High-performance Liquid Chromatography-quadrupole Time-of-flight Mass Spectrometry Research Articles

Renal protective effect of Isaria felina mycelium powder on diet and STZ-induced diabetes mice and the identification of major chemical constituents

Type 2 diabetes is a group of metabolic diseases mainly characterized by chronic hyperglycemia. Diabetic kidney disease is the most common microvascular complication of diabetes.The aim of this study was to investigate the therapeutic effect of Isaria felina mycelium powder (IFMP) on diet and streptozotocin (STZ)-induced diabetic mouse model. And we analyze the main chemical constituents of Isaria felina mycelium powder.The main chemical constituents of the alcohol-soluble IFMP were prepared and identified by high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS). The fasting blood glucose and renal index of the mouse model was determined. Serum levels of triglyceride (TG), total cholesterol (TC), serum creatinine (Scr) and blood urea nitrogen (BUN) were detected. The levels of CD2AP and nephrin proteins in the kidney were detected by ELISA kits.A total of 32 compounds were identified, including nucleosides (16 types), amino acids (13 types), sterols (one type) and glycosides (two types). The experimental results showed that compared with the diabetes model group, IFMP reduced fasting blood glucose, food intake and water intake. Biochemical analysis indicated that IFMP reduced the serum levels of TG, TC, BUN and Scr, and increased the levels of CD2AP and nephrin proteins in the kidneys.Therefore, IFMP has the potential to treat diabetes and prevent kidney damage induced by hyperglycemia.

Discovery of potential inhibitors targeting SARS-CoV-2 Mpro.

The coronavirus disease (COVID-19) pandemic, resulting from human-to-human transmission of a novel severe acute respiratory syndrome coronavirus (SARS-CoV-2), has caused a global health emergency. The lack of a specific drug or treatment strategy against this disease makes it devastating. Given that the main protease (Mpro) of SARS-CoV-2 plays an indispensable role in viral polyprotein processing, its successful inhibition prevents viral replication and constrains virus spread. Therefore, developing an effective SARS-CoV-2 Mpro inhibitor to treat COVID-19 is imperative. We employed a high-throughput screening (HTS) method based on fluorescence polarization (FP) assay and further confirmed by the fluorescence resonance energy transfer (FRET) method for the discovery of Mpro inhibitors. Then multiple approaches were taken to investigate the inhibition profiles of the hit compounds against Mpro, including 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) proliferation assay, surface plasmon resonance analysis (SPR), high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry (HPLC-Q-TOF-MS), cytopathic effect (CPE) assay, molecule docking, and the drug-likeness analysis. In this study, four Mpro inhibitors with low toxicity were selected from HTS. According to SPR, all the hit compounds had medium binding affinities toward SARS-CoV-2 Mpro. HPLC-Q-TOF-MS results revealed the non-covalent linkage of each compound with SARS-CoV-2 Mpro. Molecule docking simulated the molecule interactions between each compound and the substrate binding pocket of SARS-CoV-2 Mpro. CPE assay was used to detect their inhibitory activities against coronaviruses HCoV-OC43 and HCoV-229E. In particular, the IMB63-8G compound demonstrated the highest antiviral potency [50% effective concentration (IC50) value of 1.71 μg/mL] and selectivity against HCoV-OC43 (SI = 39), which was more than 4-fold higher than that of ribavirin (RBV). Besides, the IMB63-8G compound possessed favorable drug-likeness characteristics. Our results will highlight the therapeutic potential of these compounds for the treatment of SARS-CoV-2 infection.

Comparative analysis of UV-initiated ARPs for degradation of the emerging substitute of perfluorinated compounds: Does defluorination mean the sole factor?

