Search Results (51)

This study proposes a method for separating asphalt and aggregates in recycled asphalt pavement (RAP) materials using surfactants as solvents. This method utilizes surfactants to soften the asphalt by reducing its surface tension, separating the RAP clusters, and washing away the asphalt from the RAP. The wastewater is recycled during the emulsification–separation process without discharge. Factors affecting the separation effect of RAP, including the type of anionic surfactants, the surfactant concentration, the emulsion-to-RAP ratio, temperature, the rotation rate and time, and the RAP’s particle size, were investigated in depth, and the separation effect and its influence on the aggregate properties were evaluated. The experimental results indicate that when using the optimal process to mix and treat 13.2 mm and 9.5 mm RAP clusters, it is possible to achieve 100% separation of the coarse RAP above 4.75 mm, with a 64.58% reduction in the asphalt content. The angularity of the aggregate remained unchanged after separation. It was observed from scanning electron microscopy (SEM) images that the asphalt on the surface of the coarse aggregate had been eluted, and the morphology of the aggregate surface was completely exposed. This environmentally friendly separation method provides new possibilities for high-content RAP recycling in pavement engineering. Full article

Background/Objectives: PURE (Protein synthesis Using Recombinant Elements), an ideal system for ribosome display, has been successfully used for nanobody selection. However, its limitations in nanobody selection, especially for synthetic nanobody libraries, have not been clearly elucidated, thereby restricting its utilization. Methods: The PURE ribosome display selection process was closely monitored using RNA agarose gel electrophoresis to assess the presence of mRNA molecules in each fraction, including the flow-through, washing, and elution fractions. Additionally, a real-time validation method for monitoring each biopanning round was implemented, ensuring the successful enrichment of target protein-specific binders. The selection process was further optimized by introducing a target protein elution step prior to the EDTA-mediated disassembly, as well as by altering the immobilization surfaces. Finally, the efficiency of PURE ribosome display was enhanced by replacing the spacer gene. Results: The efficiency of PURE ribosome display was merely 4% with an unfavourable spacer gene. Using this spacer gene, EGFP- and human fatty acid-binding protein 4-specific nanobodies from a synthetic nanobody library were we successfully identified through optimizing the selection process. Choosing a spacer gene less prone to secondary structure formation increased significantly its efficiency in displaying synthetic nanobody libraries. Conclusions: Implementing a target protein elution step prior to EDTA-mediated disassembly and modifying the immobilization surfaces effectively increase selection efficiency. For PURE ribosome display, efficiency was further improved using a suitable spacer gene, enabling the display of large libraries. Full article

Dye intermediates are important industrial chemicals; there is a lack of systematic field experiments and relevant validation data regarding the remediation of groundwater contamination by dye intermediates. This study examines the eluting effects of alcohol eluting agents, non-ionic surfactants, and deionized water on 4-methoxy-2-nitroaniline (4M2N) in a contaminated aquifer medium from a historically polluted dye intermediate production site in northwest China. The findings indicate that alcohol eluting agents exhibit superior eluting effects compared to non-ionic surfactants. Under optimized conditions, including 60% n-propanol concentration, a liquid-to-solid ratio of 15:1, two eluting cycles, an elution pH of 3, and a 2 h eluting duration, the eluting concentration of 4-methoxy-2-nitroaniline reached 75.49 mg/kg, exceeding that of the composite eluting agent by two times more and deionized water by three times further. Analysis revealed that the liquid-to-solid ratio and number of eluting cycles are the primary factors influencing eluting efficiency. Field trials conducted using treated groundwater involved injecting 31,560 m³ of treated groundwater over 152 days, resulting in the extraction of 38,550 m³ and the removal of about 1887 kg of 4-methoxy-2-nitroaniline. The concentrations of contaminants in both pumping wells and monitoring wells exhibited a certain degree of increase at various times. Field applications of treated groundwater washing facilitated the release of 4-methoxy-2-nitroaniline from the aquifer medium, which significantly enhances remediation efficiency. This provides theoretical support for data analysis and the promotion of similar remediation efforts. Full article

