Acid Isomers Research Articles

Mode of Action of Retinoic Acid in the Regulation of Glucose Metabolism in Mud Crab, Scylla serrata: Evidence for the Involvement of Crustacean Hyperglycemic Hormone

In the current study, we evaluated the effect of retinoids, 9-Cis retinoic acid (9CRA) and all-trans retinoic acid (ATRA) on carbohydrate metabolism in the mud crab, Scylla serrata. Intact and eyestalk ablated male crabs (n=10 crabs per group) were selected and retinoic acid isomers, 9CRA and ATRA were injected into the crabs. Parallel controls were maintained. Significant reduction in the hemolymph sugar levels was observed in crabs subjected to eyestalk ablation (ESX) over intact crabs. Injection of 9CRA into intact but not in ESX crabs exhibited a dose- and time-dependent manner hyperglycemic response. ATRA administration did not induce hyperglycemia either in intact or ESX crabs. Injection of 9CRA showed a significant decrease in total carbohydrate content and glycogen levels in hepatopancreas and muscle tissues of intact crabs over their respective controls. Further, the activity levels of glycogen phosphorylase were also elevated in selected tissues of intact crabs administered with 9CRA. In addition, intact crabs which received 9CRA resulted in up regulation of expression of CHH mRNA from the eyestalks. Taken together, we postulate that 9CRA-induced hyperglycemia might be ascribed to neurotransmitter effects mediating the release of CHH from the eyestalk. The release of CHH eventually caused glycogenolysis in the selected hepatopancreas and muscle tissues of the mud crab Scylla serrata thereby hyperglycemia.

Generation of thiyl radicals in a spatiotemporal controlled manner by light: Applied for the cis to trans isomerization of unsaturated fatty acids/phospholipids.

Thiyl radicals are important reactive sulfur species and can cause cis to trans isomerization on unsaturated fatty acids. However, biocompatible strategies for the controlled generation of thiyl radicals are still lacking. In this work, we report the study of a series of naphthacyl-derived thioethers as photo-triggered thiyl radical precursors. Tertiary naphthacyl thioether was identified to be a suitable template that could be used to produce both aryl and alkyl thiyl radicals under ultraviolet (UV) light or sunlight. The effective cis-to trans-isomerization of unsaturated fatty acid models (methyl oleate, methyl linoleate) and a natural phospholipid (cardiolipin) using these photo-triggered substrates was demonstrated. This reaction was also proved to proceed effectively in cells to produce thiyl radicals and subsequent fatty acid isomerization. Additionally, the most promising thiyl radical precursor showed antiviral activity in a pseudotyped virus model, likely due to disrupting viral lipid membranes upon UV activation. These findings highlight the potential of thiyl radicals for both biochemical and antiviral applications.

Determination of Trends Underlying Aspartic Acid Isomerization in Intact Proteins Reveals Unusually Rapid Isomerization of Tau.

Spontaneous chemical modifications in long-lived proteins can potentially change protein structure in ways that impact proteostasis and cellular health. For example, isomerization of aspartic acid interferes with protein turnover and is anticorrelated with cognitive acuity in Alzheimer's disease. However, few isomerization rates have been determined for Asp residues in intact proteins. To remedy this deficiency, we used protein extracts from SH-SY5Y neuroblastoma cells as a source of a complex, brain-relevant proteome with no baseline isomerization. Cell lysates were aged in vitro to generate isomers, and extracted proteins were analyzed by data-independent acquisition (DIA) liquid chromatography-mass spectrometry (LC-MS). Although no Asp isomers were detected at day 0, isomerization increased over time and was quantifiable for 105 proteins by day 50. Data analysis revealed that the isomerization rate is influenced by both primary sequence and secondary structure, suggesting that steric hindrance and backbone rigidity modulate isomerization. Additionally, we examined lysates extracted under gentle conditions to preserve protein complexes and found that protein-protein interactions often slow isomerization. Base catalysis was explored as a means to accelerate Asp isomerization due to findings of accelerated asparagine deamidation. However, no substantial rate enhancement was found for isomerization, suggesting fundamental differences in acid-base chemistry. With an enhanced understanding of Asp isomerization in proteins in general, we next sought to better understand Asp isomerization in tau. In vitro aging of monomeric and aggregated recombinant tau revealed that tau isomerizes significantly faster than any similar protein within our data set, which is likely related to its correlation with cognition in Alzheimer's disease.