Due to the increasing attention for the residual of per- and polyfluorinated compounds in environmental water, Sodium p-Perfluorous Nonenoxybenzenesulfonate (OBS) have been considered as an alternative solution for perfluorooctane sulfonic acid (PFOS). However, recent detections of elevated OBS concentrations in oil fields and Frontal polymerization foams have raised environmental concerns leading to the decontamination exploration for this compound. In this study, three advanced reduction processes including UV-Sulfate (UV-SF), UV-Iodide (UV-KI) and UV-Nitrilotriacetic acid (UV-NTA) were selected to evaluate the removal for OBS. Results revealed that hydrated electrons (eaq−) dominated the degradation and defluorination of OBS. Remarkably, the UV-KI exhibited the highest removal rate (0.005 s−1) and defluorination efficiency (35 %) along with the highest concentration of eaq− (K = −4.651). Despite that nucleophilic attack from eaq− on sp2 carbon and H/F exchange were discovered as the general mechanism, high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC/Q-TOF-MS) analysis with density functional theory (DFT) calculations revealed the diversified products and routes. Intermediates with lowest fluorine content for UV-KI were identified, the presence nitrogen-containing intermediates were revealed in the UV-NTA. Notably, the nitrogen-containing intermediates displayed the enhanced toxicity, and the iodine poly-fluorinated intermediates could be a potential-threat compared to the superior defluorination performance for UV-KI.

Antibacterial activities of Miang extracts against selected pathogens and the potential of the tannin-free extracts in the growth inhibition of Streptococcus mutans.

Bacterial pathogens have remained a major public health concern for several decades. This study investigated the antibacterial activities of Miang extracts (at non-neutral and neutral pH) against Bacillus cereus TISTR 747, Escherichia coli ATCC 22595, Salmonella enterica serovar Typhimurium TISTR 292 and Streptococcus mutans DMST 18777. The potential of Polyvinylpolypyrrolidone (PVPP)-precipitated tannin-free Miang extracts in growth-inhibition of the cariogenic Streptococcus mutans DMST 18777 and its biofilms was also evaluated. The tannin-rich fermented extracts had the best bacterial growth inhibition against S. mutans DMST 18777 with an MIC of 0.29 and 0.72 mg/mL for nonfilamentous fungi (NFP) Miang and filamentous-fungi-processed (FFP) Miang respectively. This observed anti-streptococcal activity still remained after PVPP-mediated precipitation of bioactive tannins especially, in NFP and FFP Miang. Characterization of the PVPP-treated extracts using High performance liquid chromatography quadrupole-time of flight-mass spectrometry (HPLC-QToF-MS) analysis, also offered an insight into probable compound classes responsible for the activities. In addition, Crystal violet-staining also showed better IC50 values for NFP Miang (4.30 ± 0.66 mg/mL) and FFP Miang (12.73 ± 0.11 mg/mL) against S. mutans DMST 18777 biofilms in vitro. Homology modeling and molecular docking analysis using HPLC-MS identified ligands in tannin-free Miang supernatants, was performed against modelled S. mutans DMST 18777 sortase A enzyme. The in silico analysis suggested that the inhibition by NFP and FFP Miang might be attributed to the presence of ellagic acid, flavonoid aglycones, and glycosides. Thus, these Miang extracts could be optimized and explored as natural active pharmaceutical ingredients (NAPIs) for applications in oral hygienic products.

Comparative Study of Rice Wine-Processed and Unprocessed Angelica sinensis by High-Performance Liquid Chromatography–Mass Spectrometry (HPLC-MS) and Chemometrics

As a common pharmaceutical plant, Angelica sinensis (called Danggui (DG) in Chinese) is widely used in food, pharmaceuticals, and cosmetics and frequently processed with rice wine. Eight batches of rice wine-processed (WDG) samples and unprocessed DG were prepared to compare the chemical component differences. High-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS) was combined with a molecular network strategy. The 140 components were identified, thereby providing a comprehensive characterization of DG and WDG. Chemometrics was used to compare the variations in these components, and the results revealed higher abundances of certain amino acids and organic acids in WDG. These differences may explain the higher efficacy of WDG. This study shows the differences between DG and WDG included variation in the content, which provides a basis for the study of WDG and support for the standardization of Chinese medicine processing.

