An integrated approach to identify rare hemoglobin variants using capillary electrophoresis, liquid chromatography and mass spectrometry.

Comparing capillary coatings for protein separation by capillary electrophoresis.

Derivatization-free quantification of L-glutamic acid in commercial chips and sauces via capillary electrophoresis with contactless conductivity detection.

Highly sensitive laser-induced fluorescence detector for algal identification by capillary electrophoresis

A compact visible-light spectrometric detector for capillary electrophoresis.

In recent years, a new compound genetic marker, deletion/insertion polymorphism-single nucleotide polymorphism (DIP-SNP), has been applied in forensic DNA mixture analysis. DIP-SNP genotyping based on capillary electrophoresis (CE) typically requires two separate amplification reactions and is difficult to interpret in complex DNA mixtures. In this study, four candidate imprinted DIP-SNP markers were selected to establish a multiplex polymerase chain reaction (PCR) set based on the amplification refractory mutation system (ARMS) principle for a one-tube reaction. Furthermore, parentally imprinted allele (PIA) typing was applied to selectively detect parental alleles. The imprinting patterns of rs35918685-rs185148 and rs11667883-rs76183558 were confirmed to be maternally imprinted, and both markers showed imprinting consistency in saliva, vaginal fluid, and menstrual blood. In DNA mixture analysis, these two markers successfully detected a minor DNA contributor in a two-person DNA mixture at a 1:20 ratio, indicating high analytical sensitivity for low template input and imbalanced mixtures. An improved RMNE method, described in a recent study, was applied to evaluate its effectiveness in narrowing down potential contributors in DNA mixtures, and a comparison of RMNE probabilities between conventional genotyping and PIA genotyping demonstrated the improved efficiency of PIA-based interpretation. These findings highlight the high forensic potential of imprinted DIP-SNP markers, particularly in the analysis of imbalanced DNA mixtures.

Chirality plays a crucial role in pharmaceutical development, as enantiomers often display distinct therapeutic, pharmacokinetic, and safety profiles. Calcium channel blockers (CCBs), a cornerstone in antihypertensive therapy, frequently possess one or more chiral centers, making enantioselective analysis essential for pharmacological and regulatory accuracy. Despite growing evidence of stereoselective differences, most marketed CCBs remain available as racemates, underscoring the need for refined analytical approaches. This review aims to bridge that gap by critically synthesizing advances in analytical, bioanalytical, and regulatory strategies for chiral CCBs. Key separation platforms-including high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), gas chromatography (GC), and supercritical fluid chromatography (SFC)-are evaluated with emphasis on chiral stationary phases such as polysaccharides, cyclodextrins, macrocyclic antibiotics, and crown ethers. Recent innovations in hyphenated mass spectrometry, spectroscopic techniques, and eco-friendly miniaturized systems are highlighted for their ability to enhance enantioselectivity, reduce solvent consumption, and improve sensitivity in complex matrices. Case studies illustrate the pharmacokinetic and pharmacodynamic relevance of individual enantiomers, reinforcing the clinical impact of chirality in CCB therapy. The review concludes by identifying current limitations and exploring future directions, including AI-assisted selector design, green chemistry integration, and lab-on-chip platforms to advance personalized antihypertensive treatment.

Stable isotope-ratio analysis by inductively coupled plasma-mass spectrometry (ICP-MS) with a multi-collector (MC-ICP-MS) is increasingly used in life sciences, offering mechanistic insight and potential diagnostic specificity. This review asks the following: What limits biomedical isotope-ratio work, and what steps will make results comparable and fit for biomedical interpretation? Our target audience includes analytical chemists, MC-ICP-MS practitioners, liquid chromatography (LC) and capillary electrophoresis (CE) users, metrology and assurance/quality control specialists, and biomedical researchers adopting isotope-ratio workflows. We map the field from bulk delta (δ) values to chemical speciation, species-specific isotope ratios, and spatially resolved readouts. Specific choices in sampling, pre-analytics, separation, sample introduction, and instrument setup are linked to the main sources of bias and to the limits of precision and traceability. Recent progress is synthesized across automated clean-up, LC/ICP-MS and CE/ICP-MS workflows, transient-signal handling, and species-specific isotope dilution with enriched spikes. The review provides practical guidance on baseline correction, peak integration, mass-bias correction, scale realization, and uncertainty budgets. Across studies, the bottlenecks are species instability and interconversion, matrix and space-charge effects, sample-spike mismatch, spectral interferences, lack of species-specific reference materials, and inconsistent operating procedures. We close with an outlook that prioritizes (i) automation and transient-signal processing, (ii) matrix-matched species-specific reference materials, (iii) instrument advances for interference control and coupling efficiency, and (iv) harmonized data-processing standard operating procedures. Together, these steps can enable reproducible multi-site biomedical isotope-ratio workflows and support their translation towards clinical applicability by clarifying where MC-ICP-MS already adds value and where research is still needed.

