[Evaluation of efficacy of Dihuang Baoyuan Granules for diabetes mellitus and composition and in vivo distribution analysis based on UHPLC-LTQ-Orbitrap MS].

Dihuang Baoyuan Granules is a prescription endorsed by HU Tianbao, a renowned and elderly Chinese medicine practitioner from Beijing, and has demonstrated definite clinical efficacy. The composition of this prescription is intricate as it includes 7 distinct herbal medicines. This study aims to analyze the chemical composition of Dihuang Baoyuan Granules, evaluate its efficacy in the treatment of diabetes and analyze the distribution of the drug components in the plasma, liver, and kidney after administration. The findings will serve as a reference for future research on pharmacodynamic substances of this prescription. UHPLC-LTQ-Orbitrap MS was employed to analyze the main chemical components of Dihuang Baoyuan Granules. A Waters ACQUITY Premier HSS T3 column(2.1 mm×100 mm, 1.8 µm) was used for chromatographic separation with 0.1% formic acid(A)-acetonitrile(B) as the mobile phases in a gradient elution at a flow rate of 0.3 mL·min~(-1). Electrospray ionization(ESI) source was used to acquire data in positive and negative ion modes. Furthermore, a rat model of diabetes mellitus was established by feeding with a high-sugar high-fat diet, and injection with streptozocin at a dose of 35 mg·kg~(-1), and the modeled rats were then administrated with Dihuang Baoyuan Granules. The fasting blood glucose, hemoglobin A1c, and other relevant indicators were measured, and the substances present in the plasma, liver, and kidney were identified. By reference to quasi-molecular ions, MS/MS fragment ions, MS spectra of reference substances, and compound information in available reports, 191 components were identified in Dihuang Baoyuan Granules, including 29 alkaloids, 24 flavonoids, 22 organic acids, 16 amino acids, 12 terpenes, 11 steroid saponins, 9 sugars, 8 phenylethanoid glycosides, 8 nucleosides, 2 phenylpropanoids, and 49 others compounds. Eighty-three chemical components were identified in rat plasma, 109 in the liver, and 98 in the kidney. Component identification and characterization of Dihuang Baoyuan Granules in vitro and in vivo provide efficacy information and guidance for the basic research on the pharmacodynamic substances and further clinical application of this prescription.

Investigating the mechanism of cornel iridoid glycosides on type 2 diabetes mellitus using serum and urine metabolites in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Cornel iridoid glycosides (CIG) are extracted from Corni fructus, a herbal medicine used in traditional Chinese medicine to treat diabetes. However, the antidiabetic effects of CIG and the underlying metabolic mechanisms require further exploration. AIM OF THE STUDY: This study aimed to assess the antidiabetic effects and metabolic mechanism of CIG by performing metabolomic analyses of serum and urine samples of rats. MATERIALS AND METHODS: A rat model of type 2 diabetes mellitus (T2DM) was established by administering a low dose of streptozotocin (30 mg/kg) intraperitoneally after 4 weeks of feeding a high-fat diet. The model was evaluated based on several parameters, including fasting blood glucose (FBG), random blood glucose (RBG), urine volume, liver index, body weight, histopathological sections, and serum biochemical parameters. Subsequently, serum and urine metabolomics were analyzed using ultra-high-pressure liquid chromatography coupled with linear ion trap-Orbitrap tandem mass spectrometry (UHPLC-LTQ-Orbitrap-MS). Data were analyzed using unsupervised principal component analysis (PCA) and supervised orthogonal partial least squares discriminant analysis (OPLS-DA). Differential metabolites were examined by the Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathways to explore the underlying mechanisms. RESULTS: After 4 weeks of treatment with different doses of CIG, varying degrees of antidiabetic effects were observed, along with reduced liver and pancreatic injury, and improved oxidative stress levels. Compared with the T2DM group, 19 and 23 differential metabolites were detected in the serum and urine of the CIG treatment group, respectively. The key metabolites involved in pathway regulation include taurine, chenodeoxycholic acid, glycocholic acid, and L-tyrosine in the serum and glycine, hippuric acid, phenylacetylglycine, citric acid, and D-glucuronic acid in the urine, which are related to lipid, amino acid, energy, and carbohydrate metabolism. CONCLUSIONS: This study confirmed the antidiabetic effects of CIG and revealed that CIG effectively controlled metabolic disorders in T2DM rats. This seems to be meaningful for the clinical application of CIG, and can benefit further studies on CIG mechanism.

[Component profile analysis of Sanhan Huashi Formula based on UHPLC-LTQ-Orbitrap-MS, GC-MS, and UHPLC-QQQ-MS/MS].

