Integrated spatial metabolomics and network pharmacology to explore the pharmacodynamic substances and mechanism of Radix ginseng-Schisandra chinensis Herb Couple on Alzheimer's disease.

Radix ginseng and Schisandra chinensis have been extensively documented in traditional Chinese medicine (TCM) for their potential efficacy in treating dementia. However, the precise mechanism of their therapeutic effects remains to be fully elucidated. In this study, air flow-assisted desorption electrospray ionization mass spectrometry imaging (AFADESI-MSI) and network pharmacology are used to investigate the pharmacodynamics and mechanism underlying the herbal combination consisting of Radix ginseng-Schisandra chinensis (RS) in a rodent model for Alzheimer's disease (AD). Brain histopathological findings suggested that RS attenuates hippocampal damage in AD mice, making this combination a potential AD treatment. Twenty-eight biomarkers were identified by spatial metabolomics analysis, which are intricately linked to neuroinflammation, neurotransmitter imbalance, energy deficiency, oxidative stress, and aberrant fatty acid metabolism in AD. The total extract of RS (TE) affected 22 of these biomarkers, with the small molecule components of RS (SN) significantly influencing 19 and the large molecule components of RS (PR) impacting 14. Nine small molecule components are likely to dominate the pharmacodynamics of RS. We constructed a target interaction network based on the corresponding bioactivities that revealed relationships amongst 11 key biomarkers, 8 active ingredients and 12 critical targets. This research illustrates the immense potential of spatial metabolomics and network pharmacology in the study of TCM, revealing the targets and mechanisms underlying herbal formulas.

In-depth investigation of the therapeutic effect of Tribulus terrestris L. on type 2 diabetes based on intestinal microbiota and feces metabolomics.

ETHNOPHARMACOLOGICAL RELEVANCE: The fruit of Tribulus terrestris L. (TT) is extensively documented in the Tibetan medical literature 'Si Bu Yi Dian', has been used to treat diabetes mellitus for more than a thousand years. However, the underlying mechanisms and comprehensive effects of TT on diabetes have yet to be investigated. AIM OF THE STUDY: The aim of the study was to systemically elucidate the potential mechanisms of TT in treating diabetes mellitus, and further investigate the therapeutic effects of the water extract, small molecular components and saccharides from TT. MATERIALS AND METHODS: Fecal metabolomics was employed to draw the metabolic profile based on UHPLC-Q-TOF-MS/MS. The V3-V4 hypervariable regions of the bacteria 16S rRNA gene were amplified to explore the structural changes of the intestinal microbiome after TT intervention and to analyze the differential microbiota. The microbial metabolites SCFAs were determined by GC-MS, and the BAs and tryptophan metabolites were quantified by UPLC-TQ-MS. Spearman correlation analysis was carried out to comprehensively investigate the relationship among the endogenous metabolites profile, intestinal microbiota and their metabolites. RESULTS: TT exhibited remarkably therapeutic effect on T2DM rats, as evidenced by improved glucolipid metabolism and intestinal barrier integrity, ameliorated inflammation and remission in insulin resistance. A total of 24 endogenous biomarkers were screened through fecal metabolomics studies, which were mainly related to tryptophan metabolism, fatty acid metabolism, bile acid metabolism, steroid hormone biosynthesis and arachidonic acid metabolism. Investigations on microbiomics revealed that TT significantly modulated 18 differential bacterial genera and reversed the disordered gut microbial in diabetes rats. Moreover, TT notably altered the content of gut microbiota metabolites, both in serum and fecal samples. Significant correlation among microbial community, metabolites and T2DM-related indicators was revealed. CONCLUSIONS: The multiple components of TT regulate the metabolic homeostasis of the organism and the balance of intestinal microbiota and its metabolites, which might mediate the anti-diabetic capacity of TT.

Establishment of Polydopamine-Modified HK-2 Cell Membrane Chromatography and Screening of Active Components from Plantago asiatica L.