A micro-nano structure enhanced chiral separation strategy is proposed by integrating biomimetic liposome assemblies with cyclodextrin (CD) derivatives as composite chiral selectors in capillary electrophoresis (CE). The self-assembly of weakly chiral, poorly soluble phospholipids into liposomes markedly amplifies chiral discrimination when coupled with CD-based molecular recognition, enabling highly efficient separation of multiple 9-fluorenylmethoxycarbonyl (Fmoc)-amino acid enantiomers. This enhancement is proposed to be associated with the formation of interfacial pre-enrichment microzones on liposome surfaces, where locally concentrated selectors and analytes may engage in synergistic hydrogen bonding, hydrophobic interactions, and steric effects, collectively strengthening chiral recognition. The composite system exhibits good selectivity and compatibility in mixed-sample separations, supporting simultaneous enantioseparation of multiple amino acids with high reproducibility. Beyond establishing an effective CE-based approach for amino acid enantiomer separation, this work demonstrates a proof-of-concept for transforming poorly soluble, weakly chiral substances into efficient chiral selectors through micro-nano assembly, offering preliminary insights for synergistic chiral separation systems.

The study investigates the capillary electrophoretic enantioseparation of agrochemicals (dichloroprop and mecoprop) using modified cyclodextrins 2-hydroxypropyl-β-cyclodextrin (CD-1), heptakis (2,3-di-O-methyl)-β-cyclodextrin (CD-2), heptakis (2,3-di-O-methyl-6-sulfo)-β-cyclodextrin (CD-3), and heptakis (2,3,6-tri-O-methyl)-β-cyclodextrin (CD-4) in 2-(N-morpholino)ethanesulfonic acid (MES) buffer. Migration times varied depending on the selector type and inclusion complex type and stability, reflecting differences in enantiomer-selector interaction strengths. The reversal of enantiomer migration order observed among the cyclodextrins suggested that even small structural differences in the chiral selectors influenced changes in intermolecular interactions and complex stability. This reversal order is useful for understanding the mechanisms. The separations were due to inclusion complexes with neutral cyclodextrins, while electrostatic interactions were responsible for negatively charged cyclodextrins. Docking and ROESY-NMR studies were used to provide supportive structural insight into the nature of chiral recognition mechanisms. The docking provided general structural information about the potential binding modes of these systems. However, native β-cyclodextrin was used for the ROESY-NMR studies because it produced very clear spectra, allowing the comparison of the basic interaction patterns among the various β-cyclodextrin derivatives. Overall, the results demonstrated that capillary electrophoresis (CE) is a sensitive technique for investigating intermolecular interactions and gaining insight into enantioselective processes.

Next-generation sequencing (NGS) provides increased discriminatory power in forensic DNA analysis due to the detection of isoalleles. Differences in sequences between alleles allow for a second layer of differentiation between DNA contributors beyond the number of short tandem repeat (STR) repeat units. However, because NGS is a more time and resource-intensive analysis than conventional capillary electrophoresis (CE), laboratories may benefit from indicators that suggest NGS is likely to provide added value. This study examined whether CE migration offsets, measured as residuals in the OSIRIS analysis software, can differ significantly among STR isoalleles. Residuals represent the time offset between a sample allele peak and its corresponding allelic ladder peak. Paired CE and NGS data from 95 single source samples were analyzed for CE-based residual differences, as the NGS data provided the sequence information of the corresponding isoalleles. Residual values differed significantly among isoalleles at several STR loci. Statistically significant differences were identified at D16S539 and D3S1358, as well as at specific allele lengths within D12S391, D13S317, and D8S1179. These findings demonstrate that CE residual variation can reflect underlying STR sequence differences between contributors. In practice, residual-based metrics could help laboratories to identify casework reference samples where NGS is likely to provide additional discrimination, without the need for processing outside of a routine CE workflow. Due to the potentially large number of isoalleles, community wide efforts to aggregate CE residual differences versus isoallele sequences may be useful in the validation and implementation of this approach to add value to forensic DNA analyses.