This study comprehensively analyzed the active components of Sanhan Huashi Formula using qualitative and quantitative mass spectrometry techniques, laying the foundation for understanding its pharmacological substance basis. UHPLC-LTQ-Orbitrap-MS and GC-MS technologies were used to analyze and identify the volatile and non-volatile components in Sanhan Huashi Formula. UHPLC-QQQ-MS/MS technology was used to simultaneously determine the content of 27 major active components in the formula. The results showed that 308 major chemical components were identified in Sanhan Huashi Formula, among which 60 compounds were identified by comparing with reference standards, mainly including alkaloids, flavonoids, coumarins, triterpenoid saponins, amino acids, and nucleosides. GC-MS technology preliminarily identified 52 volatile compounds, with γ-eudesmol and ß-eudesmol as the main components. The quantitative results demonstrated good linearity(r>0.99) for the 27 active components, indicating the stability, simplicity, and reliability of the established method. Among them, amygdalin, nodakenin, arecoline, ephedrine, and pseudoephedrine had relatively high content and were presumably the main pharmacologically active substances. In conclusion, this study systematically and comprehensively characterized the major chemical components and patterns in Sanhan Huashi Formula, providing a basis for understanding its pharmacological mechanisms and clinical applications.

Using ultra-performance liquid chromatography with linear ion trap-electrostatic field orbitrap mass spectrometry, network pharmacology, and molecular docking to explore the constituent targets and action mechanisms of decoction of Angelica sinensis, Zingiberis Rhizoma Recens, and Mutton in the treatment of diarrhea-predominant irritable bowel syndrome.

BACKGROUND: Irritable bowel syndrome (IBS) is a chronic intestinal disorder characterized by abdominal discomfort, stool characteristics, and changes in bowel habits. Among them, diarrhea-type (diarrhea-predominant irritable bowel syndrome, abbreviated as IBS-D) is the most common. Because its pathogenesis is not understood, symptomatic treatment is currently used in clinical practice, and its long-term effect is still unclear. Decoction of Angelica sinensis, Zingiberis Rhizoma Recens, and Mutton (DAZM) is a famous traditional Chinese medicine recipe created by Zhang Zhongjing, a famous doctor in the Eastern Han Dynasty 2000 years ago, and is still in use today. Our research team has previously investigated the clinical study of DAZM in the treatment of IBS-D and conducted animal experiment research, indicating that DAZM has a significant effect on improving IBS-D. Yet, there are few reports on the specific mechanism of action of DAZM in improving the treatment of IBS and related types. Most studies discuss and verify its efficacy and protection from a clinical perspective. For this reason, this research will explore the constituent targets and mechanisms of DAZM to improve the treatment of IBS-D, provide relevant scientific evidence, and also provide reference evidence for the efficacy of food therapy decoction in improving the treatment of diseases and mechanism to open up new experimental research ideas. METHODS: Identification of drug ingredients and collection of targets for DAZM using ultra-performance liquid chromatography with linear ion trap-electrostatic field orbitrap mass spectrometry and the Bioinformatics Analysis Tool for Molecular Mechanisms of Traditional Chinese Medicine database, active ingredients were selected based on their oral bioavailability and drug-like properties. Obtained IBS-D targets using the GeneCards database, took the intersection of IBS-D targets and DAZM targets and obtained potential targets of DAZM for the treatment of IBS-D. Using Cytoscape software to draw a network diagram of "Food therapy decoction-ingredient-target-disease" and selected the ingredients with larger parameter values by topological analysis. Correlation analysis of the selected active and parametric ingredients with prominent symptoms of IBS-D using SymMap database, and selection of potential core ingredients. The construction of protein interaction networks from the String database and the selection of potential core targets. Gene Ontology functional and Kyoto Encyclopedia of Genes and Genomes signaling pathway enrichment analyses using the Metascape database, establishing the bioinformatic processes and signaling pathways involved. Molecular docking of core ingredients and potential core targets was performed using AutoDock Vina, and the results were visualized using Python molecule (PyMOL) and LigPlus+. Finally, based on the results of this research combined with previous literature reports, the discussion section of this paper summarizes in detail the key core ingredients, targets, and signaling pathways of DAZM in improving IBS-D. RESULTS: DAZM may act on eight potential core targets (threonine kinase 1, insulin, tumour necrosis factor, tumour protein p53, interleukin 6, epidermal growth factor receptor, connexin ß1, and interleukin 1ß) through eight core ingredients (Zingiberone, Shyobunone, Palmitic acid, Sebiferic acid, ß-Bisabolene, ß-Sitosterol, Stigmasterol, and Oleic acid). inhibit pro-inflammatory factors through Advanced Glycation End Products-Receptor (AGE-RAGE) signaling pathway, Cyclic Adenosine Monophosphate (cAMP) signaling pathway, MAPK signaling pathway, IL-17 signaling pathway, Calmodulin (CaM) signaling pathway, and other pathways. It can alleviate the inflammatory response, enhance the intestinal mucosal barrier and regulate intestinal motility, and play a role in the treatment and improvement of IBS-D. CONCLUSIONS: This research mainly found the mechanism of DAZM on IBS-D, which may involve multiple ingredients, multiple targets, and multiple pathways. DAZM with medicinal and edible functions can effectively improve the treatment of IBS-D. This kind of dietary therapy is suitable for long-term treatment and is worthy of promotion.