Investigating the mechanism and efficacy material basis of Xiehuo Xiaoying decoction for treating Graves' disease via thyroid cell apoptosis based on proteomics and molecular docking techniques

Ethnopharmacological relevanceFor numerous years, the Xiehuo Xiaoying decoction (XHXY), a traditional Chinese medicine formula, has demonstrated substantial promise in treating Graves' disease (GD) in clinical settings, showcasing significant potential. However, the therapeutic mechanism and efficacy material basis of XHXY remains obscure. Aim of the studyThis work aims to investigate the underlying mechanisms and to study the efficacy material basis of XHXY in anti-GD effect using a combination of TMT quantitative proteomics and molecular docking method. Materials and methodsGD model was initiated by administering Ad-TSH289. Subsequently, the mice underwent a four-week regimen that included oral gavage of XHXY at doses of 17 g/kg·d and 34 g/kg·d, along with intraperitoneal injections of Gentiopicroside (GPS). Utilizing the principles of pharmacological chemistry in traditional Chinese medicine, we employed high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (HPLC-QTOF/MS) to discern prescribed prototype composition of XHXY in serum samples from mouse. TMT proteomics research provided evidence of XHXY's putative targets and important pathways in vivo. The binding activity of probable action targets and prototype composition was detected by molecular docking. Finally, Immunohistochemistry (IHC) and TUNEL staining were used to verify the mechanism of XHXY and GPS in anti-GD. ResultsXHXY and GPS alleviated GD by ameliorating the pathological changes and reducing thyroxine and TRAb levels. In mouse serum, a total of 31 prototypical XHXY ingredients were detected, and the majority of these components were from monarch and minister medicine. Proteomics study results indicated that the XHXY may mainly regulate targets including FAS-associated death domain protein (FADD), Apolipoprotein C-III, etc. and main pathways are Apoptosis, Cholesterol metabolism, TNF signalling pathway, etc. Strong binding activity of the prototypical active ingredient and GPS towards FADD, Caspase 8, and Caspase 3 was demonstrated by molecular docking. XHXY and its primary component, GPS, elevated the expression of FADD, Caspase 8, and Caspase 3, and enhance apoptosis in thyroid cells, as lastly validated by TUNEL and IHC staining. ConclusionsXHXY exhibits a favorable therapeutic effect in treating GD by promoting apoptosis in thyroid cells through the upregulation of FADD, Caspase 8, and Caspase 3 expression. And GPS is the main efficacy material basis for its therapeutic effect in anti-GD.

Anticancer activity, phytochemical investigation and molecular docking insights of Citrullus colocynthis (L.) fruits

Cancer disease is regarded as one of the most significant public health issues, regardless of economic standards. Medicinal plants are now regarded as a natural source of anticancer medicines due to their antioxidant and anti-mutagenic actions. Cucurbitaceae is considered to be one of the most economically significant families. One family species is Citrullus colocynthis (L.), which has a high concentration of many active secondary chemical metabolites. Various C. colocynthis plant extracts showed cytotoxicity against some cancer cells. This study aims to identify the C. colocynthis fruit components and determine whether they have anticancer action against MIA PaCa-2 and A431 cells. High-Performance Liquid Chromatography/Quadrupole Time of Flight/Mass Spectrometry (HPLC/QTOF/MS); the technique was accustomed to investigate the compounds of the ethyl acetate (EtOAc) fruit extract. Anticancer activity was investigated on both MIAPaCa-2 and A-431 cell lines. DPPH assay for antioxidant activity was carried out. Molecular modelling was employed to help understand the molecular basis for the observed anticancer activity. 24 compounds were tentatively identified by comparing the extract’s fragmentation pattern in positive mode against reference compounds spectra and literature. The EtOAc extract of C. colocynthis had effective positive results on cancer cells (MIAPaCa-2 and A-431) and was characterized by slight or no harmful effect on normal (healthy) cells. For the DPPH assay, EtOAc and BuOH extracts exhibited high antioxidant activity (86 and 76%, respectively) compared with the oxidative potential of the standard compound (Caffeic acid, 98%). One of the major cucurbitacin derivatives that LC/MS tentatively identified in the EtOAc extract was Cucurbita-5(10),6,23-triene-3β,25-diol. During this study, docking experiments and MD simulations were carried out, which suggested the anti-pancreatic cancer activity of C. colocynthis extract to be attributed to EGFR inhibition by Cucurbita-5(10),6,23-triene-3β,25-diol. Therefore, expansion of this type of research should be encouraged in the hope of obtaining natural therapeutics for cancerous tumors in the future, having the advantage of being cheaper, safer, and with fewer side effects.

