Assessing the precision of a detergent-assisted cartridge precipitation workflow for non-targeted quantitative proteomics.

Detergent-based workflows incorporating sodium dodecyl sulfate (SDS) necessitate additional steps for detergent removal ahead of mass spectrometry (MS). These steps may lead to variable protein recovery, inconsistent enzyme digestion efficiency, and unreliable MS signals. To validate a detergent-based workflow for quantitative proteomics, we herein evaluate the precision of a bottom-up sample preparation strategy incorporating cartridge-based protein precipitation with organic solvent to deplete SDS. The variance of data-independent acquisition (SWATH-MS) data was isolated from sample preparation error by modelling the variance as a function of peptide signal intensity. Our SDS-assisted cartridge workflow yield a coefficient of variance (CV) of 13%-14%. By comparison, conventional (detergent-free) in-solution digestion increased the CV to 50%; in-gel digestion provided lower CVs between 14% and 20%. By filtering peptides predicting to display lower precision, we further enhance the validity of data in global comparative proteomics. These results demonstrate the detergent-based precipitation workflow is a reliable approach for in depth, label-free quantitative proteome analysis.

Liquid-liquid phase separation of amyloid-ß oligomers modulates amyloid fibrils formation.

Soluble amyloid-ß oligomers (AßOs) are proposed to instigate and mediate the pathology of Alzheimer's disease, but the mechanisms involved are not clear. In this study, we reported that AßOs can undergo liquid-liquid phase separation (LLPS) to form liquid-like droplets in vitro. We determined that AßOs exhibited an α-helix conformation in a membrane-mimicking environment of SDS. Importantly, SDS is capable of reconfiguring the assembly of different AßOs to induce their LLPS. Moreover, we found that the droplet formation of AßOs was promoted by strong hydrated anions and weak hydrated cations, suggesting that hydrophobic interactions play a key role in mediating phase separation of AßOs. Finally, we observed that LLPS of AßOs can further promote Aß to form amyloid fibrils, which can be modulated by (-)-epigallocatechin gallate. Our study highlights amyloid oligomers as an important entity involved in protein liquid-to-solid phase transition and reveals the regulatory role of LLPS underlying amyloid protein aggregation, which may be relevant to the pathological process of Alzheimer's disease.

Quantitatively measuring the cytotoxicity of viscous hydrogels with direct cell sampling in a micro scale format "MicroDrop" and its comparison to CCK8.

Tissue engineering holds promise for developing therapeutic applications using viscous materials e.g. hydrogels. However, assessing the cytotoxicity of such materials with conventional assays can be challenging due to non-specific interactions. To address this, we optimized a live/dead staining method for quantitative evaluation and compared it with the conventional CCK8 assay. Our MicroDrop method involved seeding droplets containing 5000 cells in 10 µl medium on 12-well plates. After allowing them to adhere for 4 h, various viscous samples were applied to the cells and measurements were conducted using a fluorescence microscope immediately and at daily intervals up to 72 h. A sodium dodecyl sulfate (SDS) dilution series compared the MicroDrop with the CCK8 assay. The findings revealed a cell-type specific pattern for 10 mg/ml hyaluronic acid (HA), wherein MC3T3-E1 cells maintained 95% viability until 72 h, while L929 cells experienced a gradual decline to 17%. 2 mg/ml HA exhibited consistent viability above 90% across all time points and cell lines. Similarly, fibrin demonstrated 90% viability across dilutions and time points, except for undiluted samples showing a decrease from 85% to 20%. Gelatin-methacrylol sustained viability above 70% across all time points at both 5% and 10% concentrations. The comparison of the SDS dilution series between viability (MicroDrop) and metabolic activity (CCK8) assay showed a correlation coefficient of 0.95. The study validates the feasibility of the established assay, providing researchers with an efficient tool for assessing cytotoxicity in viscous materials. Additionally, it holds the potential to yield more precise data on well-known hydrogels.

A green and highly sensitive microwell spectrofluorimetric method with high throughput for the determination of pemigatinib based on dual fluorescence enhancement by photoinduced electron transfer blocking and micellization: Application to the analysis of tablets, content uniformity testing, and human plasma.

