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Discovery of drugs rapidly and effectively is an important aspect for Alzheimer's disease (AD). In this study, a novel high-throughput screening (HTS) method aims at screening the small-molecules with amyloid-β (Aβ) binding affinity from natural medicines, based on the combinational use of biolayer interferometry (BLI) and ultra-high-performance liquid chromatography coupled with diode-array detector and quadrupole/time-of-flight tandem mass spectrometry (UHPLC−DAD-Q/TOF-MS/MS) has been firstly developed. Briefly, the components in natural medicines disassociated from biotinylated Aβ were collected to analyze their potential Aβ binding affinity by UHPLC−DAD-Q/TOF-MS/MS. Here, baicalein was confirmed to exhibit the highest binding affinity with Aβ in Scutellaria baicalensis. Moreover, polyporenic acid C (PPAC), dehydrotumulosic acid (DTA), and tumulosic acid (TA) in Kai-Xin-San (KXS) were also identified as potent Aβ inhibitors. Further bioactivity validations indicated that these compounds could inhibit Aβ fibrillation, improve the viability in Aβ-induced PC-12 cells, and decrease the Aβ content and improve the behavioral ability in Caenorhabditis elegans. The molecular docking results confirmed that PPAC, DTA, and TA possessed good binding properties with Aβ. Collectively, the present study has provided a novel and effective HTS method for the identification of natural inhibitors on Aβ fibrillation, which may accelerate the process on anti-AD drugs discovery and development.

In this study, a fully automated two-dimensional liquid chromatography (2D-LC) system was used for the investigation of the clean-up effect and was compared with a traditional Quick Easy Cheap Effective Rugged and Safe (QuEChERS) liquid chromatography (LC) method. The focus of those investigations was on negative electrospray ionization (ESI) mode. For that purpose, matrix fingerprinting profiles were created. The results allowed a comparison of both methods regarding the estimation of the number and the polarity of detected compounds. Moreover, the results of the present study were compared with the results generated in positive ESI mode (presented in a previous study). Furthermore, the two methods were compared with regard to matrix effects (ME) of 321 analytes in positive ESI mode and 96 analytes in negative ESI mode. In general, fewer compounds could be detected when 2D-LC and/or the negative ESI mode was used. Especially, very polar compounds with m/z values >1000 could be separated and could not be detected anymore when 2D-LC was applied. Furthermore, the best results were obtained for most analytes when 2D-LC was used, although the extent of ME seemed to be higher with 2D-LC.

Background: Impurities in pharmaceutical compounds can influence their clinical effects and represent a potential health risk. To ensure the safety and effectiveness of a drug, it is necessary to investigate potential impurities. Methods: In this paper, a new impurity was separated and characterized by two-dimensional high performance liquid chromatography coupled to quadrupole time-of-flight tandem mass spectrometry (2D HPLC-Q/TOF-MS) in negative electrospray ionization mode. The peak containing the new impurity, eluted from the first dimension chromatographic system, was selectively trapped by a switching valve based on its retention time and transferred to the second dimension chromatographic system, which was connected to the mass spectrometer. We obtained MET-TA by chemical synthesis, and its structure was characterized by MS/MS and further confirmed by nuclear magnetic resonance (NMR). Results: The impurity was found to be (2S, 3S)-2,3.-dihydroxy-4-((1R,2S)-1-hydroxy-1-(3-hydroxyphenyl) propan-2-yl)amino)-4-oxobutanoic acid, labelled as MET-TA. In this study, we investigated the mechanism of formation of MET-TA, and found that it was the amidation product of metaraminol and tartaric acid. Conclusions: The identification, structural elucidation, synthesis and most probable mechanism of formation of MET-TA are discussed in detail in this paper.

