Separations

18 pages, 929 KiB

Open AccessArticle

Improved Expression of Aggregation-Prone Tau Proteins Using a Spidroin-Derived Solubility Tag

by Kevin Muwonge, Bedri Yaman, Attila Mészáros, Giorgio Russo, Alexander Volkov and Peter Tompa

Separations 2024, 11(7), 198; https://doi.org/10.3390/separations11070198 - 25 Jun 2024

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Tauopathies, a group of neurodegenerative disorders, are characterized by the abnormal aggregation of microtubule-associated Tau proteins in neurons and glial cells. The process of Tau proteins transitioning from soluble, intrinsically disordered monomers to disease-associated aggregates is still unclear. Investigating these molecular mechanisms requires [...] Read more.

Tauopathies, a group of neurodegenerative disorders, are characterized by the abnormal aggregation of microtubule-associated Tau proteins in neurons and glial cells. The process of Tau proteins transitioning from soluble, intrinsically disordered monomers to disease-associated aggregates is still unclear. Investigating these molecular mechanisms requires the reconstitution of such processes in cellular and in vitro models using recombinant proteins at high purity and yield. However, the production of phase-separating or aggregation-prone recombinant proteins like Tau’s hydrophobic-rich domains or disease mutation-carrying variants on a large scale is highly challenging due to their limited solubility. To overcome this challenge, we have developed an improved strategy for expressing and purifying recombinant Tau proteins using the major ampullate spidroin-derived solubility tag (MaSp-NT*). This approach involves using NT* as a fusion tag to enhance the solubility and stability of expressed proteins by forming micelle-like particles within the cytosol of E. coli cells. We found that fusion with the NT* tag significantly increased the solubility and yield of highly hydrophobic and/or aggregation-prone Tau constructs. Our purification method for NT* fusion proteins yielded up to twenty-fold higher amounts than proteins purified using our novel tandem-tag (6xHis-SUMO-Tau-Heparin) purification system. This enhanced expression and yield were demonstrated with full-length Tau (hT40/Tau441), its particularly aggregation-prone repeat domain (Tau-MTBR), and Frontotemporal dementia (FTD)-associated mutant (Tau-P301L). These advancements offer promising avenues for the production of large quantities of Tau proteins suitable for in vitro experimental techniques such as nuclear magnetic resonance (NMR) spectroscopy without the need for a boiling step, bringing us closer to effective treatments for tauopathies. Full article

(This article belongs to the Special Issue Peptide Synthesis, Separation and Purification)

14 pages, 840 KiB

Open AccessArticle

Simultaneous LC-MS/MS Method for the Quantitation of Probenecid, Albendazole, and Its Metabolites in Human Plasma and Dried Blood Spots

by Mamunur Rashid, Yashpal S. Chhonker, Sandeep K. Singh and Daryl J. Murry

Separations 2024, 11(7), 197; https://doi.org/10.3390/separations11070197 - 25 Jun 2024

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Abstract

Millions of individuals throughout the world suffer from lymphatic filariasis (LF), which is a morbid disease caused by Wuchereria bancrofti, Brugia malayi, and Brugia timori. These infections belong to tissue-invading nematodes and are one of the major neglected tropical diseases [...] Read more.

Millions of individuals throughout the world suffer from lymphatic filariasis (LF), which is a morbid disease caused by Wuchereria bancrofti, Brugia malayi, and Brugia timori. These infections belong to tissue-invading nematodes and are one of the major neglected tropical diseases that often result in permanent and enduring disability among individuals in endemic regions. Due to combination therapy, LF eradication has drastically decreased infections globally. The development of blood micro-sampling techniques allowing precise quantitation of drugs in blood would facilitate pharmacokinetic (PK) studies in remote populations. Therefore, an LC-MS/MS bioanalytical method was utilized to analyze albendazole (ABZ), albendazole sulfone (ABZ-ON), albendazole sulfoxide (ABZ-OX), and probenecid (PR) in plasma and dried blood spots. Solid-phase extraction was utilized to extract the analyte from both plasma and blood-spiked DBS. Analytes of interest were eluted with a gradient mobile system using 0.05% formic acid in water (A) and 0.05% formic acid in methanol (B) and separated using a reversed-phase Acquity ^®BEH C18 UPLC column (100 × 2.1 mm, 1.7 µm). Precision and accuracy at each QC level were within the acceptable limit, i.e., ±15% for all analytes in both the matrices. Tests for stability under laboratory and storage conditions indicated that no notable changes were observed for plasma and DBS. The LC-MS/MS method demonstrated its capability to consistently identify all target analytes (ABZ, ABZ-ON, ABZ-OX, and PR) at low concentrations, even at the small specimen volumes obtained from DBS cards. This confirms the efficacy and durability of DBS cards as a micro-sampling technique. Moreover, it enhances collection efforts for therapeutic drug monitoring in remote locations for patients infected with lymphatic filariasis. Full article

(This article belongs to the Special Issue Advances in Chemical Fingerprint Assessment and Analytical Isolation of Natural Products and Pharmaceuticals)

21 pages, 5071 KiB

Open AccessReview

Summary of Pretreatment of Waste Lithium-Ion Batteries and Recycling of Valuable Metal Materials: A Review

by Linye Li, Yuzhang Li and Guoquan Zhang

Separations 2024, 11(7), 196; https://doi.org/10.3390/separations11070196 - 25 Jun 2024

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Abstract

The recycling of used lithium-ion batteries has become a growing concern. As a large number of rare metal elements are present in waste lithium-ion batteries, recycling them can significantly improve resource utilization and reduce the material cost of battery production. The process of [...] Read more.