Determination of furosine in liquid milk by high performance liquid chromatography-quadrupole time-of-flight mass spectrometry

Furosine is often used both domestically and internationally as an indicator of the degree of heating to evaluate milk quality. However, in actual detection, the complexity of the milk matrix may lead to the inaccurate quantification of furosine in liquid milk. Therefore, in this study, an efficient and accurate method based on high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF/MS) was established to determine furosine in liquid milk. A 2.00 mL milk sample was hydrolyzed with 5 mL 12.00 mol/L hydrochloric acid solution and 1 mL water at 110 ℃ for 12 h. After hydrolysis, vortex-mixing and filtration were performed. The filtrate was diluted six times with 6.00 g/L ammonium acetate solution and then analyzed. Gradient elution was performed with 0.20% formic acid aqueous solution and acetonitrile solution as mobile phases, followed by chromatographic separation on an AQ-C18 column (150 mm×3.5 mm, 5 μm). The data were collected by Q-TOF/MS with an electrospray ionization source operated in positive-ion mode. The accuracy of the quantification of furosine in milk was assessed by investigating the effects of the hydrochloric acid concentration (0.30, 1.25, and 3.00 mol/L) in the furosine solution on the MS response. The results showed that high hydrochloric acid concentrations inhibited the response signals. A good linear relationship was obtained in the mass concentration range of 0.05-2.00 mg/L, with a correlation coefficient (r) of 0.994. The limit of detection of the method was 0.50 mg/100 g, which meets the requirements of actual sample detection. The average recoveries of furosine ranged from 79.9% to 119.7% at three spiked levels of 1.52, 3.03, and 15.17 mg/100 g, with relative standard deviations of 1.4%-2.6%. The method was applied to detect 303 samples from 101 batches of pasteurized milk sold in the market, and the contents of furosine in these samples ranged from 5.1 to 11.9 mg/100 g. The proposed method is characterized with high efficiency, recovery, sensitivity, and accuracy. Thus, it can be used for the determination of large quantities of samples and provides technical support for the continuous promotion of the high-quality development of the whole dairy industry chain.

Integrated strategy for the screening of cyclooxygenase-2 inhibitors from triterpenoid saponins in Clematis tangutica.

Screening of novel cyclooxygenase-2 (COX-2) inhibitors from complex natural products is not an easy task. To establish an efficient and feasible strategy for screening COX-2 inhibitors from triterpenoid saponins (TPSs) in Clematis tangutica. Taking TPSs in C.tangutica as example, an optimized macroporous resin (MR) method was established for the enrichment of TPSs. High-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-QTOFMS) was performed to establish the phytochemical profiling of TPSs. Molecular docking was performed to predict the ligand-target interactions and discover the active substances. Chemometric techniques were performed to visualize the structure-effect relationships. High-speed countercurrent chromatography and preparative HPLC were performed to prepare the targets. In vitro activity experiment of COX-2 was performed to verify the virtual screening results. TPSs in C.tangutica were well enriched with the recovery rate of (80.22 ± 2.37)%. Thirty-four kinds of TPSs of oleanane type were deduced by HPLC-QTOFMS. Five TPSs of clematangoside C, clematangoside D, clematangoticoside J, hederoside H1 , and hederasaponin B showed stronger binding abilities with COX-2. The structure with more sugar groups at C-28 may be more conducive to the combination with COX-2. Targets were prepared with purities all above 98%. The IC50 values of target TPSs were 6.03 ± 0.24, 12.44 ± 0.15, 9.36 ± 0.19, 4.78 ± 0.13, and 2.59 ± 0.11 μmol/L, respectively. The integrated strategy using MR, HPLC-QTOFMS, molecular docking, chemometrics, target preparation, and in vitro verification was feasible for rapidly screening COX-2 inhibitors from TPSs in C.tangutica.