Cell membrane chromatography (CMC) has been widely recognized as a highly efficient technique for in vitro screening of active compounds. Nevertheless, conventional CMC approaches suffer from a restricted repertoire of cell membrane proteins, making them susceptible to oversaturation. Moreover, the binding mechanism between silica gel and proteins primarily relies on intermolecular hydrogen bonding, which is inherently unstable and somewhat hampers the advancement of CMC. Consequently, this investigation aimed to establish a novel CMC column that could augment protein loading, enhance detection throughput, and bolster binding affinity through the introduction of covalent bonding with proteins. This study utilizes polydopamine (PDA)-coated silica gel, which is formed through the self-polymerization of dopamine (DA), as the carrier for the CMC column filler. The objective is to construct the HK-2/SiO2-PDA/CMC model to screen potential therapeutic drugs for gout. To compare the quantity and characteristics of Human Kidney-2 (HK-2) cell membrane proteins immobilized on SiO2-PDA and silica gel, the proteins were immobilized on both surfaces. The results indicate that SiO2-PDA has a notably greater affinity for membrane proteins compared to silica gel, resulting in a significant improvement in detection efficiency. Furthermore, a screening method utilizing HK-2/SiO2-PDA/CMC was utilized to identify seven potential anti-gout compounds derived from Plantago asiatica L. (PAL). The effectiveness of these compounds was further validated using an in vitro cell model of uric acid (UA) reabsorption. In conclusion, this study successfully developed and implemented a novel CMC filler, which has practical implications in the field.

A novel strategy on the study of whole intestinal metabolic profiles for Polygalae Radix before and after processing.

INTRODUCTION: Relieving toxicity and enhancing a calming effect after processing Polygalae Radix (PR) are widely known. Aromatic carboxylic acids (ACAs) may be crucial processed products. However, due to the limited detection methods for ACAs, the whole metabolic profiles via intestinal bacteria are still not very clear. OBJECTIVE: Designing a novel strategy for the detection of ACAs and tracking the whole metabolic profiles before and after processing PR. MATERIALS AND METHODS: The stable-isotope labelling derivatisation (SILD) method based on multidimensional ultra-high performance liquid chromatography coupled with a mass spectrometer (UHPLC-MS) technology and UNIFI-pathway mode was firstly designed to systematically study the metabolisms of all the drug-derived ingredients ranging from m/z 100 to 2000 in processing PR via intestinal bacteria. Firstly, the SILD with UHPLC coupled with a triple-quadrupole MS technology was designed to trace eight ACA metabolites of the processed PR with intestinal bacteria. Additionally, the UHPLC coupled with a quadrupole time-of-flight MS with UNIFI-pathway mode was adopted to monitor relatively big metabolites. RESULTS: The metabolism mechanism of ACAs (eight kinds) and the relatively big molecular metabolites (98 kinds) were deeply traced in PR, PR with refined honey (HP), and PR with licorice (LP) via the intestinal bacteria. Totally 106 intact metabolic profiles of drug-derived ingredients were presented. Importantly, the influence of LP on the metabolism of compounds after incubation of intestinal bacteria was greater than that of HP. CONCLUSION: This research provides a comprehensive and systematic guidance for further study on in vivo metabolisms of the processed PR.

Ligand-targeted fishing of α-glucosidase inhibitors from Tribulus terrestris L. based on chitosan-functionalized multi-walled carbon nanotubes with immobilized α-glucosidase.

α-Glucosidase inhibitors in natural products are one of the promising drugs for the treatment of type 2 diabetes. However, due to the complexity of the matrix, it is challenging to comprehensibly clarify the specific pharmacodynamic substances. In this study, a novel high-throughput inhibitor screening strategy was established based on covalent binding of α-glucosidase on chitosan-functionalized multi-walled carbon nanotubes coupled with high-resolution mass spectrometry. The synthesized MWCNTs@CS@GA@α-Glu was characterized by TEM, SEM, FTIR, Raman, and TG. Performance studies showed that the microreactor exhibited stronger thermostability and pH tolerance than that of the free one while maintaining its inherent catalytic activity. Feasibility study applying a model mixture of known α-glucosidase ligand and non-ligands indicated the selectivity and specificity of the system. By integrating ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-QTOF-MS) with ion mobility mass spectrometry (IMS), 15 ligands were obtained and tentatively identified from Tribulus terrestris L., including 8 steroidal saponins, 4 flavonoids, and 3 alkaloids. These inhibitors were further validated by in vivo experiments and molecular docking simulation.