Somatic embryogenesis and micropropagation of Boswellia serrata Roxb.: a rare medicinal tropical tree species

Somatic embryogenesis was investigated in Boswellia serrata Roxb., a medicinal tree species extensively utilized in tropical regions. In this study, an in vitro protocol for somatic embryogenesis and the generation of plantlets from immature zygotic embryos of B. serrata was developed. The morphogenic potential of the zygotic embryos was assessed on Murashige and Skoog (MS) medium enriched with various cytokinins, including 6-furfurylaminopurine (KIN), N6-benzyladenine (BA), and 1-phenyl-3-(1,2,3-thiadiazol-5-yl)-urea (TDZ), and auxins such as indole-3-acetic acid (IAA), α-naphthaleneacetic acid (NAA), and 2,4-dichlorophenoxyacetic acid (2,4-D), both individually as well as in combinations. The MS medium containing 4.54 µM TDZ, 2.26 µM 2,4-D, and 200 mg/l polyvinylpyrrolidone (PVP) was found to be effective in inducing direct somatic embryos (89.63%) in the cultures. Various forms of somatic embryos, including globular, heart-shaped, torpedo-shaped, and cotyledonary shapes, were observed on the surface of the zygotic embryos. Enhanced germination and conversion rates to plantlets (73.2 ± 1.6%) were achieved upon subculturing to a medium containing gibberellic acid (GA3) alone. The survival rate of plantlets under ex vitro conditions was approximately 72%. The stability of the ploidy level in the regenerated plantlets was verified through the application of flow cytometry (FCM). The analysis of leaf samples obtained from plantlets generated through somatic embryogenesis, performed using LC-Q-Tof-MS, identified the presence of the boswellic acid isomer, 3-O-acetyl-11-keto-β-boswellic acid. The concentration of this particular isomer was found to be 0.13 µg/g of dry weight of the leaf. The propagation protocol presented in this study provides a significant micropropagation technique for this valuable plant species. Additionally, this method can be utilized for genetic transformation and the production of bioactive boswellic acids of medicinal significance.

Revealing Unpredicted Aspartic Acid Isomerization Hotspots by Probing Diagnostic Fragmentation Propensities in Top-Down and Middle-Down Mass Spectrometry.

Isomerization of aspartic acid residues is a relevant degradation pathway of protein biopharmaceuticals as it can impair their biological activity. However, the in silico prediction of isomerization hotspots and their consequences remains ambiguous and misleading. We have previously shown that all ion differential analysis (AiDA) of middle-down spectra can be used to reveal diagnostic terminal and internal fragments with more sensitivity than the conventional fragment ion mass matching methodology. In this study, we use AiDA to characterize the degradation of an antibody fragment at three aspartic acid isomerization sites including a novel DW motif directly with electron-transfer/higher-energy collisional dissociation top-down and middle-down mass spectrometry. We show that AiDA methodology is pivotal to probe diagnostic fragmentation propensities of terminal c and z fragments at the N-terminus and vicinity of isomerization sites in addition to the diagnostic c+57 terminal fragments. Furthermore, AiDA can probe remote structural changes in the loop of an antibody complementarity-determining region induced by isomerization and the succinimide intermediate, revealing interactions between residues in agreement with molecular simulations. This study shows that aspartic acid residues at noncanonical DW and DF motifs can be hotspots for isomerization despite being ranked as false positives in physics-based prediction models. We show that the enzyme-free, fast, and sensitive AiDA methodology can be used as an orthogonal technique to fractionation for online variant characterization.

In vivo Differential Effects of Extractable and Non-Extractable Phenolic Compounds from Grape Pomace on the Regulation of Obesity and Associated Metabolic Alterations.