Design and construction of a matrix with specific adsorption on the target compounds can effectively reduce the detection limit of surface-assisted laser desorption/ionization time-of-flight mass spectrometry (SALDI-TOF MS) analysis. Sulfonic acid functionalized hierarchical porous covalent organic frameworks (H-COF-SO3H) was synthesized by defect-structure and post-modification method, and then used as matrix and adsorbent for the determination of quaternary ammonium herbicides paraquat (PQ) and diquat (DQ). N2 adsorption–desorption experiments confirmed that H-COF-SO3H possesses hierarchical porosity with pore widths concentrated at 1.3,1.5, and 2.8 nm. The strong UV absorption at 200–450 nm and good thermal stability made H-COF-SO3H being a promising matrix without background interference. H-COF-SO3H can efficiently enrich PQ and DQ via electrostatic attraction, and the key role of -SO3H group on specific adsorption was confirmed by density functional theory (DFT) calculations. The limits of detection (LODs) for PQ and DQ with H-COF-SO3H enrichment were 0.5 and 0.1 ng·mL−1, respectively, which were 20 and 60 times higher than those without H-COF-SO3H enrichment, respectively. The spiked recoveries of PQ and DQ for the three food samples were 92.0–113.2% and 80.1–102.6% with RSDs of 2.2–9.2% and 2.0–8.7%, respectively. This work provides an analyte-oriented approach for fabricating SALDI-TOF MS matrix.

The aroma characteristics of Pu-erh tea were studied in this work. A total of 117 volatile compounds were identified by HS-SPME combined with GC–TOF/MS, of which 29 active aroma compounds were identified by olfactometry. The active aroma compounds of Pu-erh tea were analyzed by intensity aroma (IA), aroma extraction dilution (AEDA), sensory quantification value (MF), detection frequency (DFA) and odor activity value (OAV). 24, 21 and 23 active aroma components were identified by DFA, AEDA and OAV, while 20 by IA and MF. Recombination of the obtained active aroma components indicated that OAV method was best for determining the active aroma compounds as the aroma profile of OAV recombination model was the most similar to that of Pu-erh tea sample. Omission test of the OAV recombination model furtherly identified 19 characteristic active aroma compounds of Pu-erh tea, and linalool, 1,2,3-trimenthoxybenzebe, 1,2,4-trimethoxybenzene and ɑ-ionone were the key characteristic active aromatic compounds.

Association reactions by femtosecond laser filamentation in gaseous C2H4 were studied by time-of-flight mass spectrometry of neutral reaction products. Direct sampling from the reaction cell to a mass spectrometer via a differential pumping stage allowed the identification of various hydrocarbon molecules C n H m with n = 3-7 and m = 4-7, which includes species not observed in the previous studies. It was found that products containing three and four carbon atoms dominate the mass spectrum with smaller yields for higher-mass species, suggesting that carbon chain growth proceeds through the reaction with C2H4 in the reaction cell. The product distribution showed a clear dependence on the laser pulse energy for filamentation.

Background Red ginseng polysaccharides (RGPs) have been acknowledged for their outstanding immunomodulation and anti-tumor activities. However, their studies are still limited by the complexity of their structural features, the absence of purification and enrichment methods, and the rarity of the analytical instruments that apply to the analysis of such macromolecules. Thus, this study is an attempt to establish a new mass spectrometry (MS)-based analysis procedure for RGPs. Methods Saponin pre-excluded powder of RG (RG-SPEP, 10 mg) was treated with 200 μL of distilled water and centrifuged for 5 h at 1000 rpm and 85 °C. Ethanol-based precipitation and centrifugation were applied to obtain RGPs from the heated extracts. Further, endo-carbohydrase treatments were performed to produce specific saccharide fragments. Solid-phase extraction (SPE) processes were implemented to purify and enrich the enzyme-treated RGPs, while matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF) MS was employed for the partial structural analysis of the obtained RGPs. Results Utilizing cellulase, porous graphitized carbon (PGC), hydrophilic interaction chromatography (HILIC), and MALDI-TOF/TOF MS, the neutral and acidic RGPs were qualitatively analyzed. Hexn and Hexn−18 (cellulose analogs) were determined to be novel neutral RGPs. Additionally, the [Unknown + Hexn] species were also determined as new acidic RGPs. Furthermore, HexAn (H) was determined as another form of the acidic RGPs. Conclusion Compared to the previous methods of analysis, these unprecedented applications of HILIC-SPE and MALDI-TOF/TOF MS to analyze RGPs proved to be fairly effective for fractionating and detecting neutral and acidic components. This new procedure exhibits great potential as a specific tool for searching and determining various polysaccharides in many herbal medicines., Graphical abstract Image 1

