Analysis of Norditerpenoid Alkaloids Extracted from Aconitum sinomantanum Nakai by Electrospray Ionization Tandem Mass Spectrometry

Electrospray ionization mass spectrometry（ESI-MS） was applied simultaneously in determining norditerpenoid alkaloids from the roots of Aconitum sinomantanum Nakai （RAS） based on molecular mass information. The tandem mass spectra（ ESI-MS^n） provided the alkaloidal structural information, through which the existence of these alkaloids was further confirmed. Accordingly, six known norditerpenoid alkaloids were simultaneously determined on the basis of their ESI-MS^n spectra. Furthermore, based on the diagnostic fragmentation pathways of alkaloidal MS^n, a rapid method for direct detection and characterization of alkaloids from an ethanolic extract of RAS was described.

Studies on the Basic Conformation of Lysozyme by Electrospray Ionization-Mass Spectrometry(ESI-MS)

In the present work, 2-mercaptoethanol and dithiothreitol were used to hydrogenise the internal disulfide bond in lysozyme. The experimental results indicate that the charge distribution of the proteins are different in the reaction process. From the calculated molecular weight, the reduction process of the disulfide bond in the molecules can be described, and the number of the disulfide bonds in the molecule can also be determined.

The Study on the Noncovalent Complexes of Silver Ion with Xanthone Diglycosides by Electrospray Ionization with Tandem Mass Spectrometry

The silver ionic complexes of xanthone glycosides were studied by ESI-MS/MS in the positive ion mode. The fragmentation pathways of silver ionic complexes under collisioninduced dissociation （CID） were investigated and the differences in MS/MS spectra of different silver ionic complexes of xanthones were correlated to the characterization of saccharide and the coordination pattern of silver ion with xanthones, including the glycosilation position and linkage type of disaccharide （1-2 and 1-6 linkages）.

Fast Screening of Chicken Egg Lysozyme in White Wine Products by Extractive Electrospray Ionization Mass Spectrometry

Fast detection of trace lysozyme,one of the most important food allergens in white wine samples,was achieved by extractive electrospray ionization mass spectrometry without sample pretreatment in this study.The multiply-charged ions of m/z1587 were chosen for the quantitative detection of lysozyme in white wine,showing linear dynamic signal responses in a range of 5―75μg/mL with a linearity coefficient of 0.999 and an acceptable relative standard deviation（RSD） of 8.0%―15.0% for directly measuring lysozyme in the complex food samples.The limit of detection for lysozyme in white wine sample was calculated to be 5 μg/mL,which was lower than the amounts that can provoke allergic reactions（oral test with 3 mg or labial test with 1 mg/mL）.A single sample analysis was completed within 1 min.The data demonstrate that extractive electrospray ionization mass spectrometry is a useful tool for fast screening lysozyme in the complex matrix,showing promising application in the rapid detection of food allergen.

Studies on Fragmentation Pathways of N-Ethoxy(phenyl) phosphoryl Amino Acids by Electrospray Ionization Tandem Mass Spectrometry

The positive and negative ESI-MS/MS spectra of N-ethoxy（phenyl） phosphoryl amino acids（EPP-AA） were investigated by electrospray ionization（ESI） ion trap mass spectrometry. The fragmentation pathways of [ M ＋ Na]^＋ and [ M - H]^- ions are proposed and rationalized. The observation may have some potential applications in the interpretation of the MS/MS spectra of novel N-phosphoryl compounds. The complexity of MS/MS spectra of EPP-AA [ M ＋ Na]^＋ ions is decreased compared with that of N-dialkyloxyphosphoryl amino acid. Therefore, the new phosphonamidate method may be considered one of the superior methods that can be used in sequencing peptides and proteins extensively.

Effects of Temperature and Energy on Stability of Oligomeric Enzyme Probed on Electrospray Ionization Mass Spectrometry

Escherichia coli 3-Deoxy-D-manno-octulosonate 8-phosphate（KDO8P） synthase catalyzed the condensation reaction between D-arabinose 5-phosphate（A5P） and phosphoenolpyruvate（PEP） to form KDO8P and inorganic phosphate（Pi）. The noncovalent tetrameric association ofKDO8P synthase was observed and dissociated in gas phase by means of electrospray ionization mass spectrometry under the very ＂soft＂ conditions. The results indicate that PEP-bound enzyme generated abundant tetrameric species as well as monomeric species at the ＂soft＂ conditions, whereas, the unbound enzyme favored the formation of a dimeric species. The mass spectra of the mixture of the enzyme with one of substrates, PEP, and A5P or one of products, KDO8P and Pi show that the complex of the unbound enzyme with PEP or Pi was prone to the formation of a monomeric species, whereas, that of the unbound enzyme with A5P or KDO8P was similar to the unbound enzyme. The intensity of the dimeric species increased with the increase of temperature at a collision voltage of 10 V. Taken together, the results presented here suggest that mass spectrometry will be a powerful tool to explore subtile conformational changes and/or subunit-subunit interactions of multiprotein assembly induced by ligand-binding and/or the changes of environmental conditions.