Stabilization of Labile Lysozyme-Ligand Interactions in Native Electrospray Ionization Mass Spectrometry.

Flavonoids are polyphenolic secondary metabolites with extensive biological activities and pharmacological effects. Exploring the interactions of flavonoids with proteins may be helpful for understanding their biological processes. Electrospray ionization mass spectrometry (ESI-MS) is a powerful tool to characterize the noncovalent protein-ligand (PL) complexes. However, some protein-flavonoid complexes are labile during electrospray ionization. Here, the labile lysozyme-flavonoid (rutin, icariin, and naringin) complexes were determined by direct ESI-MS without derivation. It has been found that low amounts of N-methylpyrrolidinone and dimethylformamide can protect labile lysozyme-flavonoid complexes away from dissociation during electrospray ionization process. The intact lysozyme-flavonoid complexes were specifically observed in mass spectra, and the measured binding affinities by ESI-MS were matched with the fluorescence data. The effects of additives on the analysis of lysozyme-flavonoid complexes were investigated by ESI-MS, combined with the molecular docking and fluorescence. This strategy was helpful to investigate the labile PL interactions by direct ESI-MS.

Native Mass Spectrometry Coupled to Spectroscopic Methods to Investigate the Effect of Soybean Isoflavones on Structural Stability and Aggregation of Zinc Deficient and Metal-Free Superoxide Dismutase.

The deficiency or wrong combination of metal ions in Cu, Zn-superoxide dismutase (SOD1), is regarded as one of the main factors causing the aggregation of SOD1 and then inducing amyotrophic lateral sclerosis (ALS). A ligands-targets screening process based on native electrospray ionization ion mobility mass spectrometry (ESI-IMS-MS) was established in this study. Four glycosides including daidzin, sophoricoside, glycitin, and genistin were screened out from seven soybean isoflavone compounds and were found to interact with zinc-deficient or metal-free SOD1. The structure and conformation stability of metal-free and zinc-deficient SOD1 and their complexes with the four glycosides was investigated by collision-induced dissociation (CID) and collision-induced unfolding (CIU). The four glycosides could strongly bind to the metal-free and copper recombined SOD1 and enhance the folding stability of these proteins. Additionally, the ThT fluorescence assay showed that these glycosides could inhibit the toxic aggregation of the zinc-deficient or metal-free SOD1. The competitive interaction experiments together with molecular docking indicate that glycitin, which showed the best stabilizing effects, binds with SOD1 between ß-sheet 6 and loop IV. In short, this study provides good insight into the relationship between inhibitors and different SOD1s.

Treatment effects of Radix ginseng-Schisandra chinensis herb pair on Alzheimer's disease: An investigation of MS-based metabolomics investigation.

Traditional Chinese medicine (TCM) plays a synergistic and comprehensive pharmacodynamic role of multi-channel and multi-target through its multi-components, showing unique therapeutic advantages in chronic and multi-gene complex diseases. Herb pair is a unique combination of two relatively fixed herbs, which embodies the integrity of TCM theory. In this study, untargeted fecal metabolomics based on MS was used to investigate the action mechanism of Radix ginseng and Schisandra chinensis (GS) herb pair on the complex disease of Alzheimer's disease (AD), and further analyze the therapeutic effects of small molecular components and saccharides of GS on AD. Quantitative analysis of bile acids (BAs) and short-chain fatty acids (SCFAs) further verified the conclusion of untargeted metabolomics. The results of the pharmacodynamics evaluation showed that the AD model was successfully constructed, and each TCM group had a different degree of improvement compared with the AD group. PCA analysis based on untargeted fecal metabolomics showed that the metabolic disorders in AD rats changed significantly over time, and there were different degrees of callback in each TCM group. The result indicated that the GS herb pair can regulate metabolic disorders of AD. Further analysis of therapeutic biomarkers showed that GS mainly regulated the metabolism of bile acid biosynthesis, sphingolipid metabolism, porphyrin and chlorophyll metabolism, etc. to treat AD. This study will help to further understand the pathogenesis of AD from metabolomics, and provide beneficial support for the further study of GS and the clinical treatment of complex diseases with TCM.