Chromatographic and electrophoretic methods reliably distinguish the heterozygous and homozygous sickle cell states with hemoglobin variants appearing at consistent percentages with specific retention or migration times. However, co-migrating variants alter the overall hemoglobin profile, complicating interpretation in compound heterozygous conditions such as βS/βD-Punjab since these variants electrophoretically migrate close to each other. These cases often require hematological correlation or molecular confirmation. As molecular testing is not available in all laboratories, mass spectrometry (MS) may serve as an alternate approach for examining complex compound heterozygous sickle cell disorders. Hemoglobin extracted from K2 EDTA blood samples (n = 154) was analyzed using flow-injection triple quadrupole MS. Summed intensities of three charge states each of βS and β globin monomers enabled calculation of the βS/β ratio to differentiate sickle cell trait (SCT) from sickle cell disease (SCD). Additional globin ratios (α/β, γ/β, δ/α (%)) were evaluated to subclassify SCD into Group 1 (βSβE, βSβD-Punjab, βSβC, βSLepore-BW), Group 2 (βSβ0), and Group 3 (βSβS). Statistical analysis was performed using SPSS v27. The βS/β ratio effectively differentiated SCT from SCD with an optimal cut-off value of 0.71 yielding 90.3% sensitivity and 98.4% specificity. The ratios α/β, γ/β, δ/α (%), βS/β, significantly discriminated against Group 1 from Group 3 (p < 0.001). While γ/β ratio significantly differentiated Group 1 from Group 2 (p < 0.001), the δ/α (%) ratio was superior for distinguishing Group 2 from Group 3 (p < 0.001). A rapid, cost-effective MS approach distinguishes SCT from SCD while γ/β and δ/α (%), ratios refine SCD subclassification alongside βS/β. Combined with capillary electrophoresis (CE), this approach enhances diagnostic resolution and may reduce the need for molecular testing.

Low-molecular-weight proteins (LWPs, <30 kDa) are crucial in the identification of tumor markers and disease diagnosis. However, current methods for analyzing LWPs typically involve complex workflows, high sample consumption, poor automation, and prolonged reaction times. In this study, immobilized enzyme microreactors (IMERs) were prepared by immobilizing trypsin encapsulated in zeolitic imidazolate frameworks (ZIF-L) within a capillary, which was then integrated with capillary electrophoresis (CE) to develop a novel strategy for the pretreatment and assay of LWPs. Compared to traditional LWP analysis protocols, which typically involved a three-step process of sequential separation, denaturation, and enzymatic digestion, the proposed strategy achieved the LWP analysis in a single step within just 4 min in a 10 μL sample, significantly reducing sample pretreatment procedures and analytical time while offering advantages such as high efficiency, rapid processing, automation, and low consumption. The prepared trypsin@ZIF-L@IMER exhibited superior activity, enhanced affinity, remarkable stability, and excellent reusability when compared with free enzyme. In addition, the trypsin@ZIF-L@IMER demonstrated high selectivity toward LWPs after the pretreatment of single, binary, quaternary, and septenary model proteins. Additionally, the development method demonstrated ultrahigh sensitivity (0.05 nM cytochrome c (CYC)) and excellent anti-interference capability (CYC/bovine serum albumin (BSA) = 1:1000), outperforming previously reported methods. Finally, the developed strategy based on trypsin@ZIF-L@IMER was used for LWP analysis in human serum. The results showed that trypsin@ZIF-L@IMER possessed higher selectivity and pretreatment capability toward LWPs in human serum when compared with other methods and free trypsin, and the identified LWPs played critical functions and roles in serum-related biological processes by Gene Ontology analysis. The developed method provides a novel strategy for efficient protein pretreatment analysis, which can be further extended to the pretreatment analysis of other proteins.

Ultrafiltration of low-concentration chymosin hydrolysis of skim milk: Impact on membrane processing performance and milk protein concentrate functional properties.

Carbohydrate-deficient transferrin (CDT) is a specific biomarker for chronic excessive alcohol consumption. The IFCC Working Group on CDT Standardization recommends 2-sialo-transferrin (2-sialo-Tf) as the sole measurand, with a decision threshold of 2.0 % CDTIFCC using HPLC. This study evaluates the impact of implementing Sebia's conversion equation on CDT measurements by capillary zone electrophoresis (CZE) for IFCC-alignment. 8,309 serum samples from a driver's license regranting program and 226 samples from patients tested forcongenital disorders of glycosylation (negative controls) were analyzed. Deming regression compared the original Sebia method with IFCC-aligned results following conversion. The upper limit of normal (ULN) was determined using control and population-based analyses. Age- and sex-dependent 0-sialo-Tf breakpoints were also evaluated. Sebia's conversion equation resulted in an upward shift in CDT within the 0-4 % range, increasing the proportion of positive samples by ∼80 % compared with the original Sebia method using a 2.3 % cut-off; applying the IFCC 2.0 % threshold still yielded a ∼50 % increase. The ULN was consistently 1.6 %, biological and analytical uncertainty correction supported the 2.0 % threshold. Although age- and sex-dependent 0-sialo-Tf breakpoints were statistically significant, exclusion of 0-sialo-Tf did not affect positivity, supporting 2-sialo-Tf as standalone marker. Longitudinal iQC monitoring confirmed reproducible quantification but revealed calibration-related positive shifts attributable to the conversion equation. Sebia's conversion equation increases the proportion of positive CDT results irrespective of the applied cut-off. However, our study demonstrates that Sebia's converted CDT, at a 2.0 % cut-off, is a robust marker for CDT measurement and achieves more harmonized CDT reporting.

