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Exosomes, a subset of the extracellular vesicle (EV) group of organelles, hold great potential for biomarker detection, therapeutics, disease diagnosis, and personalized medicine applications. The promise and potential of these applications are hindered by the lack of an efficient means of isolation, characterization, and quantitation. Current methods for exosome and EV isolation (including ultracentrifugation, microfiltration, and affinity-based techniques) result in impure recoveries with regard to remnant matrix species (e.g., proteins, genetic material) and are performed on clinically irrelevant time and volume scales. To address these issues, a polyethylene terephthalate (PET) capillary-channeled polymer (C-CP) fiber stationary phase is employed for the solid-phase extraction (SPE) of EVs from various matrices using a micropipette tip-based format. The hydrophobic interaction chromatography (HIC) processing and a spin-down workflow are carried out using a table-top centrifuge. Capture and subsequent elution of intact, biologically active exosomes are verified via electron microscopy and bioassays. The performance of this method was evaluated by capture and elution of exosome standards from buffer solution and three biologically relevant matrices: mock urine, reconstituted non-fat milk, and exosome-depleted fetal bovine serum (FBS). Recoveries were evaluated using UV-Vis absorbance spectrophotometry and ELISA assay. The dynamic binding capacity (50%) for the 1-cm-long (~ 5 μL bed volume) tips was determined using a commercial exosome product, yielding a value of ~ 7 × 1011 particles. The novel C-CP fiber spin-down tip approach holds promise for the isolation of exosomes and other EVs from various matrices with high throughput, low cost, and high efficiency.

Presence of plastic debris in marine and freshwater ecosystems is increasingly reported. Previous research suggested plastic debris had a strong affiliation for many pollutants, such as polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), and heavy metals. In this study, the sorption behavior of pharmaceuticals and personal care products (PPCPs), including carbamazepine (CBZ), 4-methylbenzylidene camphor (4MBC), triclosan (TCS), and 17α-ethinyl estradiol (EE2), to polyethylene (PE) debris (250 to 280 μm) was investigated. The estimated linear sorption coefficients ( K d ) are 191.4, 311.5, 5140, and 53,225 L/kg for CBZ, EE2, TCS, and 4MBC, and are related to their hydrophobicities. Increase of salinity from 0.05 to 3.5 % did not affect the sorption of 4MBC, CBZ, and EE2 but enhanced the sorption of TCS, likely due to the salting-out effect. Increase of dissolved organic matter (DOM) content using Aldrich humic acid (HA) as a proxy reduced the sorption of 4MBC, EE2, and TCS, all of which show a relatively strong affiliation to HA. Results from this work suggest that microplastics may play an important role in the fate and transport of PPCPs, especially for those hydrophobic ones.

This study investigates key factors affecting the adsorption behavior of proteins on the multimodal chromatographic adsorbent Capto MMC, aiming to enhance selective protein separation strategies. Batch equilibrium experiments were conducted using six model proteins to explore the combined effects of pH, ionic strength, and the nature of salts (kosmotropic and chaotropic) on protein–ligand interactions. Given that the Capto MMC ligand supports multiple interaction modes beyond cation exchange, particular focus was placed on acidic proteins (pI 4–5), which exhibited binding even at moderately elevated pH values—conditions ineffective for conventional cation exchangers. Hydrophobic interactions were identified as critical for stable binding of proteins like BSA and fetuin, while hydrophilic proteins such as ovalbumin showed minimal adsorption. Chromatographic column experiments were performed to evaluate elution performance under various buffer conditions, revealing that prolonged adsorption phases can reduce recovery yields for proteins with less stable tertiary structures. The findings highlight how salt type, pH, and protein hydrophobicity interplay to modulate multimodal binding mechanisms, providing practical insights for the design of tailored purification protocols.

