Glycopeptide enrichment is a crucial step in glycoproteomic analysis, often achieved through solid-phase extraction (SPE) on polar stationary phases in hydrophilic interaction liquid chromatography (HILIC). This study explores the potential of polyaniline (PANI)-coated silica gel for enriching human immunoglobulin G (IgG). Experimental conditions were varied to assess their impact on glycopeptide enrichment efficiency, comparing PANI-cotton wool SPE with conventional cotton wool as SPE sorbents. Two formic acid concentrations (0.1% and 1%) in elution solvent were tested, revealing that higher concentrations led to earlier elution of studied glycopeptides, especially for sialylated glycopeptides. Substituting formic acid with acetic acid increased the interaction of neutral glycopeptides with the PANI-modified sorbent, while sialylated glycopeptides showed no significant change in enrichment efficiency. Acetonitrile concentration in the elution solvent (5%, 10%, and 20%) affected the enrichment efficiency with most glycopeptides eluting at the lowest acetonitrile concentration. The acetonitrile concentration in conditioning and washing solutions (65%, 75%, and 85%) played a crucial role; at 65% acetonitrile, glycopeptides were least retained on the stationary phase, and neutral glycopeptides were even detected in the flow-through fraction. This study shows the potential of in-house-prepared PANI-modified sorbents for SPE-HILIC glycopeptide enrichment, highlighting the crucial role of tuning experimental conditions in sample preparation to enhance enrichment efficiency and selectivity.

• Klíčová slova
• Glycopeptide enrichment, HILIC, Immunoglobulin G, Polyaniline, Solid-phase extraction,
• MeSH
• acetonitrily MeSH
• aniliny * MeSH
• chromatografie kapalinová metody   MeSH
• extrakce na pevné fázi * metody   MeSH
• formiáty * MeSH
• glykopeptidy * chemie   MeSH
• hydrofobní a hydrofilní interakce MeSH
• lidé MeSH
• rozpouštědla MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrily MeSH
• aniliny * MeSH
• formiáty * MeSH
• formic acid MeSH Prohlížeč
• glykopeptidy * MeSH
• polyaniline MeSH Prohlížeč
• rozpouštědla MeSH

About 30% of the FDA approved drugs in 2021 were protein-based therapeutics. However, therapeutic proteins can be unstable and rapidly eliminated from the blood, compared to conventional drugs. Furthermore, on-target but off-tumor protein binding can lead to off-tumor toxicity, lowering the maximum tolerated dose. Thus, for effective treatment therapeutic proteins often require continuous or frequent administration. To improve protein stability, delivery and release, proteins can be encapsulated inside drug delivery systems. These drug delivery systems protect the protein from degradation during (targeted) transport, prevent premature release and allow for long-term, sustained release. However, thus far achieving high protein loading in drug delivery systems remains challenging. Here, the use of protein desolvation with acetonitrile as an intermediate step to concentrate monoclonal antibodies for use in drug delivery systems is reported. Specifically, trastuzumab, daratumumab and atezolizumab were desolvated with high yield (∼90%) into protein nanoparticles below 100 nm with a low polydispersity index (<0.2). Their size could be controlled by the addition of low concentrations of sodium chloride between 0.5 and 2 mM. Protein particles could be redissolved in aqueous solutions and redissolved antibodies retained their binding activity as evaluated in cell binding assays and exemplified for trastuzumab in an ELISA.