Protein purification is a crucial step for various downstream applications like drug development, antibody preparation, and structure determination. The constant pursuit is for methods that are more efficient and cost-effective. We propose a novel approach using an elastin-like polypeptide (ELP) as an aggregation core that serves as an anchor between the beads in a chromatography column. In this method, a chilled sample containing a [target protein type] fusion protein is loaded onto a pre-equilibrated IMAC (immobilized metal affinity chromatography) column with a low-salt buffer. The column is then washed with a warm buffer containing high salt to remove impurities. Here, the key step involves warming the column above the ELP’s transition temperature (Tt), which triggers its aggregation. This aggregation is expected to trap the target protein tightly between the beads. Subsequently, a harsh wash with high salt and high imidazole can be applied to remove even persistent contaminants, achieving high protein purity. Finally, the temperature is lowered, and a cold, low-salt buffer is introduced to reverse the aggregation and elute the purified target protein. This method has the potential to eliminate the need for sophisticated chromatography systems while still achieving high protein purity. Full article

In this study, adsorbents based on molecularly imprinted polymers (MIPs) in two solid-phase extraction application forms, pipette tip and magnetic extraction, were used for the selective extraction of coumarins. The pipette-tip solid-phase extraction reduced solvent volumes; the magnetic MIP extraction was simple and effective for phase separation. Parameters affecting extraction, such as the amount of adsorbent, type of washing solvent, volume of the elution solvent, and extraction times for magnetic extraction, were optimized. The MIP-based adsorbents displayed high selectivity and extraction efficiency, resulting in recoveries ranging from 70.3 to 102.0% (RSD % less than 5.5%) for five coumarins under study, 6,7-dihydroxycoumarin-6-β-D-glucoside, coumarin, 7-methoxycoumarin, 6-methylcoumarin, and dicoumarol. The extracts were analyzed by high-performance liquid chromatography with diode array (DAD) and fluorescence (FLD) detectors, reaching limits of quantification of 0.5 and 0.9 µg·mL⁻¹ for coumarin and dicoumarol detected by DAD and 0.001–0.012 µg·mL⁻¹ for the other prohibited simple coumarins when used as a fragrance (detected by FLD). The proposed method was validated and its applicability was shown for the analysis of cosmetic samples like shower gel and perfume. Full article

Fatty acid analysis is an essential step in evaluating the potential of macroalgae for biodiesel production. An extraction method was developed to simultaneously analyze up to five types of biodiesel-fuel-related fatty acids (myristic acid, palmitic acid, cis-palmitvaccenic acid, stearic acid, and oleic acid) in macroalgae using liquid chromatography and tandem mass spectrometry (LC–MS/MS). Lypophilization and solid-phase extraction (SPE) techniques were applied to improve the extraction efficiency and effectively purify samples. The optimal conditions for SPE were set by comparing the recoveries according to the various solvent conditions for each step (loading, washing, and elution). In addition, the introduction of trimethylaminoethyl (TMAE) derivatives to the hydroxyl group of the target analyte increased the ionization efficiency and sensitivity. The derivatized samples were analyzed using the LC–MS/MS method with electrospray ionization in the positive and multiple-reaction monitoring modes. The target analytes were separated and detected within 13.5 min using a CAPCELL PAK C18 MGII S3 column. Gradient elution was performed using distilled water and acetonitrile containing 5 mM ammonium acetate. This method offers a reliable and sensitive tool for the analysis of macroalgae samples for their potential use in biodiesel production. To the best of our knowledge, this is the first report on the simultaneous determination of fatty acids in macroalgae using LC–MS/MS with TMAE derivatization. Full article

