Journal of separation science最新文献

Endogenous glycopeptides in human serum are valuable candidates for disease biomarker discovery, but their low abundance and structural complexity pose significant analytical challenges. In this study, we developed and optimized a robust workflow for enriching and in-depth characterizing the endogenous glycopeptidome using a novel hydrophilic interaction liquid chromatography (HILIC) sorbent. The HILIC-based strategy effectively enriched glycopeptides from mono-glycosylated IgG and avidin digests, as well as multi-glycosylated horseradish peroxidase (HRP), demonstrating high selectivity, femtomolar-level sensitivity (down to 1 fmol), reproducibility (RSD > 1), and reusability for up to four cycles. Comprehensive analysis by mass spectrometry (MS) identified 334 endogenous intact N-linked glycopeptides, including 318 N-glycosites from 289 glycoproteins. In addition, 242 endogenous intact O-linked glycopeptides from 38 glycoproteins are identified. These represent one of the most extensive endogenous glycopeptide datasets to date. N-glycans were primarily complex biantennary types, while Core 1 structures dominated O-glycans. Motif analysis revealed a strong preference for canonical N-glycosylation motifs (NXT/NXS) and a nearly equal distribution of O-glycosylation on serine and threonine residues. Gene Ontology analysis indicated functional enrichment in membrane-associated, catalytic, and immune-related processes. This optimized HILIC-MS platform enables sensitive and reproducible glycoproteomic profiling, providing a valuable tool for biomarker discovery.

High-performance affinity microcolumns with entrapped humic acid were utilized to investigate interactions between this natural carrier agent and several classes of antibiotics that are common emerging environmental contaminants, or micropollutants. Aldrich humic acid was used as a general model for this type of binding agent. Chromatographic studies under various temperature and mobile phase conditions were used to characterize interactions of the humic acid with the antibiotics sulfadiazine and sulfamethoxazole (sulfonamides), clarithromycin (a macrolide), and lincomycin (a lincosamide). It was determined by this approach that sulfadiazine and sulfamethoxazole had moderate affinities for the humic acid at pH 7.0 and 25°C, with distribution equilibrium constants (K_D) of ∼2-3 × 10¹ L/kg and global affinities (nK'_a) of ∼0.8-1.0 × 10³ M^-1. Lincomycin and clarithromycin had stronger binding, with K_D and nK'_a values of 3.8-7.5 × 10² L/kg and 1.3-2.6 × 10⁴ M^-1. All the antibiotics had a negative $\Delta G^0$ for this binding, representing spontaneous reactions, and a negative change in enthalpy; however, the change in free energy due to entropy was positive in some cases but negative in others. The binding strength decreased in each case as the ionic strength increased. A change in pH also affected binding, as was consistent with the presence of significant electrostatic interactions from some of the antibiotics. These experiments demonstrated how affinity microcolumns could be employed to study such interactions quickly and with only small amounts of binding agent. The fundamental information obtained through this analytical technique should be valuable in characterizing the transport and activity of these antibiotics in the environment and in adapting this approach to the study of other binding agents and micropollutants that may be found in water.

A chromatography/mass spectrometry method is presented for the determination of the weight percent of aromatic compounds in a heavy marine fuel oil. This information facilitates the estimation of the stability, engine performance, and environmental as well as health-related consequences related to heavy fuel oils. These fuel oils are complex mixtures of many different compound types. Thus, a previously reported method was used to first separate the oil sample via distillation, precipitation, and fractionation into asphaltenes, heavy saturated hydrocarbons, alkylaromatic hydrocarbons, aromatic and heteroaromatic, as well as polar compounds. For the fractionation, both flash column chromatography and solid-phase extraction techniques were utilized to ensure proper separation of the compound classes in the maltenes fraction of the sample. High-temperature two-dimensional gas chromatography coupled with high-resolution electron ionization time-of-flight mass spectrometry was used to determine the overall compound compositions of the obtained fractions (other than the asphaltenes) and to classify the detected compounds. The results indicate that the saturated compound fraction contained only alkanes, while all the other fractions contained aromatic compounds. Combining the weight percent of these fractions was used to determine the weight percent of the aromatic compounds in the oil sample.