• MeSH
• acetonitrily MeSH
• chlorid sodný terapeutické užití   MeSH
• lidé MeSH
• nádory * farmakoterapie   MeSH
• nanočástice * MeSH
• systémy cílené aplikace léků MeSH
• trastuzumab terapeutické užití   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrily MeSH
• chlorid sodný MeSH
• trastuzumab MeSH

The sample preparation step is pivotal in glycoproteomic analysis. An effective approach in glycoprotein sample preparation involves enriching glycopeptides by solid-phase extraction (SPE) using polar stationary phases in hydrophilic interaction liquid chromatography (HILIC) mode. The aim of this work is to show how different experimental conditions influence the enrichment efficiency of glycopeptides from human immunoglobulin G (IgG) on an aminopropyl-modified SPE column. Different compositions of the elution solvent (acetonitrile, methanol, and isopropanol), along with varying concentrations of elution solvent acidifiers (formic and acetic acid), and different concentrations of acetonitrile for the conditioning and washing solvents (65%, 75%, and 85% acetonitrile) were tested to observe their effects on the glycopeptide enrichment process. Isopropanol proved less effective in enriching glycopeptides, while acetonitrile was the most efficient, with methanol in between. Higher formic acid concentrations in the elution solvent weakened the ionic interactions, particularly with sialylated glycopeptides. Substituting formic acid with acetic acid led to earlier elution of more glycopeptides. The acetonitrile concentration in conditioning and washing solutions played a key role; at 65% acetonitrile, glycopeptides were not retained on the SPE column and were detected in the flow-through fraction. Ultimately, it was proven that the enrichment method was applicable to human plasma samples, resulting in a significant decrease in the abundances of non-glycosylated peptides. To the best of our knowledge, this study represents the first systematic investigation into the impact of the mobile phase on glycopeptide enrichment using an aminopropyl-modified SPE column in HILIC mode. This study demonstrates the substantial impact of even minor variations in experimental conditions, which have not yet been considered in the literature, on SPE-HILIC glycopeptide enrichment. Consequently, meticulous optimization of these conditions is imperative to enhance the specificity and selectivity of glycoproteomic analysis, ensuring accurate and reliable quantification.

• Klíčová slova
• Glycopeptide enrichment, Glycoproteomics, Hydrophilic interaction liquid chromatography, Immunoglobulin G, Solid-phase extraction,
• MeSH
• 2-propanol MeSH
• acetáty MeSH
• acetonitrily MeSH
• chromatografie kapalinová metody   MeSH
• extrakce na pevné fázi metody   MeSH
• formiáty * MeSH
• glykopeptidy * chemie   MeSH
• hydrofobní a hydrofilní interakce MeSH
• imunoglobulin G chemie   MeSH
• lidé MeSH
• methanol * MeSH
• rozpouštědla MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 2-propanol MeSH
• acetáty MeSH
• acetonitrily MeSH
• formiáty * MeSH
• formic acid MeSH Prohlížeč
• glykopeptidy * MeSH
• imunoglobulin G MeSH
• methanol * MeSH
• rozpouštědla MeSH

We developed a fast, selective, and sensitive method for the determination of various neutral and acidic phytocannabinoids with an emphasis on the separation of structurally related compounds. Optimized ultra-high performance supercritical fluid chromatography (UHPSFC) allowed the separation of 2 groups of structural isomers, including isomers of m/z 357: cannabidiolic and Δ9-tetrahydrocannabinolic acid, and isomers of m/z 315: cannabichromene, Δ9-tetrahydrocannabinol, Δ8-tetrahydrocannabinol, cannabicyclol, and cannabidiol only in mere 3.5 min followed by 1.5 min equlibration. The 2-ethylpyridine functionalized stationary phase and gradient elution using mobile phase comprising carbon dioxide and methanol: acetonitrile (25:75) + 5% water mixture were selected after the optimization. Tandem mass spectrometry (MS/MS) with electrospray ionization in positive and negative modes with methanol + 5% water as a make-up solvent provided adequate selectivity and sensitivity needed for analysis of phytocannabinoids in complex matrices. The limits of quantification were in the range 0.01-0.5 ng/mL for most of the monitored cannabinoids. The optimized UHPSFC-MS/MS method was then used for the determination of cannabinoids in various products, such as dietary supplements, nutraceuticals, and cosmetics. Solvent extraction methods were optimized for the cosmetic and nutraceutical products with the accuracy in the range 80.4-120.6% and precision 0.5-18.9%. To extract cannabinoids from the herbal infusion matrix, supercritical fluid extraction (SFE) and pressurized liquid extraction (PLE) methods were developed using environmentally friendly solvents water, ethanol, and carbon dioxide. The detailed optimization of extraction solvent composition, temperature, and pressure was carried out with the emphasis on avoiding the thermal degradation of cannabinoids. Optimized SFE and PLE methods were compared and applied to different herbal infusions to confirm declared cannabinoids content.