Extracellular vesicles (EVs) have garnered much interest due to their fundamental role in intracellular communication and their potential utility in clinical diagnostics and as biotherapeutic vectors. Of particular relevance is the subset of EVs referred to as exosomes, ranging in size from 30 to 150 nm, which contain incredible amounts of information about their cell of origin, which can be used to track the progress of disease. As a complementary action, exosomes can be engineered with therapeutic cargo to selectively target diseases. At present, the lack of highly efficient methods of isolation/purification of exosomes from diverse biofluids, plants, and cell cultures is a major bottleneck in the fundamental biochemistry, clinical analysis, and therapeutic applications. Equally impactful, the lack of effective in-line means of detection/characterization of isolate populations, including concentration and sizing, is limiting in the applications. The method presented here couples hydrophobic interaction chromatography (HIC) performed on polyester capillary-channeled polymer (C-CP) fiber columns followed by in-line optical absorbance and multi-angle light scattering (MALS) detection for the isolation and characterization of EVs, in this case present in the supernatant of Chinese hamster ovary (CHO) cell cultures. Excellent correlation was observed between the determined particle concentrations for the two detection methods. C-CP fiber columns provide a low-cost platform (< $5 per column) for the isolation of exosomes in a 15-min workflow, with complementary absorbance and MALS detection providing very high-quality particle concentration and sizing information.

We have investigated the dispersion state of a perfluorosulfonated ionomer (PFSI; Nafion®) in aqueous dispersion and the effect of methanol (MeOH) added to the aqueous dispersion by small-angle X-ray scattering (SAXS) as well as static and dynamic light scattering (SLS and DLS, respectively). Although both electrostatic and hydrophobic interactions of PFSI are expected to be strong in the dispersions, SAXS profiles obtained were satisfactorily fitted by the spherical particle model of a bimodal molar mass distribution. The rod-like aggregate model proposed in previous papers was denied at least for the present PFSI dispersion. Although the SAXS profiles exhibited a weak peak and the auto-correlation functions of DLS showed a log-time decay by the “repulsive cage effect” due to the long-ranged electrostatic interaction among PFSI particles, the concentration dependence of SLS results was probably normal because the cancellation of the electrostatic and hydrophobic interactions. The addition of MeOH into the aqueous dispersion of PFSI weakened both the hydrophobic and electcrostatic interactions of PFSI, and it is rather difficult to classify whether MeOH is a good or poor solvent (dispersant) for PFSI.

The study of intermolecular interactions between ethanol (E-OH), cinnamaldehyde (CAD) with anionic surfactant sodium dodecyl sulfate (SDS) in non-aqueous media has been examined by utilizing conductometric and spectroscopic techniques. The critical micelle concentration (CMC) values have been determined. The experimental conductance data were analyzed against temperature and concentration using standard relations. The pseudo phase separation model has been adopted to calculate various thermodynamic parameters like standard free energy, ∆G°mic, enthalpy, ∆H°mic, and entropy, ∆S°mic, of micelle formation. Fourier transforms infrared analysis (FTIR), and Fluorescence spectra were taken out to assess the possible interactions prevailing in the micellar systems. The findings demonstrated that the presence of SDS, and the composition of CAD + ethanol might affect the thermodynamic parameters. The discrepancy in these parameters with the surfactant concentration or with the temperature change indicates the manifestation of different interactions prevailing in the studied systems.

Polarization and charge-transfer interactions play an important role in ligand–receptor complexes containing metals, and only quantum mechanics methods can adequately describe their contribution to the binding energy. In this work, we selected a set of benzenesulfonamide ligands of human Carbonic Anhydrase II (hCA II)—an important druggable target containing a Zn2+ ion in the active site—as a case study to predict the binding free energy in metalloprotein–ligand complexes and designed specialized computational methods that combine the ab initio fragment molecular orbital (FMO) method and GRID approach. To reproduce the experimental binding free energy in these systems, we adopted a machine-learning approach, here named formula generator (FG), considering different FMO energy terms, the hydrophobic interaction energy (computed by GRID) and logP. The main advantage of the FG approach is that it can find nonlinear relations between the energy terms used to predict the binding free energy, explicitly showing their mathematical relation. This work showed the effectiveness of the FG approach, and therefore, it might represent an important tool for the development of new scoring functions. Indeed, our scoring function showed a high correlation with the experimental binding free energy (R2 = 0.76–0.95, RMSE = 0.34–0.18), revealing a nonlinear relation between energy terms and highlighting the relevant role played by hydrophobic contacts. These results, along with the FMO characterization of ligand–receptor interactions, represent important information to support the design of new and potent hCA II inhibitors.