Synthesis, Structure Confirmation of Deuterium-substituted Kynurenine and the Conformation Analysis of Kynurenine in Daptomycin

Background: After the hydrolysis of daptomycin in deuterated hydrochloric acid, the deuterium-substituted kynurenine was found, but the structure of deuterium-substituted kynurenine has not been reported. Introduction: The deuterium-substituted kynurenines were simply synthesized and confirmed to be tri- and tetra-substituted products by high resolution mass spectrum and NMR. In further, the deuterium-substituted kynurenines were used to determine the conformation of kynurenine to be L-type in daptomycin through conformation analysis combined with derivation and high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLCQ/TOF-MS). Methods: In the present study, a simple synthesis method was developed for deuteriumsubstituted kynurenine, and its structure was confirmed by high resolution mass spectrometry and NMR. L-kynurenine was mixed with the deuterated hydrochloric acid and heated at 110 o C for 7 h. The hydrogen/deuterium exchange products of L-kynurenine were obtained through the hydrogen/deuterium exchange method. After the derivation of deuterium-substituted L-kynurenine by Marfey’s reagent, the conformation of kynurenine in daptomycin was deduced by HPLC-Q/TOFMS. Results: The deuterium-substituted kynurenines were confirmed to be tri- and tetra-substituted products by high resolution mass spectrum. Further, Hydrogen NMR spectrum indicated that the deuterium-substitution positions were β-position on amino acid and 3’ and 5’ positions on the benzene ring. Thus, the tri-deuterium-substituted product was L-[β, 3’, 5’- 2 H3] kynurenine-d3, while the tetra-deuterium-substituted product was L-[β, β, 3’, 5’- 2 H4] kynurenine-d4. Furthermore, the deuterium-substituted kynurenines were used to determine the conformation of kynurenine to be L-type in daptomycin through conformation analysis combined with derivation and HPLC-Q/TOF-MS. Conclusion: The synthesis, structures, and application of tri- or tetra- deuterium-substituted kynurenine were reported in this study.

Comparative analysis of lipid components in fresh Crassostrea Hongkongensis (raw) and its dried products by using high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (HPLC/Q-TOF-MS).

The lipids of the oyster (Crassostrea hongkongensis) have a special physiological activity function, which is essential to maintain human health. However, comprehensive research on their lipids species and metabolism is not so common. In our study, based on the high-performance liquid chromatography/quadrupole time-of-flight mass spectrometer (HPLC/Q-TOF-MS), the non-targeted lipidomics research of Crassostrea hongkongensis fresh and dried products was determined. Meanwhile, we analyzed its lipid outline, screened the differences between the lipid molecules of Crassostrea hongkongensis fresh and dried products, and determined the lipid metabolic pathway. Results showed that 1,523 lipid molecules were detected, in which polyunsaturated fatty acids mostly existed in such lipids as phosphoglyceride. Through the multivariate statistical analysis, according to the conditions of P < 0.05, FC > 2 or FC < 0.05, and VIP > 1.2, 239 different lipid molecules were selected, including 37 fatty acids (FA), 60 glycerol phospholipids (GP), 20 glycerin (GL), 38 sheath lipids (SP), 31 steroid lipids (ST), 36 polyethylene (PK), and 17 progesterone lipids (PR). Combined with the Kyoto Encyclopedia of Genes and Genomes (KEGG), the differential lipid molecules were analyzed to mainly determine the role of the glycerin phospholipid metabolic pathway. As a whole, the results of this study provide the theoretical basis for the high-value utilization of oysters and are helpful to the development of oysters' physiological activity functions and deep utilization.

Improvement of antioxidant activity and active ingredient of Dendrobium officinale via microbial fermentation.

This study used brewer's yeast to ferment Dendrobium officinale and single-factor and orthogonal experiments were conducted to determine the optimal fermentation conditions. The antioxidant capacity of Dendrobium fermentation solution was also investigated by in vitro experiments, which showed that different concentrations of fermentation solution could effectively enhance the total antioxidant capacity of cells. The fermentation liquid was found to contain seven sugar compounds including glucose, galactose, rhamnose, arabinose, and xylose using gas chromatography-mass spectrometry (GC-MS) and high performance liquid chromatography-quadrupole-time of flight mass spectrometry (HPLC-Q-TOF-MS), with the highest concentrations of glucose and galactose at 194.628 and 103.899 μg/ml, respectively. The external fermentation liquid also contained six flavonoids with apigenin glycosides as the main structure and four phenolic acids including gallic acid, protocatechuic acid, catechol, and sessile pentosidine B.

