Quantitative characterization of glutaminolysis in human plasma using liquid chromatography-tandem mass spectrometry.

Glutaminolysis is the metabolic pathway that lyses glutamine to glutamate, alanine, citrate, aspartate, and so on. As partially recruiting reaction steps from the tricarboxylic acid (TCA) cycle and the malate-aspartate shuttle, glutaminolysis takes essential place in physiological and pathological situations. We herein developed a sensitive, rapid, and reproducible liquid chromatography-tandem mass spectrometry method to determine the perturbation of glutaminolysis in human plasma by quantifying 13 involved metabolites in a single 20-min run. A pHILIC column with a gradient elution system consisting of acetonitrile-5 mM ammonium acetate was used for separation, while an electrospray ionization source (ESI) operated in negative mode with multiple reaction monitoring was employed for detection. The method was fully validated according to FDA's guidelines, and it generally provided good results in terms of linearity (the correlation coefficient no less than 0.9911 within the range of 0.05-800 µg/mL), intra- and inter-day precision (less than 18.38%) and accuracy (relative standard deviation between 89.24 and 113.4%), with lower limits of quantification between 0.05 and 10 µg/mL. The new analytical approach was successfully applied to analyze the plasma samples from 38 healthy volunteers and 34 patients with type 2 diabetes (T2D). Based on the great sensitivity and comprehensive capacity, the targeted analysis revealed the imperceptible abnormalities in the concentrations of key intermediates, such as iso-citrate and cis-aconitate, thus allowing us to obtain a thorough understanding of glutaminolysis disorder during T2D. Graphical abstract ᅟ.

Investigation of chondroitin sulfate D and chondroitin sulfate E as novel chiral selectors in capillary electrophoresis.

Various chiral selectors have been utilized successfully in capillary electrophoresis (CE); however, the number of polysaccharides used as chiral selectors is still small and the mechanism of enantiorecognition has not been fully elucidated. Chondroitin sulfate D (CSD) and chondroitin sulfate E (CSE), belonging to the group of glycosaminoglycans, are linear, sulfated polysaccharides with large mass. In this paper, they were investigated for the first time for their potential as chiral selectors by CE. The effect of buffer composition and pH, chiral selector concentration, and applied voltage were systematically examined and optimized. A variety of drug enantiomers were resolved in the buffer pH range of 2.8-3.4 using 20 mM Tris/H3PO4 buffer with 5.0 % CSD or CSE and 20 kV applied voltage. A central composite design was used to validate the optimized separation parameters and satisfactory uniformity was obtained. As observed, CSE allowed satisfactory separation of the enantiomers of amlodipine, laudanosine, nefopam, sulconazole, and tryptophan methyl ester, as well as partial resolution of citalopram, duloxetine, and propranolol under the optimized conditions. CSD allowed partial or nearly baseline separation of amlodipine, laudanosine, nefopam, and sulconazole. The results indicated that CSE has a better enantiorecognition capability than CSD toward the tested drugs.

Identify production area, growth mode, species, and grade of Astragali Radix using metabolomics "big data" and machine learning.

BACKGROUND: Astragali Radix (AR) is a widely used herbal medicine. The quality of AR is influenced by several key factors, including the production area, growth mode, species, and grade. However, the markers currently used to distinguish these factors primarily focus on secondary metabolites, and their validation on large-scale samples is lacking. PURPOSE: This study aims to discover reliable markers and develop classification models for identifying the production area, growth mode, species, and grade of AR. METHODS: A total of 366 batches of AR crude slices were collected from six provinces in China and divided into learning (n = 191) and validation (n = 175) sets. Three ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) methods were developed and validated for determining 22 primary and 10 secondary metabolites in AR methanol extract. Based on the quantification data, seven machine learning algorithms, such as Nearest Neighbors and Gradient Boosted Trees, were applied to screen the potential markers and build the classification models for identifying the four factors associated with AR quality. RESULTS: Our analysis revealed that secondary metabolites (e.g., astragaloside IV, calycosin-7-O-ß-D-glucoside, and ononin) played a crucial role in evaluating AR quality, particularly in identifying the production area and species. Additionally, fatty acids (e.g., behenic acid and lignoceric acid) were vital in determining the growth mode of AR, while amino acids (e.g., alanine and phenylalanine) were helpful in distinguishing different grades. With both primary and secondary metabolites, the Nearest Neighbors algorithm-based model was constructed for identifying each factor of AR, achieving good classification accuracy (>70%) on the validation set. Furthermore, a panel of four metabolites including ononin, astragaloside II, pentadecanoic acid, and alanine, allowed for simultaneous identification of all four factors of AR, offering an accuracy of 86.9%. CONCLUSION: Our findings highlight the potential of integrating large-scale targeted metabolomics and machine learning approaches to accurately identify the quality-associated factors of AR. This study opens up possibilities for enhancing the evaluation of other herbal medicines through similar methodologies, and further exploration in this area is warranted.