A Strategy for Identification and Structural Characterization of Compounds from Plantago asiatica L. by Liquid Chromatography-Mass Spectrometry Combined with Ion Mobility Spectrometry.

Plantago asiatica L. (PAL) as a medicinal and edible plant is rich in chemical compounds, which makes the systematic and comprehensive characterization of its components challenging. In this study, an integrated strategy based on three-dimensional separation including AB-8 macroporous resin column chromatography, ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF MS), and ultra-high performance liquid chromatography-mass spectrometry with ion-mobility spectrometry (UHPLC-IM-MS) was established and used to separate and identify the structures of compounds from PAL. The extracts of PAL were firstly separated into three parts by AB-8 macroporous resin and further separated and identified by UHPLC-Q-TOF MS and UHPLC-IM-MS, respectively. Additionally, UHPLC-IM-MS was used to identify isomers and coeluting compounds, so that the product ions appearing at the same retention time (RT)can clearly distinguish where the parent ion belongs by their different drift times. UNIFI software was used for data processing and structure identification. A total of 86 compounds, including triterpenes, iridoids, phenylethanoid glycosides, guanidine derivatives, organic acids, and fatty acids, were identified by using MS information and fragment ion information provided by UHPLC-Q-TOF MS and UHPLC-IM-MS. In particular, a pair of isoforms of plantagoside from PAL were detected and identified by UHPLC-IM-MS combined with the theoretical calculation method for the first time. In conclusion, the AB-8 macroporous resin column chromatography can separate the main compounds of PAL and enrich the trace compounds. Combining UHPLC-IM-MS and UHPLC-Q-TOF MS can obtain not only more fragments but also their unique drift times and RT, which is more conducive to the identification of complex systems, especially isomers. This proposed strategy can provide an effective method to separate and identify chemical components, and distinguish isomers in the complex system of traditional Chinese medicine (TCM).

Effect of Qishen granules on isoproterenol-induced chronic heart failure in rats evaluated by comprehensive metabolomics.

Qishen granules (QSG), a Chinese herbal formula, has been widely used in the treatment of myocardial ischemic chronic heart failure (CHF) for many years, but its mechanism of action is still unclear. In this study, comprehensive metabolomics was used to investigate the underlying protective mechanisms of QSG in an isoproterenol-induced CHF rat model. A total of 14 biomarkers were identified in serum and 34 biomarkers in urine, which were mainly related to fatty acid metabolism, bile acid metabolism, amino acid metabolism, purine metabolism, vitamin metabolism, and inflammation. Finally, 22 markers were selected for quantitative analysis of serum, urine, and fecal samples to verify the reliability of the results of untargeted metabolomics, and the results were similar to those of untargeted metabolomics. The correlation analysis showed that the targeted quantitative endogenous metabolites and CHF-related indexes were closely related. QSG might alleviate myocardial inflammatory response, oxidative stress, and amino acid metabolism disorder in CHF by regulating the level of endogenous metabolites. This study revealed QSG could regulate potential biomarkers and correlated metabolic pathway, which provided support for the further application of QSG.

Comprehensive chemical profiling and potential chemical marker's evaluation of Tribulus terrestris by UPLC-QTOF-MS in combination with ion mobility spectrometry.

Analyzing the chemical components of traditional Chinese medicines containing multiple isomers is thorny. Here, an analytical strategy by using ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry coupled ion mobility spectrometry was applied to characterize the main compounds from the fruits of Tribulus terrestris, which significantly boosted the separation efficiency and broadened the capacity of chromatographic column. A total of 155 chemicals including 120 steroidal saponins, 13 flavonoids, 20 alkaloids and 2 ferulic acids were identified or tentatively identified using the propounded method. An α-glucosidase inhibition test was conducted to compare whether the aptness of Tribulus terrestris differed from different origins. The results indicated that the activity of Tribulus terrestris from Inner Mongolia was better than that from Jilin and Hebei. Principal component analysis was next employed to seek out the potential chemical markers among these Tribulus terrestris from three origins. There were 13 substances exist significant differences in content of Tribulus terrestris from the three producing areas. These significant differences involve 11 steroidal saponins and 2 alkaloids. Among them, Inner Mongolia possessed the highest contention of all the 11 saponins. This suggested steroidal saponins may emerge huge potential of α-glucosidase inhibition activity. In conclusion, the present study furnished the identification of chemical components of medical herbs with various isomers as well as disclosed the latent diabetes treatment potential of steroidal saponins in Tribulus terrestris.