Variation in the chemical profiles of three foxglove species in the central Balkans.

The aim of this study was to determine intra- and interspecies variation in the qualitative and quantitative composition of methanol-soluble metabolites in the leaves of three Digitalis species (D. lanata, D. ferruginea, and D. grandiflora) from the central Balkans. Despite the steady use of foxglove constituents for human health as valuable medicinal products, populations of the genus Digitalis (Plantaginaceae) have been poorly investigated to describe their genetic and phenetic variation. Following untargeted profiling using UHPLC-LTQ Orbitrap MS, by which we identified a total of 115 compounds, 16 compounds were quantified using the UHPLC(-)HESI-QqQ-MS/MS approach. In total, 55 steroid compounds, 15 phenylethanoid glycosides, 27 flavonoids, and 14 phenolic acid derivatives were identified across the samples with D. lanata and D. ferruginea showing a great similarity, while 15 compounds were characteristic only for D. grandiflora. The phytochemical composition of methanol extracts, considered here as complex phenotypes, are further examined along multiple levels of biological organization (intra- and interpopulation) and subsequently subjected to chemometric data analysis. The quantitative composition of the selected set of 16 chemomarkers belonging to the classes of cardenolides (3 compounds) and phenolics (13 compounds) pointed to considerable differences between the taxa studied. D. grandiflora and D. ferruginea were found to be richer in phenolics as compared to cardenolides, which otherwise predominate in D. lanata over other compounds. PCA revealed lanatoside C, deslanoside, hispidulin, and p-coumaric acid to be the main compounds contributing to the differences between D. lanata on one side and D. grandiflora and D. ferruginea on the other, while p-coumaric acid, hispidulin, and digoxin contribute to the diversification between D. grandiflora and D. ferruginea. However, quantitative variation in the metabolite content within species was faint with mild population diversification visible in D. grandiflora and particularly in D. ferruginea. This pointed to the highly conserved content and ratio of targeted compounds within the analyzed species, which was not severely influenced by the geographic origin or environmental conditions. The presented metabolomics approach might have, along with morphometrics and molecular genetics studies, a high information value for further elucidation of the relationships among taxa within the genus Digitalis.

[Effect of processing method on chemical constituents of Rehmanniae Radix: based on UHPLC-LTQ-Orbitrap MS].

This study aims to explore the chemical composition of Rehmanniae Radix braised with mild fire and compare the effect of processing method on the chemical composition of Rehmanniae Radix. To be specific, ultra-high performance liquid chromatography with linear ion trap-orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) was used to screen the chemical constituents of Rehmanniae Radix. The chemical constituents were identified based on the relative molecular weight and fragment ions, literature information, and Human Metabolome Database(HMDB). The ion peak area ratio of each component before and after processing was used as the index for the variation. SIMCA was employed to establish principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) models of different processed products. According to the PCA plot, OPLS-DA plot, and VIP value, the differential components before and after the processing were screened out. The changes of the content of differential components with the processing method were analyzed. A total of 66 chemical components were identified: 57 of raw Rehmanniae Radix, 55 of steamed Rehmanniae Radix, 55 of wine-stewed Rehmanniae Radix, 51 of repeatedly steamed and sundried Rehmanniae Radix Praeparata, 62 of traditional bran-braised Rehmanniae Radix, and 63 of electric pot-braised Rehmanniae Radix. Among them, the 9 flavonoids of braised Rehmanniae Radix were from Citri Reticulatae Pericarpium. PCA suggested significant differences in the chemical composition of Rehmanniae Radix Praeparata prepared with different processing methods. OPLS-DA screened out 32 chemical components with VIP value >1 as the main differential components. Among the differential components, 9 were unique to braised Rehmanniae Radix(traditional bran-braised, electric pot-braised) and the degradation rate of the rest in braised(traditional bran-braised, electric pot-braised) or repeatedly steamed and sundried Rehmanniae Radix was higher than that in the steamed or wine-stewed products. The results indicated the chemical species and component content of Rehmanniae Radix changed significantly after the processing. The 32 components, such as rehmapicrogenin, martynoside, jionoside D, aeginetic acid, hesperidin, and naringin, were the most important compounds to distinguish different processed products of Rehmanniae Radix. The flavonoids introduced by Citri Reticulatae Pericarpium as excipient may be the important material basis for the effectiveness of braised Rehmanniae Radix compared with other processed products.