• Klíčová slova
• Cannabis, Mass spectrometry, Phytocannabinoids, Supercritical fluid chromatography, Supercritical fluid extraction,
• MeSH
• acetonitrily MeSH
• Cannabis * chemie   MeSH
• ethanol MeSH
• kanabidiol * analýza   MeSH
• kanabinoidy * analýza   MeSH
• methanol chemie   MeSH
• oxid uhličitý chemie   MeSH
• rozpouštědla analýza   MeSH
• superkritická fluidní chromatografie * metody   MeSH
• tandemová hmotnostní spektrometrie metody   MeSH
• tetrahydrokanabinol analýza   MeSH
• voda MeSH
• vysokoúčinná kapalinová chromatografie metody   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrily MeSH
• ethanol MeSH
• kanabidiol * MeSH
• kanabinoidy * MeSH
• methanol MeSH
• oxid uhličitý MeSH
• rozpouštědla MeSH
• tetrahydrokanabinol MeSH
• voda MeSH

Determination of cholesterol in food matrices is essential for quality control concerning the health of consumers. Herein, a simple electrochemical approach for cholesterol quantitation in dairy products is evaluated. The newly developed differential pulse voltammetric method using acetonitrile-perchloric acid mixture as a supporting electrolyte is statistically compared to GC-MS and HPLC-UV. Oxidation signals of cholesterol at +1.5 V and +1.4 V (vs. Ag/AgNO3 in acetonitrile) provide detection limits of 4.9 µM and 6.1 µM on boron-doped diamond and glassy carbon electrodes, respectively. A simple liquid-liquid extraction procedure from dairy products into hexane resulted in a recovery rate of (74.8 ± 3.8)%. The method provides results in close agreement (at a 95% confidence level) with GC-MS, while HPLC-UV resulted in a significant difference in estimated cholesterol concentrations for all samples. This newly developed method is a simpler, faster and cheaper alternative to instrumentally demanding MS-based methods and clearly outperforms HPLC-UV.

• Klíčová slova
• Boron-doped diamond, Cholesterol, Dairy products, Differential pulse voltammetry, GC–MS, Oxidation,
• MeSH
• acetonitrily MeSH
• bor * MeSH
• cholesterol MeSH
• elektrody MeSH
• mléčné výrobky * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrily MeSH
• bor * MeSH
• cholesterol MeSH

Sensitive electrophoretic determination of 3-hydroxybutyrate (3HB) as an indicator of human ketogenesis is performed in fused silica capillary covalently coated by an anionic copolymer of poly(acrylamide-co-sodium-2-acrylamido-2-methylpropanesulphonate) (PAMAMPS). Baseline separation of 3HB from other components of human serum is achieved in a 20 μm capillary with an effective length of 17 cm covered by 4% PAMAMPS, which generates a cathodic EOF with a mobility of 8.30 ± 0.00 · 10-9 m2/V.s in 80 mM MES/His as background electrolyte. 3HB migrates in counter-current electrophoretic mode against EOF, that effectively improving electrophoretic resolution. Sample pre-treatment is based on adding of 45 μL acetonitrile to 15 μL serum and, after shaking, a 28 mm long zone of supernatant is injected into the capillary, and sharpened after turning on a separation voltage of 20 kV using the technique of large volume sample stacking, where the EOF forces the residual acetonitrile from the capillary. When combined with universal contactless conductivity detection, the achieved LOD and LOQ are 0.43 μM and 1.44 μM, respectively, that are sufficiently low for monitoring the physiological 3HB level. The performed clinical study subsequently showed that serum 3HB increases from a concentration of 71 μM, corresponding to normal food, to level of 1924 μM after 60 h of fasting and returns to the normal physiological concentration 48 h after commencing consumption of high-saccharide food.