Pemigatinib (PGT) is a recently FDA-approved small molecule kinase inhibitor used for the treatment of relapsed or refractory myeloid/lymphoid neoplasms in adults. This study introduces the development of a first microwell spectrofluorimetric method (MW-SFM) for quantifying PGT in FDA-approved tablets and plasma samples. The method utilized the enhancement of PGT's weak native fluorescence by blocking photoinduced electron transfer (PET) and micellization with sodium lauryl sulfate (SLS). The MW-SFM was performed in 96-microwell plates, and fluorescence signals were measured using a fluorescence microplate reader with excitation at 290 nm and emission at 350 nm. The method exhibited a linear range of 2-250 ng mL-1, and a limit of quantitation was 6.5 ng mL-1. The accuracy and precision of the method were confirmed with recovery rates ranging from 96.5% to 102.8% and relative standard deviations of 1.52% to 3.51%. The MW-SFM successfully analyzed Pemazyre® tablets, assessed content uniformity, and analyzed PGT-spiked human plasma samples. The greenness of the MW-SFM was verified using three different metric tools. In conclusion, the proposed MW-SFM is a valuable tool in supporting quality assessment of dosage forms, conducting pharmacokinetic studies, and monitoring therapeutic outcomes.

Utility of green chemistry for feasible spectrofluorometric determination of trifluridine via metal complexation: Application to dosage forms.

The goal of the current research was to establish a quick and practical fluorometric technique for trifluridine analysis. The approach relied on the drug's complex formation with the zinc ion to produce a high-fluorescence product. The fluorescence was further enhanced by adding sodium dodecyl sulfate, and it was observed at 450 nm following excitation at 400 nm. With a determination coefficient of 0.9994, the association between emission intensity and trifluridine concentration was linear between 1 and 125 ng mL-1. The quantitation limit was 0.987 ng mL-1 while 0.333 ng mL-1 was the detection limit. The buffer type, pH and concentration, type of surfactant and concentration, and finally the diluting solvent were among the reaction conditions that were closely examined. With great precision and reliability, the drug in question was quantified using this method in dosage formulations. The proposed method's level of greenness was assessed using two methodologies: the analytical greenness metric (AGREE) and the Green Analytical Procedure Index (GAPI).

Decellularized kidney capsule as a three-dimensional scaffold for tissue regeneration.

Tissue regeneration is thought to have considerable promise with the use of scaffolds designed for tissue engineering. Although polymer-based scaffolds for tissue engineering have been used extensively and developed quickly, their ability to mimic the in-vivo milieu, overcome immunogenicity, and have comparable mechanical or biochemical properties has limited their capability for repair. Fortunately, there is a compelling method to get around these challenges thanks to the development of extracellular matrix (ECM) scaffolds made from decellularized tissues. We used ECM decellularized sheep kidney capsule tissue in our research. Using detergents such as Triton-X100 and sodium dodecyl sulfate (SDS), these scaffolds were decellularized. DNA content, histology, mechanical properties analysis, attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), biocompatibility, hemocompatibility and scanning electron microscope (SEM) imaging were measured. The results showed that the three-dimensional (3D) structure of the ECM remained largely intact. The scaffolds mentioned above had several hydrophilic properties. The best biocompatibility and blood compatibility properties were reported in the SDS method of 0.5%. The best decellularization scaffold was introduced with 0.5% SDS. Therefore, it can be proposed as a scaffold that has ECM like natural tissue, for tissue engineering applications.

Dose-dependent effects of sodium dodecyl sulfate and hydrogen peroxide treatments on methane emission from pig manure during storage.

