Carbon and graphene quantum dots based architectonics for efficient aqueous decontamination by adsorption chromatography technique - Current state and prospects.

Aquatic ecosystems and potable water are being exploited and depleted due to urbanization and the encouragement of extensive industrialization, which induces the scarcity of pure water. However, current decontamination methods are limited and inefficient. Various innovative remediation strategies with novel nanomaterials have recently been demonstrated for wastewater treatment. Carbon dots (C-dots) and graphene quantum dots (GQ-dots) are the most recent frontiers in carbon nanomaterial-based adsorption studies. C-dots are extremely small (1-10 nm) quasi-spherical carbon nanoparticles (mostly sp3 hybridized carbon), whereas GQ-dots are fragments of graphene (1-20 nm) composed of primarily sp2 hybridized carbon. This article highlights the function of C-dots and GQ-dots with their specifications and characteristics for the efficient removal of organic and inorganic contaminants in water via adsorption chromatography. The alteration of adsorption attributes with the hybrid blending of these dots has been critically analyzed. Moreover, various top-down and bottom-up approaches for synthesizing C-dots and GQ-dots, which ultimately affect their morphology and structure, are described in detail. Finally, we review the research deficit in the adsorption of diverse pollutants, fabrication challenges, low molecular weight, self-agglomeration, and the future of the dots by providing research prospects and selectivity and sensitivity perspectives, the importance of post-adsorption optimization strategies and the path toward scalability at the tail of the article.

Rapid indirect separation of glutamine enantiomers by micellar electrokinetic chromatography. Analysis of dietary supplements.

Glutamine is the most abundant free proteinogenic α-amino acid. It is naturally produced in the organism and acts as a precursor for the synthesis of different biologically important molecules (such as proteins or nucleotides). However, under stressful conditions, the organism is unable to produce it in enough amounts to function properly. Thus, glutamine (Gln)-based supplements have become increasingly popular over the last decade. Since legal regulations establish that amino acid-based dietary supplements must contain only the L-enantiomer and not the racemate, adequate chiral methodologies are required to achieve their quality control. In this work, an analytical methodology based on the use of micellar electrokinetic chromatography is proposed for the rapid enantiomeric determination of DL-Gln in dietary supplements. Using (+)-1-(9-fluorenyl)-ethyl chloroformate as a derivatizing agent and ammonium perfluorooctanoate as separation medium, the Gln diastereoisomers formed under optimal conditions were separated in 8 min with a resolution of 2.8. The analytical characteristics of the method were evaluated in terms of linearity, precision, accuracy, and limits of detection/quantitation, and they were found appropriate for the analysis of L-Gln-based dietary supplements.

Factors affecting therapeutic protein purity and yield during chromatographic purification.

Therapeutic proteins are recombinant proteins generated through recombinant DNA technology and have attracted a great deal of interest in numerous applications, including pharmaceutical, cosmetic, human and animal health, agriculture, food, and bioremediation. Producing therapeutic proteins on a large scale, mainly in the pharmaceutical industry, necessitates a cost-effective, straightforward, and adequate manufacturing process. In industry, a protein separation technique based mainly on protein characteristics and modes of chromatography will be applied to optimize the purification process. Typically, the downstream process of biopharmaceutical operations may involve multiple chromatography phases that require the use of large columns pre-packed with resins that must be inspected before use. Approximately 20% of the proteins are assumed to be lost at each purification stage during the production of biotherapeutic products. Hence, to produce a high quality product, particularly in the pharmaceutical industry, the correct approach and understanding of the factors influencing purity and yield during purification are necessary.

Evaluation of Physicochemical Properties of Ipsapirone Derivatives Based on Chromatographic and Chemometric Approaches.

Drug discovery is a challenging process, with many compounds failing to progress due to unmet pharmacokinetic criteria. Lipophilicity is an important physicochemical parameter that affects various pharmacokinetic processes, including absorption, metabolism, and excretion. This study evaluated the lipophilic properties of a library of ipsapirone derivatives that were previously synthesized to affect dopamine and serotonin receptors. Lipophilicity indices were determined using computational and chromatographic approaches. In addition, the affinity to human serum albumin (HSA) and phospholipids was assessed using biomimetic chromatography protocols. Quantitative Structure-Retention Relationship (QSRR) methodologies were used to determine the impact of theoretical descriptors on experimentally determined properties. A multiple linear regression (MLR) model was calculated to identify the most important features, and genetic algorithms (GAs) were used to assist in the selection of features. The resultant models showed commendable predictive accuracy, minimal error, and good concordance correlation coefficient values of 0.876, 0.149, and 0.930 for the validation group, respectively.