Characterization and identification of chemical constituents in Corni Fructus and effect of storage using UHPLC-LTQ-Orbitrap-MS.

Corni Fructus (CF) has been widely used as both traditional medicine and food; however, systematic studies on its chemical profile and the impact of storage periods on the indicative components are lacking. In this study, UHPLC-LTQ-Orbitrap-MS was used to investigate the fragmentation behaviors of multiple compounds from CF and the content variety of its indicative components for different storage periods. The major basic components of CF were determined to be iridoid glucosides, pentacyclic triterpenoids, phenolic acids, tannins and flavonoids. The characteristic cleavage pathways of the iridoid glucosides, pentacyclic triterpenoids, phenolic acids, tannins and flavonoids were further investigated and elaborated, which could assist in identifying the structures of similar components of other Chinese herbal medicines. Using accurate mass measurements for each precursor ion and the subsequent fragmented ions, and then comparing with standards and literature data, a total of 130 components, including 69 iridoid glucosides, 9 pentacyclic triterpenoids, 16 phenolic acids, 20 tannins and 16 flavonoids, 47 of which are potentially new compounds, were identified. The storage period studies indicated that the contents of 19 indicative components in CF changed differently with the prolongation of the storage period. Among them, morroniside, loganin, sweroside, cornuside, gallic acid, oleanolic acid and ursolic acid were the most important. These results provide abundant information for the identification and improved understanding of the chemical constituents in CF to clarify the content variety of its indicative components for different storage periods.

Protective Effects of Arbutus unedo L. Honey in the Alleviation of Irinotecan-Induced Cytogenetic Damage in Human Lymphocytes-An In Vitro Study.

Strawberry tree (Arbutus unedo L.) honey (STH) has been used since ancient times as a folk medicine remedy, especially in certain Mediterranean countries. This honey, rich in phenolic content, is well recognized for its antioxidant, anti-inflammatory, and antimicrobial activities, and is used for the treatment of skin lesions as well as gastrointestinal and respiratory disorders. This study investigated whether STH alleviates genome damage in human peripheral blood lymphocytes produced by the cytotoxic drug irinotecan. The phenolic profile of STH was previously estimated by ultra-high-performance liquid chromatography coupled to a linear ion trap-Orbitrap hybrid mass spectrometer. The effects of STH were evaluated at three concentrations (1×, 5×, and 10×), based on the daily consumption of the honey by an adult person. After 2 h of in vitro exposure, standard lymphocyte cultures for the analysis of chromosome aberrations and the cytokinesis-block micronucleus cytome assay were established. Our results demonstrate that STH offered remarkable geno- and cytoprotection when administered with irinotecan. These findings are relevant for drawing preliminary conclusions regarding the in vitro safety of the tested honey. However, further studies are needed with the application of more complex experimental models.

The profiles of durian (Durio zibethinus Murr.) shell phenolics and their antioxidant effects on H2O2-treated HepG2 cells as well as the metabolites and organ distribution in rats.

Durian is a nutritious tropical fruit with potent antioxidant, anti-inflammatory, antibacterial and anti-cancer effects. However, the durian shell was mainly discarded as waste, while there were few studies on the characterization of its phenolic profiles, antioxidant activities, and in vivo metabolites. In the present study, a total of 17 compounds were identified in durian shell extract (DSE) by using an ultra-high-performance liquid chromatography coupled with linear ion trap quadrupole Orbitrap mass spectrometry (UHPLC-LTQ-Orbitrap-MS/MS), while 33 metabolites were found in rats' plasma, urine and organ. Moreover, DSE could effectively reduce H2O2-induced oxidative damage in HepG2 cells, reduce the expression of Reactive Oxygen Species (ROS), Malondialdehyde (MDA) and Lactate Dehydrogenase (LDH) and inhibit apoptosis by regulating the expression of Bcl-2-Associated X (BAX), B-Cell Lymphoma 2 (BCL-2), Caspase-3 and Caspase-9 genes and proteins related to mitochondrial pathway apoptosis. This is the first comprehensive report on Durian shell phenolics, their metabolic profiles and underlying mechanisms of the in vitro antioxidant activities.