Grape pomace (GP) is a by-product rich in phytochemicals, including extractable polyphenols (EPPs) and non-extractable polyphenols (NEPPs), which have distinct metabolic fates that may affect their biological activities. The benefits of GP have been reported in relation to obesity and its comorbidities, particularly when administered preventively focusing on EPPs. Therefore, the aim of this study was to investigate the effects of EPPs and NEPPs from GP as a treatment for obesity and its associated metabolic alterations. A previous comprehensive characterization of the selected GP revealed the most relevant individual compounds in the EPPs fraction (malvidin hexoside, (-)-epicatechin, quercetin, and procyanidin dimer B2 isomer II), as well as in the NEPPs fraction (hydroxybenzoic acid isomers I-II). The experiment was performed in obese rats with insulin resistance, treated for 8 weeks with EPPs or NEPPs grape pomace fractions (100mg/kg). After the intervention, the HFFD + EPP group showed a significantly lower weight gain (9.6%) and body mass index (9.7%) compared to the HFFD group. While liver triglyceride levels were only significantly reduced in the HFFD + NEPP group (47%) compared to the HFFD group. Neither treatment resulted in a reduction of insulin resistance. Therefore, the supplementation with grape pomace phenolic fractions to an animal model of obesity exerted differential beneficial effects on body weight and liver lipid accumulation, overall contributing to an amelioration of some the metabolic alterations present in obesity, although not to aspects such as glycemic homeostasis.

PROSPECTS OF USING SOME SPECIES OF CHRYSANTHEMUMS AS A SOURCE FOR OBTAINING BIOLOGICALLY ACTIVE SUBSTANCES

Chrysanthemums are widely known all over the world as ornamental plants. However, in Asian countries since ancient times they are considered as dual-purpose crops and along with floristic use have a long history of use as food and medicinal plants. Chrysanthemum morifolium Ramat. and Chry-santhemum indicum L. are the most studied species of the genus Chrysanthemum L., included in the pharmacopoeias of China, Japan and the USA. At the same time, garland (Glebionis coronaria L. (Cass. ex Spach.)) is cultivated in Russia as a food plant, which is also a potential source of plant raw materials for obtaining therapeutic and prophylactic agents. In this connection, the purpose of the present work was to analyze the literature data on the chemical composition and pharmacological properties of G. coronaria in comparison with C. indicum and C. morifolium to assess the prospects of its use in domestic medical practice. According to literature data, all three species of chrysanthemums have similar chemical composition and contain flavonoids (aglycones of luteolin, apigenin, acacetin, quercetin, kaempferol, isoramnetin, hesperitin, diosmetin, their mono- and diglycosides, as well as malonyl and acetyl derivatives of glycosides); phenolic acids (isomers of caffeoyl- and dicaffeoylquinic acids and their methylated derivatives); sesquiterpenoids. Carotenoids were found in flowers; in the above-ground part - fatty acids and their derivatives, sterols and polyacetylenes; in stems, roots and leaves - anthraquinones. Among the mineral elements, the predominant content of potassium is noted. The results of published pharmacological studies indicate that these chrysanthemum species exhibit antioxidant, anti-inflammatory, antimicrobial, nephro-, hepato-, cardio- and neuroprotective effects. Thus, G. coronaria has a similar set of metabolites to C. indicum and C. morifolium, but is characterized by a smaller diversity of identified substances, which opens the prospect of its further phytochemical study. Taking into account the closeness of chemical composition, G. coronaria may exhibit the same types of biological activity, which are identified for other species of chrysanthemums. Analysis of literature data has shown that along with more studied species, G. coronaria can also serve as a source of biologically active substances. The availability of registered varieties and agrotechnologies has the potential to create a stable raw material base for obtaining domestic medicines and prophylactic products.

Comparative metabolomic study of twelve Acacia species by UHPLC-q-tof-ESI-MS coupled with chemometrics in correlation with antibacterial activity.