Isolation of Extracellular Outer Membrane Vesicles (OMVs) from Escherichia coli Using EVscore47 Beads.

Outer membrane vesicles (OMVs) are attractive for biomedical applications based on their intrinsic properties in relation to bacteria and vesicles. However, their widespread use is hampered by low yields and purities. In this study, EVscore47 multifunctional chromatography microspheres were synthesized and used to efficiently isolate functional OMVs from Escherichia coli. Through this technology, OMV loss can be kept to a minimum, and OMVs can be harvested using EVscore47 at 11-fold higher yields and ~13-fold higher purity than those achieved by means of ultracentrifugation. Based on the results presented here, we propose a novel EVscore47-based isolation of OMVs that is fast and scalable.

Advancements and greenification potential of magnetic molecularly imprinted polymers for chromatographic analysis of veterinary drug residues in milk.

Milk, as a widely consumed nutrient-rich food, is crucial for bone health, growth, and overall nutrition. The persistent application of veterinary drugs for controlling diseases and heightening milk yield has imparted substantial repercussions on human health and environmental ecosystems. Due to the high demand, fresh consumption, complex composition of milk, and the potential adverse impacts of drug residues, advanced greener analytical methods are necessitated. Among them, functional materials-based analytical methods attract wide concerns. The magnetic molecularly imprinted polymers (MMIPs), as a kind of typical functional material, possess excellent greenification characteristics and potencies, and they are easily integrated into various detection technologies, which have offered green approaches toward analytes such as veterinary drugs in milk. Despite their increasing applications and great potential, MMIPs' use in dairy matrices remains underexplored, especially regarding ecological sustainability. This work reviews recent advances in MMIPs' synthesis and application as efficient sorbents for veterinary drug extraction in milk followed by chromatographic analysis. The uniqueness and effectiveness of MMIPs in real milk samples are evaluated, current limitations are addressed, and greenification opportunities are proposed. MMIPs show promise in revolutionizing green analytical procedures for veterinary drug detection, aligning with the environmental goals of modern food production systems.

[Ion chromatography for the determination of n-butylamine in the workplace air].

Objective: To establish a method for the determination of n-butylamine in the air of the workplace by ion chromatography. Methods: In February 2022, on-site sampling was carried out using an atmospheric sampler. N-butylamine was adsorbed by a neutral silica gel tube and then performed for qualitative and quantitative determination by ion chromatography after ultrasonic desorption with 10 mmol/L sulfuric acid solution. Results: The linear range of the method was 0.0375-100.0 µg/ml, the linear equation of the standard curve was y=0.0713x-0.0327, the correlation coefficient was 0.9992. The detection limit of the method was 11.25 µg/L, and the lower limit of quantification was 37.50 µg/L, the lowest quantitative concentration was 0.025 mg/m(3) (in term of sampling 7.5 L). The average desorption efficiency of the method was 91.50%-95.38%, the precision was 1.10%-2.30%, the standard recovery was 83.83%-100.02%, sampling efficiency was 100.00%. Conclusion: This method is fast, sensitive and accurate, and can be used for the determination of n-butylamine in the air of workplace.

A versatile multimodal chromatography strategy to rapidly purify protein nanostructures assembled in cell lysates.