Aspartame-Sweetened Tap Water: Transformation Products and 2,6-Dichloro-1,4-Benzoquinone Formation.

Aspartame (APM), a dipeptide of aspartic acid (ASP) and phenylalanine (PHE), is a widely used artificial sweetener in beverages. It is unclear whether residual chlorine in tap water can react with APM to form disinfection byproducts (DBPs). Therefore, we investigated the formation of DBPs from the reaction of APM with residual chlorine in authentic tap water. APM and a commercial sweetener (CS) packet containing APM were studied under authentic and simulated tap water conditions. Eight chlorinated products of APM were detected using solid-phase extraction (SPE) and high performance liquid chromatography quadrupole time-of-flight mass spectrometry (HPLC-QTOF-MS). These new chloro-products were tentatively identified based on accurate masses, isotopic patterns of 35,37Cl, and MS/MS spectra. Furthermore, we identified APM as a precursor to 2,6-dichloro-1,4-benzoquinone (DCBQ). DCBQ significantly increased to 2.3-12 ng/L with the addition of APM or CS in tap waters collected from different locations compared to 1.4-1.8 ng/L in the same tap water samples without sweetener. DCBQ and two of the chlorinated transformation products were identified in cold prepared tea containing APM. DCBQ formation was eliminated when the residual chlorine in tap water was reduced by ascorbic acid or boiling prior to the addition of APM or CS. This study found that eight new DBPs and DCBQ were produced by the reactions of residual chlorine with APM and CS. These findings show an unintended exposure source of emerging DBPs via APM sweetened beverages.

Quantitative Bottom-Up Glycomic Analysis of Polysaccharides in Food Matrices Using Liquid Chromatography-Tandem Mass Spectrometry.

Carbohydrates are the most abundant biomolecules in nature, and specifically, polysaccharides are present in almost all plants and fungi. Due to their compositional diversity, polysaccharide analysis remains challenging. Compared to other biomolecules, high-throughput analysis for carbohydrates has yet to be developed. To address this gap in analytical science, we have developed a multiplexed, high-throughput, and quantitative approach for polysaccharide analysis in foods. Specifically, polysaccharides were depolymerized using a nonenzymatic chemical digestion process followed by oligosaccharide fingerprinting using high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-QTOF-MS). Both label-free relative quantitation and absolute quantitation were done based on the abundances of oligosaccharides produced. Method validation included evaluating recovery for a range of polysaccharide standards and a breakfast cereal standard reference material. Nine polysaccharides (starch, cellulose, β-glucan, mannan, galactan, arabinan, xylan, xyloglucan, chitin) were successfully quantitated with sufficient accuracy (5-25% bias) and high reproducibility (2-15% CV). Additionally, the method was used to identify and quantitate polysaccharides from a diverse sample set of food samples. Absolute concentrations of nine polysaccharides from apples and onions were obtained using an external calibration curve, where varietal differences were observed in some of the samples. The methodology developed in this study will provide complementary polysaccharide-level information to deepen our understanding of the interactions of dietary polysaccharides, gut microbial community, and human health.

Identification of Novel Parishin Compounds from the Twig of Maclura tricuspidata and Comparative Analysis of Parishin Derivatives in Different Parts