Comprehensive fecal metabolomics and gut microbiota for the evaluation of the mechanism of Panax Ginseng in the treatment of Qi-deficiency liver cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: Qi deficiency liver cancer (QDLC) is an important part of liver cancer research in traditional Chinese medicine (TCM). In the course of its treatment, Panax ginseng is often selected as the main Chinese herbal medicine, and its function has special significance in the tumor treatment of Qi deficiency constitution. However, its mechanism is not clear. AIM OF THE STUDY: The research tried to evaluate the mechanism of Panax ginseng in the treatment of QDLC through fecal metabonomics and gut microbiota on the basis of previous pharmacodynamic evaluation. MATERIALS AND METHODS: Firstly, biomarkers and related metabolic pathways were screened and identified by metabonomics and Kyoto Encyclopedia of Genes and Genomes (KEGG) database. Then, 16S rRNA sequencing technique was used to investigate the composition, ß diversity and key differences of gut microbiota. Finally, the relationship among phenotypes, gut microbiota and fecal metabolites was comprehensively analyzed by spearman correlation coefficient. RESULTS: 31 pharmacodynamic potential biomarkers and 20 synergistic potential biomarkers of effective parts of Panax ginseng on QDLC were screened and identified by fecal metabonomics. And then, 6 major metabolic pathways were searched, including bile acid biosynthesis, unsaturated fatty acid biosynthesis, tryptophan metabolism, arachidonic acid metabolism, pyrimidine metabolism, vitamin B6 metabolism. In the study of gut microbiota, at the genus level, 25 species of bacteria with significant differences of effective parts on QDLC and 23 species of bacteria with significant differences of synergistic action of ginsenosides and polysaccharides were screened. In addition, Spearman correlation analysis showed that there was a complex potential relationship among phenotype, gut microbiota and fecal metabolites during the development of QDLC and Panax ginseng intervention, which was mainly reflected in the close potential relationship between bacteria and fecal metabolites such as bile acids, unsaturated fatty acids and indole compounds. CONCLUSION: Through the changes of fecal endogenous metabolites and intestinal bacteria, the mechanism of Panax ginseng on QDLC were preliminarily clarified.

Boronate Affinity-Based Oriented and Double-Shelled Surface Molecularly Imprinted Polymers on 96-Well Microplates for a High-Throughput Pharmacokinetic Study of Rutin and Its Metabolites.

The boronate affinity-based oriented and double-shelled surface molecularly imprinted polymers on 96-well microplates (BDMIPs) were designed and applied to high-specific and high-throughput pharmacokinetic (PK) study of rutin and its metabolites from rat plasma without concentration and redissolution. It integrated the advantages of covalent effects-based boronate affinity, noncovalent effects of ethylene imine polymer (PEI) dendrimer, multiple cavities-based double-shelled layers, and multiparallel wells-based 96-well microplates. Furthermore, ultrahigh-performance liquid chromatography triple quadrupole tandem mass spectrometry (UHPLC-MS/MS) was used to accurately quantify targets. It showed lower limits of detection (LODs) up to 100-fold than the conventional method. And PKs of rutin and trace isoquercetin (IQC) were first reported at the same time. The platform can provide a fast, simple, low-cost, high-selective, high-effective, and high-throughput methodological reference for analysis of large-scale samples in the fields of agriculture and food.

Pharmacokinetics and tissue distribution study of 18 bioactive components in healthy and chronic heart failure rats after oral administration of Qi-Shen-Ke-Li formula using ultra-high-performance liquid chromatography/triple quadrupole mass spectrometry.