A novel stability-indicating HPLC method for the quantitative determination of impurities in meropenem raw material was established and fully validated for the further improvement of the official monograph in Pharmacopeia. The method was carried out on a Shim-Pack CLC-ODS column with UV detection at 220 nm. The influences of different band columns, composition ratio of mobile phase, gradient elution procedure, and pH value of mobile phase on separation of impurities in meropenem were investigated. The parameters specificity, precision, linearity, range, accuracy, limit of detection, limit of quantitation, and robustness were studied according to the regulatory guidelines recommended by the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH). The linear regression analysis results of EP impurity A and EP impurity B show that there is a good linear relationship between response and concentration, and the correlation coefficients are 0.9999 and 1.0000, respectively. The accuracy of EP impurity A and EP impurity B is between 90 and 104%. Through analysis, the LOD and LOQ values of EP impurities A and B were measured and found to be 0.22 and 0.74 µg mL-1, respectively. Moreover, three unknown degradation impurities in meropenem were identified by two-dimensional liquid chromatography tandem with high-resolution ion trap/time-of-flight mass spectrometry in electrospray ionization mode. Compared with the current method in Pharmacopeia, the developed method had greatly improved the separation and detection ability of impurities. The established method was highly specific, sensitive, and accurate for reliable routine quality control of meropenem raw material. The structures of unknown impurities were characterized based on the MS/MS data.

Background: Indobufen is a drug that hinders the aggregation of platelets by reversibly repressing the cyclooxygenase enzyme, further bringing about diminished thromboxane production. During quality control of indobufen tablets, an unknown impurity was detected. Objective: To characterize an unknown impurity in indobufen tablets. Methods: A new method compatible with mass spectrometry detection was set up. A C18 column at 35 °C with a mobile phase consisting of aqueous buffer (including ammonium formate) and methanol (35: 65, v/v) was used at a flow rate of 1.0 mL/min at 228 nm. High-performance liquid chromatography quadrupole time-of-flight mass spectrometry mass spectrometry (HPLC-Q-TOF MS) was used to identify the impurity with the electrospray ionization (ESI) source in the positive ionization mode. Results: The results of HPLC-Q-TOF MS analysis indicated that the protonated molecule ions [M + H]+ of the unknown impurity was at m/z 312. Preparative LC method was put into practice with a Prep- C18 column with a mobile phase consisting of water and methanol (20: 80, v/v) at a flow rate of 20.0 mL/min at 228 nm. The assignment of the 1D and 2D NMR signals was performed for the unknown impurity. In addition, possible formation of the novel impurity was also studied. Conclusion: An unknown impurity in indobufen tablets was characterized. The impurity was assigned as 2-(4-(1-hydroxy-3-oxoisoindolin-2-yl) phenyl) butanoic acid.

Lipophilic marine biotoxins azaspiracids (AZAs) are produced by dinoflagellates Azadinium and Amphidoma. Recently, several strains of Azadinium poporum were isolated from Japanese coastal waters. In our present study, AZA analogues in two strains (mdd421 and HM536) of A. poporum were analyzed by several detection techniques on the liquid chromatography-tandem mass spectrometry (LC-MS/MS) and liquid chromatography-quadrupole time of flight mass spectrometry (LC-QTOFMS). The dominant AZA analogue in the Japanese A. poporum strains was AZA2. Other known AZA analogues were AZA11, AZA35, AZA2 methyl ester and AZA2 phosphate ester. Besides these AZAs, thirteen new AZA analogues were discovered in the two strains. A putative AZA analogue (Compound 1) with the smallest molecular weight ever found in nature was also discovered in the two strains. This is the first report describing detailed AZA profiles in Japanese isolates of A. poporum.