Authenticity assessment of cultivated berries via phenolic profiles of seeds.

Considering the health-benefits of berry fruits consumption and increased market demands for food authenticity as one of the most important quality assurances, phenolic profiling by high-performance thin layer chromatography and ultra-high-performance liquid chromatography hyphenated with mass spectrometry was combined with multivariate analysis for phytochemical characterization and intercultivar discrimination of cultivated berry seeds. The phenolic profiles of 45 berry seeds from nine genuine Serbian cultivated fruit species (strawberry, raspberry, blackberry, black currant, blueberry, gooseberry, cape gooseberry, chokeberry, and goji berry) revealed a good differentiation according to botanical origin. In order to determine biomarkers responsible for the classification, a total of 103 phenolic compounds were identified, including 53 phenolic acids and their derivatives, 26 flavonoids and 24 glycosides. Biomarkers derived from the phenolic profile of berry seeds proved to be a powerful tool in the authentication of botanical origin, and may be useful in detection of frauds in berry-based seed-containing product.

Establishment of Specific Chromatogram and TLC Identification for Qingxin Lianziyin / 中国实验方剂学杂志

ObjectiveTo establish the specific chromatogram and thin layer chromatography（TLC） of Qingxin Lianziyin（QXLZY） benchmark samples， in order to clarify the key quality attributes and provide a reference for the quality evaluation of QXLZY. MethodHigh performance liquid chromatography（HPLC） specific chromatogram of QXLZY benchmark samples was developed by using a YMC Hydrosphere C18 column（4.6 mm×250 mm， 5 μm） with the mobile phase of acetonitrile（A）-0.2% formic acid aqueous solution（B） for gradient elution（0-10 min， 5%-20%A； 10-20 min， 20%A； 20-25 min， 20%-24%A； 25-40 min， 24%-30%A； 40-55 min， 30%-50%A； 55-65 min， 50%-100%A； 65-75 min， 100%A； 75-75.1 min， 100%-5%A； 75.1-90 min， 5%A）， and the detection wavelength was 360 nm. Ultra-high performance liquid chromatography-linear ion trap/orbitrap mass spectrometry（UHPLC-LTQ-Orbitrap MS） with electrospray ionization（ESI） was used to identify the components of QXLZY benchmark samples by accurate relative molecular weight and multilevel MS fragment ion information， the detection conditions were positive and negative ion modes and data dependency scanning mode. TLC identification methods for Ophiopogonis Radix， Lycii Cortex， Nelumbinis Semen， Poria， Astragali Radix and Ginseng Radix et Rhizoma in QXLZY were established. ResultA total of 15 characteristic peaks were identified from Glycyrrhizae Radix et Rhizoma， Plantaginis Semen and Scutellariae Radix， and the relative standard deviations of the retention times of 15 characteristic peaks in 15 batches of QXLZY benchmark samples were≤3% with peak 8（baicalin） as the reference peak. A total of 100 compounds， including flavonoids， organic acids， saponins， amino acids and others， were identified in the benchmark samples by UHPLC-LTQ-Orbitrap MS. The established TLC had good separation and was suitable for the identification of Ophiopogonis Radix， Lycii Cortex， Nelumbinis Semen， Poria， Astragali Radix and Ginseng Radix et Rhizoma in QXLZY. ConclusionThe material basis of QXLZY benchmark samples is basically determined by MS designation and source attribution. The established specific chromatogram and TLC of QXLZY are simple， stable and reproducible， which can provide a reference for the development and quality control of QXLZY.

Effect of processing method on chemical constituents of Rehmanniae Radix: based on UHPLC-LTQ-Orbitrap MS / 中国中药杂志