Mitigation of methane (CH4) emissions from slurry pits within pig barns can be achieved through treatment of residual slurry left after frequent flushing of the slurry pits. In this study, dosages of additives such as sodium dodecyl sulfate (SDS) and hydrogen peroxide (H2O2) were optimized to achieve reduction in CH4 emissions from residual pig slurry during storage. In addition, the effects on emissions when both the treatments were combined and the effects of SDS treatment on slurry acidified with sulfuric acid (H2SO4) were studied in order to reduce CH4 and ammonia (NH3) emissions from residual pig slurry storage. A maximum of 98% and 70% reduction in CH4 emissions were achieved with SDS and H2O2 treatments, respectively. The combination of SDS and H2O2 did not increase efficiency in reducing CH4 emissions compared to SDS treatment alone. Whereas the application of SDS to slurry acidified with H2SO4 (pH 6.2) increased the CH4 mitigation efficiency by 15-30% compared to treating slurry with only SDS. The combined treatment (SDS + H2SO4) reduced NH3 emissions by 20% compared to treating slurry with H2SO4 (pH 6.2) alone. Hereby, combined treatment (SDS + H2SO4) can reduce both CH4 and NH3 emissions, with a reduced amount of chemicals required for the treatment. Hence, application of SDS at concentrations <2 g kg-1 to acidified slurry is recommended to treat residual pig manure in pig barns.

The investigation of the binding ability between sodium dodecyl sulfate and Cu (II) in urban stormwater runoff.

The simultaneous presence of heavy metals and surfactants in runoff induces complexation and ecological harm during migration. However, interactions between these pollutants are often overlooked in past studies. Thus, investigating heavy metal-surfactant complexes in runoff is imperative. In this work, Cu (II) and sodium dodecyl sulfate (SDS) were selected to investigate the interaction between heavy metals and surfactants due to the higher detected frequency in runoff. Through 1H NMR and FTIR observation of hydrogen atom nuclear displacement and functional group displacement of SDS, the change of SDS and Cu (II) complexation was obtained, and then the complexation form of Cu (II) and SDS was verified. The results showed that solution pH values and ionic strength had significant effects on the complexation of Cu (II). When the pH values increase from 3.0 to 6.0, the complexation efficiency of SDS with Cu (II) increased by 12.12% at low concentration of SDS, which may be attributed to the excessive protonation in the aqueous solution at acidic condition. The increase of ionic strength would inhibit the complexation reaction efficiency by 19.57% and finally reached the platform with concentration of NaNO3 was 0.10 mmol/L, which was mainly due to the competitive relationship between Na (I) and Cu (II). As a general filtering material in stormwater treatment measures, natural zeolite could affect the interaction between SDS and Cu (II) greatly. After the addition of SDS, the content of free Cu (II) in the zeolite-SDS-Cu (II) three-phase mixed system was significantly reduced, indicating that SDS had a positive effect on the removal of Cu (II) from runoff. This study is of great significance for investigating the migration and transformation mechanism of SDS and Cu (II) in the future and studying the control technology of storm runoff pollution.

Effect of surfactant's charge properties on behavior, physiology, and biochemistry and the release of microcystins of Microcystis aeruginosa.

Surfactant pollution is escalatitheng in eutrophic waters, but the effect of surfactant charge properties on the physiological and biochemical properties of toxin-producing microalgae remains inadequately explored. To address this gap, this study explores the effects and mechanisms of three common surfactants-cetyltrimethylammonium bromide (CTAB, cationic), sodium dodecyl sulfate (SDS, anionic), and Triton X-100 (nonionic)-found in surface waters, on the agglomeration behavior, physiological indicators, and Microcystin-LR (MC-LR) release of Microcystis aeruginosa (M. aeruginosa) by using UV-visible spectroscope, Malvern Zetasizer, fluorescence spectrometer, etc. Results suggest that charge properties significantly affect cyanobacterial aggregation and cellular metabolism. The CTAB-treated group demonstrates a ∼5.74 and ∼9.74 times higher aggregation effect compared to Triton X-100 and SDS (300 mg/L for 180 min) due to strong electrostatic attraction. Triton X-100 outperforms CTAB and SDS in polysaccharide extraction, attributed to its higher water solubility and lower critical micelle concentration. CTAB stimulates cyanobacteria to secrete proteins, xanthohumic acid, and humic acids to maintain normal physiological cells. Additionally, the results of SEM and ion content showed that CTAB damages the cell membrane, resulting in a ∼90% increase in the release of intracellular MC-LR without cell disintegration. Ionic analyses confirm that all three surfactants alter cell membrane permeability and disrupt ionic metabolic pathways in microalgae. This study highlights the relationship between the surface charge properties of typical surfactants and the dispersion/agglomeration behavior of cyanobacteria. It provides insights into the impact mechanism of exogenous surfactants on toxic algae production in eutrophic water bodies, offering theoretical references for managing surfactant pollution and treating algae blooms.

