Molecular Characterization of Ongoing Enzymatic Reactions in Raw Garlic Cloves Using Extractive Electrospray Ionization Mass Spectrometry

Characterization of enzymatic reactions occurring in untreated biological samples is of increasing interest. Herein, the chemical conversion of alliin to allicin, catalyzed by allinase, in raw garlic cloves has been followed in vivo by internal extractive electrospray ionization mass spectrometry (iEESI-MS). Both precursors and products of the enzymatic reaction were instantaneously extracted by infused solution running throughout the tissue and directly electrospray ionized on the edge of the bulk sample for online MS analysis. Compared to the room-temperature (+25 °C) scenario, the alliin conversion in garlic cloves decreased by (7.2 ± 1.4) times upon heating to +80 °C and by (5.9 ± 0.8) times upon cooling to −16 °C. Exposure of garlic to gentle ultrasound irradiation for 3 h accelerated the reaction by (1.2 ± 0.1) times. A 10 s microwave irradiation promoted alliin conversion by (1.6 ± 0.4) times, but longer exposure to microwave irradiation (90 s) slowed the reaction by (28.5 ± 7.5) times compared to the reference analysis. This method has been further employed to monitor the germination process of garlic. These data revealed that over a 2 day garlic sprouting, the allicin/alliin ratio increased by (2.2 ± 0.5) times, and the averaged degree of polymerization for the detected oligosaccharides/polysaccharides decreased from 11.6 to 9.4. Overall, these findings suggest the potential use of iEESI-MS for in vivo studies of enzymatic reactions in native biological matrices

Stabilization of Proteins and Noncovalent Protein Complexes during Electrospray Ionization by Amino Acid Additives

Ionization of proteins and noncovalent protein complexes with minimal disturbance to their native structure presents a great challenge for biological mass spectrometry (MS). In living organisms, the native structure of intracellular proteins is commonly stabilized by solute amino acids (AAs) accumulated in cells at very high concentrations. Inspired by nature, we hypothesized that AAs could also pose a stabilizing effect on the native structure of proteins and noncovalent protein complexes during ionization. To test this hypothesis, here we explored MS response for various protein complexes upon the addition of free AAs at mM concentrations into the electrospray ionization (ESI) solution. Thermal activation of ESI droplets in the MS inlet capillary was employed as a model destabilizing factor during ionization. Our results indicate that certain AAs, in particular proline (Pro), pose considerable positive effect on the stability of noncovalent protein complexes in ESI-MS without affecting the signal intensity of protein ions and original protein–ligand equilibrium, even when added at the 20 mM concentration. The data suggest that the degree of protein stabilization is primarily determined by the osmolytic and ampholytic characteristics of AA solutes. The highest stability and visibility of noncovalent protein complexes in ESI-MS are achieved using AA additives with neutral isoelectric point, moderate proton affinity, and unfavorable interaction with the native protein state. Overall, our results indicate that the simple addition of free amino acids into the working solution can notably improve the stability and accuracy of protein analysis by native ESI-MS

Simultaneous Preconcentration and Desalting of Organic Solutes in Aqueous Solutions by Bubble Bursting

Significant efforts are being made to develop more practical and versatile approaches for the preconcentration and purification of complex chemical samples. Inspired by the naturally occurring enrichment of organic compounds in sea aerosols, in this study we demonstrate the potential of induced bubble bursting as an approach for the preconcentration of organic solutes in various aqueous solutions. Apart from the preconcentration of organics, notable decrease in the concentration of metal salt components was discovered for the first time. On the basis of a series of model experiments, the phenomenon has been attributed to intermolecular competition at the surface interface of rising bubbles. Overall, our results indicate the high versatility and simplicity of the bubble bursting approach for the simultaneous preconcentration and desalting of organic solutes in aqueous solutions for mass spectrometry, chromatography, optical detection, and other fields of analysis

Real Time Online Correction of Mass Shifts and Intensity Fluctuations in Extractive Electrospray Ionization Mass Spectrometry