Nanokit coupled electrospray ionization mass spectrometry for analysis of angiotensin converting enzyme 2 activity in single living cell

Angiotensin-converting enzyme 2(ACE2) is not only an enzyme but also a functional receptor on cell membrane for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2). Here, the activity of ACE2 in single living cell is firstly determined using a nanokit coupled electrospray ionization mass spectrometry(nanokit-ESI-MS). Upon the insertion of a micro-capillary into the living h ACE2-CHO cell and the electrochemical sorting of the cytosol, the target ACE2 enzyme hydrolyses angiotensin II inside the capillary to generate angiotensin 1-7. After the electrospray of the mixture at the tip of the capillary, the product is differentiated from the substrate in molecular weight to achieve the detection of ACE2 activity in single cells. The further measurement illustrates that the inflammatory state of cells does not lead to the significant change of ACE2 catalytic activity, which elucidates the relationship between intracellular ACE2 activity and inflammation at single cell level. The established strategy will provide a specific analytical method for further studying the role of ACE2 in the process of virus infection, and extend the application of nanokit based single cell analysis.

Real-time toxicity prediction of Aconitum stewing system using extractive electrospray ionization mass spectrometry

Due to numerous obstacles such as complex matrices,real-time monitoring of complex reaction systems(e.g.,medicinal herb stewing system)has always been a challenge though great values for safe and rational use of drugs.Herein,facilitated by the potential ability on the tolerance of complex matrices of extractive electrospray ionization mass spectrometry,a device was established to realize continuous sampling and real-time quantitative analysis of herb stewing system for the first time.A complete analytical strategy,including data acquisition,data mining,and data evaluation was proposed and implemented with overcoming the usual difficulties in real-time mass spectrometry quantification.The complex Fuzi(the lateral root of Aconitum)-meat stewing systems were real-timely monitored in150 min by qualitative and quantitative analysis of the nine key alkaloids accurately.The results showed that the strategy worked perfectly and the toxicity of the systems were evaluated and predicated accordingly.Stewing with trotters effectively accelerated the detoxification of Fuzi soup and reduced the overall toxicity to 68%,which was recommended to be used practically for treating rheumatic arthritis and enhancing immunity.The established strategy was versatile,simple,and accurate,which would have a wide application prospect in real-time analysis and evaluation of various complex reaction systems.

Study of the Transient Reactive Pd(Ⅳ) Intermediate in the Pd(OAc)2-Catalyzed Oxidative Coupling Reaction System by Electrospray Ionization Tandem Mass Spectrometry

Pd-catalyzed oxidative coupling reaction was of great importance in the aromatic C--H activation and the for-mation of new C-O and C--C bonds. Sanford has pioneered practical, directed C-H activation reactions em-ploying Pd（OAc）2 as catalyst since 2004. However, until now, the speculated reactive Pd（Ⅳ） transient intermedi-ates in these reactions have not been isolated or directly detected from reaction solution. Electrospray ionization tandem mass spectrometry （ESI-MS/MS） was used to intercept and characterize the reactive Pd（Ⅳ） transient inter-mediates in the solutions of Pd（OAc）2-catalyzed oxidative coupling reactions. In this study, the Pd（IV） transient in-termediates were detected from the solution of Pd（OAc）2-catalyzed oxidative coupling reactions by ESI-MS and the MS/MS of the intercepted Pd（IV） transient intermediate in reaction system was the same with the synthesized au-thentic Pd（Ⅳ） complex. Our ESI-MS（/MS） studies confirmed the presence of Pd（Ⅳ） reaction transient intermedi-ates. Most interestingly, the MS/MS of Pd（Ⅳ） transient intermediates showed the reductive elimination reactivity to Pd（Ⅱ） complexes with new C-O bond formation into product in gas phase, which was consistent with the proposed reactivity of the Pd（Ⅳ） transient intermediates in solution.

