RESUMO
In this work, we compare two structurally near-amorphous rigid-rod polymersâpoly(indacenodithiophene-co-benzothiadiazole), p(IDT-BT), and poly(indacenodithiophene-co-benzopyrollodione), p(IDT-BPD)âwith orders of magnitude different mobilities to understand the effect charge carrier intrachain delocalization has on electronic transport. Quantum chemical calculations show that p(IDT-BPD) has a barrier to torsion that is significantly lower than that of p(IDT-BT) and is thus more likely to have reduced conjugation lengths. We utilize absorption and photoluminescence spectroscopy to characterize energetic disorder and show that p(IDT-BPD) has higher energetic disorder. Charge modulation spectroscopy (CMS) and model calculations are used to show that charge carriers are substantially delocalized in p(IDT-BT) and occupy near-uniform energetic environments. We find that mobility activated hopping barriers are similar in these two materials. Electronic structure calculations show that both intrachain and interchain couplings of monomer units are poor enough in p(IDT-BPD) that charge carriers collapse to single IDT units and transport via a through-space tunneling mechanism. This work highlights the remarkable charge transport properties of p(IDT-BT) by showing that high mobilities are achievable on device-relevant length scales with only 1D carrier delocalization.
RESUMO
Artificial nanozymes (enzyme-mimics), specifically metallic nanomaterials, have garnered significant attention recently due to their reduced preparation cost and enhanced stability in a wide range of environments. The present investigation highlights, for the first time, a straightforward green synthesis of biogenic platinum nanoparticles (PtNPs) from a natural resource, namely Prunella vulgaris (Pr). To demonstrate the effectiveness of the phytochemical extract as an effective reducing agent, the PtNPs were characterized by various techniques such as UV-vis spectroscopy, High-resolution Transmission electron microscopy (HR-TEM), zeta-potential analysis, Fourier-transform infrared spectroscopy (FTIR), and Energy dispersive spectroscopy (EDS). The formation of PtNPs with narrow size distribution was verified. Surface decoration of PtNPs was demonstrated with multitudinous functional groups springing from the herbal extract. To demonstrate their use as viable nanozymes, the peroxidase-like activity of Pr/PtNPs was evaluated through a colorimetric assay. Highly sensitive visual detection of H2O2 with discrete linear ranges and a low detection limit of 3.43 µM was demonstrated. Additionally, peroxidase-like catalytic activity was leveraged to develop a colorimetric platform to quantify glutamate biomarker levels with a high degree of selectivity, the limit of detection (LOD) being 7.00 µM. The 2,2-Diphenyl-1-picrylhydrazyl (DPPH) test was used to explore the scavenging nature of the PtNPs via the degradation of DPPH. Overall, the colorimetric assay developed using the Pr/PtNP nanozymes in this work could be used in a broad spectrum of applications, ranging from biomedicine and food science to environmental monitoring.
Assuntos
Antioxidantes , Ácido Glutâmico , Peróxido de Hidrogênio , Nanopartículas Metálicas , Platina , Prunella , Platina/química , Nanopartículas Metálicas/química , Peróxido de Hidrogênio/química , Peróxido de Hidrogênio/análise , Antioxidantes/química , Antioxidantes/análise , Antioxidantes/farmacologia , Prunella/química , Ácido Glutâmico/análise , Ácido Glutâmico/química , Extratos Vegetais/químicaRESUMO
This study provides the first experimental polarized intermolecular and intramolecular optical absorption components of field-induced polarons in regioregular poly(3-hexylthiophene-2,5-diyl), rr-P3HT, a polymer semiconductor. Highly aligned rr-P3HT thin films were prepared by a high temperature shear-alignment process that orients polymer backbones along the shearing direction. rr-P3HT in-plane molecular orientation was measured by electron diffraction, and out-of-plane orientation was measured through series of synchrotron X-ray scattering techniques. Then, with molecular orientation quantified, polarized charge modulation spectroscopy was used to probe mid-IR polaron absorption in the âω = 0.075 - 0.75 eV range and unambiguously assign intermolecular and intramolecular optical absorption components of hole polarons in rr-P3HT. This data represents the first experimental quantification of these polarized components and allowed long-standing theoretical predictions to be compared to experimental results. The experimental data is discrepant with predictions of polaron absorption based on an adiabatic framework that works under the Born-Oppenheimer approximation, but the data is entirely consistent with a more recent nonadiabatic treatment of absorption based on a modified Holstein Hamiltonian. This nonadiabatic treatment was used to show that both intermolecular and intramolecular polaron coherence break down at length scales significantly smaller than estimated structural coherence in either direction. This strongly suggests that polaron delocalization is fundamentally limited by energetic disorder in rr-P3HT.