Washing machines are one of the tools that bring great convenience to people’s daily lives. However, washing machines that have been used for a long time often develop issues such as odor and mold, which can pose health hazards to consumers. There exists a conspicuous gap in our understanding of the microorganisms that inhabit the inner workings of washing machines. In this study, samples were collected from 22 washing machines in Shanghai, China, including both water eluted from different parts of washing machines and biofilms. Quantitative qualitative analysis was performed using fluorescence PCR quantification, and microbial communities were characterized by high-throughput sequencing (HTS). This showed that the microbial communities in all samples were predominantly composed of bacteria. HTS results showed that in the eluted water samples, the bacteria mainly included Pseudomonas, Enhydrobacter, Brevibacterium, and Acinetobacter. Conversely, in the biofilm samples, Enhydrobacter and Brevibacterium were the predominant bacterial microorganisms. Correlation analysis results revealed that microbial colonies in washing machines were significantly correlated with years of use and the type of detergent used to clean the washing machine. As numerous pathogenic microorganisms can be observed in the results, effective preventive measures and future research are essential to mitigate these health problems and ensure the continued safe use of these household appliances. Full article

Tissue fibrosis is characterized by chronic fibroblast activation and consequently excessive accumulation of collagen-rich extracellular matrix. In vitro microplate-based assays are essential to investigate the underlying mechanism and the effect of antifibrotic drugs. In this study, in the absence of a gold-standard method, we optimized a simple, cost-effective, Sirius Red-based colorimetric measurement to determine the collagen production of fibroblasts grown on 96-well tissue culture plates. Based on our findings, the use of a serum-free medium is recommended to avoid aspecific signals, while ascorbate supplementation increases the collagen production of fibroblasts. The cell-associated collagens can be quantified by Sirius Red staining in acidic conditions followed by alkaline elution. Immature collagens can be precipitated from the culture medium by acidic Sirius Red solution, and after subsequent centrifugation and washing steps, their amount can be also measured. Increased attention has been paid to optimizing the assay procedure, including incubation time, temperature, and solution concentrations. The resulting assay shows high linearity and sensitivity and could serve as a useful tool in fibrosis-related basic research as well as in preclinical drug screening. Full article

Nucleic acid amplification testing facilitates the detection of disease through specific genomic sequences and is attractive for point-of-need testing (PONT); in particular, the early detection of microorganisms can alert early response systems to protect the public and ecosystems from widespread outbreaks of biological threats, including infectious diseases. Prior to nucleic acid amplification and detection, extensive sample preparation techniques are required to free nucleic acids and extract them from the sample matrix. Sample preparation is critical to maximize the sensitivity and reliability of testing. As the enzymatic amplification reactions can be sensitive to inhibitors from the sample, as well as from chemicals used for lysis and extraction, avoiding inhibition is a significant challenge, particularly when minimising liquid handling steps is also desirable for the translation of the assay to a portable format for PONT. The reagents used in sample preparation for nucleic acid testing, covering lysis and NA extraction (binding, washing, and elution), are reviewed with a focus on their suitability for use in PONT. Full article

In the development of bioanalytical LC-MS methods for the determination of drugs in plasma samples in a clinical setting, adequate sample preparation is of utmost importance. The main goals are to achieve the selective extraction of the analytes of interest and attain thorough matrix removal while retaining acceptable ecological properties, cost-effectiveness, and high throughput. Solid-phase extraction (SPE) offers a versatile range of options, from the selection of an appropriate sorbent to the optimisation of the washing and elution conditions. In this work, the first SPE method for the simultaneous extraction of six anticancer drugs used in novel therapeutic combinations for advanced breast cancer treatment—palbociclib, ribociclib, abemaciclib, anastrozole, letrozole, and fulvestrant—was developed. The following sorbent chemistries were tested: octylsilyl (C8), octadecylsilyl (C18), hydrophilic–lipophilic balance (HLB), mixed-mode cation-exchange (MCX and X-C), and mixed-mode weak cation-exchange (WCX), with different corresponding elution solvents. The samples were analysed using LC-MS/MS, with a phenyl column (150 × 4.6 mm, 2.5 μm). The best extraction recoveries (≥92.3%) of all analytes were obtained with the C8 phase, using methanol as the elution solvent. The optimised method was validated in the clinically relevant ranges, showing adequate precision (inter-day RSD ≤ 14.3%) and accuracy (inter-day bias −12.7–13.5%). Finally, its applicability was successfully proven by the analysis of samples from breast cancer patients. Full article