Novel Rearrangement of Small Peptides in Electrospray Ionization Tandem Mass Spectrometry

Electrospray ionization mass spectrometry (ESI-MS) is a powerful method for sequencing peptides. A novel fragmentation pattern with the loss of a neutral fragment of 45 Da was observed with the dipeptides, tripeptides, tetrapeptides and pentapeptides containing phenylalanine or histidine residues. A novel rearrangement reaction with the extrusion of a formamide piece was studied and the rearrangement mechanism was proposed and confirmed by deuterium labeling experiments with ESI-MSn and high-resolution mass spectrometry. These findings are poten-tially helpful in identifying the specific sequence pattern in the peptide sequencing.

Liquid chromatography electrospray ionization tandem mass spectrometry(LC/ESI-MS/MS) method for quantitative estimation of moxifloxacin in human plasma

A rapid and sensitive liquid chromatography tandem mass spectrometry(LC-MS/MS) method has been developed and validated for the determination of moxifloxacin(MOXI) in human plasma. After a simple protein precipitation using acetonitrile, the post treatment samples were analysed on a C18 column interfaced with a Triple Quadropole Tandem Mass Spectrometer. Positive electrospray ionization was employed as the ionization source. The mobile phase consisted of 0.1% formic acid–acetonitrile(60:40, v/v). Ciprofloxacin(CIPRO) was used as an internal standard. The analyte and internal standard(CIPRO) were monitored in the multiple reaction monitoring mode(MRM). The mass transition ion-pair has been followed as m/z 402→358.2 for MOXI and 332→288.1 for CIPRO. The method was linear in the concentration range of 25–5000 ng/mL. The lower limit of quantification was 25 ng/mL. The intra- and inter-day precision(relative standard deviation) and accuracy(relative error) values were within 12.4%. Each plasma sample was analyzed within 3 min.

Elucidation of O-Phosphoryl and N-Phosphoryl Amino Acids by Electrospray Ionization Tandem Mass Spectrometry

Mass spectroscopic characteristics of phosphoryl amino acids were studied in detail by positive and negative electrospray ionization mass spectrometry (ESI-MS) in conjunction with tandem mass spectrometry (MS/MS). Be- sides N-diisopropyloxyphosphoryl amino acids (N-DIPP-AA), O-phospho- and O-diisopropyloxyphosphoryl amino acids (O-DIPP-AA) were studied and compared to N-DIPP-AA. The fragmentation pathways of [M+H]+, [M+Na]+ and [M-H]- ions of phosphoryl amino acids were summarized. In addition to several similar patterns, each of them showed its characteristic fragmention.

Identification,structure elucidation and origin of a common pyridinium-thiocyanate intermediate in electrospray mass spectrometry among the benziamidazole-class proton pump inhibitors

During the analysis of benziamidazole-class irreversible proton pump inhibitors,an unusual mass spectral response with the mass-to-charge ratio at[Mt10]t intrigued us,as it couldn't be assigned to any literature known relevant structure,intermediate or adduct ion.Moreover,this mysterious mass pattern of[Mt10]t has been gradually observed by series of marketed proton pump inhibitors,viz.omeprazole,pantoprazole,lansoprazole and rabeprazole.All the previous attempts to isolate the corresponding component were unsuccessful.The investigation of present work addresses this kind of signal to a pyridinium thiocyanate mass spectral intermediate(10),which is the common fragment ion of series of labile aggregates.The origin of such aggregates can be traced to the reactive intermediates formed by acid-promoted degradation.These reactive intermediates tend to react with each other and give raise series of complicated aggregates systematically in a water/acetonitrile solution by electrospray ionization.The structure of the corresponding pyridinium thiocyanate species of omeprazole(10a)has been eventually characterized with the help of synthetic specimen(10a′).Our structural proposal as well as its origin was supported by in situ nuclear magnetic resonance,chemical derivatization and colorimetric experiments.

Comparison of different approaches for direct coupling of solid-phase microextraction to mass spectrometry for drugs of abuse analysis in plasma

The direct coupling of solid-phase microextraction(SPME)to mass spectrometry(MS)(SPME-MS)has proven to be an effective method for the fast screening and quantitative analysis of compounds in complex matrices such as blood and plasma.In recent years,our lab has developed three novel SPME-MS techniques:SPME-microfluidic open interface-MS(SPME-MOI-MS),coated blade spray-MS(CBS-MS),and SPME-probe electrospray ionization-MS(SPME-PESI-MS).The fast and high-throughput nature of these SPME-MS technologies makes them attractive options for point-of-care analysis and anti-doping testing.However,all these three techniques utilize different SPME geometries and were tested with different MS instruments.Lack of comparative data makes it difficult to determine which of these methodologies is the best option for any given application.This work fills this gap by making a comprehensive comparison of these three technologies with different SPME devices including SPME fibers,CBS blades,and SPME-PESI probes and SPME-liquid chromatography-MS(SPME-LC-MS)for the analysis of drugs of abuse using the same MS instrument.Furthermore,for the first time,we developed different desorption chambers for MOI-MS for coupling with SPME fibers,CBS blades,and SPME-PESI probes,thus illustrating the universality of this approach.In total,eight analytical methods were developed,with the experimental data showing that all the SPME-based methods provided good analytical performance with R^(2)of linearities larger than 0.9925,accuracies between 81%and 118%,and good precision with an RSD%≤13%.