RESUMO
As cases of multidrug resistant bacterial infections increase, scientists and clinicians around the world are increasingly turning to bacteriophages as alternatives to antibiotics. Even though our understanding of phage has increased significantly since the early days of its discovery, over a century ago, the currently used tools and technologies for phage purification for therapeutic applications are severely limited. Bacteriophages are produced by bacterial cultures, and impurities such as endotoxins must therefore be removed before clinical use. We present an anion exchange bind-and-elute membrane chromatographic method for purifying T7 bacteriophage from Escherichia coli culture supernatant that removes undesirable impurities, while ensuring a high viable phage count in the purified product. Our method does not involve the use of chemicals such as organic solvents and caesium chloride that could typically leave residual toxicity in the final product. It also does not require expensive equipment, such as an ultracentrifuge. Using our method, that is based on an in-house designed membrane module, 65% of viable T7 phage was recovered, and up to 94% endotoxins could be removed. The method, which took approximately 15 min, is rapid and scalable, and produces quite pure bacteriophage samples in a single step. It therefore potentially represents a major improvement over the status quo, and shows the way ahead for streamlining phage manufacturing for therapeutic use.
Assuntos
Bacteriófagos , Cromatografia , Endotoxinas , Ânions , SolventesRESUMO
Flow uniformity within the device is critically important in membrane chromatography. Recent studies have shown that the design of the device has a significant impact on flow uniformity, and thereby on separation efficiency. The main premise of this work is that computational fluid dynamics (CFD) could serve as a fast and inexpensive tool for preliminary optimization of the design of a membrane chromatography device. CFD also helps in identifying factors that affect flow uniformity. In this paper, CFD is used to compare the fluidic attributes of conventional membrane chromatography devices such as the stacked disc and radial flow devices with those of more recently developed ones such as the different versions of the laterally-fed membrane chromatography (LFMC) device. These are compared based on pulse tracer solute dispersion, which is a useful metric for measuring flow uniformity, and is thereby a good predictor of chromatographic separation performance. The poor separation performance typically observed with conventional membrane chromatography devices could be attributed to the high degree of solute dispersion within these devices. CFD is then used to analyze the impact of factors such as membrane aspect ratio, and channel dimensions on the performance of z2-laterally-fed membrane chromatography (z2LFMC) devices. The results discussed in the paper demonstrate that CFD could indeed serve as a powerful optimization and performance prediction tool for membrane chromatography.
Assuntos
Cromatografia , Hidrodinâmica , Cromatografia/métodos , Simulação por ComputadorRESUMO
Chronic pain is now included in the designation of chronic diseases, such as cancer, diabetes, and cardiovascular disease, which can impair quality of life and are major causes of death and disability worldwide. Pain can be treated using cannabinoids such as Δ9-tetrahydrocannabinol (Δ9-THC) and cannabidiol (CBD) due to their wide range of therapeutic benefits, particularly as sedatives, analgesics, neuroprotective agents, or anti-cancer medicines. While little is known about the pharmacokinetics of these compounds, there is increasing interest in the scientific understanding of the benefits and clinical applications of cannabinoids. In this review, we study the use of nanomaterial-based electrochemical sensing for detecting Δ9-THC and CBD. We investigate how nanomaterials can be functionalized to obtain highly sensitive and selective electrochemical sensors for detecting Δ9-THC and CBD. Additionally, we discuss the impacts of sensor pretreatment at fixed potentials and physiochemical parameters of the sensing medium, such as pH, on the electrochemical performance of Δ9-THC and CBD sensors. We believe this review will serve as a guideline for developing Δ9-THC and CBD electrochemical sensors for point-of-care applications.