This study introduces an innovative method for continuous multistage liquid–liquid extraction. Addressing challenges like coalescence and settling times that limit efficiency makes centrifugal partition extraction a promising technique for continuous product separation. It combines the benefits of a mixer-settler cascade with centrifugal extractors in one compact apparatus. The working principle is based on centrifugal partition chromatography, where two immiscible liquid phases are contacted inside a rotor, one serving as the mobile phase and the other as the stationary phase. In centrifugal partition extraction, both phases are mobile. The rotating apparatus allows dispersion and reduces settling time compared with static settlers, with its 300 times higher acceleration. To overcome constructional challenges, a novel concept, termed counter-current centrifugal extraction, is introduced in this work. Thereby, the primary objective is to validate the hypothesis that the process is feasible with a modified rotor design consisting of 10 chambers connected by individual ducts. Combined with alternating pump directions, this enables a pseudo-continuous counter-current flow. A proof of concept is investigated using computational fluid dynamics. The methodologies employed yield valuable insights into hydrodynamic performance. Based on these simulations, an optimized geometry for the flow area is developed, where 100% of the total chamber height is utilized as a dispersion zone compared with the previous 45%. The improvements result in elliptical chambers inclined at 35° to the radial direction, with a height of 6 mm and a volume of 1.43 mL. Additionally, baffles are placed inside the chambers opposite to the two inlets, enhancing dispersion of the inflowing phases. Finally, during alternating operation mode, it is demonstrated that ducts with a volume of 0.17 mL lead to an alternating phase ratio between 0.48 and 0.57 inside the chambers, which was repeatable over five simulated switches. This confirms the feasibility of the concept proposed based on hydrodynamics.

This study examined an improved and simplified method for solid-phase extraction that provides rapid and accurate determination and identification of 116 pharmaceutical and personal care products (PPCPs) in an aqueous environment using ultra-high performance liquid chromatography-tandem mass spectrometry. The common active compounds include 22 sulfonamides, 18 quinolones, 8 macrolides, 18 β-agonists, 6 sedative-hypnotics, 3 antipyretic-analgesics, 3 antihypertensives, 2 antidiabetic drugs, 3 antihistamines, 8 sex hormones, 2 antivirals, 6 nitroimidazoles, 8 glucocorticoids, and 3 amphenicols, lincomycin, pimaricin, levothyroxine sodium, bisphenol A, aldosterone, and melamine in water samples. Key parameters of tandem mass spectrometry, ultra-high performance liquid chromatography, and solid-phase extraction were optimized to enhance the analytical performance. The calibration curves were accomplished at seven concentration levels, and a satisfactory linear relationship (R > 0.99) was observed within the range of 5–800 ng/mL. Results showed varying limits of detection (0.0136–13.3 ng/L for different analytes based on signal-to-noise (S/N) = 3) and limits of quantitation (0.0452–44.4 ng/L). Recoveries of the spiked samples ranged from 53.1% to 116.5% with relative standard deviation lower than 9.9%. This approach effectively minimized matrix interference, improved extraction efficiency, and enhanced detection sensitivity, enabling more accurate PPCP residue analysis in water. The validated method was applied to raw water, treated water, and river water samples from Hangzhou, detecting 47 compounds at concentrations ranging from nondetected to 359 ng/L. Our findings provided critical technical support for the preliminary establishment of an environmental monitoring system targeting emerging pollutants. Notably, to the best of our knowledge, this study represented the first reported detection of melamine, loratadine, aldosterone, and levothyroxine sodium in aquatic environments. In particular, melamine was detected in aquatic environment for the first time, thus expanding the understanding of PPCPs’ pollution status.