The recycling of used lithium-ion batteries has become a growing concern. As a large number of rare metal elements are present in waste lithium-ion batteries, recycling them can significantly improve resource utilization and reduce the material cost of battery production. The process of recycling used lithium-ion batteries involves three main technology parts: pretreatment, material recovery, and cathode material recycling. Pretreatment includes discharge treatment, uniform crushing, and removing impurities. Material-recovery technology mainly involves traditional pyrometallurgical and hydrometallurgical technologies, as well as the developing biometallurgy technology. Analysis of existing data shows that pretreatment technology is crucial for the recycling of used lithium-ion batteries. Hydrometallurgical technology and pyro-hydrometallurgical technology are expected to be the most suitable industrialization technology paths in the future, with biometallurgical technology and direct recycling technology providing a low-pollution development direction. This article summarizes the different pretreatment techniques and valuable metal-recovery pathways. The advantages and disadvantages of each method were evaluated. The economic costs, environmental benefits, and degree of industrialization of each method were assessed. The possible development directions of various methods are summarized to provide reference for future research. Full article

(This article belongs to the Section Analysis of Energies)

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30 pages, 2013 KiB

Open AccessReview

Puerariae lobatae Radix: Progress in Extraction, Separation, and Pharmacological Activities Research

by Erjian Gao, Wei Wang, Yuanyuan Huang, Zhijie Luo, Bangzheng Chen, Siqiu Xiao and Dewen Li

Separations 2024, 11(7), 195; https://doi.org/10.3390/separations11070195 - 24 Jun 2024

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Abstract

Kudzu root (Puerariae lobatae Radix) is the tuberous root of Pueraria lobata, family Leguminosae. Kudzu root contains a variety of beneficial active ingredients such as puerarin, daidzin, daidzein, genistenin, 3′-hydroxy puerarin, β-sitosterol, stigmasterol, arachidic acid, and so on. Modern medical research [...] Read more.

Kudzu root (Puerariae lobatae Radix) is the tuberous root of Pueraria lobata, family Leguminosae. Kudzu root contains a variety of beneficial active ingredients such as puerarin, daidzin, daidzein, genistenin, 3′-hydroxy puerarin, β-sitosterol, stigmasterol, arachidic acid, and so on. Modern medical research shows that active ingredients in kudzu root are widely used clinically as raw materials for the treatment of hyperglycemia, non-alcoholic fatty liver disease, myocardial infarction, alcohol addiction, oxidative stress, inflammatory response, and retinal blockage due to their various pharmacological effects such as improving cardiovascular circulation, lowering blood lipids, lowering blood pressure, lowering blood sugar, being antipyretic, being estrogen-like, and relieving alcohol. China has rich resources of kudzu root, and active ingredients are usually extracted before it is made into a preparation, so whether the extraction and separation process is reasonable will directly affect the ease of preparation and the efficacy of the treatment. This paper reviews the process methods for the extraction and separation of active ingredients in kudzu root and its common pharmacological activities. The aim is to provide some references for readers to compare the advantages and disadvantages of various extraction and separation methods as well as understand the active ingredients and pharmacological activities of kudzu root. Full article

(This article belongs to the Special Issue Advances in Chemical Fingerprint Assessment and Analytical Isolation of Natural Products and Pharmaceuticals)

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15 pages, 2119 KiB

Open AccessArticle

Rapid Discovery of Antimicrobial and Antimalarial Agents from Natural Product Fragments

by Jianying Han, Xueting Liu, Lixin Zhang, Wesley C. Van Voorhis, Ronald J. Quinn and Miaomiao Liu

Separations 2024, 11(7), 194; https://doi.org/10.3390/separations11070194 - 23 Jun 2024

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Abstract

Fragment-based drug discovery (FBDD) focuses on small compounds, known as fragments, typically with a molecular weight of less than 300 Da. This study highlights the benefits of employing a pure natural product library for FBDD, contrasting with the predominant use of synthetic libraries. [...] Read more.