Soil washing is a well-established remediation technology for treating soil contaminated with heavy metals. It involves the separation of contaminants from the soil using acidic washing agents. Nevertheless, the application of washing agents at high concentrations may lead to soil acidification and the destruction of the clay structure. To avert this problem, recently, a soil washing variant has been presented, which solely employs high-pressure water without any chemical solvents. However, the fine soil generated from soil washing at a high-pressure contains high levels of heavy metals and requires proper treatment. This study examines the use and applicability of natural aquaculture materials as stabilizing agents for treating heavy metals (Cu, Pb, and Zn) in fine soil generated by high-pressure soil washing. Three aquaculture materials were assessed, namely, cockle shells (CKS), scallop shells (SLS), and Asterias amurensis starfish (ASF). Each material was processed to yield three types of stabilizing agents: natural type (-#10 mesh), natural type (-#20 mesh), and calcined(C) type (-#10 mesh). Each stabilizing agent was added to the contaminated soil at a ratio of 0 to 10 wt%, and then, mixed with an appropriate amount of water. After wet curing for 28 days, the stabilization efficiency of Cu, Pb, and Zn was evaluated using 0.1 N HCl solution. The elution of heavy metals showed a decreasing trend with higher dosages of stabilizing agents. The calcined type (-#10) showed the highest stabilization efficiency, followed by the natural type (-#20) and natural type (-#10). In addition, a comparison of the efficiency of the different stabilizing agents showed that calcined ASF (CASF) had the highest stabilization efficiency, followed by calcined SLS (CSLS), calcined CKS (CCKS), natural ASF (NASF), natural SLS (NSLS), and natural CKS. Finally, analysis of samples exhibiting the highest stabilization efficiency by scanning electron microscopy–energy dispersive X-ray spectrometry (SEM–EDX) confirmed that the pozzolanic reaction contributed to the stabilization treatment. The results of this study demonstrate that heavy metal-contaminated fine soil, generated by high-pressure washing, can be remediated by stabilizing Cu, Pb, and Zn using waste aquaculture materials (CKS, SLS, and ASF), which are often illegally dumped into the sea or landfills and cause environmental damage. Full article

4-Vinylpyridine molecularly imprinted polymer (4-VPMIP) microparticles for mandelic acid (MA) metabolite as a major biomarker of exposure to styrene (S) were synthesized by bulk polymerization with a noncovalent approach. A common mole ratio of 1:4:20 (i.e., metabolite template: functional monomer: cross-linking agent, respectively) was applied to allow the selective solid-phase extraction of MA in a urine sample followed by high-performance liquid chromatography–diode array detection (HPLC-DAD). In this research, the 4-VPMIP components were carefully selected: MA was used as a template (T), 4-Vinylpyridine (4-VP) as a functional monomer (FM), ethylene glycol dimethacrylate (EGDMA) as a cross-linker (XL), and azobisisobutyronitrile (AIBN) as an initiator (I) and acetonitrile (ACN) as a porogenic solvent. Non-imprinted polymer (NIP) which serves as a “control” was also synthesized simultaneously under the same condition without the addition of MA molecules. Fourier transform infrared (FT-IR) spectroscopy and scanning electron microscopy (SEM) were used to characterize the imprinted and nonimprinted polymer to explain the structural and morphological characteristics of the 4-VPMIP and surface NIP. The results obtained from SEM depicted that the polymers were irregularly shaped microparticles. Moreover, MIPs surfaces had cavities and were rougher than NIP. In addition, all particle sizes were less than 40 µm in diameter. The IR spectra of 4-VPMIPs before washing MA were a little different from NIP, while 4-VPMIP after elution had a spectrum that was almost identical to the NIP spectrum. The adsorption kinetics, isotherms, competitive adsorption, and reusability of 4-VPMIP were investigated. 4-VPMIP showed good recognition selectivity as well as enrichment and separation abilities for MA in the extract of human urine with satisfactory recoveries. The results obtained in this research imply that 4-VPMIP might be used as a sorbent for MA solid-phase extraction (MISPE), for the exclusive extraction of MA in human urine. Full article