Assuntos
Canabidiol , Canabinoides , Dor Crônica , Humanos , Canabidiol/uso terapêutico , Dronabinol , Dor Crônica/tratamento farmacológico , Qualidade de VidaRESUMO
Photoactive organic and hybrid organic-inorganic materials such as conjugated polymers, covalent organic frameworks (COFs), metal-organic frameworks (MOFs), and layered perovskites, display intriguing photophysical signatures upon interaction with light. Elucidating structure-photophysics-property relationships across a broad range of functional materials is nontrivial and requires our fundamental understanding of the intricate interplay among excitons (electron-hole pair), polarons (charges), bipolarons, phonons (vibrations), inter-layer stacking interactions, and different forms of structural and conformational defects. In parallel with electronic structure modeling and data-driven science that are actively pursued to successfully accelerate materials discovery, an accurate, computationally inexpensive, and physically-motivated theoretical model, which consistently makes quantitative connections with conceptually complicated experimental observations, is equally important. Within this context, the first part of this perspective highlights a unified theoretical framework in which the electronic coupling as well as the local coupling between the electronic and nuclear degrees of freedom can be efficiently described for a broad range of quasiparticles with similarly structured Holstein-style vibronic Hamiltonians. The second part of this perspective discusses excitonic and polaronic photophysical signatures in polymers, COFs, MOFs, and perovskites, and attempts to bridge the gap between different research fields using a common theoretical construct - the Multiparticle Holstein Formalism. We envision that the synergistic integration of state-of-the-art computational approaches with the Multiparticle Holstein Formalism will help identify and establish new, transformative design strategies that will guide the synthesis and characterization of next-generation energy materials optimized for a broad range of optoelectronic, spintronic, and photonic applications.
RESUMO
PEGylated proteins are usually purified using chromatographic methods, which are limited in terms of both speed and scalability. In this paper, we describe a microfiltration membrane-based hybrid method for purifying PEGylated proteins. Polyethylene glycol (or PEG) is a lower critical solution temperature polymer which undergoes phase transition in the presence of a lyotropic salt and forms micelle-like structures which are several microns in size. In the proposed hybrid method, the PEGylated proteins are first converted to their micellar form by the addition of a lyotropic salt (1.65 M ammonium sulfate). While the micelles are retained using a microfiltration membrane, soluble impurities such as the unmodified protein are washed out through the membrane. The PEGylated proteins thus retained by the membrane are recovered by solubilizing them by removing the lyotropic salt. Further, by precisely controlling the salt removal, the different PEGylated forms of the protein, i.e., mono-PEGylated and di-PEGylated forms, are fractionated from each other. Hybrid separation using two different types of microfiltration membrane devices, i.e., a stirred cell and a tangential flow filtration device, are examined in this paper. The membrane-based hybrid method for purifying PEGylated proteins is both fast and scalable.
RESUMO
Conventional hollow fiber hemodialyzers have a cylindrical shell-and-tube design. Due to their circular cross-section and radial flow distribution and collection in the headers, the flow of blood in the header as well as in the hollow fiber membranes is non-uniform. The creation of high shear stress and high shear rate zones or stagnation zones could result in problems, such as cell lysis and blood clotting. In this paper, a novel cuboid hemodialyzer design is proposed as an alternative to the conventional cylindrical hemodialyzer. The primary motivation behind the proposed design is to create uniform flow conditions and thereby minimize some of the above-mentioned adverse effects. The most salient feature of the proposed design is a cuboid shell within which the hollow fiber membrane bundle is potted. The lumen of the fibers is fed from one side using a flow distributor consisting of embedded primary and secondary channels, while the fibers are drained from the other side using a flow collector, which also has embedded primary and secondary channels. The flow characteristics of the lumen side of the cuboid hemodialyzer were compared with those of a conventional hemodialyzer based on computational fluid dynamics (CFD) simulations. The results of CFD simulations clearly indicated that the flow of liquid within the cuboid dialyzer was significantly more uniform. Consequently, the shear rate and shear stress were also more uniform. By adopting this new design, some of the problems associated with the conventional hemodialyzer design could potentially be addressed.