BACKGROUND: Protein nanostructures produced through the self-assembly of individual subunits are attractive scaffolds to attach and position functional molecules for applications in biomaterials, metabolic engineering, tissue engineering, and a plethora of nanomaterials. However, the assembly of multicomponent protein nanomaterials is generally a laborious process that requires each protein component to be separately expressed and purified prior to assembly. Moreover, excess components not incorporated into the final assembly must be removed from the solution and thereby necessitate additional processing steps. RESULTS: We developed an efficient approach to purify functionalized protein nanostructures directly from bacterial lysates through a type of multimodal chromatography (MMC) that combines size-exclusion, hydrophilic interaction, and ion exchange to separate recombinant protein assemblies from excess free subunits and bacterial proteins. We employed the ultrastable filamentous protein gamma-prefoldin as a material scaffold that can be functionalized with a variety of protein domains through SpyTag/SpyCatcher conjugation chemistry. The purification of recombinant gamma-prefoldin filaments from bacterial lysates using MMC was tested across a wide range of salt concentrations and pH, demonstrating that the MMC resin is robust, however the optimal choice of salt species, salt concentration, and pH is likely dependent on the protein nanostructure to be purified. In addition, we show that pre-processing of the samples with tangential flow filtration to remove nucleotides and metabolites improves resin capacity, and that post-processing with Triton X-114 phase partitioning is useful to remove lipids and any remaining lipid-associated protein. Subsequently, functionalized protein filaments were purified from bacterial lysates using MMC and shown to be free of unincorporated subunits. The assembly and purification of protein filaments with varying amounts of functionalization was confirmed using polyacrylamide gel electrophoresis, Förster resonance energy transfer, and transmission electron microscopy. Finally, we compared our MMC workflow to anion exchange chromatography with the purification of encapsulin nanocompartments containing a fluorescent protein as a cargo, demonstrating the versatility of the protocol and that the purity of the assembly is comparable to more traditional procedures. CONCLUSIONS: We envision that the use of MMC will increase the throughput of protein nanostructure prototyping as well as enable the upscaling of the bioproduction of protein nanodevices.

A Pragmatic Guide to Enrichment Strategies for Mass Spectrometry-Based Glycoproteomics.

Glycosylation is a prevalent, yet heterogeneous modification with a broad range of implications in molecular biology. This heterogeneity precludes enrichment strategies that can be universally beneficial for all glycan classes. Thus, choice of enrichment strategy has profound implications on experimental outcomes. Here we review common enrichment strategies used in modern mass spectrometry-based glycoproteomic experiments, including lectins and other affinity chromatographies, hydrophilic interaction chromatography and its derivatives, porous graphitic carbon, reversible and irreversible chemical coupling strategies, and chemical biology tools that often leverage bioorthogonal handles. Interest in glycoproteomics continues to surge as mass spectrometry instrumentation and software improve, so this review aims to help equip researchers with the necessary information to choose appropriate enrichment strategies that best complement these efforts.

Large-scale crystallization as an intermediate processing step in insulin downstream process: explored advantages and identified tool for process intensification.

The rising global prevalence of diabetes and increasing demand for insulin, calls for an increase in accessibility and affordability of insulin drugs through efficient and cost-effective manufacturing processes. Often downstream operations become manufacturing bottlenecks while processing a high volume of product. Thus, process integration and intensification play an important role in reducing process steps and time, volume reduction, and lower equipment footprints, which brings additional process efficiencies and lowers the production cost. Manufacturers thrive to optimize existing unit operation to maximize its benefit replacing with simple but different efficient technologies. In this manuscript, the typical property of insulin in forming the pH-dependent zinc-insulin complex is explored. The benefit of zinc chloride precipitation/crystallization has been shown to increase the in-process product purity by reducing the product and process-related impurities. Incorporation of such unit operation in the insulin process has also a clear potential for replacing the high cost involved capture chromatography step. Same time, the reduction in volume of operation, buffer consumption, equipment footprint, and capabilities of product long time storage brings manufacturing flexibility and efficiencies. The data and capabilities of simple operation captured here would be significantly helpful for insulins and other biosimilar manufacturer to make progresses on cost-effective productions.

Symmetrization of Peaks in Chiral Chromatography with an Area-Invariant Resolution Enhancement Method.

A majority of enantiomeric separations show some degree of peak asymmetry, which is detrimental to quantitative and semiquantitative chiral analysis. This paper presents a simple and rapid peak symmetrization algorithm for the correction or reduction of peak tailing or fronting in exponentially modified Gaussians. Raw chromatographic data can be symmetrized by adding a correct fraction of the first derivative to the chromatogram. The area remains invariant since the area under the first derivative is zero for a pure Gaussian and numerically close to zero for asymmetric peaks. A method of easily extracting the distortion parameter is provided, as well as insight into how pre-smoothing the data with the "perfect smoother" algorithm can suppress high frequencies effectively. The central difference method is also used to compute the first derivative, reducing root-mean-square noise by up to 28% compared to the standard forward difference method. A survey of 40 chiral separations is presented, demonstrating the range of asymmetry observed in chiral separations. Examples of symmetrization of the peaks from enantiomers in comparable and disproportionate concentrations are also provided. Artifacts of deconvolution are discussed, along with methods to mitigate such artifacts.