This study aims to explore the chemical composition of Rehmanniae Radix braised with mild fire and compare the effect of processing method on the chemical composition of Rehmanniae Radix. To be specific, ultra-high performance liquid chromatography with linear ion trap-orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) was used to screen the chemical constituents of Rehmanniae Radix. The chemical constituents were identified based on the relative molecular weight and fragment ions, literature information, and Human Metabolome Database(HMDB). The ion peak area ratio of each component before and after processing was used as the index for the variation. SIMCA was employed to establish principal component analysis(PCA) and orthogonal partial least squares discriminant analysis(OPLS-DA) models of different processed products. According to the PCA plot, OPLS-DA plot, and VIP value, the differential components before and after the processing were screened out. The changes of the content of differential components with the processing method were analyzed. A total of 66 chemical components were identified: 57 of raw Rehmanniae Radix, 55 of steamed Rehmanniae Radix, 55 of wine-stewed Rehmanniae Radix, 51 of repeatedly steamed and sundried Rehmanniae Radix Praeparata, 62 of traditional bran-braised Rehmanniae Radix, and 63 of electric pot-braised Rehmanniae Radix. Among them, the 9 flavonoids of braised Rehmanniae Radix were from Citri Reticulatae Pericarpium. PCA suggested significant differences in the chemical composition of Rehmanniae Radix Praeparata prepared with different processing methods. OPLS-DA screened out 32 chemical components with VIP value >1 as the main differential components. Among the differential components, 9 were unique to braised Rehmanniae Radix(traditional bran-braised, electric pot-braised) and the degradation rate of the rest in braised(traditional bran-braised, electric pot-braised) or repeatedly steamed and sundried Rehmanniae Radix was higher than that in the steamed or wine-stewed products. The results indicated the chemical species and component content of Rehmanniae Radix changed significantly after the processing. The 32 components, such as rehmapicrogenin, martynoside, jionoside D, aeginetic acid, hesperidin, and naringin, were the most important compounds to distinguish different processed products of Rehmanniae Radix. The flavonoids introduced by Citri Reticulatae Pericarpium as excipient may be the important material basis for the effectiveness of braised Rehmanniae Radix compared with other processed products.

Component profile analysis of Sanhan Huashi Formula based on UHPLC-LTQ-Orbitrap-MS, GC-MS, and UHPLC-QQQ-MS/MS / 中国中药杂志

This study comprehensively analyzed the active components of Sanhan Huashi Formula using qualitative and quantitative mass spectrometry techniques, laying the foundation for understanding its pharmacological substance basis. UHPLC-LTQ-Orbitrap-MS and GC-MS technologies were used to analyze and identify the volatile and non-volatile components in Sanhan Huashi Formula. UHPLC-QQQ-MS/MS technology was used to simultaneously determine the content of 27 major active components in the formula. The results showed that 308 major chemical components were identified in Sanhan Huashi Formula, among which 60 compounds were identified by comparing with reference standards, mainly including alkaloids, flavonoids, coumarins, triterpenoid saponins, amino acids, and nucleosides. GC-MS technology preliminarily identified 52 volatile compounds, with γ-eudesmol and β-eudesmol as the main components. The quantitative results demonstrated good linearity(r>0.99) for the 27 active components, indicating the stability, simplicity, and reliability of the established method. Among them, amygdalin, nodakenin, arecoline, ephedrine, and pseudoephedrine had relatively high content and were presumably the main pharmacologically active substances. In conclusion, this study systematically and comprehensively characterized the major chemical components and patterns in Sanhan Huashi Formula, providing a basis for understanding its pharmacological mechanisms and clinical applications.

Differentiation of three Asparagus species by UHPLC-MS/MS based molecular networking identification and chemical profile analysis.

Asparagi Radix (AR), a traditional Chinese medicine, is the dried roots of Asparagus cochinchinensis (Lour.) Merr. Modern pharmacological studies have shown that AR has various excellent bioactivities, such as antioxidative, antitumor, antibacterial, anti-inflammatory, and hypoglycemic effects. However, the quality control method of AR is incomplete and there are various AR adulterants in markets due to their similar morphological characters. Here, holistic and practical quality evaluation methods were developed to chemically distinguish three common Asparagus species in markets, including Asparagus cochinchinensis (Lour.) Merr., Asparagus officinalis L., and Asparagus lycopodineus (Baker) F.T.Wang & Tang. The chemical constituents of three species were rapidly tentatively annotated using a combination of ultra-high pressure liquid chromatography-linear ion trap-orbitrap high resolution mass spectrometry (UHPLC-LTQ-Orbitrap-MS) and molecular networking (MN). Fifty-six steroidal saponins were annotated, including common and characteristic chemical constituents of the three Asparagus species. Besides, to establish holistic and practical methods to differentiate three Asparagus species, an HPLC-ELSD (evaporative light scattering detector) was applied for fingerprint analysis and content determination of the sum of protoneodioscin and protodioscin of twenty samples. Each Asparagus species showed characteristic chemical profile and AR showed much higher level of the sum of protoneodioscin and protodioscin than that in the others. The above analyses showed that the three Asparagus species mainly contain steroidal saponins and the developed HPLC-ELSD profile of saponin can be used to differentiate them. In conclusion, this study reveals the different chemical constituents of three Asparagus species and provides relatively feasible quality evaluation methods for them which are essential for the rational utilization of these Asparagus species.

Polyphenol Content of Green Pea (Pisum sativum L.) Hull under In Vitro Digestion and Effects of Digestive Products on Anti-Inflammatory Activity and Intestinal Barrier in the Caco-2/Raw264.7 Coculture Model.