Monoclonal antibodies (mAbs), such as infliximab, are important treatment options for different diseases. Immunogenicity is a major risk, resulting in anti-drug antibodies (ADAs), being associated with adverse events and loss of response, influencing long-term outcomes. The development of ADAs against infliximab is primarily measured by immunoassays like radioimmunoassay (RIA). Although liquid chromatography-tandem mass spectrometry (LC-MS/MS) is increasingly utilized across different fields, this technique is currently not used for ADAs against infliximab measurements. Therefore, we developed the first LC-MS/MS method. Stable isotopically labeled infliximab antigen-binding fragments (SIL IFX F(ab’)₂) were used to bind and measure ADAs indirectly. Protein A magnetic beads were used to capture IgG, including ADAs, whereafter SIL IFX F(ab’)₂ was added for labeling. After washing, internal standard addition, elution, denaturation and digestion samples were measured by LC-MS/MS. Internal validation showed good linearity between 0.1 and 16 mg/L (R² > 0.998). Sixty samples were used for cross-validation with RIA, and no significant difference between ADA concentrations was found. The methods had high correlation (R = 0.94, p < 0.001) and excellent agreement, intraclass correlation coefficient = 0.912 (95% confidence interval 0.858–0.947, p < 0.001). We present the first ADA against the infliximab LC-MS/MS method. The method is amendable for quantifying other ADAs, making it applicable as a template for future ADA methods. Full article

(+)-catechin is one category of flavonoids in cocoa shell waste and it has been reported to have many health benefits. In order to isolate it from aqueous extracted solution of cocoa shell waste by solid phase extraction (SPE), a series of dual ionic liquids@ZIF8-covered silica were prepared as the sorbents. Regarding the operation conditions of SPE and the characteristic structure of (+)-catechin, ZIF8-covered silica was synthesized to establish a stable and porous substrate, and various dual ionic liquids with multiple properties were immobilized on substrate to obtain a high adsorption capacity. Different adsorption conditions were investigated and the highest adsorption capacity (58.0 mg/g) was obtained on Sil@ZIF8@EIM-EIM at 30 °C during 60.0 min. When the sorbent was applied in the SPE process, 96.0% of the total amount of (+)-catechin from cocoa shell waste can be isolated after several washing and elution steps. The satisfactory recoveries of 97.5–100.2% and RSDs of 1.3–3.2% revealed that the SPE process was accurate and precise. The stability of Sil@ZIF8@EIM-EIM was tested in water and the reusability was tested using repeated adsorption/desorption process. The results revealed that Sil@ZIF8@EIM-EIM as an efficient sorbent can isolate (+)-catechin from cocoa shell waste. Full article

This paper describes the synthesis of novel molecularly imprinted magnetic nano-beads for the selective extraction (MISPE) of zearalenone mycotoxin in river and tap waters and further analysis by high-performance liquid chromatography (HPLC) with fluorescence detection (FLD). A semi-covalent imprinting approach was achieved for the synthesis of the molecularly imprinted polymers (MIP). The nanoparticles were prepared by covering the starting Fe₃O₄ material with a first layer of tetraethyl orthosilicate (TEOS) and then with a second layer using cyclododecyl 2-hydroxy-4-(3-triethoxysilylpropylcarbamoyloxy) benzoate. The last was used with a dual role, template and functional monomer after the extraction of the template molecule. The material was characterized by transmission electron microscopy (TEM), X-ray diffraction (XRD) and Fourier transform infrared spectroscopies (FT-IR). The solid phase extraction was optimized in all the steps: loading, washing and elution. The optimal conditions allowed the determination of zearalenone in trace levels of 12.5, 25 and 50 µg L⁻¹ without significant differences between the fortified and found level concentrations. Full article