50-Fold Reduction of Separation Time in Open-Channel Hydrodynamic Chromatography via Lateral Vortices.

Despite its relatively long history, open-channel hydrodynamic chromatography (OC-HDC) still represents a niche technique for determining the size distribution of particle suspensions. Practical limitations of this separation method ultimately arise from the low eluent velocity that is necessary to contain the adverse increase of analyte bandwidth caused by Taylor-Aris dispersion. Because of the micrometric size of the channel cross section, the low eluent velocity translates into order of pL-per-minute flow rates, which introduce a challenge for both the injection and the detection systems. In this article, we provide theoretical/numerical evidence illustrating how a sizable reduction of the Taylor-Aris effect can be obtained by triggering cross-sectional vortices alongside the main pressure-driven axial flow. As a case study, we consider a square channel geometry where the lateral vortices are created by DC-induced electroosmosis. The analysis of particle separation is based on the classical excluded-volume macrotransport approach, which allows to derive the average particle velocity and the axial dispersion coefficient from the solution of two stationary advection-diffusion problems defined onto the channel cross section. We find that lateral vortices can enhance the separation efficiency quantitatively, e.g., by reducing the separation time of a two-species mixture by a 50-fold factor compared to standard OC-HDC.

Shape-Enhanced Open-Channel Hydrodynamic Chromatography.

Hydrodynamic chromatography (HDC) is a well-established analytical separation method for the size separation of nano- and microparticles and large molecular weight solutes such as synthetic polymers and proteins. We report on a theoretical study showing that the separation resolution of open-tubular HDC can be significantly enhanced by changing the cross-sectional shape of the separation channel. By enforcing Brenner's macro-transport approach, we provide theoretical/numerical evidence showing how the shape of the cross section influences quantitatively both the selectivity and the axial dispersion of the suspended particles in HDC. The separation performance of square-, triangle-, and star-shaped channel cross sections is compared to that of a cylindrical capillary over three decades of the particle Péclet number in terms of the minimal separation length and time to obtain the unit resolution of a two-particle mixture. Enhancement factors up to 400% are found in the case of triangular shapes, with the best performing case being the 70.6° angle, which can be obtained by KOH etching of bulk silicon.

Systematic assessment of process analytical technologies for biologics.

The application of process analytical technology (PAT) for biotherapeutic development and manufacturing has been employed owing to technological, economic, and regulatory advantages across the industry. Typically, chromatographic, spectroscopic, and/or mass spectrometric sensors are integrated into upstream and downstream unit operations in in-line, on-line, or at-line fashion to enable real-time monitoring and control of the process. Despite the widespread utility of PAT technologies at various unit operations of the bioprocess, a holistic business value assessment of PAT has not been well addressed in biologics. Thus, in this study, we evaluated PAT technologies based on predefined criteria for their technological attributes such as enablement of better process understanding, control, and high-throughput capabilities; as well as for business attributes such as simplicity of implementation, lead time, and cost reduction. The study involved an industry-wide survey, where input from subject matter industry experts on various PAT tools were collected, assessed, and ranked. The survey results demonstrated on-line liquid Chromatography (LC), in-line Raman, and gas analysis techniques are of high business value especially at the production bioreactor unit operation of upstream processing. In-line variable path-length UV/VIS measurements (VPE), on-line LC, multiangle light scattering (MALS), and automated sampling are of high business value in Protein A purification and polishing steps of the downstream process. We also provide insights, based on our experience in clinical and commercial manufacturing of biologics, into the development and implementation of some of the PAT tools. The results presented in this study are intended to be helpful for the current practitioners of PAT as well as those new to the field to gauge, prioritize and steer their projects for success.

Dihydrolucilactaene, a Potent Antimalarial Compound from Fusarium sp. RK97-94.

A recently discovered secondary metabolism regulator, NPD938, was used to alter the secondary metabolite profile in Fusarium sp. RK97-94. Three lucilactaene analogues were detected via UPLC-ESI-MS analysis in NPD938-treated culture. The three metabolites were successfully purified and identified as dihydroNG391 (1), dihydrolucilactaene (2), and 13α-hydroxylucilactaene (3) via extensive spectroscopic analyses. DihydroNG391 (1) exhibited weak in vitro antimalarial activity (IC50 = 62 µM). In contrast, dihydrolucilactaene (2) and 13α-hydroxylucilactaene (3) showed very potent antimalarial activity (IC50 = 0.0015 and 0.68 µM, respectively). These findings provide insight into the structure-activity relationship of lucilactaene and its analogues as antimalarial lead compounds.