• Klíčová slova
• Capillary electrophoresis, Clinical analysis, Coating, Contactless conductivity detection, Ketogenesis, Stacking,
• MeSH
• acetonitrily MeSH
• akrylové pryskyřice MeSH
• alkylsulfonany MeSH
• elektroforéza kapilární * metody   MeSH
• kyselina 3-hydroxymáselná MeSH
• lidé MeSH
• omezení příjmu potravy * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrily MeSH
• akrylové pryskyřice MeSH
• alkylsulfonany MeSH
• kyselina 3-hydroxymáselná MeSH
• poly(acrylamide-co-2-acrylamido-2-methyl-1-propanesulfonate) MeSH Prohlížeč

A method for purity control of newly synthesized lactic acid-based liquid crystals has been developed. The electrokinetic chromatography proved to be suitable for the separation of these electroneutral substances from their impurities. The separations were performed in an acidic acetonitrile-based background electrolyte (BGE) with a pseudostationary phase formed by a cationic surfactant. During the optimization step, appropriate concentrations of cetyltrimethylammonium bromide, acetic acid, and water were seeked. In the optimized method, separations were carried out in acetonitrile with 1-mol/L acetic acid, 80-mmol/L cetyltrimethylammonium bromide, and 6% (v/v) water. Interesting positive effects of a small water content in the BGE on electroosmotic flow and resolution of liquid crystal substances from their impurities were observed and discussed. Samples of five liquid crystal substances, both pure and containing impurities from synthesis, were analyzed. The identification of analytes was based on a comparison of relative migration times related to the migration time of mesityl oxide. For all five samples, impurities were separated from the liquid crystals and the method thus showed its viability. To the best of our knowledge, this method is used for the first time for the purity control of newly synthesized liquid crystals. This method can be used to confirm or complement the results obtained by commonly used high-performance liquid chromatography and supercritical fluid chromatography methods. Furthermore, the electrokinetic chromatography method requires very small amounts of sample, solvents, and buffer constituents. Overall, its operational costs are significantly lower.

• Klíčová slova
• capillary electrophoresis, electrokinetic chromatography, liquid crystals, nonaqueous,
• MeSH
• acetonitrily chemie   MeSH
• cetrimonium MeSH
• chromatografie micelární elektrokinetická kapilární * metody   MeSH
• elektrolyty MeSH
• kapalné krystaly * MeSH
• superkritická fluidní chromatografie * MeSH
• voda chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• acetonitrily MeSH
• cetrimonium MeSH
• elektrolyty MeSH
• voda MeSH

Irradiation of coumarin-3-carboxylic acid in acetonitrile and methanol solutions at 355 nm results in complex multistep photochemical transformations, strongly dependent on the solvent properties and oxygen content. A number of reaction intermediates, which themselves undergo further (photo)chemical reactions, were identified by steady-state and transient absorption spectroscopy, mass spectrometry, and NMR and product analyses. The triplet excited compound in acetonitrile undergoes decarboxylation to give a 3-coumarinyl radical that traps molecular oxygen to form 3-hydroxycoumarin as the major but chemically reactive intermediate. This compound is oxygenated by singlet oxygen, produced by coumarin-3-carboxylic acid sensitization, followed by a pyrone ring-opening reaction to give an oxalic acid derivative. The subsequent steps lead to the production of salicylaldehyde, carbon monoxide, and carbon dioxide as the final products. When 3-coumarinyl radical is not trapped by oxygen in degassed acetonitrile, it abstracts hydrogen from the solvent and undergoes triplet-sensitized [2 + 2] cycloaddition. The reaction of 3-coumarinyl radical with oxygen is largely suppressed in aerated methanol as a better H-atom donor, and coumarin is obtained as the primary product in good yields. Because coumarin derivatives are used in many photophysical and photochemical applications, this work provides detailed and sometimes surprising insights into their complex phototransformations.