Real time online calibration of mass shift and intensity fluctuation to improve the accuracy of measurements for identification and quantitation in trace mass spectrometric analysis was demonstrated using extractive electrospray ionization mass spectrometry (EESI-MS). The signals of authentic compounds (e.g., lysine (Lys), proline (Pro), and histidine (His)) spiked into the extractive solution for the EESI process were used as the references to calibrate the signal of analytes (e.g., methionine (Met)) in the untreated sample solution. The intensity of the analyte signal was recorded simultaneously with the reference signals. The analyte signals at a given time point were calibrated on the basis of these correlation factors and real time signal response of the reference. The calibrated signal of Met at 10 μg L^–1 was improved with a better signal-to-noise ratio (S/N from 2.3 to 4.3), better linearity (<i>R</i>² from 0.9758 to 0.9980), and reduced relative standard deviation (RSD from 9.8% to 6.0%). The shift of mass-to-charge ratio of Met signal between the detected and theoretical values was decreased from 247 ± 133 to −7 ± 167 ppm for 50 min of detection using a linear ion trap mass analyzer and was reduced from −0.27 ± 0.60 to −0.12 ± 0.23 ppm for 50 min of detection using an Orbitrap mass analyzer (<i>P</i> = 95%). This method has been validated using a certified standard amino acids solution (GBW­(E)­100062) and applied for quantitative detection of amino acids in chicken feed, urine, nutritional drink, and facial mask samples, showing that the method is useful to improve the accuracy of mass spectrometric analysis

DataSheet_1_Epidemiological evidence for associations between variants in CHRNA genes and risk of lung cancer and chronic obstructive pulmonary disease.pdf

BackgroundGenetic studies have previously reported that single-nucleotide polymorphisms (SNPs) in CHRNA genes (such as CHRNA3, CHRNA4, CHRNA5, or CHRNA3-CHRNA5-CHRNB4 clusters) are linked to the risk of neoplastic and non-neoplastic diseases. However, these conclusions were controversial and no systematic research synopsis has been available. We aimed to synthesize current knowledge of variants in the CHRNA genes on the risk of diseases.MethodsWe systematically searched for publications using PubMed, Medline, and Web of Science on or before 25 August 2021. A total of 1,818 publications were identified, of which 29 were deemed eligible for inclusion that could be used to perform meta-analysis based on at least three data sources to assess whether the morbidity associated with neoplastic and non-neoplastic diseases can be attributed to SNPs in CHRNA genes. To further evaluate the authenticity of cumulative evidence proving significant associations, the present study covered the Venice criteria and false-positive report probability tests. Through the Encyclopedia of DNA Elements (ENCODE) project, we created functional annotations for strong associations.ResultsMeta-analyses were done for nine genetic variants with two diseases {chronic obstructive pulmonary disease (COPD) and lung cancer (LC)}that had at least three data sources. Interestingly, eight polymorphisms were significantly related to changes in the susceptibility COPD and LC (p ConclusionsOur study summarized comprehensive evidence showing that common mutations in CHRNA genes are strongly related to LC and COPD risk. The study also elucidated the vital function of CHRNA genes in genetic predispositions to human diseases.</p

Quantitative Determination of Bulk Molecular Concentrations of β‑Agonists in Pork Tissue Samples by Direct Internal Extractive Electrospray Ionization-Mass Spectrometry

Rapid quantitative determination of bulk molecular concentration in solid samples without sample pretreatment is demonstrated using the internal extractive electrospray ionization-mass spectrometry (iEESI-MS) analysis of six β-agonists, including salbutamol (Sal), clenbuterol (Cle), ractopamine (Rac), terbutaline (Ter), tulobuterol (Tul), brombuterol (Bro), in pork tissue samples. Single sample analysis only required 1 min. The linear range of detection was about 0.01–1000 μg/kg (<i>R</i>² > 0.9994). The limit-of-detection (LOD) varied from 0.002 μg/kg for Sal to 0.006 μg/kg for Tul. Relative standard deviation (RSD) of quantitation was in the range 6.5–11.3%. The analytical results were validated by gas chromatography–mass spectrometry (GC–MS) and high-performance liquid chromatography–mass spectrometry (LC–MS), showing the accuracy rates of 92–105%. The current study extends the power of ambient MS as a method for the quantification of molecules at the surface of solid samples (e.g., in μg/cm² units) toward the quantification of molecules in bulk sample volume (i.e., in μg/kg units), which is commonly required in food safety control, biomedical analysis, public security, and many other disciplines

Direct Assessment of Phytochemicals Inherent in Plant Tissues Using Extractive Electrospray Ionization Mass Spectrometry