Evaluation of the enantioselectivity of carbon nanoparticles-modified chiral separation systems using dextrin as chiral selector by capillary electrokinetic chromatography.

Nanoparticles (NPs) can be used as pseudostationary phases (PSPs) in EKC, which is similar to the use of micelle additives as applied in MEKC. To date, the use of NPs to enhance enantiomeric separation by EKC with ß-CD or its derivative as chiral selector has been reported only in two papers. However, to the best of our knowledge, there has been no prior effort to use NPs for achieving enantioseparation with polysaccharides as chiral selector. This paper describes for the first time the use of carbon nanoparticles (CNPs) as PSPs to modify chiral separation system employing dextrin as chiral selector for the enantioseparations of several basic drugs in capillary EKC. Three different types of CNPs, including carbogenic nanoparticles (NPs), carboxylated single-walled carbon nanotubes, and carboxylated multiwalled carbon nanotubes, were used as running buffer additives, respectively. The potential of the PSPs and the effects of dextrin concentration, buffer pH, and buffer concentration on the enantioseparations were evaluated. Four pairs of tested enantiomers were successfully resolved in less than 15 min with the resolution values in the range of 1.41-4.52 under optimized conditions. Compared to the buffer without NPs, the introduction of NPs into the buffer enhanced the separation of the enantiomers.

[Transmembrane transport and metabolism of diammonium glycyrrhizinate across rat small intestine in Ussing Chamber].

OBJECTIVE: To explore the transmembrane transport and metabolism of diammonium glycyrrhizinate in intestines of rats. METHOD: An Ussing Chamber model were used to investigate the transmembrane transport of diammonium glycyrrhizinate (GZ), the concentrations of diammonium glycyrrhizinate and its two metabolites were determined by HPLC. RESULT: The permeability coefficients of GZ in difference intestinal mucous membranes were ranged from 0.3 x 10(-6) cm x s(-1) to 1.1 x 10(-6) cm x s(-1). The metabolism of GZ in enterocytes during its transport process was negligible. The concentration of diammonium glycyrrhizinate and pH had limit effects on the transport amount and the permeability coefficients of GZ. CONCLUSION: GZ is a low permeability drug, but it can be absorbed at all segments of the small intestine in rats. Ileum is the major absorption region of GZ.

Investigation of the synergistic effect with amino acid-derived chiral ionic liquids as additives for enantiomeric separation in capillary electrophoresis.

Recently, chiral ionic liquids (ILs) have drawn more and more attention in chiral separation by capillary electrophoresis (CE). In this paper, two chiral ILs based on amino acid derivatives, L-alanine and L-valine tert butyl ester bis (trifluoromethane) sulfonimide, were applied for the first time in CE to evaluate their potential synergistic effects with classical chiral selectors (ß-cyclodextrin derivatives) for enantiomeric separation. As observed, improved separation of tested drug enantiomers was obtained with the presence of chiral ILs compared to the conventional ß-cyclodextrin derivatives separation system. Parameters such as type and proportion of organic modifier, type and concentration of chiral ILs, concentration of chiral selector, buffer pH and applied voltage were systematically investigated with Me-ß-CD/chiral ILs as model system to optimize the novel synergistic system, and the best results were obtained when 15 mM chiral ILs were introduced into the 30 mM sodium citrate/citric acid (20% organic modifier included) buffer solution containing 20 mM Me-ß-CD at pH 5.0 with a 20 kV applied voltage for naproxen, pranoprofen and warfarin.