Size separation of sodium dodecyl sulfate-proteins by capillary electrophoresis in dilute and ultra-dilute dextran solutions.

SDS capillary gel electrophoresis is a widely used in the biopharma and the biomedical fields for rapid size separation of proteins. However, very limited information is available on the use of dilute and ultra-dilute sieving matrices for SDS-protein analysis. Here, background electrolytes (BGEs) containing 1%-0% dextran were used in borate-based BGE to separate a protein sizing ladder (PSL) ≤225 kDa and the intact and subunit forms of a therapeutic monoclonal antibody (mAb). The separation performance for the PSL and mAb components differed significantly with decreasing dextran concentration. Ferguson and reptation plots were used to elucidate the separation mechanism. Highly diluted dextran solutions resulted in linear Ferguson plots for both solute types (cf. Ogston theory) in spite of this model assumes a rigid pore structure, thus cannot describe the separation mechanism in ultra-dilute polymer solutions with no reticulations. The saddle differences between the resolution of the PSL and the intact/subunit mAb forms in ultra-dilute dextran-borate matrices suggested the importance of shape selectivity, manifested by the adequate separation of the SDS covered intact as well as light and heavy chain subunits of the therapeutic mAb even at zero dextran concentration.

Development of an Amorphous Solid Dispersion Formulation for Mitigating Mechanical Instability of Crystalline Form and Improving Bioavailability for Early Phase Clinical Studies.

In this work, an amorphous solid dispersion (ASD) formulation was systematically developed to simultaneously enhance bioavailability and mitigate the mechanical instability risk of the selected crystalline form of a development drug candidate, GDC-0334. The amorphous solubility advantage calculation was applied to understand the solubility enhancement potential by an amorphous formulation for GDC-0334, which showed 2.7 times theoretical amorphous solubility advantage. This agreed reasonably well with the experimental solubility ratio between amorphous GDC-0334 and its crystalline counterpart (∼2 times) in buffers of a wide pH range. Guided by the amorphous solubility advantage, ASD screening was then carried out, focusing on supersaturation maintenance and dissolution performance. It was found that although the type of polymer carrier did not impact ASD performance, the addition of 5% (w/w) sodium dodecyl sulfate (SDS) significantly improved the GDC-0334 ASD dissolution rate. After ASD composition screening, stability studies were conducted on selected ASD powders and their hypothetical tablet formulations. Excellent stability of the selected ASD prototypes with or without tablet excipients was observed. Subsequently, ASD tablets were prepared, followed by in vitro and in vivo evaluations. Similar to the effect of facilitating the dissolution of ASD powders, the added SDS improved the disintegration and dissolution of ASD tablets. Finally, a dog pharmacokinetic study confirmed 1.8 to 2.5-fold enhancement of exposure by the developed ASD tablet over the GDC-0334 crystalline form, consistent with the amorphous solubility advantage of GDC-0334. A workflow of developing an ASD formulation for actual pharmaceutical application was proposed according to the practice of this work, which could provide potential guidance for ASD formulation development in general for other new chemical entities.

Ultrathin Micellar Foam Films of Sodium Caseinate Protein Solutions.