Advancements in cofactor regeneration for efficient UDP-GlcNAc and UDP-GalNAc synthesis.

Intracellular membrane proteins represent a vast and pharmacologically essential class of therapeutic targets, yet their characterization within native pathological environments remains experimentally challenging. Here, we report what is, to the best of our knowledge, the first direct affinity measurements performed at the surface of Golgi microsomes isolated from cancer cells. By integrating affinity capillary electrophoresis (ACE) with a refined mathematical framework, we overcome long-standing obstacles associated with membrane protein analysis, including the need for target modification, intrinsic enzymatic activity, labeled probe, or large protein excess. The microfluidic format further minimizes sample consumption while enabling both direct and indirect detection modalities, thereby expanding applicability to impure compounds and ligands lacking charge or UV absorbance. Applying this strategy to UGCG, a Golgi-resident membrane protein of oncological relevance, we demonstrate quantitative ligand-binding measurements on fairly monodisperse vesicular fractions (∼150 nm) that preserve native intracellular architecture. Collectively, this work establishes a robust, broadly applicable platform for target engagement studies in complex biological systems and highlights the potential of ACE to accelerate drug discovery efforts involving intracellular or extracellular membrane-bound targets.

The metabolic landscape of biliary tract cancer (BTC) remains poorly characterized. This study aimed to identify tumor-specific metabolic alterations in BTC using paired tumor and adjacent normal tissues. Metabolomic profiling was performed on paired tumor and adjacent normal tissues from 71 patients with BTC using capillary electrophoresis time-of-flight mass spectrometry. Differential metabolites were identified using paired statistical analysis with false discovery rate correction. Pathway enrichment analysis was conducted using the Kyoto Encyclopedia of Genes and Genomes database. Seventeen metabolites were significantly altered between tumor and normal tissues. Pathway analysis identified glycerophospholipid metabolism as the most enriched pathway, driven by water-soluble precursor and intermediate metabolites, including phosphorylcholine, CDP-choline, and ethanolamine phosphate. Hierarchical clustering demonstrated partially distinct metabolic patterns between tumor and normal tissues, with substantial inter-sample variability observed among tumor samples. Metabolites related to amino sugar and nucleotide sugar metabolism were also increased in tumor tissues. Additional pathways, including nicotinate and nicotinamide metabolism and arginine and proline metabolism, were also enriched. Principal component analysis showed partial separation between tumor and normal samples, indicating global metabolic differences between the two groups. These findings indicate metabolic alterations across multiple pathways in BTC. Paired tissue metabolomics revealed coordinated metabolic alterations in BTC involving choline phospholipid precursor metabolism, amino sugar and nucleotide sugar metabolism, and additional amino acid-related pathways. These results highlight the presence of broad metabolic reprogramming in BTC and underscore the importance of tissue-based metabolomic profiling for characterizing tumor metabolism.

Erlotinib (ERL), a first-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), continues to be a mainstay treatment for EGFR-driven cancers, especially non-small cell lung cancer. It is analytically challenging to determine ERL in pharmaceutical products and biological matrices with accuracy and reliability due to its limited therapeutic window, variable bioavailability, substantial metabolism, and low plasma concentrations. Therefore, quality control, pharmacokinetic analysis, stability evaluation, and therapeutic drug monitoring all depend on strong analytical techniques. The analytical methods for ERL determination that have been published over the last 20 years are critically compiled and assessed in this review. The spectroscopic, chromatographic, hyphenated, and electrophoretic techniques such as capillary electrophoresis, high-performance liquid chromatography (HPLC), high-performance thin-layer chromatography (HPTLC), UV-visible spectrophotometry, and liquid chromatography-tandem mass spectrometry (LC-MS/MS) are methodically examined. Alongside new developments like automation, green analytical chemistry, and sophisticated sample-preparation techniques, current analytical issues including matrix interference, solvent usage, and method scalability are discussed. This review offers a thorough analytical perspective intended to guide future development and technique selection for ERL analysis in clinical and pharmaceutical settings.

α-Glucosidase immobilization on cellulose filter paper coated with a blend of chitosan and polyvinyl alcohol for inhibitor screening from traditional Chinese medicines.