Mutan is an extracellular sticky polymer having α-1-3 and α-1-6 glycosidic linkages with a large diversity in molecular weights and structures depending on the source. These compounds are reported to be highly thermostable and also have potential physiochemical and biological applications. The main aim of this review is to provide an overview of glucosyltransferases and their role in mutan synthesis. The production strategies and structural properties of bacterial mutans are discussed with a goal to improve production efficiency. The physicochemical features, chemical modifications, potential industrial applications and future prospects are also discussed. According to data, mutan and its derivatives will play a larger role in medicinal sectors and as thermoplastics in the near future.Abbreviations: ABTS: 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid; BHI: Brain heart infusion broth; 13C (HSQC) NMR: Heteronuclear Single Quantum Coherence NMR; CBMs: Carbohydrate binding modules; DPPH: 2,2-diphenyl-1-picrylhydrazyl; FTIR: Fourier-transform infrared spectroscopy; GC-MS: Gas chromatography-mass spectrometry; GPC: Gel permeation chromatography; Gtfs: Glucosyltransferases; 1H (DQF-COSY): Double-quantum filtered correlation spectroscopy; HPAEC-PAD: High-performance anion exchange chromatography with pulsed amperometric detection; HPLC: High performance liquid chromatography; HPSEC-RI: High-performance size exclusive chromatography coupled with refractive index; HPSEC-MALLS: High-performance size exclusive chromatography with multi-angle laser light scattering detection; MALDI-TOF: Matrix-Assisted Laser Desorption/Ionization-Time of Flight mass spectrometry; Mw: Weight-average molecular weight; MWD: Molecular weight distribution; NMR: Nuclear magnetic resonance spectroscopy; TEM: Transmission electron microscopy; THB: Todd Hewitt Broth; TTY: Tryticase tryptose yeast extract broth.

Honey varies depending on plant origin and environmental conditions. Chemical constituents can be used to determine the origins, characteristics, and storage period of a specific honey. Styrax japonica (snowbell) honey has excellent flavor, but very few studies have been carried out on its chemical constituents. Therefore, the present study was aimed to identify chemical constituents containing in snowbell honey and to screen candidate marker compounds for predicting the storage period of snowbell honey using the identified compounds. Twelve compounds including three new compounds were purified and isolated from the snowbell honey. Three new compounds were elucidated as 1α,3β,5β-bornetriol, 7-hydroxy-2,2,6-trimethyl-1-oxaspiro[2.5]oct-5-en-4-one, and 2-benzyl-2,3-dihydroxybutanoic acid, based on ESI-MS and NMR experiments. 6,7-Dihydroxylinalool and 6,7-dihydroxygeraniol isolated as known compounds in this study were identified for the first time in snowbell honey as well as in other honeys. The isolated compounds were qualified and quantified by LC-ESI-Q-TOF-MS/MS analysis. All compounds were detected in snowbell honeys of different production years. The levels of tyrosol, 7-hydroxy-2,2,6-trimethyl-1-oxaspiro[2.5]oct-5-en-4-one, and (–)-dihydroxyphaseic acid were lower in snowbell honey with long storage period than in short storage period, which is suggested to be used as candidate marker compounds for predicting the storage period of snowbell honey.

The aggregation process of α-synuclein (α-syn) is substantial in the pathogenesis of Parkinson's disease. Indolinone derivatives are inhibitors of α-syn aggregates and can be used as PET-based radiotracers for imaging α-syn fibrils. However, no investigations on the metabolism of indolinone derivatives have been reported until now. In the present research, a 13C and 15N isotope labeling strategy was developed to synthesize compound [13C2,15N]-(Z)-1-(4-aminobenzyl)-3-((E)-(3-phenyl)allylidene)indolin-2-one (M0'), which was then used in a study of metabolism in hepatocytes. The metabolites were characterized using accurate mass and characteristic ion measurements. In the metabolic system, compound M0' was the main component (accounting for 97.5% of compound-related components) after incubation in hepatocytes for 3 h, which indicated that compound M0' possessed great metabolic stability. Seven metabolites have been successfully verified by UPLC/Q TOF MS in metabolic studies, including hydroxyl M0' (M1'), hydroxyl and methylated M0' (M2'), N-acetylated M0' (M3'), sulfate of hydroxyl M0' (M4'), the glucose conjugate of M0' (M5'), glucuronide conjugate of M0' (M6'), and glucuronide conjugate of hydroxyl M0' (M7'). The study on metabolism provides the important information to develop effective α-syn aggregate inhibitors and new PET-tracer-related indolinone derivatives.