The extraction of membrane proteins remain a challenge due to innate hydrophobicity, dynamic discrepancy, and restrain effect of membrane lipids. Nanomaterials with high surface area have competency of hydrophobic-hydrophobic lipid interactions. It is shown here that both graphene and graphene oxide dissolved in solubilization buffer are viable sorbents for efficient extraction of membrane proteins. LC-MS/MS analysis further revealed that graphene (50–200 nm) and graphene oxide (50–200 nm) can enrich more kinds of membrane proteins than a commercially available kit. Graphene was further applied to the enrichment of membrane proteins of normal cells as well as cancer cells, and 1079 and 872 proteins were identified, respectively, among which 56.5% and 60.5% were membrane proteins. In particular, 241 proteins were significantly regulated in cancer cells. Gene expression of 15 proteins was verified by qRT-PCR, and 4 of them were further quantified by immunoassay. These data collectively demonstrate that graphene has great potential to improve membrane protein extractions and thus can serve downstream cancer proteomics. Graphical abstract Two dimensional carbon nanomaterials, including graphene and graphene oxide, were employed as solid matrix to avoid lipid bilayer interference and enhance the extraction efficiency of membrane and membrane associated proteins. The strategy will benefit downstream membrane proteomics analysis.

Carrier flotation is a technique that can recover fine particles by using coarse carrier particles during the flotation process. In heterogeneous carrier flotation, coarse mineral particles of different minerals are used as carriers to recover fine mineral particles. By using Cu2+-treated pyrite particles as carriers, fine chalcopyrite particles recovery could be improved. However, a disadvantage of this heterogeneous carrier flotation is that it requires a post-flotation separation process to improve the grade of the final Cu concentrate. This study tested mechanical and chemical treatments to detach finely ground chalcopyrite (D50~3.5 µm) particles from Cu2+-treated coarse pyrite particles (−125 + 106 µm) after flotation. The results showed that the ultrasonic treatment was not effective to detach chalcopyrite fines from Cu2+-treated pyrite particles. However, acid treatment was effective to detach chalcopyrite fines from coarse pyrite particles. At pH 2, approximately 96% of chalcopyrite fines were detached from coarse Cu2+-treated pyrite particles. The acid treatment of flotation froth (mixture of chalcopyrite fines and Cu2+-treated pyrite particles) decomposed the collector KAX (potassium amyl xanthate) and dissolved the Cu precipitates adsorbed on the pyrite surface. This weakened the hydrophobic attraction force between the chalcopyrite fines and coarse pyrite particles, thus promoting the detachment of chalcopyrite fines from Cu2+-treated coarse pyrite particles.

The site-specific antibody-drug conjugates (ADCs), particularly those utilizing the engineered cysteine in Fc fragments of mAbs (THIOMAB™ antibodies), have emerged as a novel class of biotherapeutics for cancer treatment. The engineered cysteine residues in these antibodies are capped by cysteine or glutathione through a disulfide bond. Prior to conjugation with linker-payloads, these caps need to be removed through a reduction process. However, monitoring the efficiency of the decapping process has been challenging due to the lack of effective analytical methods. Intact reversed-phase liquid chromatography-mass spectrometry and hydrophobic interaction chromatography methods failed to separate decapped and capped intact THIOMAB™ mAbs in our study. Instead the fragmentation of mAbs provided a novel strategy to analyze the decapping effiency. After cleavage using a hinge specific enzyme, the generated Fc fragments with and without cysteine and/or glutathione caps displayed different hydrophobicity and were well separated by RPLC, allowing quantitative determination of the decapping efficiency. Enzymes that cleave both above and below the hinge disulfide bonds were tested. The use of FabALATICA can determine percentages of molecules with 0, 1, and 2 cysteine and/or glutathione caps, respectively, regardless of whether the antibody contains the hinge LALA mutations. On the other hand, FabRICATOR enzyme can only be utilized for antibodies without LALA mutations for the overall decapping percentage and cannot be used to estimate intact antibody each with 0, 1, and 2 caps. Therefore, FabALACTICA cleavage followed by RPLC provides a wider application of monitoring the decapping efficiency of all antibodies with the engineered cysteine in Fc.