Genus Acacia comprises around 1500 species. They are widely used to treat inflammation as well as bacterial and fungal infections as they are enriched in phytochemicals, especially phenolics. The aim of this study was to evaluate the antibacterial activity of leaves' methanolic extracts of twelve Acacia species growing in Egypt against Vibrio parahaemolyticus, Salmonella enterica, Listeria monocytogens, Klebsiella pnemoniae, Bacillus aquimaris, Bacillus subtilis, and Escherichia coli. These species are Acacia nilotica (wild and cultivated), Acacia seyal, Acacia auriculiformis, Acacia saligna, Acacia xanthophloea, Acacia tortilis subsp. raddiana (Gabal Elba and Aswan), Acacia tortilis, Acacia laeta (wild and cultivated), and Acacia albida. Furthermore, to study the metabolomic composition and variation among these species using ultra-high-performance liquid chromatography-electrospray ionization quadrupole time of flight mass spectrometry (UHPLC-q-tof-ESI-MS) coupled with multivariate statistical analysis and correlate it to the antibacterial potential. Results showed that Acacia nilotica (AN) has superior antibacterial activity over the other species. In addition, it exhibited a distinct segregation in Principal component analysis (PCA) and partial least square discriminant analysis (PLS-DA). Full profiling of AN using UHPLC-ESI-q-tof-MS revealed 42 phenolics mainly catechins. It was further subjected to bio-guided fractionation and revealed the presence of methyl gallic acid, gallic acid, catechin gallate, and digallate isomers in its most bioactive fraction. These compounds were identical to the compounds annotated as VIPs and were responsible for the segregation of AN in both PCA and PLS-DA analyses. Hence, this study sheds light on the use of chemometrics as an early tool for the detection of bioactive compounds.

Mass spectrometry differentiation of sciadonic acid isomers and estimate of long-chain polyunsaturated fatty acid intake from fresh and processed duck eggs

Mass spectrometry differentiation of sciadonic acid isomers and estimate of long-chain polyunsaturated fatty acid intake from fresh and processed duck eggs

An accurate analytical method based on gas chromatography-mass spectrometry for quantifying all oleic, linoleic, and linolenic acid isomers, including trans-form and cis–trans-mixed isomers, in food

An accurate analytical method based on gas chromatography-mass spectrometry for quantifying all oleic, linoleic, and linolenic acid isomers, including trans-form and cis–trans-mixed isomers, in food

Stability of sugars in yogurts with simple and complex microbial composition during refrigerated shelflife.

There is a large and growing market for natural sweeteners with low glycemic index. Of particular interest are the "rare sugars" (e.g., D-tagatose). Rare sugars could be applied in dairy foods, such as yogurt, as a sucrose replacement. Yogurt contains live and active cultures and consequently their enzymes, including those capable of sugar hydrolysis or isomerization which could undermine the nutritional benefits of rare sugars. The purpose of this research was to evaluate sugar stability and impact of added sugars during storage of commercial nonfat Greek yogurts having simple and complex microbial compositions. Two commercial nonfat Greek yogurt products with different bacterial profiles (simple: Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus; complex: S. thermophilus, L. delbrueckii ssp. bulgaricus, Bifidobacterium lactis, Lactobacillus acidophilus, and Lacticaseibacillus rhamnosus) were supplemented with sugars (fructose, galactose, glucose, lactose, sucrose and D-tagatose) at 10% (wt/wt) in duplicate and sampled every 2 weeks. Individual sugar concentrations and lactic acid were measured using a combination of enzyme kits (Megazyme) and high-performance liquid chromatography (HPLC). Sugar (lactose, glucose, galactose) and lactic acid isomer concentrations were significantly different between the 2 commercial products (p-value < 0.05) but neither product changed significantly during refrigerated storage (p-value > 0.05). Each added sugar (10%) was stable throughout the 6-week refrigerated storage period (p-value > 0.05). The pH of all yogurts decreased slightly (-0.1 pH) during storage, but with no corresponding change in lactic acid concentration. These results provide foundational data on the stability of various sugars in cultured dairy products throughout refrigerated shelflife.