Process development exploiting competitive adsorption-based displacement effects in monoclonal antibody aggregate removal-A new high-throughput screening procedure for membrane chromatography.

High-throughput screening (HTS) approaches are commonly used to accelerate downstream process development. Although most HTS approaches use batch isothermal data (KP screen) or bind and elute mode as screening procedure, different or new process designs are rarely investigated. In this paper, a mechanistic model case study for the separation of two different two-component solutions was conducted and confirmed prior evidence. With these outcomes, a novel HTS screening procedure was developed including the determination of competitive adsorption-based displacement effects and key parameter identification. The screening procedure employing an overload bind and elute (OBE) mode is presented in a case study dealing with IgG aggregate removal in a typical monoclonal antibody purification step, applying a Sartobind® S membrane adsorber (MA). Based on a MA scale down device, the OBE mode allows the determination of classical process parameters and dynamic effects, such as displacement effects. Competitive adsorption-based displacement effects are visualized by introducing a displacement identifier leading to a displacement process map. Based on this map, the approach is transferred to and confirmed by the OBE recycle experiments with 4.6 and 8.2 ml benchtop scsale devices resulting in 45% reduced IgG monomer and 88% increased higher molecular weight species binding capacities.

Ultrahigh-speed, ultrahigh-resolution preparative separation of protein biopharmaceuticals using membrane chromatography.

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.

Recent forensic applications of enhanced chromatographic separation methods.

This study reviews the recent applications of enhanced separation methods employed in forensic analysis utilizing gas chromatography, liquid chromatography, and supercritical fluid chromatography published between 2015 to 2020, except papers previously covered in relevant review articles. Applications of enhanced chromatographic separation methods to arson investigations, environmental forensics, sexual assault investigations, drug analysis, and toxicology are discussed. Future directions for enhanced chromatographic separation methods in forensic science are also explored.

A fast, single column high-performance anion exchange chromatography with pulsed amperometric detection method for the determination of saccharides in atmospheric aerosol samples.

Saccharides, especially anhydro sugars present in atmospheric aerosols, can be used as tracers to track sources of atmospheric aerosols. High-performance anion-exchange chromatography with pulsed amperometric detection is a commonly used technique for determining these saccharides, but the reported methods suffer from three drawbacks. First, to achieve separation of the complete set of atmospheric saccharides, run times are very long, typically longer than 60 minutes. Second, some methods require two columns to achieve the desired separation. Finally, in an era when electrolytic eluent preparation allows for excellent precision and accuracy, these methods require manually prepared eluents, which can lead to separation inconsistency for closely eluting analytes. These drawbacks make existing methods difficult to automate. To address this issue, we developed a fast method that uses only a single column for separation and electrolytically generated eluent that resolves 12 key atmospheric aerosol saccharides in 20 minutes. The resolved saccharides include anhydro sugars (levoglucosan, galactosan, and mannosan), sugar alcohols (erythritol, xylitol, and mannitol), and mono-/disaccharides (arabinose, galactose, glucose, mannose, fructose, and sucrose). To our knowledge, this report is the first instance of achieving such a significant reduction in run time with good resolution for this set of saccharides.

Current approaches for the purification of antibody-drug conjugates.

In the past two decades, antibody-drug conjugates have gained increasing attention because they expand the therapeutic index when compared with that of traditional chemotherapies. Antibody-drug conjugates are highly complex structures consisting of antibodies covalently conjugated with small-molecule cytotoxic drugs. The complex structure of antibody-drug conjugates makes chemistry, manufacturing, and control difficult. In contrast to antibody production, distinct purification methods following conjugation of antibodies with drug-linkers are required for the manufacturing. For process development of antibody drug conjugates, the drug-to-antibody ratio, free drug-linkers, and aggregates are critical quality attributes that must be strictly controlled and removed by appropriate purification techniques. In this review, features of various purification methods used to purify antibody drug conjugates are described and evaluated. The future landscape of the antibody-conjugates field is also discussed briefly.