Parishin compounds are rare polyphenolic glucosides mainly found in the rhizome of the traditional Chinese medicinal plant, Gastrodia elata. These constituents are reported to have several biological and pharmacological activities. In the present study, two novel parishin derivatives not previously reported as plant-based phytochemicals were identified from a twig of Maclura tricuspidata (MT) and two new compounds were elucidated as 1-(4-(β-d-glucopyranosyloxy)benzyl)-3-hydroxy-3-methylpentane-1,5-dioate (named macluraparishin E) and 1,3-bis(4-(β-d-glucopyranosyloxy)benzyl)-3-hydroxy-3-methylpentane- 1,5-dioate (macluraparishin C), based on the experimental data obtained by UV-Visible (UV-Vis) spectroscopy, high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-QTOF-MS) and nuclear magnetic resonance (NMR) spectroscopy. Additionally, gastrodin, parishin A and parishin B were positively identified by spectroscopic evidence and the comparison of HPLC retention time with the corresponding authentic standards. Gastrodin, parishin A and parishin B, macluraparishin E and macluraparishin C were found to be the most abundant constituents in the MT twig. The compositions and contents of these constituents were found to vary depending on the different parts of the MT plant. In particular, the contents of parishin A, parishin B, macluraparishin C and macluraparishin E were higher in the twig, bark and root than in the leaves, xylem and fruit.

An efficient mixed enzymes-assisted mechanical bio-extraction of polysaccharides from Dendrobium officinale and determination of monosaccharides by HPLC-Q-TOF/MS

The mixed enzymes-assisted mechanical bio-extraction method was first used to extract polysaccharides from Dendrobium officinale. Different parameters including the ratio of enzyme, the amount of enzyme, the grinding time, the extraction time and the solid/liquid ratio were investigated by single factor experiments and multifactorial experiments. Through the response surface methodology the optimal extraction conditions were obtained with the ratio of cellulase to pectinase was 2: 1 and total amount of enzyme was 0.23 mg, the grinding time of 11.48 min, the extraction time of 5.99 min. The obtained polysaccharide extracts were hydrolyzed and derivatized and then injected into high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF/MS) for monosaccharide composition analysis. After optimization of the chromatographic conditions (including mobile phase and column), twelve monosaccharides were successfully determined within 20 min. The proposed method provided satisfactory linearity with the correlation coefficients higher than 0.99, suitable recoveries (81.46–114.92 %), acceptable reproducibility ranging from 0.06 % to 4.77 %, low limits of detection (0.70–45.45 ng/mL). Compared with other methods, this method makes the extraction efficiency much higher and has the advantages of simple operation, environmental friendliness and mild extraction conditions. Therefore, this method can be used for the extraction of polysaccharides from plants and the determination of monosaccharides and has the potential to be used in more areas.

Structure Analysis of Unsaturated Polymyxin E Components Based on High-Performance Liquid Chromatography - Quadrupole/ Time of Flight Tandem Mass Spectrometry and Photochemical Reaction

Background: Polymyxin E (PME), which is a complex of cationic cyclic lipodecapeptides, is used to treat multidrug-resistant gram-negative bacterial infections. Besides the main components PME1 and PME2, polymyxin containing unsaturated fatty acyl (FA) group with lower contents is hardly to determine the structure without chromatographic preparations and NMR. Introduction: The peptide sequences of PME components has been carried out based on high performance liquid chromatography-quadrupole / time-of-flight mass spectrometry (HPLC-Q/TOF-MS). However, the components with double bond on the FA, such as 2’, 3’-dehydro PME1, were difficult to be determined or easily misjudged by MS/MS. The transformation of such unsaturated components to be epoxidized components or di-hydroxylated components can promote the acquisition of more fragment ions in the MS/MS, so as to assist in judging the position of double bonds on FA. Methods: In this paper, the PME mixtures were dissolved in an equal proportion of 20% ACN aqueous solution and 2-acetylpyridine. The above PME solution was transferred to a quartz cuvette and irradiated with the ultraviolet lamp at 254 nm for 8h. The dehydro PME components were converted to be epoxy PMEs and dihydroxy PMEs. A fragmentation pathway of epoxidized components or di-hydroxylated components based on Q/TOF-MS/MS was proposed for the first time. Results: According to the characteristic ions of epoxidized components and di-hydroxylated components, 2’, 3’-epoxy PME1/E2 and 2’, 3’-dihydroxy PME1/E2 were confirmed. It can be inferred that the double bond is located at the 2’, 3’-position of FA. Conclusion: The structure of unsaturated PME component with double bond on the FA is elucidated by HPLC-Q/TOF-MS combined with photochemical reaction. This strategy is applicable to other lipopeptides containing unsaturated FA chain.