Impact of raw materials on the nutritional value of confections

The techniques for determining the nutritional value of confectionery products, namely, determining the mass fraction of protein, fat, water-soluble vitamins B1 and B2, omega-3 fatty acids, and cholesterol in fat-containing confectionery products, have been developed. Data on the content of protein, fat, saturated fatty acids and trans isomers of unsaturated fatty acids, B vitamins, omega-3 fatty acids,and cholesterol in fat-containing confectionery products have been obtained. A methodology to monitor the content of vitamins, omega-3 fatty acids, and cholesterol in fat-containing confectionery products has been developed.

Bioassay-guided evaluation of antimicrobial properties and profile of bioactive compounds from leaf, peel and mesocarp of four apple cultivars (Malus domestica Borkh.) grown in Serbia: Application of HPTLC-EDA and UHPLC Q-ToF MS techniques

The aim of this study was to evaluate the antimicrobial properties and profile of bioactive compounds from mesocarp, peel and leaves of four autochthonous apple cultivars against human pathogens, Escherichia coli, Staphylococcus aureus and Bacillus subtilis and the apple pathogen Erwinia amylovora by direct detection on HPTLC plates and subsequent chemometric analysis. UHPLC Q-ToF MS was used for detailed characterization of the bioactive compounds with antimicrobial properties. Leaf extracts showed the highest antibacterial activity against all bacterial strains, followed by peel extracts, while the mesocarp extracts showed only weak and selective inhibition zones for E. coli and B. subtillis. The apple cultivars, ‘Kadumana’ and ‘Kopaoničanka’ showed the best antimicrobial properties. The active compounds were triterpenoids, coumaroylquinic acid and caffeoylquinic acid isomers and methyl derivatives, naringenin, kaempferol and quercetin aglycones and glycosides, phloretin and its glycosides and acylated derivatives that detected for the first time in the apple leaves and fruits.

Anti-inflammatory Activity of Chlorogenic Acid and Call for Further Research

In recent years, phenolic acids have garnered considerable interest on account of their diverse biological, practical, and pharmacological effects. Chlorogenic Acid (CGA), presently referred to as 3-CQA, is the most prevalent isomer among caffeoylquinic acid isomers (3-, 4-, and 5-CQA). Among the naturally occurring phenolic acid compounds found in green coffee extracts and tea, it is one of the most readily accessible acids. Recent studies have demonstrated that chlorogenic acid exerts its anti-inflammatory effects via various mechanisms, including the inhibition of inflammatory mediators, demonstration of antioxidant activity, modulation of signaling pathways, and regulation of the immune system. Each of these mechanisms is essential for immune system regulation. Chlorogenic acid mitigates tissue injury and impedes the inflammatory response through mechanisms, including inhibition of pro-inflammatory chemical synthesis and activity, scavenging of free radicals, and modulation of immune cell functionality. Throughout this review article, we have methodically examined the advantageous contributions of CGA in relation to its origin, assimilation, metabolism, molecular mechanisms, mechanistic effects, and impact on various forms of inflammation.

Dissociation Chemistry of Protonated α-, β- and Other Amino Acids Using Multiple Stage Tandem Mass Spectrometry.

To discriminate amino acid isomers by multiple stage tandem mass spectrometry (MSn), the fragmentation of protonated amino acids were investigated by MSn with collision-induced dissociation (CID) and density functional theory calculations. The CID of protonated α-amino acids results in a loss of 46 Da, corresponding to H2O and CO, and iminium ions appear as resultant fragments. The CID of protonated β-amino acids also produces iminium ions, but the corresponding loss is 60 Da instead of 46 Da. H2O loss initiates the fragmentation of protonated β-amino acids, producing protonated β-lactams as an intermediate. Subsequently, protonated β-lactams are easily converted to iminium ions and CH2CO. By contrast, H2O loss from the protonated forms of γ- and ε-amino acids provides protonated lactams with 5- and 7-membered rings, respectively. Protonated lactams with more than 5-membered rings provide stable fragments and do not undergo further degradation during CID. In addition, protonated forms of γ- and ε-amino acids undergo NH3 loss as a competitive fragmentation pathway of H2O loss, producing protonated lactones. Because the fragmentation of protonated amino acid by CID depends on the position of amino and carboxyl groups, the tandem mass spectrometry with CID can discriminate α-, β-, and other amino acids.