20-Hydroxyecdysone (20E) derived from plants has a wide range of physiological and pharmacological effects on animals and humans, and rapid and sensitive methods for screening of the endogenous 20E in plants are thus required. Herein, a matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry (MALDI-TOF/TOF MS) method is described for rapid and sensitive determination of endogenous 20E in plants. It is based on the use of the (3-(acridin-9-ylamino) phenyl) boronic acid (AYPBA) as the mass tag to assist the MS and tandem MS (MSn) analysis of 20E on MALDI-TOF/TOF MS. Good linearity was obtained with a determination coefficient (R2) larger than 0.99 in the range of 0.025–2.5 μΜ. The limit of detection (LOD) was 2.4 fmol. Acceptable precision and accuracy were gained by intra- and inter-day analysis with relative standard deviations less than 19.5% and relative recoveries ranging from 85.7 to 105.2%. In addition, the AYPBA labeled 20E produced abundant characteristic fragment ions under the high energy collision-induced dissociation, which facilitated the identification of 20E by MS2 analysis on MALDI-TOF/TOF MS. Using the method, we enabled the identification and quantification of endogenous 20E in four herbs including Cyanotis arachoidea, Achyranthes bidentata, Spinacia oleracea and Chenopodium quinoa willd., demonstrating the feasibility of the proposed method for screening of the endogenous 20E in plants.

Influence of Fe2O3 on the pyrolysis products distribution of demineralized Hongshaquan (HSQ) and Daliuta (DLT) coal was investigated by a novel in-situ pyrolysis vacuum ultraviolet single photon ionization time-of-flight mass spectrometry. The experiment samples were obtained by mechanical mixing of Fe2O3 and demineralized coal from acid elution with the mass ratio of 10:1, 5:1 and 2:1. Due to the characteristics of in-situ sampling, soft ionization and high vacuum environment, the initial pyrolysis volatiles including alkenes, aromatic hydrocarbons, phenols, bi-phenols and a small amount of sulfur and nitrogen-containing substances can be detected. The results show that Fe2O3 has similar influence on the products distribution of two demineralized coal, and it is relatively stronger on DLT coal than that on HSQ. With the increase of Fe2O3 content, the light fraction content is promoted such as alkenes and aromatic hydrocarbons, but bi-phenols are obviously inhibited. Correspondingly, the heavy products are also decreased, and the evolution peak temperatures of the product increase. Fe2O3 is successively reduced to FeO and Fe while transforming heavy components into light components. The increase of aliphatic carbon (or aromatic hydrocarbon substituted carbon) during coal pyrolysis process is mainly responsible for the decrease pyrolysis peak temperature.

Rhizoma Anemarrhenae is a well-known herbal medicine with saponins as its commonly regarded major bioactive components. It is essential to classify the properties of saponins which are associated with their toxicity and efficacy. In this study, 25 compounds were identified by HPLC-Q-TOF/MS in the extract of Rhizoma Anemarrhenae and 8 saponins were detected in rat plasma by HPLC-MS/MS after oral administration of this extract. These were neomangiferin, mangiferin, timosaponin E1, timosaponin E, timosaponin B-II, timosaponin B-III, timosaponin A-III and timosaponin A-I. A sensitive and accurate HPLC-MS/MS method was developed and successfully applied to a pharmacokinetic study of the abovementioned eight saponins after oral administration of the Rhizoma Anemarrhenae extract to rats. The method validation, including specificity, linearity, precision, accuracy, recovery, matrix effect and robustness, met the requirements of the intended use. The pharmacokinetic parameter, T max value, ranged from 2 to 8 h for these eight saponins whereas their elimination half-life (t 1/2) ranged from 4.06 to 9.77 h, indicating slow excretion. The plasma concentrations of these eight saponins were all very low, indicating a relatively low oral bioavailability. All these results provide support for further clinical studies.