Sodium caseinates (NaCas), derived from milk proteins called caseins, are often added to food formulations as emulsifiers, foaming agents, and ingredients for producing dairy products. In this contribution, we contrast the drainage behavior of single foam films made with micellar NaCas solutions with well-established features of stratification observed for the micellar sodium dodecyl sulfate (SDS) foam films. In reflected light microscopy, the stratified SDS foam films display regions with distinct gray colors due to differences in interference intensity from coexisting thick-thin regions. Using IDIOM (interferometry digital imaging optical microscopy) protocols we pioneered for mapping nanotopography of foam films, we showed that drainage via stratification in SDS films proceeds by the expansion of flat domains that are thinner than surrounding by a concentration-dependent step-size, and nonflat features (nanoridges and mesas) form at the moving front. Furthermore, stratifying SDS foam films show stepwise thinning, such that the step-size and terminal film thickness decrease with concentration. Here we visualize the nanotopography in protein films with high spatiotemporal resolution using IDIOM protocols to address two long-standing questions. Do protein foam films formulated with NaCas undergo drainage via stratification? Are thickness transitions and variations in protein foam films determined by intermicellar interactions and supramolecular oscillatory disjoining pressure? In contrast with foam films containing micellar SDS, we find that micellar NaCas foam films display just one step, nonflat and noncircular domains that expand without forming nanoridges and a terminal thickness that increases with NaCas concentration. We infer that the differences in adsorbing and self-assembling unimers triumph over any similarities in the structure and interactions of their micelles.

Drug-ionic surfactant interactions: density, sound speed, spectroscopic, and electrochemical studies.

The failure of antibiotics against infectious diseases has become a global health issue due to the incessant use of antibiotics in the community and a lack of entry of new antibacterial drugs onto the market. The limited knowledge of biophysical interactions of existing antibiotics with bio-membranes is one of the major hurdles to design and develop more effective antibiotics. Surfactant systems are the simplest biological membrane models that not only mimic the cell membrane functions but are also used to investigate the biophysical interactions between pharmaceutical drugs and bio-membranes at the molecular level. In this work, volumetric and acoustic studies were used to investigate the molecular interactions of moxifloxacin (MXF), a potential antibacterial drug, with ionic surfactants (dodecyl-tri-methyl-ammonium bromide (DTAB), a cationic surfactant and sodium dodecyl sulfate (SDS), an anionic surfactant) under physiological conditions (phosphate buffer, pH 7.4) at T = 298.15-313.15 K at an interval of 5 K. Various volumetric and acoustic parameters were computed from the density and sound speed data and interpreted in terms of MXF-ionic surfactant interaction using electrostriction effect and co-sphere overlap model. Absorption spectroscopy and cyclic voltammetry were further used to determine the binding, partitioning, and related free energies of MXF with ionic micelles.

Characterisation of cytotoxicity and immunomodulatory effects of glycolipid biosurfactants on human keratinocytes.

Skin irritation and allergic reactions associated with the use of skincare products formulated with synthetically derived surfactants such as sodium lauryl ether sulphate (SLES) have encouraged the search for naturally derived and biocompatible alternatives. Glycolipid biosurfactants such as sophorolipids (SL) and rhamnolipids (RL) offer a potential alternative to SLES. However, most studies on the bioactive properties of microbial glycolipids were determined using their mixed congeners, resulting in significant inter-study variations. This study aims to compare the effects of highly purified SL (acidic and lactonic) and RL (mono-RL and di-RL) congeners and SLES on a spontaneously transformed human keratinocyte cell line (HaCaT cells) to assess glycolipids' safety for potential skincare applications. Preparations of acidic SL congeners were 100% pure, lactonic SL were 100% pure, mono-RL were 96% pure, and di-RL were 97% pure. Cell viability using XTT assays, cell morphological analyses, and immunoassays revealed that microbial glycolipids have differing effects on HaCaT cells dependent on chemical structure. Compared with SLES, acidic SL and mono-RL have negligible effects on cell viability, cell morphology, and production of pro-inflammatory cytokines. Furthermore, at non-inhibitory concentrations, di-RL significantly attenuated IL-8 production and CXCL8 expression while increasing IL-1RA production and IL1RN expression in lipopolysaccharide-stimulated HaCaT cells. Although further studies would be required, these results demonstrate that as potential innocuous and bioactive compounds, microbial glycolipids could provide a substitute to synthetic surfactants in skincare formulations and perform immunopharmacological roles in topical skin infections such as psoriasis. KEY POINTS: • Purified glycolipid congeners have differing effects on human keratinocytes. • Compared with SLES, acidic sophorolipids and mono-rhamnolipids have innocuous effects on keratinocytes. • Di-rhamnolipids and mono-rhamnolipids modulate cytokine production in lipopolysaccharide stimulated human keratinocytes.