Green pea hulls are a byproduct of the processing of green pea and are rich in phenolic substances. In the present study, in vitro digestion, human colonic adenocarcinoma cell line (Caco-2) monolayer, and the Caco-2/macrophage cell lines of the murine origin (Raw264.7) coculture model were established to investigate the release of polyphenols, absorption, and transport of digestive products and their effects on inflammation and intestinal barrier. During the digestive process, polyphenols were constantly released from the pea hulls, reaching the maximum amount in the small intestine (total phenolic content (TPC): 5.41 ± 0.04 mg gallic acid (GAE)/g dry weight (DW)), and the digestive products (800 µg/mL) could reduce the secretion of NO (50.9%), IL-6 (50.6%), and TNF-α (24.6%) and inhibit the mRNA expression of cyclooxygenase-2 (COX-2) (37.2%) and inducible nitric oxide synthase (iNOS) (91.1%) compared with the lipopolysaccharide (LPS) group. A total of 12 phenolic components were quantified by ultraperformance liquid chromatography-linear ion trap orbitrap tandem mass spectrometry (UHPLC-LTQ-OrbiTrap-MS) technology. Kaempferol trihexoside in digestive products could be absorbed and transported (1.25 ± 0.13 ng quercetin/mL). The digestive products could promote the expression of claudin-1 (210.8%), occludin (64.9%), and zonulin occludin-1 (ZO-1) (52.0%) compared with the LPS group and exert anti-inflammatory effects after being absorbed. The results indicated that pea hull polyphenols could be continuously released and absorbed to play a positive role in protecting the intestinal barrier and anti-inflammatory activity.

Six phenanthrenes from the roots of Cymbidium faberi Rolfe. and their biological activities.

A new phenanthrene compound, 7-(4-hydroxybenzyl)-8-methoxy-9,10- dihydrophenanthrene-2,5-diol (HMD), along with five known compounds (Coelonin, DD, Shancidin, HDP and MDD) were isolated from the roots of Cymbidium faberi Rolfe. (CFR). Their structures were identified using various spectroscopic methods. These compounds were reported for the first time in the genus. All isolated compounds were tested by radical-scavenging ability against 1,1-diphenyl-2-picryl-hydrazyl (DPPH), cytotoxic activity against three human cancer cell lines and inflammatory activity. Among them, Shancidin exhibited the stronger DPPH-scavenging activity (IC50=6.67 ± 0.84 µΜ) and cytotoxic activity against three tumour cell lines. Except for HDP, all compounds dose-dependently suppressed production of NO, TNF-α, IL-6 in LPS induced mouse primary peritoneal macrophage and showed anti-inflammatory activity. Moreover, 18 compounds were identified by UHPLC-LTQ-Orbitrap-MS combined with MS database, which provides a basis for further research.[Formula: see text].

The metabolite profiling of 3,4-dicaffeoylquinic acid in Sprague-Dawley rats using ultra-high performance liquid chromatography equipped with linear ion trap-Orbitrap MS.

3,4-Dicaffeoylquinic acid (3,4-DiCQA) is a dicaffeoylquinic acid that possesses antioxidant, anti-inflammatory, antibacterial, antiviral, anticancer, hypoglycemic, hypotensive, and hepatoprotective activities. This study developed a rapid and reliable method using ultra-high performance liquid chromatography equipped with linear ion trap-Orbitrap MS to identify the metabolites of 3,4-DiCQA in rat plasma, urine, feces, and tissues. The metabolic profile of 3,4-DiCQA was determined after an oral administration of 200 mg/kg to rats. A strategy of full scan-parent ions list acquisition coupled to diagnostic product ion analysis for screening and identification of target metabolites was used. A total of 67 metabolites, combined with accurate mass measurement, diagnostic ions, neutral losses, and reference standards, were observed and characterized for the first time. The results indicated that hydrolysis, methylation, hydrogenation, hydration, dehydroxylation, dehydrogenation, sulfate conjugation, and glucuronide conjugation were the major metabolic reactions of 3,4-DiCQA in vivo.

A triple combination strategy of UHPLC-MSn, hypolipidemic activity and transcriptome sequencing to unveil the hypolipidemic mechanism of Nelumbo nucifera alkaloids.