Identification of anti-inflammatory components in Dioscorea nipponica Makino based on HPLC-MS/MS, quantitative analysis of multiple components by single marker and chemometric methods

The rhizome of Dioscorea nipponica Makino (RDN) is a widely used herbal medicine, which has significant anti-inflammatory activities on various inflammatory diseases. However, the bioactive compositions responsible for the anti-inflammatory activity of RDN are still unknown. This study aimed to identify the anti-inflammatory bioactive compounds in RDN using high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-Q/TOF-MS), quantitative analysis of multiple components by single marker (QAMS) and chemometric methods. Firstly, an HPLC-Q/TOF-MS method was employed for identification of bioactive steroidal saponins in RND, and a total of twelve steroid saponins were identified. Then, QAMS method was employed to determine the contents of seven bioactive steroidal saponins, including protodioscin, protogracillin, methyl protodioscin, pseudoprotodioscin, pseudoprogracillin, dioscin and gracillin in RND samples using dioscin as the reference analyte. The anti-inflammatory effects of RDN samples were then evaluated by inhibition of NO production in LPS-induced RAW264.7 cells. Furthermore, chemometric methods, including Pearson correlation analysis and partial least squares regression (PLSR) were employed to investigate the correlations between chemical components and anti-inflammatory activities, and explore the potential anti-inflammatory bioactive compounds of RDN. The results indicated that protodioscin, dioscin and gracillin were selected as the major anti-inflammatory compounds in RND. The further verification experiments showed that protodioscin, dioscin and gracillin exhibited great inhibition on NO production with IC50 values (the half maximal inhibitory concentration) of 0.712 μM, 0.469 μM and 0.815 μM, respectively. They also significantly reduced the levels of TNF-α, IL-1β, and IL-6 in LPS-induced RAW264.7 cells. The present study provided evidences for the anti-inflammatory activity of RND and identification of the anti-inflammatory components in RDN.

A case of carboxyatractyloside intoxication by ingestion of the cocklebur (Xanthium strumarium) in a Korean native cow

A 19-month-old Korean native cow died the following day after consuming new silage. Grossly, the liver showed enlargement, redness, and haemorrhages in all the lobes. Additionally, many of the bur-shaped fruits of the cocklebur (Xanthium strumarium) were found in the gastric contents. The histological evaluation confirmed centrilobular hepatic necrosis. Additionally, carboxyatractyloside (CATR), a material fatal to animals found in the cocklebur, was detected in the gastric contents using high-performance liquid chromatography-quadrupole-time of flight mass spectrometry (HPLC-Q-TOF-MS). Based on the pathological findings and analytical confirmation, CATR intoxication was diagnosed. Therefore, careful feeding and elimination of the cocklebur is essential for minimising economic loss.

Antidepressant-like activity, active components and related mechanism of Hemerocallis citrina Baroni extracts.

Hemerocallis citrina Baroni [Asphodelaceae], which is traditional herbal medicine, has been widely used for treating depressive disorders in Eastern-Asia countries. However, the active compounds and corresponding mechanism of anti-depression are not yet completely clarified. In this study, the anti-depressive activities of six H. citrina extracts were primarily evaluated. The results showed that the water extract of H. citrina flowers (HCW) displays significant anti-depressive activity. A total of 32 metabolites were identified from HCW by high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS) and nuclear magnetic resonance (NMR). And then, the anti-depressive activity of the high-level compound (rutin) in HCW was also estimated. The results indicated that rutin displayed significant anti-depressive activity and was one of the main active ingredients. Finally, the anti-depressive mechanisms of HCW and rutin were investigated based on the intestinal microorganisms. The results showed that HCW and rutin increase the diversity and richness of the intestinal flora and regulate the specific intestinal microorganisms such as Bacteroides and Desulfovibrio genera in depressed mice. This work marks the first comprehensive study of the active components, anti-depressive activities and corresponding mechanisms of different H. citrina extracts, which provide a potential possibility for developing new antidepressants.