RESUMO
Recombinant SARS-CoV-2 trimeric spike protein produced by mammalian cell culture is a potential candidate for a COVID-19 vaccine. However, this protein is much larger than most typical biopharmaceutical proteins and its large-scale manufacture is therefore challenging. Particularly, its purification using resin-based chromatography is difficult as the diffusive transport of this protein to and from its binding site within the pores of the stationary phase particles is slow. Therefore, very low flow rates need to be used during binding and elution, and this slows down the purification process. Also, due to its large size, the binding capacity of this protein on resin-based media is low. Membrane chromatography is an efficient and scalable technique for purifying biopharmaceuticals. The predominant mode of solute transport in a membrane is convective and hence it is considered better than resin-based chromatography for purifying large proteins. In this paper, we propose a membrane chromatography-based purification method for fast and scalable manufacture of recombinant SARS-CoV-2 trimeric spike protein. A combination of cation exchange z2 laterally-fed membrane chromatography and size exclusion chromatography was found to be suitable for obtaining a homogeneous spike protein sample from mammalian cell culture supernatant. The proposed method is both fast and scalable and could be explored as a method for manufacturing vaccine grade spike protein.
Assuntos
Proteínas Recombinantes , Glicoproteína da Espícula de Coronavírus , Vacinas contra COVID-19 , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/biossíntese , Glicoproteína da Espícula de Coronavírus/isolamento & purificação , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/isolamento & purificaçãoRESUMO
The fractionation of positional isomers of a PEGylated protein is quite challenging as these have similar molecular weight, and only very slightly different surface charge. In this study, cation exchange z2 laterally-fed membrane chromatography (z2LFMC), which has been shown to be suitable for high-speed, high-resolution protein purification, was used to fractionate positional isomers of mono-PEGylated lysozyme. The performance of the z2LFMC device was compared with a commercial preparative cation exchange column having the same volume and ligand. PEGylated lysozyme purification experiments showed that while the positional isomers of mono-PEGylated lysozyme could not be satisfactorily resolved using the preparative commercial cation exchange column, almost baseline resolution of these could be achieved using the z2LFMC device. Moreover, the z2LFMC device-based process was faster by an order of magnitude. The results discussed in this paper demonstrate that z2LFMC is a superior alternative to column-based chromatography for challenging protein separations, such as the one discussed here, both in terms of speed and resolution.
Assuntos
Muramidase , Polietilenoglicóis , Cromatografia por Troca Iônica/métodos , Peso Molecular , Polietilenoglicóis/química , Proteínas/químicaRESUMO
Non-polarizable empirical potentials have been proven to be incapable of capturing the mixing of methane-water mixtures at elevated pressures. Although density functional theory-based ab initio simulations may circumvent this discrepancy, they are limited in terms of the relevant time and length scales associated with mixing phenomena. Here, we show that the many-body MB-nrg potential, designed to reproduce methane-water interactions with coupled cluster accuracy, successfully captures this phenomenon up to 3 GPa and 500 K with varying methane concentrations. Two-phase simulations and long time scales that are required to fully capture the mixing, affordable due to the speed and accuracy of the MBX software, are assessed. Constructing the methane-water equation of state across the phase diagram shows that the stable mixtures are denser than the sum of their parts at a given pressure and temperature. We find that many-body polarization plays a central role, enhancing the induced dipole moments of methane by 0.20 D during mixing under pressure. Overall, the mixed system adopts a denser state, which involves a significant enthalpic driving force as elucidated by a systematic many-body energy decomposition analysis.
RESUMO
Extending on the previous work by Riera et al. [J. Chem. Theory Comput. 16, 2246-2257 (2020)], we introduce a second generation family of data-driven many-body MB-nrg models for CO2 and systematically assess how the strength and anisotropy of the CO2-CO2 interactions affect the models' ability to predict vapor, liquid, and vapor-liquid equilibrium properties. Building upon the many-body expansion formalism, we construct a series of MB-nrg models by fitting one-body and two-body reference energies calculated at the coupled cluster level of theory for large monomer and dimer training sets. Advancing from the first generation models, we employ the charge model 5 scheme to determine the atomic charges and systematically scale the two-body energies to obtain more accurate descriptions of vapor, liquid, and vapor-liquid equilibrium properties. Challenges in model construction arise due to the anisotropic nature and small magnitude of the interaction energies in CO2, calling for the necessity of highly accurate descriptions of the multidimensional energy landscape of liquid CO2. These findings emphasize the key role played by the training set quality in the development of transferable, data-driven models, which, accurately representing high-dimensional many-body effects, can enable predictive computer simulations of molecular fluids across the entire phase diagram.