ETHNOPHARMACOLOGICAL RELEVANCE: Nelumbo nucifera (N. nucifera), a kind of edible Chinese herbal, has been studied in treating hyperlipidemia. However, the hypolipidemic mechanism of N. nucifera remains unknown. Aims of this review: We aimed to screen the effective constituent of N. nucifera alkaloids and elucidated the potential mechanism for treating hyperlipidemia. A triple combination strategy of UHPLC-MSn, hypolipidemic activity and transcriptome sequencing was built to unveil the hypolipidemic mechanism of Nelumbo nucifera alkaloid. MATERIALS AND METHODS: We comprehensively investigated the characterization of N. nucifera alkaloids by using UHPLC-LTQ-Orbitrap MSn. And the hypolipidemic activity of candidate active ingredients were evaluated on sodium oleate-induced HepG2 cell. Finally, O-nornuciferine and N. nucifera alkaloid extraction were analyzed by RNA sequence (RNA-seq) to decipher the underlying hypolipidemic mechanism and were verified by qRT-PCR. RESULTS: 35 compounds were identified from N. nucifera alkaloid extraction by UHPLC-LTQ-Orbitrap MSn. Among them, O-nornuciferine and N. nucifera alkaloid extraction which showed significant hypolipidemic activity were analyzed by transcriptome sequencing. After the intervention of O-nornuciferine and N. nucifera alkaloid extraction, 1 and 158 differentially expressed genes (DEGs) were identified, severally. The enrichment analysis indicated that the hypolipidemic effect was adjusted by the expression of numerous key DEGs involved in bile secretion, glycerolipid and sphingolipid metabolism, PPAR signaling pathway. CONCLUSIONS: O-nornuciferine and N. nucifera alkaloids had exibited significant effects in hyperlipidemia. The candidate genes were LDLR, LPL and ANGPTL4, etc. It was most likely that they adjusted lipid metabolism by modulating expression levels of various key factors which were involved in bile secretion, glycerolipid metabolism, sphingolipid metabolism and PPAR signaling pathway, and so on. This study clarified the hypolipidemic mechanism of the alkaloids in N. nucifera, and laid a foundation for the subsequent development of clinical application and better quality of N. nucifera.

Analysis of flavonoids and phenylethanoid glycosides in the Tibetan herb Lagotis brevituba Maxim based on UHPLC-LTQ-orbitrap-MS / 药学学报

Ultra high performance liquid chromatography tandem linear ion trap orbitrap mass spectrometry (UHPLC-LTQ-orbitrap-MS) was applied to analyze and identify flavonoids and phenylethanoid glycosides in the Tibetan herb Lagotis brevituba Maxim. A method of data-dependent scan coupling with dynamic exclusion was developed for analyzing flavonoids and phenylethanoid glycosides under positive and negative ion mode of electrospray ionization (ESI). The compounds of Lagotis brevituba Maxim. were systematically identified through exact molecular mass, fragmentation patterns, retention time and reported references. A total of 167 compounds were detected, of which 84 were flavonoids and 83 were phenylethanoid glycosides, which greatly enriched the number and types of flavonoids and phenylethanol glycosides in Lagotis genus medicinal plants. Baohuoside Ⅰ, 4 disaccharide O-glycoside flavonoids (composed of deoxyhexose and glucuronic acid), 9 C-glycoside flavonoids, 15 tetrasaccharide phenylethanoid glycosides and 5 phenylethanoid glycosides with substituents on the β-position of the phenylethyl group were identified in Lagotis genus medicinal plants for the first time. This study provides scientific support for elucidating the material basis and improving the quality control of Lagotis brevituba Maxim.

Green Pea (Pisum sativum L.) Hull Polyphenol Extracts Ameliorate DSS-Induced Colitis through Keap1/Nrf2 Pathway and Gut Microbiota Modulation.

As a processing by-product, green pea hull (GPH) was found to be rich in phenolic components in our previous studies. In this study, UHPLC-LTQ-OrbiTrap-MS (Ultra performance liquid chromatography-linear ion trap orbitrap tandem mass spectrometry) technique was used to quantify polyphenols, and DSS (sodium dextran sulfate)-induced colitis mouse model was established to explore the effect of GPH extracts on colitis. The results showed that quercetin and its derivatives, kaempferol trihexanside and catechin and its derivatives were the main phenolic substances in the extract, reaching 2836.57, 1482.00 and 1339.91 µg quercetin/g GPH extract, respectively; GPH extracts can improved inflammatory status, repaired colonic function, regulated inflammatory factors, and restored oxidative balance in mice. Further, GPH extracts can activate Keap1-Nrf2-ARE signaling pathway, regulate downstream antioxidant protease and gut microbiota by increasing F/B value and promoting the growth of Lactobacillaceae and Lachnospiraceae, and improve the level of SCFAs (short-chain fatty acids) to relieve DSS-induced colitis in mice. Therefore, GPH may be a promising dietary resource for the treatment of ulcerative colitis.