Dinor-12-oxo-phytodienoic acid conjugation with amino acids inhibits its phytohormone bioactivity in Marchantia polymorpha.

Jasmonates (JAs) are important phytohormones that regulate plant tolerance to biotic and abiotic stresses, and developmental processes. Distinct JAs in different plant lineages activate a conserved signaling pathway that mediates these responses: dinor-12-oxo-phytodienoic acid (dn-OPDA) isomers in bryophytes and lycophytes, and JA-Ile in most vascular plants. In many cases, the final responses triggered by these phytohormones depend on the accumulation of specialized metabolites. To identify compounds regulated by the dn-OPDA pathway in the liverwort Marchantia polymorpha, untargeted metabolomic analyses were carried out in response to wounding, a stress that activates the dn-OPDA pathway. A previously unreported group of molecules was identified from these analyses: dn-OPDA-amino acid conjugates (dn-OPDA-aas). Their accumulation after wounding and herbivory was confirmed by targeted metabolic profiling in Marchantia and in all species in which we previously detected dn-iso-OPDA. Mutants in GRETCHEN-HAGEN 3A (MpGH3A) failed to accumulate dn-OPDA-aa conjugates and showed a constitutive activation of the OPDA pathway and increased resistance to herbivory. Our results show that dn-iso-OPDA bioactivity is reduced by amino acid conjugation. Therefore, JA conjugation in land plants plays dichotomous roles: jasmonic acid conjugation with isoleucine (Ile) produces the bioactive JA-Ile in tracheophytes, whereas conjugation of dn-iso-OPDA with different amino acids deactivates the phytohormone in bryophytes and lycophytes.

Capsular polysaccharide structure of Acinetobacter baumannii K58 from clinical isolate MRSN31468

Capsular polysaccharides (CPS) of Acinetobacter baumannii is a virulence factor with diverse structures. CPS are produced by the CPS biosynthesis gene cluster in their K locus (KL). However, CPS variations may occur due to insertion of additional genes from external sources, e.g., prophages. Recently, the CPS structure from a clinical isolate, BAL062 which includes KL58 locus, was found to have a pseudaminic acid isomer (8ePse5NAc7NAc) as a result of prophage inserted epaA/epaB genes. Here, we report a CPS structure produced by A. baumannii strain MRSN31468 which also belongs to a KL58 type. The K58 CPS structure was determined by 1D and 2D NMR analysis of the oligosaccharides derived from the CPS by a phage depolymerase, and supported by the sugar composition analysis. The K58 CPS structure has the following tetra saccharide repeating unit. [Display omitted] The K58 CPS differs from the CPS from BAL062 only by replacing 8-epimerized β-8ePse5NAc7NAc with β-Pse5NAc7NAc.

Integrating Epoxidation, High-Resolution Mass Spectrometry and Ultraviolet Spectroscopy to Unravel the Complex Profile of Boswellic Acids and Related Compounds in the Boswellia serrata Gum Resin Extract

The chemical characterization of natural products is often a complex task that demands powerful analytical techniques. Liquid chromatography with high-resolution tandem mass spectrometry (HRMS/MS) is often employed, yet it can face hard challenges when isomeric species are present, and reference standards are lacking. In such cases, the confidence level in compound identification can be significantly improved by the collection of orthogonal information on target analytes. In this work, 23 key compounds in Boswellia serrata extract (BSE), 12 of which correspond to boswellic acids (BAs) and 11 to triterpenoidic acid isomers, were identified by combining RPLC followed by serial UV and ESI(-)-FTMS and FTMS/MS detections with the evaluation of the reactivity towards C=C bond epoxidation with meta-chloroperoxybenzoic acid (m-CPBA), proposed as a fast chemical tool to gather information about C=C bond steric hindrance, a key structural feature of BAs and related compounds. The interpretation of UV spectra acquired after chromatographic separation corroborated the identification of the substitution patterns of enonic and dienic residues in ketoboswellic and dehydroboswellic acids. Moreover, MS/MS based on higher-energy collision-induced dissociation (HCD) unveiled new fragmentation pathways, providing important structural details on target analytes. The integrated approach developed during this study might pave the way for a deeper understanding of the BSE bioactive properties. Moreover, it can be considered an example of a more general strategy for the analysis of complex mixtures of natural compounds including also isomeric species.