电喷雾解吸电离质谱法直接测定蔬菜表面的莠去津残留

A direct determination method for the atrazine residue on the vegetable was developed by using desorption electrospray ionization mass spectrometry (DESI MS) without any sample pretreatment.Acetonitrile-water (1:1,v/v),which contained 0.1% formic acid,was used as the spray solvent.The working conditions,such as ESI gas inlet pressure,ESI flow rate,ESI spray voltage,spray-to-sample distance,spray-to-cone-hole distance and the collision induced dissociation (CID) voltage for MS/MS,were optimized for both DESI and esquires 6 000 mass spectrometer.The linear range of atrazine on cabbage leaves was 25.25-2 525 pg/mm2,the R2 was 0.991 6,and the relative standard deviations were between 3.37% and 26.17%.The LOD of atrazine calculated by S/N=3 was 2.50 pg/mm2.

β-环糊精与蛋白质非共价复合物的电喷雾质谱研究

电喷雾质谱(ESI-MS)已经广泛应用于非共价复合物的检测和研究[1,2].ESI是一种极软的离子化过程,它不仅能在不断裂共价键的情况下使分子离子化,而且可以在离子化过程中保持分子间弱的非共价键作用.……

葛根素-7-磷酰二烷基酯的电喷雾质谱研究

Puerarin,an isoflavone compound,is the bioactive component of traditional Chinese medicine.A novel dialkyl puerarin-7-yl phosphate was synthesized and investigated by positive ion electrospray ionization ion trap mass spectrometry (ESI MS)in conjunction with tandem mass spectrometry.The fragmentation pathways of dialkyl puerarin-7-yl phosphates were investigated.

1-(N-苄氧羰基-氨基)-烷基膦酸单酯的正离子电喷雾电离多级质谱特征碎裂

作为一类重要的酶抑制剂,膦酸酯可被认为是酶的过渡态类似物.由于它不易被水解因而常被用作酶的抑制剂,或者作为具有酰胺或酯酶活性的催化抗体.鉴于其特殊的生物活性,目前已经报道了很多此类物质的合成方法[1～3].……

Metabolomic profiling for identification of metabolites and relevant pathways for taurine in hepatic stellate cells

AIM To develop a reliable and simple method to identify important biological metabolites and relevant pathways for taurine in hepatic stellate cells(HSCs), in order to provide more data for taurine therapy.METHODS All the biological samples were analyzed by using highperformance liquid chromatography-time electrospray ionization/quadrupole-time of flight mass spectrometry. Principal component analysis and partial least squares discriminant analysis were used to identify statistically different metabolites for taurine in HSCs, and metabolomic pathway analysis was used to do pathway analysis for taurine in HSCs. The chemical structure of the related metabolites and pathways was identified by comparing the m/z ratio and ion mode with the data obtained from free online databases.RESULTS A total of 32 significant differential endogenous metabolites were identified, which may be related to the mechanism of action of taurine in HSCs. Among the seven relevant pathways identified, sphingolipid metabolism pathway, glutathione metabolism pathway and thiamine metabolism pathway were found to be the most important metabolic pathways for taurine in HSCs.CONCLUSION This study showed that there were distinct changes in biological metabolites of taurine in HSCs and three differential metabolic pathways including sphingolipid pathway, glutathione pathway and thiamine metabolism pathway might be of key importance in mediating the mechanism of action of taurine in HSCs.

2',3'-双脱氢-2',3'双脱氧胸苷5'-亚磷酸苄酯的ESI-MS^n中的一种新颖的重排反应

爱滋病(AIDS)是一种严重威胁人类健康的传染性疾病.研究和开发新的、高疗效的抗AIDS的药物已成了生物学家、化学家和药物学家的主要的研究方向之一.2′,3′-双脱氢-2′,3′-双脱氧胸苷(d4T)是HIV逆转录酶抑制剂[1,2],它们是美国FDA通过的可用于治疗AIDS的一类药物[3].……