RESUMO
This paper discusses ultrahigh-speed, ultrahigh-resolution preparative protein separation using an in-house designed membrane chromatography device. The performance of the membrane chromatography device was systematically compared with an equivalent resin-packed preparative column. Experiments carried out using model proteins showed that membrane chromatography gave more than four times greater resolution than the preparative column, while at the same time being more than 19 times faster. Membrane chromatography was therefore a better option, not only in terms of higher productivity but also in terms of higher product purity. Membrane chromatography was also superior in terms of resolving and presenting tracer impurity peaks in the chromatogram. Experiments carried out using monoclonal antibody samples showed that membrane chromatography was suitable for ultrahigh speed, and ultrahigh resolution fractionation of charge variants. This paper highlights and explains the need for proper device design for enabling the use of membrane chromatography for the efficient purification of protein biopharmaceuticals.
Assuntos
Produtos Biológicos , Anticorpos Monoclonais , Cromatografia/métodosRESUMO
We describe and discuss a simple dry-compression technique for preparing a flat cuboid chromatography device containing a shallow packed-bed of crystalline hydroxyapatite nanoparticles. We then discuss the use of this device for fast protein separation in the bind-and-elute mode. Such separation could be carried out at quite low pressures, making it possible to use inexpensive low pressure chromatography systems. In the flow rate range examined in this study, the pressure-drop across the device increased linearly with flow rate, indicating negligible media compaction during use. Using this device, binary protein mixtures could be separated in about a minute. Contrary to that observed in most packed-bed chromatographic separations, the width of the flow through and eluted peaks decreased with increase in flow rate. Therefore, both productivity and purity could be simultaneously increased by increasing flow rate. The suitability of this device for preparative protein separations was demonstrated by carrying out purification of a monoclonal antibody (Trastuzumab) from mammalian cell culture supernatant. This study opens up the possibility of developing dry-compression based flat cuboid packed-bed chromatography devices for fast preparative protein separation.
Assuntos
Durapatita , Nanopartículas , Animais , Anticorpos Monoclonais , Cromatografia/métodos , PressãoRESUMO
Engineered T cell therapies have revolutionized modern oncology, however processes for manufacturing T cell therapies vary and the impact of manufacturing processes On the cell product is poorly understood. Herein, we have used a commercially available hollow fiber membrane bioreactor (HFMBR) operated in a novel mode to demonstrate that T cells can be engineered with lentiviruses, grown to very high densities, and washed and harvested in a single, small volume bioreactor that is readily amenable to automation. Manufacturing within the HFMBR dramatically changed the programming of the T cells and yielded a product with greater therapeutic potency than T cells produced using the standard manual method. This change in programming was associated with increased resistance to cryopreservation, which is beneficial as T cell products are typically cryopreserved prior to administration to the patient. Transcriptional profiling of the T cells revealed a shift toward a glycolytic metabolism, which may protect cells from oxidative stress offering an explanation for the improved resistance to cryopreservation. This study reveals that the choice of bioreactor fundamentally impacts the engineered T cell product and must be carefully considered. Furthermore, these data challenge the premise that glycolytic metabolism is detrimental to T cell therapies.
Assuntos
Reatores Biológicos , Linfócitos T , HumanosRESUMO
We employ the Holstein model for polarons to investigate the relationship among defects, topology, Coulomb trapping, and polaron delocalization in covalent organic frameworks (COFs). We find that intrasheet topological connectivity and π-column density can override disorder-induced deep traps and significantly enhance polaron migration by several orders of magnitude in good agreement with recent experimental observations. The combination of percolation networks and micropores makes trigonal COFs ideally suited for charge transport followed by kagome/tetragonal and hexagonal structures. By comparing the polaron spectral signatures and coherence numbers of large three-dimensional frameworks having a maximum of 180 coupled chromophores, we show that controlling nanoscale defects and the location of the counteranion is critical for the design of new COF-based materials yielding higher mobilities. Our analysis establishes design strategies for enhanced conductivity in COFs that can be readily generalized to other classes of conductive materials such as metal-organic frameworks and perovskites.
