Picoplatin binding to proteins: X-ray structures and mass spectrometry data on the adducts with lysozyme and ribonuclease A.

The reactivity of the anticancer drug picoplatin (cis-amminedichlorido(2-methylpyridine)platinum(II) complex) with the model proteins hen egg white lysozyme (HEWL) and bovine pancreatic ribonuclease (RNase A) was investigated by electrospray ionisation mass spectrometry (ESI MS) and X-ray crystallography. The data were compared with those previously obtained for the adducts of these proteins with cisplatin, carboplatin and oxaliplatin under the same experimental conditions. ESI-MS data show binding of Pt to both proteins, with fragments retaining the 2-methylpyridine ligand and, possibly, a chloride ion. X-ray crystallography identifies different binding sites on the two proteins, highlighting a different behaviour of picoplatin in the absence or presence of dimethyl sulfoxide (DMSO). Metal-containing fragments bind to HEWL close to the side chains of His15, Asp18, Asp119 and both Lys1 and Glu7, whereas they bind to RNase A on the side chain of His12, Met29, His48, Asp53, Met79, His105 and His119. The data suggest that the presence of DMSO favours the loss of 2-methylpyridine and alters the ability of the Pt compound to bind to the two proteins. With both proteins, picoplatin appears to behave similarly to cisplatin and carboplatin when dissolved in DMSO, whereas it behaves more like oxaliplatin in the absence of the coordinating solvent. This study provides important insights into the pharmacological profile of picoplatin and supports the conclusion that coordinating solvents should not be used to evaluate the biological activities of Pt-based drugs.

In vitro study on the competitive reactions between arsenite and selenite with glutathione.

Exposure to arsenic can cause various biological effects by increasing the production of reactive oxygen species (ROS). Selenium acts as a beneficial element by regulating ROS and limiting heavy metal uptake and translocation. There are studies on the interactive effects of As and Se in plants, but the antagonistic and synergistic effects of these elements based on their binding to glutathione (GSH) molecules have not been studied yet. In this study, we aimed to investigate the antagonistic or synergistic effects of As and Se on the binding mechanism of Se and As with GSH at pH 3.0, 5.0, or 6.5. The interaction of As and Se in Se(SG)2 + As(III) or As(SG)3 + Se(IV) binary systems and As(III) + Se(IV) + GSH ternary system were examined depending on their ratios via liquid chromatography diode array detector/electrospray mass spectrometry (LC-DAD/MS) and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). The results showed that the formation of As(GS)3 was not detected in the As(III) + Se(SG)2 binary system, indicating that As(III) did not affect the stability of Se(SG)2 complex antagonistically. However, in the Se(IV) + As(SG)3 binary system, the addition of Se(IV) to As(SG)3 affected the stability of As(SG)3 antagonistically. Se(IV) reacted with GSH, disrupting the As(SG)3 complex, and consequently, Se(SG)2 formation was measured using LC-MS/DAD. In the Se(IV) + GSH + As(III) ternary system, Se(SG)2 formation was detected upon mixing As(III), Se(IV), and GSH. The increase in the concentration of As(III) did not influence the stability of the Se(SG)2 complex. Additionally, Se(IV) has a higher affinity than As(III) to the GSH, regardless of the pH of the solution. In both binary and ternary systems, the formation of the by-product glutathione trisulfide (GSSSG) was detected using LC-ESI-MS/MS.

HPLC-ESI/MS-MS metabolic profiling of white pitaya fruit and cytotoxic potential against cervical cancer: Comparative studies, synergistic effects, and molecular mechanistic approaches.