• Klíčová slova
• Coumarin, Photochemistry, Photooxidation, Singlet oxygen,
• MeSH
• acetonitrily chemie   MeSH
• kumariny * MeSH
• kyslík MeSH
• methanol * MeSH
• rozpouštědla chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrily MeSH
• coumarin-3-carboxylic acid MeSH Prohlížeč
• kumariny * MeSH
• kyslík MeSH
• methanol * MeSH
• rozpouštědla MeSH

Double and triple bonds have significant effects on the biological activities of lipids. Determining multiple bond positions in their molecules by mass spectrometry usually requires chemical derivatization. This work presents an HPLC/MS method for pinpointing the double and triple bonds in fatty acids. Fatty acid methyl esters were separated by reversed-phase HPLC with an acetonitrile mobile phase. In the APCI source, acetonitrile formed reactive species, which added to double and triple bonds to form [M + C3H5N]+• ions. Their collisional activation in an ion trap provided fragments helpful in localizing the multiple bond positions. This approach was applied to fatty acids with isolated, cumulated, and conjugated double bonds and triple bonds. The fatty acids were isolated from the fat body of early-nesting bumblebee Bombus pratorum and seeds or seed oils of Punicum granatum, Marrubium vulgare, and Santalum album. Using the method, the presence of the known fatty acids was confirmed, and new ones were discovered.

• Klíčová slova
• acetonitrile-related adducts, acetylenic lipids, double and triple bond localization, in-source derivatization, mass spectrometry,
• MeSH
• acetonitrily chemie   MeSH
• estery chemie   izolace a purifikace   MeSH
• hmotnostní spektrometrie MeSH
• mastné kyseliny chemie   izolace a purifikace   MeSH
• molekulární struktura MeSH
• včely chemie   MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrile MeSH Prohlížeč
• acetonitrily MeSH
• estery MeSH
• mastné kyseliny MeSH

The advantages of using mixtures of organic solvents for the separation of labeled oligosaccharides on the amide stationary phase under hydrophilic interaction liquid chromatography conditions are presented. The effect of the type of buffer as well as solvent or their mixtures on retention of uracil, saccharide labeling reagents (2-aminobenzoic acid, 2-aminobenzamide, ethyl 4-aminobenzoate, procainamide), and corresponding labeled saccharides were evaluated. The successful isocratic separation of labeled isomeric trisaccharides (maltotriose, panose, and isomaltotriose) was achieved in the mobile phase consisting of a 90% (v/v) mixture of organic solvents (methanol/acetonitrile 60:40) and 10% (v/v) 30 mM ammonium formate, pH 3.3. Changing the volume ratio between methanol/acetonitrile from 60:40 to 50:50 (v/v) allowed to obtain the separation of di-, tri-, and tetrasaccharides labeled by ethyl 4-aminobenzoate in less than 10.5 min.

• Klíčová slova
• HILIC, Mass spectrometry, Oligosaccharide, Saccharide/glycan labeling, Selectivity,
• MeSH
• acetonitrily chemie   MeSH
• amidy chemie   MeSH
• chemické techniky analytické přístrojové vybavení   metody   MeSH
• chromatografie kapalinová * MeSH
• formiáty chemie   MeSH
• hydrofobní a hydrofilní interakce MeSH
• isomerie MeSH
• oligosacharidy izolace a purifikace   MeSH
• ortoaminobenzoáty chemie   MeSH
• rozpouštědla chemie   MeSH
• sacharidy chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetonitrile MeSH Prohlížeč
• acetonitrily MeSH
• amidy MeSH
• anthranilamide MeSH Prohlížeč
• anthranilic acid MeSH Prohlížeč
• formiáty MeSH
• formic acid MeSH Prohlížeč
• oligosacharidy MeSH
• ortoaminobenzoáty MeSH
• rozpouštědla MeSH
• sacharidy MeSH