RESUMO
Understanding the underlying physical mechanisms that govern charge transport in two-dimensional (2D) covalent organic frameworks (COFs) will facilitate the development of novel COF-based devices for optoelectronic and thermoelectric applications. In this context, the low-energy mid-infrared absorption contains valuable information about the structure-property relationships and the extent of intra- and inter-framework "hole" polaron delocalization in doped and undoped polymeric materials. In this study, we provide a quantitative characterization of the intricate interplay between electronic defects, domain sizes, pore volumes, chemical dopants, and three dimensional anisotropic charge migration in 2D COFs. We compare our simulations with recent experiments on doped COF films and establish the correlations between polaron coherence, conductivity, and transport signatures. By obtaining the first quantitative agreement with the measured absorption spectra of iodine doped (aza)triangulene-based COF, we highlight the fundamental differences between the underlying microstructure, spectral signatures, and transport physics of polymers and COFs. Our findings provide conclusive evidence of why iodine doped COFs exhibit lower conductivity compared to doped polythiophenes. Finally, we propose new research directions to address existing limitations and improve charge transport in COFs for applications in functional molecular electronic devices.
RESUMO
PEGylated proteins comprise a class of value-added biopharmaceuticals. High-resolution separation techniques are required for the purification of these molecules. In this study, we discuss the application of a newly developed z2 laterally-fed membrane chromatography (or z2LFMC) device for carrying out high-resolution purification of a PEGylated protein drug. The device used in the current study contained a stack of anion exchange (Q) membranes. The membrane bed-height of this z2LFMC device being small, it could be operated at very high flow rates, at relatively low back pressures. The primary goal was to speedily and efficiently separate a mono-PEGylated protein from impurities present in the PEGylation reaction mixture. A resin-based anion exchange column having the same ligand and bed-volume was used as the control device. The purification performance of the z2LFMC device and the control column were compared terms of resolution, recovery and purity. The z2LFMC device outperformed the control column in terms of every metric compared in this study. Higher purity (85.4% as opposed to 77.9%) and higher recovery (28% greater) of the target mono-PEGylated protein were obtained using the z2LFMC device at 20-time higher speed. These results clearly demonstrate that the z2LFMC device could be a faster and more efficient alternative to resin-based columns for purification of biopharmaceuticals.
Assuntos
Química Farmacêutica , Cromatografia , Polietilenoglicóis , Proteínas , Produtos Biológicos/isolamento & purificação , Química Farmacêutica/métodos , Polietilenoglicóis/química , Proteínas/química , Proteínas/isolamento & purificaçãoRESUMO
Simultaneously reducing the bed-height and increasing the area of cross-section, while keeping the bed-volume the same, would substantially reduce the pressure drop across a process chromatography column. This would minimize problems such as resin compaction and non-uniformity in column packing, which are commonly faced when using soft chromatographic media. However, the increase in macroscale convective dispersion due to the increase in column diameter, and the resultant loss in resolution would far outweigh any potential benefit. Cuboid-packed bed devices have lower macroscale convective dispersion compared to their equivalent cylindrical columns. In this paper, we discuss how and why a flat cuboid chromatography device having a short bed-height gives better protein separation, at a significantly lower pressure drop, than a taller column having the same bed-volume. First, we explored this option based on computational fluid dynamic (CFD) simulations. Depending on the flow rate, the pressure drop across the flat cuboid device was lower than that in the tall column by a factor of 6.35 to 6.4 (i.e. less than 1/6th the pressure). The CFD results also confirmed that the macroscale convective dispersion within the flat cuboid device was significantly lower. Head-to-head separation experiments using a 1 mL flat cuboid device having a bed-height of 10 mm, and a 1 mL tall column having a bed-height of 25.8 mm, both packed with the same chromatographic media, were carried out. The number of theoretical plates per unit bed-height was on an average, around 2.5 time times greater with the flat cuboid device, while the total number of theoretical plates in the two devices were comparable. At any given superficial velocity, the height equivalent of a theoretical plate in the tall column was on an average, higher by a factor 2.5. Binary protein separation experiments showed that at any given flow rate, the resolution obtained using the flat cuboid device was significantly higher than that obtained with the tall column. This work opens up the possibility of designing and developing short bed-height chromatography devices for carrying out high-resolution biopharmaceutical purifications, at very low pressures.