Natural approach became a high demand for the prevention and treatment of such diseases for their proven safety and efficacy. This study is aimed to perform comparative phytochemical analysis of white pitaya (Hylocereus undatus) peel, pulp and seed extracts via determination of total flavonoid content, phenolic content, and antioxidant capacity, coupled with HPLC-ESI/MS-MS analysis. Further, we evaluated the synergistic cytotoxic potential with Cisplatin against cervical cancer cells with investigation of underlying mechanism. The highest content of phenolics and antioxidants were found in both seed and peel extracts. The HPLC-ESI/MS-MS revealed identification of flavonoids, phenolic acids, anthocyanin glycosides, lignans, stilbenes, and coumarins. The cytotoxicity effects were evaluated by MTT assay against prostate, breast and cervical (HeLa) and Vero cell lines. The seed and peel extracts showed remarkable cytotoxic effect against all tested cell lines. Moreover, the selectivity index confirmed high selectivity of pitaya extracts to cancer cells and safety on normal cells. The combined therapy with Cisplatin effectively enhanced its efficacy and optimized the treatment outcomes, through the apoptotic ability of pitaya extracts in HeLa cells, as evaluated by flow cytometry. Besides, RT-PCR and western blotting analysis showed downregulation of Bcl-2 and overexpression of P53, BAX among HeLa cells treated with pitaya extracts, which eventually activated apoptosis process. Thus, pitaya extract could be used as adjuvant therapy with cisplatin for treatment of cervical cancer. Furthermore, in-vivo extensive studies on the seed and peel extracts, and their compounds are recommended to gain more clarification about the required dose, and side effects.

Improved Detection of Tryptic Peptides from Tissue Sections Using Desorption Electrospray Ionization Mass Spectrometry Imaging.

DESI-MSI is an ambient ionization technique used frequently for the detection of lipids, small molecules, and drug targets. Until recently, DESI had only limited use for the detection of proteins and peptides due to the setup and needs around deconvolution of data resulting in a small number of species being detected at lower spatial resolution. There are known differences in the ion species detected using DESI and MALDI for nonpeptide molecules, and here, we identify that this extends to proteomic species. DESI MS images were obtained for tissue sections of mouse and rat brain using a precommercial heated inlet (approximately 450 °C) to the mass spectrometer. Ion mobility separation resolved spectral overlap of peptide ions and significantly improved the detection of multiply charged species. The images acquired were of pixel size 100 µm (rat brain) and 50 µm (mouse brain), respectively. Observed tryptic peptides were filtered against proteomic target lists, generated by LC-MS, enabling tentative protein assignment for each peptide ion image. Precise localizations of peptide ions identified by DESI and MALDI were found to be comparable. Some spatially localized peptides ions were observed in DESI that were not found in the MALDI replicates, typically, multiply charged species with a low mass to charge ratio. This method demonstrates the potential of DESI-MSI to detect large numbers of tryptic peptides from tissue sections with enhanced spatial resolution when compared to previous DESI-MSI studies.

A comprehensive HRMS methodology using LC-(ESI)-/GC-(APCI)-QTOF MS complementary platforms for wide-scope target screening of >750 pesticides in olive oil.

This study presents the development and validation of a comprehensive high-resolution mass spectrometry (HRMS) methodology for the detection of 771 pesticides in olive oil, using liquid chromatography with electrospray ionization, operating in positive and negative mode, and gas chromatography with atmospheric-pressure chemical ionization in positive mode, both coupled to quadrupole-time-of-flight mass spectrometry (LC-(ESI)-/GC-(APCI)-QTOF MS). Special reference is made to the post-acquisition evaluation step, in which all LC/GC-HRMS analytical evidence (i.e. mass accuracy, retention time, isotopic pattern, MS/MS fragmentation) is taken into account in order to successfully identify the compounds. The sample preparation of the method involves a QuEChERS-based protocol, common for both techniques, differentiated only on the reconstitution step, making the method highly applicable in routine analysis. A smart evaluation of method's performance was carried out, with 65 representative analytes comprising the validation set. The method was validated in terms of linearity, accuracy, matrix effect and precision, while the limits of detection and quantification of the method were estimated. Finally, twenty Greek olive oil samples were analysed in both analytical platforms and the findings included the pesticides lambda-cyhalothrin, chlorpyrifos, phosphamidon, pirimiphos-methyl and esprocarb at low ng g-1 level.

Detection and Imaging of Exposure-Related Metabolites and Xenobiotics in Hard Tissues by Laser Sampling and Mass Spectrometry.