Objectives The present study was conducted to evaluate the usefulness of capillary electrophoresis (CE), cation-exchange high-performance liquid chromatography (HPLC), and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) for the screening of hemoglobin (Hb) variants prevalent in southern China. Methods A total of 102 types of Hb variants in 1,083 variant carriers were identified over a 5-year period. These variants were analyzed by a CE method (Capillarys 3 TERA), a cation-exchange HPLC analyzer (Variant II Turbo 2.0), and a MALDI-TOF MS system (QuanTOF). Results The presence of 85 (83.3%, 85/102), 84 (82.4%, 84/102), and 62 (60.8%, 62/102) Hb variants was detected by Capillarys 3 TERA, Variant II Turbo 2.0, and QuanTOF, respectively. Of the three methods, only Capillarys 3 TERA recognized all 10 of the most frequent Hb variants in southern China. There were six, two, and three Hb variants that can only be detected by Capillarys 3 TERA, Variant II Turbo 2.0, and QuanTOF, respectively. The detection limit of mass difference for QuanTOF was approximately 11 to 20 Da. Conclusions MALDI-TOF MS is suitable for use as an auxiliary method rather than a stand-alone method for the screening of Hb variants prevalent in southern China.

Aim To purify Cryptosporidium spp. oocysts from clinical stool samples and evaluate using an up-to-date mass spectrometry protocol producing high-quality reference spectra. Methods and results A refined purification protocol was developed for oocysts from stools, involving salt flotation and potassium bromide density centrifugation. Purified oocysts were prepared for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) by formic acid extraction, and the extracts analysed using the Bruker MALDI Biotyper system. Individual spectral markers were identified by their specific mass peaks. Cryptosporidium parvum oocysts (Iowa strain) propagated in vivo, and C. parvum (n = 2) and Cryptosporidium hominis (n = 1) oocysts from clinical stool samples produced distinct spectra that were considered specific to Cryptosporidium spp. with no evidence of contamination. Conclusions The production of distinct spectra demonstrated the utility of the purification method for oocysts from clinical stool samples and provided reference spectra. Significance and impact of the study The use of MALDI-TOF MS and other mass spectrometry techniques has been limited previously to C. parvum oocysts propagated in vivo. Appropriate purification of oocysts can achieve sufficient biomass, enabling analysis by MALDI-TOF MS and potentially other mass spectrometry platforms, facilitating peptide and protein discovery and identification of biomarkers from a much wider range of Cryptosporidium spp. from natural infections.

Analysis and identification of seized doping-related products are important tasks for customs or forensic laboratories in order to prevent potentially dangerous and illegal compounds to go into circulation. At the Section of Forensic Chemistry in Copenhagen, we have a workflow consisting of four complimentary validated methods to identify common doping-related substances: liquid chromatography-ultraviolet (LC-UV), LC coupled with time of flight mass spectrometry (LC-TOF-MS), the colorimetric Bradford assay, and an immunoassay. The Bradford assay screens for peptide or proteins in the sample, and the immunoassay confirmed human chorionic gonadotropin (hCG). LC-UV was carried out with a C4 protein column for identification of peptides and proteins from a standard reference library, based on retention times and ratios between peak areas at 220, 254, and 280 nm. LC-TOF-MS was performed using a C18 column, and identification was based on comparison of the retention time and the accurate mass with those of reference standards. In 2019, we received 36 samples for peptide/protein analysis, all of which were tested using the LC-UV, LC-TOF-MS, and colorimetric method, and samples suspected of containing hCG were confirmed with an immunoassay. We found a total of 15 samples containing an illegal doping substance, 12 samples containing substances not prohibited by the Danish Doping List, and nine samples containing no peptides or proteins. In conclusion, the four complimentary methods constitute a suitable approach for identifying common peptide/protein doping substances in the day-to-day routine of a forensic laboratory, with limited sample preparation and interpretation of data.