Surfactant induced gelation of TEMPO-oxidized cellulose nanofibril dispersions probed using small angle neutron scattering.

In this work, we studied TEMPO-oxidized cellulose nanofibril (OCNF) suspensions in the presence of diverse surfactants. Using a combination of small angle neutron scattering (SANS) and rheology, we compared the physical properties of the suspensions with their structural behavior. Four surfactants were studied, all with the same hydrophobic tail length but different headgroups: hexaethylene glycol mono-n-dodecyl ether (C12EO6, nonionic), sodium dodecyl sulfate (SDS, anionic), cocamidopropyl betaine (CapB, zwitterionic), and dodecyltrimethylammonium bromide (DTAB, cationic). Contrast variation SANS studies using deuterated version of C12EO6 or SDS, or by varying the D2O/H2O ratio of the suspensions (with CapB), allowed focusing only on the structural properties of OCNFs or surfactant micelles. We showed that, in the concentration range studied, for C12EO6, although the nanofibrils are concentrated thanks to an excluded volume effect observed in SANS, the rheological properties of the suspensions are not affected. Addition of SDS or CapB induces gelation for surfactant concentrations superior to the critical micellar concentration (CMC). SANS results show that attractive interactions between OCNFs arise in the presence of these anionic or zwitterionic surfactants, hinting at depletion attraction as the main mechanism of gelation. Finally, addition of small amounts of DTAB (below the CMC) allows formation of a tough gel by adsorbing onto the OCNF surface.

SCASP: A Simple and Robust SDS-Aided Sample Preparation Method for Proteomic Research.

SDS is widely used in sample preparation for proteomic research. However, SDS is incompatible with LC and electrospray ionization. SDS depletion is therefore required ahead of LC-MS analysis. Most of current SDS removal strategies are time consuming, laborious, and have low reproducibility. Here, we describe a method, SDS-cyclodextrin (CD)-assisted sample preparation, by which CD can bind to SDS and form CD-SDS complexes in solutions, allowing for direct tryptic digestion. We demonstrate that SDS-CD-assisted sample preparation is a simple, fast, and robust SDS-based sample preparation method for proteomics application.

Mimicry of a biophysical pathway leads to diverse pollen-like surface patterns.

A ubiquitous structural feature in biological systems is texture in extracellular matrix that gains functions when hardened, for example, cell walls, insect scales, and diatom tests. Here, we develop patterned liquid crystal elastomer (LCE) particles by recapitulating the biophysical patterning mechanism that forms pollen grain surfaces. In pollen grains, a phase separation of extracellular material into a pattern of condensed and fluid-like phases induces undulations in the underlying elastic cell membrane to form patterns on the cell surface. In this work, LCE particles with variable surface patterns were created through a phase separation of liquid crystal oligomers (LCOs) droplet coupled to homeotropic anchoring at the droplet interface, analogously to the pollen grain wall formation. Specifically, nematically ordered polydisperse LCOs and isotropic organic solvent (dichloromethane) phase-separate at the surface of oil-in-water droplets, while, different LCO chain lengths segregate to different surface curvatures simultaneously. This phase separation, which creates a distortion in the director field, is in competition with homeotropic anchoring induced by sodium dodecyl sulfate (SDS). By tuning the polymer chemistry of the system, we are able to influence this separation process and tune the types of surface patterns in these pollen-like microparticles. Our study reveals that the energetically favorable biological mechanism can be leveraged to offer simple yet versatile approaches to synthesize microparticles for mechanosensing, tissue engineering, drug delivery, energy storage, and displays.

Effect of Solubility Improvement via Formation of an Amorphous Composite of Indomethacin and Sulindac on Membrane Permeability.