RATIONALE: Qi-Shen-Ke-Li (QSKL) is a traditional Chinese formula used in clinical practice to treat chronic heart failure (CHF) in humans. To rationalize the use of this formula in clinical practice, the pharmacokinetics and tissue distribution in rats after oral administration of QSKL were investigated using ultra-high-performance liquid chromatography/triple quadrupole mass spectrometry (UHPLC/TQ-MS). METHODS: The CHF model was induced by intraperitoneal injection of isoprenaline (ISO; also known as isoproterenol) and evaluated by HE staining and brain natriuretic peptide (BNP) measurement. The UHPLC/TQ-MS method was then applied to determine the concentrations of 18 bioactive components in rat plasma and tissues of heathy and CHF rats after oral administration of QSKL. This was followed by investigating the pharmacokinetics and tissue distribution profiles of these bioactive compounds in the heathy and CHF rats. RESULTS: The pharmacokinetics results showed that the duration time of two compounds was prolonged, the absorption rate of four compounds was accelerated, and the bioavailability of four compounds was increased in the CHF rats compared with the healthy rats. Meanwhile, the tissue distribution results showed that the QSKL formula could be distributed rapidly and widely in different rat tissues. The bioavailability of eight compounds in the liver was enhanced in CHF rats. This suggested that the drug/toxic effects should be considered in clinical practice, as drug-drug interactions might occur in liver metabolism during the drug combination. CONCLUSIONS: The pharmacokinetic profiles and tissue distribution of 18 bioactive compounds in QSKL are altered by the CHF status. This study provides insight for better clinical applications of this formula in the future and lays the foundation for the development of a new drug for chronic heart failure based on the QSKL formula.

Stable isotope labeling derivatization combined with multiple-mass spectrometry technologies to monitor metabolites of tenuifoliside A incubated with intestinal bacteria incubation model.

Aromatic carboxylic acids (ACAs), play important roles in preventive and therapeutic effects for some diseases. However, complex matrix effect and poor detection sensitivity make it difficult and even rare to detect ACAs in complex bio-samples. Herein, a stable isotope labeling derivatization (SILD) method based on one-pot synthesis of carboxylic amides by aniline (AN) and aniline-d5 (AN-d5) was firstly designed for quantitatively monitoring ACAs under mild conditions. The detection sensitivity was improved up to 500 folds. Importantly, when taking the trace tenuifoliside A (TA) containing p-hydroxyl-benzoyl- (HB) and 3, 4, 5-trimethoxylcinnamoyl- (TC) unit as a special example via intestinal bacteria incubation, the metabolites ACAs and whole metabolic profiles of TA were firstly accurately and systematically monitored by applying the SILD method combined with multiple-mass spectrometry (MMS) technologies. It provides a convenient, universal, high-sensitivity and high-recovery methodological tool for the systematically metabolic study of trace drugs in vitro and in vivo.

Investigation of interactions between cytochrome c and ginsenosides by native mass spectrometry and molecular docking simulations.

RATIONALE: Ginsenosides are considered to be the main functional components in ginseng and possess various important pharmacological activities. The study of the interactions between ginsenosides and proteins is indispensable for understanding the pharmacological activities of ginsenosides. In this work, the interactions of ginsenosides with cytochrome c (cyt c) were investigated by native mass spectrometry and molecular docking simulations. METHODS: The interactions of four ginsenosides (Rb1 , Rb3 , Rf, Rg1 ) and cyt c in NH4 OAc solution were investigated by electrospray ionization linear ion trap mass spectrometry (ESI-LTQ-MS). Molecular docking simulations of cyt c complexes were carried out by AutoDock. RESULTS: The native mass spectrometry results showed that the four ginsenosides were directly bound to cyt c, with stoichiometric ratios of 1:1 and 2:1 in NH4 OAc. The order of relative binding abilities of ginsenosides to cyt c obtained by ESI-MS was Rb1 > Rb3 > Rf > Rg1 , which was consistent with the docking results. Moreover, molecular docking simulations also indicated potential binding sites of cyt c and ginsenosides. Hydrogen-bond interaction played a very important role in cyt c binding with ginsenosides. CONCLUSIONS: It has been demonstrated that native MS is a useful tool to investigate the interactions of ginsenosides with cyt c. Molecular docking is a good complement to ESI analysis, and can provide information on potential binding sites of cyt c-ginsenoside complexess. This strategy will be helpful to further understand the interactions of proteins and small molecules.

Trace determination and characterization of ginsenosides in rat plasma through magnetic dispersive solid-phase extraction based on core-shell polydopamine-coated magnetic nanoparticles.