Graphical abstract Traditional Chinese medicines played an important role in the treatment of COVID-19 in 2020. Ephedra sinica, one of the major constituent herbs of multi-component herbal formula, has been widely used to treat COVID-19 in China. However, its active components are still unclear. The objectives of this study are to screen and evaluate active components from the traditional Chinese medicine Ephedra sinica for the treatment of COVID-19. In our study, we established an ACE2/CMC bioaffinity chromatography model, and then developed an ACE2/CMC-HPLC-IT-TOF-MS system for the active compounds screening and identification from Ephedra sinica extract. We performed molecular docking and surface plasmon resonance (SPR) assays to assess the binding characteristics (binding mode and KD value). We used CCK-8 staining to assess the toxicity of screened compounds, and also used SARS-CoV-2 pseudovirus to observe the viropexis effect of screened compounds in ACE2h cells. In this current work, one fraction was fished out, separated and identified as ephedrine (EP), pseudoephedrine (PEP), and methylephedrine (MEP). Binding assays showed that the three compounds could bind with ACE2 in a special way to some amino acid residues, similar to the way SARS-CoV-2 bound with ACE2. Additionally, the three compounds, especially EP, can inhibit the entrance of SARS-CoV-2 spike pseudovirus into ACE2h cells because they can reduce the entrance ratio of pseudovirus in the pseudovirus model. Overall, the ACE2/CMC-HPLC-IT-TOF-MS system was established and verified to be suitable for ACE2-targeted bioactive compound screening. EP, PEP, and MEP with ACE2-binding features were screened out from Ephedra sinica, and acted as blockers inhibiting SARS-CoV-2 spike pseudovirus entering ACE2h cells. Supplementary Information The online version contains supplementary material available at 10.1007/s00216-021-03233-7.

Pu-erh tea is believed to be a beneficial beverage for health due to its many kinds of pharmacological effects. Nevertheless, detailed information related to differences in metabolites of Pu-erh raw tea from different geographical origins remains scarce. In this study, 43 elements were found in water-soluble components of Pu-erh raw tea by highly sensitive ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (U-HPLC/Q-TOF-MS). The characteristic groups of 29 metabolites from nondestructive proton nuclear magnetic resonance (1 H-NMR) spectroscopy were assigned. The variables contributed largely to the origin classification, mainly including valine, threonine, chlorogenic acid, quinic acid, epiafzelechin, and gallic acid ester, were screened out by sparse partial least squares discriminant analysis (sPLS-DA) method. This study provided a feasible and rapid technique for distinguishing Pu-erh tea from different areas by 1 H-NMR combined with sPLS-DA.

Syneilesis palmata (S. palmata) is a common Korean therapeutic herb extensively used to treat pain. The species Paris verticillata (P. verticillata) is morphologically similar to S. palmata but wit...

1. Chemical sensing in vertebrates is crucial in their lives, and efforts are undertaken towards deciphering their chemical language. Volatile organic compounds (VOCs) are a group of chemicals believed to play an essential role in a wide variety of animal interactions. Therefore, understanding what animals sense themselves and untangling the ecological role of their volatile cues can be accomplished by analysing VOC emissions. A proton-transfer-reaction time-of-flight mass spectrometer (PTR-TOF-MS) is an instrument that measures VOCs in real-time in an air sample. Since this technique acts as a hyper-sensitive 'nose' it has a similar potential in deciphering the chemical language of vertebrates. 2. Here, we validate the use of PTR-TOF-MS as a tool to measure VOCs from vertebrates, which in turn will help resolve vertebrate interactions through VOCs. The instrument monitors and records the full spectrum of VOCs emitted by an individual with a high accuracy and low detection limit, including transient VOC emissions. We propose and test diverse measuring configurations that allow for measurement of VOC emissions from different vertebrates and their exudates: full body, specific parts of the body, urine and femoral pores. In addition, we test configurations for sudden and short-lasting processes as VOCs emitted during adder skin shedding as well as the emissions of skin secretions upon mechanical and physiological stimulation in amphibia. Our configurations work in tandem with gas chromatography-mass spectrometry (GC-MS) to allow compound structure verification. 3. We discuss the configurations and methodologies used and conclude with recommendations for further studies, such as the choice of chamber size and flow. We also report the results of the measurements on vertebrates-that are novel to science-and discuss their ecological meaning. 4. We argue that PTR-TOF-MS has a high potential to resolve important unanswered questions in vertebrate chemical ecology with great adaptability to a wide range of experimental set-ups. If combined with a structure verification tool, such as GC-MS, the creative deployment of PTR-TOF-MS in various future study designs will lead to the identification of ecologically relevant VOCs.