Inhibition of LPS-induced inflammatory response in RAW264.7 cells by natural Chlorogenic acid isomers involved with AKR1B1 inhibition

Inflammation is the physiological response of the immune system to injury or infection, typically manifested by local tissue congestion, swelling, heat, and pain. Prolonged or excessive inflammation can lead to tissue damage and the development of many diseases. The anti-inflammatory effects of natural ingredients have been extensively researched and confirmed. This study investigated the effects of Chlorogenic acid (CGA) isomers —— 3-Caffeolyquninic acid (3-CQA), 4-Caffeolyquninic acid (4-CQA), and 5-Caffeolyquninic acid (5-CQA) —— on the inflammatory response and oxidative stress reaction induced by LPS in RAW264.7 cells. Overall, 3-CQA exhibited the most significant reduction in levels of TNF-α, IL-6, NO, and ROS. 4-CQA showed superior inhibition of TNF-α compared to 5-CQA (p < 0.05), while no significant difference in other parameters. We further used DARTS and CETSA to demonstrate that CGA isomers have stable affinity with AKR1B1. As a positive control, the AKR1B1 antagonist epalrestat exhibited similar effects to the CGA isomers. 3-CQA having the smallest half-inhibitory concentration (IC50) for AKR1B1, while 4-CQA and 5-CQA have similar values. AutoDock simulations of the docking conformations revealed minimal differences in the average binding energies of the CGA isomers. The main differences were that VAL47 formed a hydrogen bond with 3-CQA, whereas GLN49 formed hydrogen bonds with 4-CQA and 5-CQA. Additionally, the number of hydrophobic bonds involving PHE122 and LEU300 varies. Our conclusion is that differences in non-covalent interactions result in the varying inhibitory abilities of CGA isomers on AKR1B1, which further affect the anti-inflammatory and antioxidant effects of CGA isomers.

Rapid separation of bile acid isomers via ion mobility mass spectrometry by complexing with spiramycin.

Bile acid (BA) is one of the main active components of bile and has multiple isomers, the structure or content of its isomers often changes due to diseases and other health problems; thus, the accurate detection of BA isomers is very important. In this study, two groups of BA isomers of glycine-conjugated BAs and taurine-conjugated BAs were simultaneously separated and quantitatively analyzed by ion mobility mass spectrometry (IM-MS). Especially, baseline mobility separation between the isomers was achieved by the formation of binary complexes via simple interaction with spiramycin (SPM), for which a separation resolution (Rp-p) of 1.96was reached. Moreover, BA isomers were quantitatively analyzed, and the limit of detection (LOD) of absolute quantification for TCDCA/TUDCA and GUDCA/GCDCA/GHDCA was 0.514 and 0.611ng∙mL-1, respectively; the LODs for molar ratio ranges of relative quantification for TCDCA/TUDCA, GUDCA/GHDCA, and GCDCA/GHDCA were 1:18-30:1, 1:18-21:1, and 1:19-21:1, respectively. Additionally, BA isomers analyzed in pig bile powder and bear bile powder were measured, which were in good consistency with those labeled, revealing the differences in BA composition and content between the two powders. Finally, BA detection and recovery analyses were performed on serum samples, with a recovery rate of ≥73.69%, RSD of ≤6.8%, and SR (standard deviation of recoveries, the degree of difference between measured values and average recovery) of ≤1.27. Due to the simple, rapid, and lack of need for complex sample preparation and chromatographic separation, the proposed method can be an effective method for BA detection in practical samples.