An ambient pressure ionization mass spectrometric strategy called internal extractive electrospray ionization mass spectrometry (iEESI-MS) has been developed and applied for direct profiling of labile phytochemicals inherent in various native plant tissues, including leaves, roots, and fruits. By passing the electrospray solvent through the plant tissue, a variety of phytochemicals, such as amino acids, sugars (e.g., glucose, sucrose, polysaccharides, etc.), and alkaloids, were continuously extracted from the sample interior, driven toward the natural/cut electro-spraying tip, and vaporized into gaseous ions for mass spectrometric interrogation. Phytochemical patterns obtained by iEESI–MS permit a rapid differentiation between various species of ginkgo plant and strawberry maturity stages, as well as characterization of physiological/pathologic conditions of chlorophytum comosum. Our experimental results further demonstrate that the established iEESI–MS approach is potentially useful for direct phytochemomics studies with minimal biodegradation, allowing elucidation of plant metabolism with high speed, specificity, and simplicity of analysis

Direct Assessment of Phytochemicals Inherent in Plant Tissues Using Extractive Electrospray Ionization Mass Spectrometry

An ambient pressure ionization mass spectrometric strategy called internal extractive electrospray ionization mass spectrometry (iEESI-MS) has been developed and applied for direct profiling of labile phytochemicals inherent in various native plant tissues, including leaves, roots, and fruits. By passing the electrospray solvent through the plant tissue, a variety of phytochemicals, such as amino acids, sugars (e.g., glucose, sucrose, polysaccharides, etc.), and alkaloids, were continuously extracted from the sample interior, driven toward the natural/cut electro-spraying tip, and vaporized into gaseous ions for mass spectrometric interrogation. Phytochemical patterns obtained by iEESI–MS permit a rapid differentiation between various species of ginkgo plant and strawberry maturity stages, as well as characterization of physiological/pathologic conditions of chlorophytum comosum. Our experimental results further demonstrate that the established iEESI–MS approach is potentially useful for direct phytochemomics studies with minimal biodegradation, allowing elucidation of plant metabolism with high speed, specificity, and simplicity of analysis

Differentiation Using Microwave Plasma Torch Desorption Mass Spectrometry of Navel Oranges Cultivated in Neighboring Habitats

The molecular fingerprinting of intact fruit samples combined with statistical data analysis can allow the assessment of fruit quality and location of origin. Herein, microwave plasma torch desorption ionization mass spectrometry (MPT-MS) was applied to produce molecular fingerprints for the juice sac and exocarp of navel oranges cultivated in three closely located habitats, and the mass spectrometric fingerprints were differentiated by principal component analysis (PCA). Because of the relatively high temperature and high ionization efficiency of MPT, the volatile aroma compounds and semivolatile chemicals in the navel oranges were sensitively detected and confidently identified by collision induced dissociation (CID). The limit of detection (LOD) of MPT-MS for vanillin was 0.119 μg/L, with the relative standard deviation (RSD, <i>n</i> = 10) of 1.7%. The results showed that MPT-MS could be a powerful analytical platform for the sensitive molecular analysis of fruits at molecular level with high chemical specificity, allowing differentiation between the same sorts grown in neighboring habitats

DataSheet_1_Predicting effect of anti-PD-1/PD-L1 inhibitors therapy for hepatocellular carcinoma by detecting plasma metabolite based on UHPLC-MS.docx

IntroductionAnti-PD-1/PD-L1 inhibitors therapy has become a promising treatment for hepatocellular carcinoma (HCC), while the therapeutic efficacy varies significantly among effects for individual patients are significant difference. Unfortunately, specific predictive biomarkers indicating the degree of benefit for patients and thus guiding the selection of suitable candidates for immune therapy remain elusive.no specific predictive biomarkers are available indicating the degree of benefit for patients and thus screening the preferred population suitable for the immune therapy. MethodsUltra-high-pressure liquid chromatography-mass spectrometry (UHPLC-MS) considered is an important method for analyzing biological samples, since it has the advantages of high rapid, high sensitivity, and high specificity. Ultra-high-pressure liquid chromatography-mass spectrometry (UHPLC-MS) has emerged as a pivotal method for analyzing biological samples due to its inherent advantages of rapidity, sensitivity, and specificity. In this study, potential metabolite biomarkers that can predict the therapeutic effect of HCC patients receiving immune therapy were identified by UHPLC-MS. ResultsA partial least-squares discriminant analysis (PLS-DA) model was established using 14 glycerophospholipid metabolites mentioned above, and good prediction parameters (R2 = 0.823, Q2 = 0.615, prediction accuracy = 0.880 and p DiscussionThis study reveals that glycerophospholipid metabolites play a crucial role in predicting the efficacy of immune therapy for HCC.</p