In the present work, four, well-studied, model peptides (e.g., substance P, bradykinin, angiotensin I and AT-Hook 3) were used to correlate structural information provided by ion mobility and ECD/CID fragmentation in a TIMS-q-EMS-ToF MS/MS platform, incorporporating an electromagnetostatic cell (EMS). The structural heterogeneity of the model peptides was observed by i) multi-component ion mobility profiles (high ion mobility resolving power, R ~ 115–145), and ii) fast online characteristic ECD fragmentation patterns per ion mobility band (~ 0.2 min). Particularly, it was demonstrated that all investigated species were probably conformers, involving cis/trans-isomerizations at X-Pro peptide bond, following the same protonation schemes, in good agreement with previous ion mobility and single point mutation experiments. The comparison between ion mobility selected ECD spectra and traditional FT-ICR ECD MS/MS spectra showed comparable ECD fragmentation efficiencies but differences in the ratio of radical (•)/prime (ʹ) fragment species (H• transfer), which were associated with the differences in detection time after the electron capture event. The analysis of model peptides using online TIMS-q-EMS-ToF MS/MS provided complementary structural information on the intramolecular interactions that stabilize the different gas-phase conformations to those obtained by ion mobility or ECD alone.

Staining of biological cells with heavy metals can increase their visibility in mass spectrometry. In this study, the potential of ruthenium red (RR) as a staining agent for single-cell analysis by inductively coupled plasma time-of-flight mass spectrometry (SC-ICP-TOF-MS) is explored using two different yeast strains and one algal species. Time-of-flight mass spectrometry allows the simultaneous detection of Ru and multiple intrinsic elements in single cells. Ru has a better correlation with Mg than with P in Saccharomyces cerevisiae (S. cerevisiae) cells. For the three tested strains, the staining efficiency of RR exceeded 96%; the staining strengths were 30-32 ag μm-2 for the yeast cells and 59 ag μm-2 for the algal cells. By deriving the cell volume of single cells from their Ru mass, the concentration of Mg and P in individual cells of S. cerevisiae can be calculated. Elemental concentrations of Mg and P were highly variable in the cell individuals, with their 25-75 percentile values of 0.10-0.19 and 0.76-2.07 fg μm-3, respectively. RR staining has several advantages: it is fast, does not affect cell viability and is highly efficient. Provided that the shape of the individual cells of a culture is similar, Ru staining allows the elemental content to be directly correlated with the cell volume to accurately calculate the intracellular concentration of target elements in single cells. Therefore, RR can be a promising cell staining agent for future application in SC-ICP-TOF-MS research.

Chinese herbal medicines (CHMs) are classified as dietary supplements. Interactions with western medications, the presence of contaminants or adulterants, or a mis-labeled or mis-used CHM may lead to toxicological emergencies that can be undetected in death investigations. Laboratories must be able to efficiently analyze cases in which CHMs are suspected. Five extractions were evaluated for their ability to extract pharmacologically active compounds from herbal matrices: water, ethanol, microwave-assisted (MAE), ethanol : chloroform, and acid-wash. Anticonvulsive and other pharmacologically active compounds in Gou Teng, Tian Ma, and Jiang Can purchased from Beijing, China and New York were compared in the powder and the extracts using Direct Analysis in Real Time-Mass Spectrometry (DART-MS). Approximately 0.25 g of macerated herb was used per extraction. The water and ethanol extractions were simple liquid extractions. For the MAE, powdered herb was soaked in 65% ethanol, microwaved, and concentrated. The ethanol : chloroform extraction involved soaking in 1 : 1 ethanol : chloroform, sonication, and concentration. In the acid-wash extraction, powdered herb was soaked in acetic acid, followed by addition of sodium hydroxide, hexane extraction, and reconstitution in ethyl acetate. The powdered herbs and extracts were analyzed using a Jeol JMS T100LC AccuTOF DART-MS in positive and negative mode. Of the evaluated methods, no single extraction worked for all active compounds from the three CHMs. The MAE extract contained the most pharmacologically active compounds, while the acid-wash contained the least for the three products. Gou Teng purchased from different sources did exhibit a difference in pharmacologically active compounds, potentially from different species., With the popularity of natural products, rapid analysis of pharmacologically active analytes from natural matrices is critical for public health. Microwave-assisted extraction with DART-MS provides efficient analysis of analytes from herbal matrices.