The importance of permeability as well as solubility of the drug has been recognized in improving the solubility of poorly water-soluble drugs. This study investigated the impact of amorphous composites of indomethacin (IMC) and sulindac (SLD) on the membrane permeability of drugs. The IMC/SLD (1/1) formulation prepared by dry grinding was amorphous with a single glass transition temperature. The Fourier transform IR spectra and Raman spectra revealed formation of hydrogen bonds between the OH group of IMC and the carbonyl group of SLD. These results suggest that an amorphous composite was formed between IMC and SLD through hydrogen bonding. The amount of dissolved IMC and SLD from the amorphous composite of IMC/SLD (1/1) was higher than that of the untreated IMC or SLD in the dissolution test. The permeated amounts and permeation rates of both drugs were enhanced by increasing the solubility of the amorphous composite. Conversely, the apparent membrane permeability coefficients (Papp) were almost same for untreated drugs and amorphous composites. In the case of hydroxypropyl-ß-cyclodextrin and sodium dodecyl sulfate, Papp of the drugs decreased with the addition of these compounds, although the drug solubility was enhanced by the solubilization effect. This study revealed that an amorphous composite formed through hydrogen bonding is an attractive pharmaceutical way to enhance the permeated amount and permeation rate without changing the Papp of both the drugs.

Boosting the Separation of Adeno-Associated Virus Capsid Proteins by Liquid Chromatography and Capillary Electrophoresis Approaches.

The purity of the three capsid proteins that make up recombinant adeno-associated virus (rAAV) is considered a critical quality attribute of gene therapy products. As such, there is a clear need to develop separation methods capable of rapidly characterizing these three viral proteins (VPs). In this study, the potential benefits and limitations of different electrophoretic and chromatographic methods were evaluated, including capillary electrophoresis-sodium dodecyl sulfate (CE-SDS), reversed phase liquid chromatography (RPLC), hydrophilic interaction chromatography (HILIC), and hydrophobic interaction chromatography (HIC), for the analysis of VPs obtained from different serotypes (i.e., AAV2, AAV5, AAV8, and AAV9). CE-SDS is considered to be the reference method and provides a suitable separation of VP1-3 proteins using generic conditions and laser induced fluorescence detection. However, the characterization of post-translational modifications (i.e., phosphorylation, oxidation) remains difficult, and species identification is almost impossible due to the lack of compatibility between CE-SDS and mass spectrometry (MS). In contrast, RPLC and HILIC were found to be less generic than CE-SDS and require tedious optimization of the gradient conditions for each AAV serotype. However, these two chromatographic approaches are inherently compatible with MS, and were shown to be particularly sensitive in detecting capsid protein variants resulting from different post-translational modifications. Finally, despite being non-denaturing, HIC offers disappointing performance for viral capsid proteins characterization.

Non-Steroidal Anti-Inflammatory Drugs Loaded to Micelles for the Modulation of Their Water Solubility.

The low water solubility of aspirin (ASPH) is well known, creating research challenges regarding both its composition and its delivery. Therefore, the development of new aspirin-based formulations that are water soluble is a research, technological, and financial issue. With the aim to improve the water solubility of ASPH, the micelle of formula SLS@ASPH (SLS = Sodium Lauryl Sulfate) was formed. The Critical Micelle Concentration (CMC) of SLS in the presence of ASPH was determined by ultrasonic velocity, complementary, and transient birefringence measurements. The SLS@ASPH was characterized by the melting point (m.p.), attenuated total reflection spectroscopy (FT-IR-ATR), and X-ray fluorescence spectroscopy (XRF) in a solid state and in a solution by ultraviolet-visible (UV-Vis) and 1H NMR spectroscopies. The SLS/ASPH molar ratio was determined to be 5/1 in SLS@ASPH. The inhibitory activity of SLS@ASPH towards lipoxygenase (LOX), an enzyme that takes part in the inflammation mechanism, was studied. The inhibitory activity of SLS@ASPH against LOX is 3.5-fold stronger than that of free SLS. The in vitro toxicity of the SLS@ASPH was tested on immortalized human keratinocyte (HaCaT) cells.