Fragment-based drug discovery (FBDD) focuses on small compounds, known as fragments, typically with a molecular weight of less than 300 Da. This study highlights the benefits of employing a pure natural product library for FBDD, contrasting with the predominant use of synthetic libraries. Practical methods for rapidly constructing such libraries from crude extracts were demonstrated across various plant and microbial samples. Twenty-nine (29) natural product fragments, including a new compound (20), were identified. Antimicrobial activities were assessed for a subset of the isolated compounds, revealing potent fragments (MICs 4–8 μg/mL) against Mycobacterium bovis bacille Calmette-Guérin (BCG), Staphylococcus aureus (SA), and methicillin-resistant S. aureus (MRSA). Furthermore, a native mass spectrometry technique was introduced to rapidly identify non-competitive fragments against malarial proteins. As a result, two pairs of non-competitive fragments, lepiotin C (31) and 7-amino deacetoxy cephalosporanic acid (32) binding to dynein light chain 1, methyl gallate (33) and β-santanin (34) binding to dUTPase, were identified, serving as promising starting points for developing potent malarial protein inhibitors. Full article

(This article belongs to the Section Analysis of Natural Products and Pharmaceuticals)

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11 pages, 658 KiB

Open AccessArticle

A Novel Solvent Microextraction Lab-in-Syringe System Coupled with Atomic Absorption Spectrometry for Thallium Determination in Water Samples

by Arina Skok, Natalia Manousi, Yaroslav Bazel, Andriy Vishnikin and Aristidis Anthemidis

Separations 2024, 11(7), 193; https://doi.org/10.3390/separations11070193 - 21 Jun 2024

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Abstract

Thallium is an accumulative highly toxic metal, that can be present in environmental samples due to industrial pollution and is dangerous for living organisms. Thus, its determination at trace levels is necessary. The lab-in-syringe (LIS) is considered to be a simple, functional, and [...] Read more.

Thallium is an accumulative highly toxic metal, that can be present in environmental samples due to industrial pollution and is dangerous for living organisms. Thus, its determination at trace levels is necessary. The lab-in-syringe (LIS) is considered to be a simple, functional, and versatile, technique that combines operational concepts and flow and sequential injection analysis. In this study, a liquid-phase microextraction LIS system was developed as a front-end to flame atomic absorption spectrometry (FAAS) for the determination of thallium in water samples. The proposed approach is based on the formation of Tl(III) ammonium–pyrrolidine–dithiocarbamate complex followed by its extraction using di-isobutyl-ketone. These procedures take place within the syringe barrel of the LIS system. The limit of detection of the developed method was 2.1 µg L⁻¹ with a linear range from 7.0 to 400 µg L⁻¹. The relative standard deviation (RSD) was 3.9% (at 50.0 µg L⁻¹ Tl(I)), demonstrating good precision. Moreover, good method accuracy was obtained since the relative recovery values were within the range of 93.4–101.2%. Finally, reliable method applicability and green merits were demonstrated using the blue applicability grade index and green analytical procedure index, respectively. The proposed method was used for the analysis of environmental water samples. Full article

(This article belongs to the Section Purification Technology)

13 pages, 4485 KiB

Open AccessArticle

Study on the Efficiency of Fine Particle Removal in a Single-Tower Dual-Cycle Desulfurization Process Utilizing Heterogeneous Condensation

by Rui Zhang, Zulpher Ahmad Mnipela, Linjun Yang and Xiaodong Si

Separations 2024, 11(7), 192; https://doi.org/10.3390/separations11070192 - 21 Jun 2024

Viewed by 317

Abstract

This study investigated a new method for controlling the emission of fine particles through heterogeneous condensation. Specifically, the research focuses on the application of single-tower double-cycle desulfurization technology in the wet flue gas desulfurization process. The establishment of a supersaturation environment necessary for [...] Read more.

This study investigated a new method for controlling the emission of fine particles through heterogeneous condensation. Specifically, the research focuses on the application of single-tower double-cycle desulfurization technology in the wet flue gas desulfurization process. The establishment of a supersaturation environment necessary for heterogeneous condensation was achieved by reducing the temperature of desulfurization slurry in the oxidation zone. Numerical simulations were used to study the distribution of the supersaturation degree and fluid dynamics characteristics in the desulfurization tower after the cooling of desulfurization slurry. Furthermore, the impact of single-tower double-cycle technology on the removal efficiency of fine particles was examined. The results of the numerical simulations indicate that cooling the desulfurization slurry in the absorption zone could establish a supersaturated vapor environment, with the supersaturation degree and region increasing as the slurry temperature decreases. Under typical operating conditions, a temperature drop of approximately 8~10 °C was found to be most suitable for the desulfurization slurry. Moreover, lowering the temperature of the desulfurization slurry in the absorption zone increases the supersaturation degree from 0.93 to 1.85. Additionally, the use of single-tower double-cycle desulfurization technology is shown to significantly enhance the removal efficiency of fine particles, particularly those within the particle size range of 0.1~1 μm. Ultimately, this method could increase the removal efficiency of fine particles from 39.9% to 57.9%. Full article

(This article belongs to the Collection Feature Paper Collection in Section 'Purification Technology')

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Separations, Volume 11, Issue 7 (July 2024) – 7 articles

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