPE. A single mixed-mode SPE two-step elution protocol using a stronger elution solvent may be feasible. Anticonvulsants had mixed results. Some would require a stronger elution solvent. Some could be included in the other two panels to maximize recovery, but some had low recovery under all conditions tested. The method development plate screened 12 SPE methods for 47 analytes in a single experiment, greatly reducing the time and labor required to identify the best sorbent and starting conditions for SPE extraction optimization for drug analysis. This workflow could be implemented for other SPE method development projects.

Sustainable ferritin from bovine by-product liver as a potential resource: Ultrasound assisted extraction and physicochemical, structural, functional, and stable analysis

Ferritin is an iron-containing protein that is widely present in all organisms and has the function of regulating the metabolic balance of iron in organisms. It can be separated from animal by-product tissues and converted into value-added components to promote the development of a circular economy. This study established a method for extracting bovine liver ferritin (BLFer) using ultrasound-assisted buffer system. The optimal extraction conditions were determined through single factor and response surface optimization experiments as follows: ultrasound temperature of 70 °C, extraction time of 25 min, ultrasonic power of 400 W and solvent-to-solid ratio of 2 mL/g, the experimental value of the ferritin yield was 32.18 ± 0.12 g/kg. Compared with traditional methods, Ultrasound-assisted extraction improved the ferritin yield and enhanced its structural stability. Bioinformatics analysis revealed that BLFer is a hydrophilic protein with strong thermal stability. The BLFer has a ferrous oxidase active center, which plays an important role in the oxidative precipitation and reductive release of iron. It can dissociate under strong acids or bases but maintains good stability after heat treatment. These findings will help improve the comprehensive utilization of animal by-products, and promote the potential application of animal by-product ferritin in food and other industries.

Cell cycle visualization tools to study cardiomyocyte proliferation in real-time.

Cardiomyocytes in the adult human heart are quiescent and those lost following heart injury are not replaced by proliferating survivors. Considerable effort has been made to understand the mechanisms underlying cardiomyocyte cell cycle exit and re-entry, with view to discovering therapeutics that could stimulate cardiomyocyte proliferation and heart regeneration. The advent of large compound libraries and robotic liquid handling platforms has enabled the screening of thousands of conditions in a single experiment but success of these screens depends on the appropriateness and quality of the model used. Quantification of (human) cardiomyocyte proliferation in high throughput has remained problematic because conventional antibody-based staining is costly, technically challenging and does not discriminate between cardiomyocyte division and failure in karyokinesis or cytokinesis. Live cell imaging has provided alternatives that facilitate high-throughput screening but these have other limitations. Here, we (i) review the cell cycle features of cardiomyocytes, (ii) discuss various cell cycle fluorescent reporter systems, and (iii) speculate on what could improve their predictive value in the context of cardiomyocyte proliferation. Finally, we consider how these new methods can be used in combination with state-of-the-art three-dimensional human cardiac organoid platforms to identify pro-proliferative signalling pathways that could stimulate regeneration of the human heart.