Enrichment of trace bioactive constituents and metabolites from complex biological samples is challenging. This study presented a one-pot synthesis of magnetic polydopamine nanoparticles (Fe3O4@SiO2@PDA NPs) with multiple recognition sites for the magnetic dispersive solid-phase extraction (MDSPE) of ginsenosides from rat plasma treated with white ginseng. The extracted ginsenosides were characterized by combining an ultra-high-performance liquid chromatography coupled to a high-resolution mass spectrometry with supplemental UNIFI libraries. Response surface methodology was statistically used to optimize the extraction procedure of the ginsenosides. The reusability of Fe3O4@SiO2@PDA NPs was also examined and the results showed that the recovery rate exceeded 80% after recycling 6 times. Furthermore, the proposed method showed greater enrichment efficiency and could rapidly determine and characterize 23 ginsenoside prototypes and metabolites from plasma. In comparison, conventional methanol method can only detect 8 ginsenosides from the same plasma samples. The proposed approach can provide methodological reference for the trace determination and characterization of different bioactive ingredients and metabolites of traditional Chinese medicines and food.

In situ analysis of single cell and biological samples with rGO-Cu functional probe ESI-MS spectrometry.

At present, probe electrospray ionization mass spectrometry is widely used in the single cell analysis. As the continuous expansion of requirements for detecting more metabolite species from single cells, the sensitivity and accuracy of the instrument and the enrichment capacity of the probes are in high demand. Therefore, a high efficiency functional probe electrospray ionization mass spectrometry (FPESI-MS) analysis platform is established. In this platform, nitrogen aggregation/gas heating system and reduced graphene oxide functional copper probe (rGO-Cu probe) are firstly reported. The nitrogen aggregation/gas heating system can promote desorption of the analytes from probe and aggregate the product ions into the ion cluster to increase the ion transport capacity and the detection efficiency. rGO-Cu probe formed huge specific surface area to enrich the analytes without sacrificing the electrical conductivity. The probe substrates were prepared by electrochemical etching, after rGO-functionalization, the tip diameter of all kinds of the probes were measured to be less than 100 nm, which is much smaller than the PC12 single cells. Graphene oxide cannot be completely reduced by electrochemical methods, thus the rGO-Cu probe can enrich the analytes through the huge surface area of the graphene thin layer, as well as the hydrogen bond and the electrostatic absorption of the surface oxygen-containing groups. Seven AD related neurotransmitters and sixteen biomarkers in PC12 single cells and twelve metabolites in serum of AD rats were successfully analyzed by using FPESI-MS, and this method is expected to be used in the rapid detection of metabolites in biological samples and living organism.

A metabolomic study of the urine of rats with Alzheimer's disease and the efficacy of Ding-Zhi-Xiao-Wan on the afflicted rats.

As a well-known traditional Chinese medicine formula, Ding-Zhi-Xiao-Wan has long been used for the routine treatment of Alzheimer's disease. However, the mechanism of Ding-Zhi-Xiao-Wan in treating Alzheimer's disease is unclear. Therefore, a nontargeted metabolomics method based on ultrahigh performance liquid chromatography with quadrupole time-of-flight mass spectrometry has been established to explore the metabolic variations in the urine of Alzheimer's disease rats and investigate the therapeutic mechanism of Ding-Zhi-Xiao-Wan on Alzheimer's disease. To develop a better rat model of Alzheimer's disease, amyloid ß25-35 was injected into the bilateral hippocampus of Sprague-Dawley rats. Multivariate analysis approaches were applied to differentiate the urine components between the four groups. Thereafter, a targeted metabolomics method was used to verify the identified endogenous metabolites and determine the mechanism of action of Ding-Zhi-Xiao-Wan. Altogether, 26 potential biomarkers were found, of which 15 biomarkers (10 of which are potential biomarkers found in nontargeted metabolomics) were identified. The results show that Ding-Zhi-Xiao-Wan mainly affects the pathways of taurine and hypotaurine metabolism, tryptophan metabolism, and phenylalanine metabolism. Ding-Zhi-Xiao-Wan might play a role in the treatment of Alzheimer's disease by mediating antioxidative stress, regulation of energy metabolism, improvement of intestinal microbes, and protection of nerve cells.