The utility of two novel laser-based methods, laser ablation electrospray ionization (LAESI) and laser desorption ionization (LDI) from silicon nanopost array (NAPA), is explored via local analysis and mass spectrometry imaging (MSI) of hard tissues (tooth and hair) for the detection and mapping of organic components. Complex mass spectra are recorded in local analysis mode from tooth dentin and scalp hair samples. Nicotine and its metabolites (cotinine, hydroxycotinine, norcotinine, and nicotine) are detected by LAESI-MS in the teeth of rats exposed to tobacco smoke. The intensities of the detected metabolite peaks are proportional to the degree of exposure. Incorporating ion mobility separation in the LAESI-MS analysis of scalp hair enables the detection of cotinine in smoker hair along with other common molecular species, including endogenous steroid hormones and some lipids. Single hair strands are imaged by MALDI-MSI and NAPA-LDI-MSI to explore longitudinal variations in the level of small molecules. Comparing spectra integrated from NAPA-LDI-MSI and MALDI-MSI images reveals that the two techniques provide complementary information. There were 105 and 82 sample-related peaks for MALDI and NAPA, respectively, with an overlap of only 16 peaks, indicating a high degree of complementarity. Enhanced molecular coverage and spatial resolution offered by LAESI-MS and NAPA-LDI-MSI can reveal the distributions of known and potential biomarkers in hard tissues, facilitating exposome research.

Removal of isobaric interference using pseudo-multiple reaction monitoring and energy-resolved mass spectrometry for the isotope dilution quantification of a tryptic peptide.

Energy-resolved mass spectrometry (ERMS) and an isotopically labelled internal standard were successfully combined to accurately quantify a tryptic peptide despite the presence of an isobaric interference. For this purpose, electrospray ionisation tandem mass spectrometry (ESI-MS/MS) experiments were conducted into an ion trap instrument using an unconventional 8 m/z broadband isolation window, which encompassed both the tryptic peptide and its internal standard. Interference removal was assessed by determining an excitation voltage that was high enough to maintain a constant value for the analyte/internal standard peaks intensity ratio, thus ensuring accurate quantification even in the presence of isobaric contamination. Pseudo-multiple reaction monitoring (MRM) was employed above this excitation voltage to quantify the trypic peptide. The internal standard calibration model showed no lack of fit and exhibited a linear dynamic range from 0.5 µM up to 2.5 µM. The detection limit was 0.08 µM. The accuracy of the method was evaluated by quantifying the tryptic peptide of three reference samples intentionally contaminated with the isobaric interference. All the reference samples were accurately quantified with ∼1% deviation despite the isobaric contamination. Furthermore, we have demonstrated that this methodology can also be applied to quantify the isobaric peptide by standard additions down to 0.2 µM. Finally, liquid chromatography ERMS (LC ERMS) experiments yielded similar results, suggesting the potential of the proposed methodology for analysing complex samples.

Identification and quantification of chain-pairing variants or mispaired species of asymmetric monovalent bispecific IgG1 monoclonal antibody format using reverse-phase polyphenyl chromatography coupled electrospray ionization mass spectrometry.

Developing a knob-into-hole asymmetric bispecific IgG1 monoclonal antibody (mAb) poses manufacturing challenges due to the expression of chain pairing variants, also called mispaired species, in the desired product. The incorrect pairing of light and heavy chains could result in heterogeneous mispaired species of homodimers, heterodimers, light chain swapping, and low molecular weight species (LMWS). Standard chromatography, capillary electrophoretic, or spectroscopic methods poorly resolve these from the main variants. Here, we report a highly sensitive reverse-phase polyphenyl ultra-high-performance liquid chromatography (RP-UHPLC) method to accurately measure mispaired species of Duet mAb format, an asymmetric IgG1 bispecific mAb, for both process development and quality control analytical tests. Coupled with electrospray ionization mass spectrometry (ESI-MS), it enabled direct online characterization of mispaired species. This single direct assay detected diverse mispaired IgG-like species and LMWS. The method resolved eight disulfide bonds dissociated LMWS and three mispaired LMWS. It also resolved three different types of IgG-like mispaired species, including two homodimers and one heterodimer. The characterization and quantification simultaneously enabled the cell line selection that produces a lesser heterogeneity and lower levels of mispaired species with the desired correctly paired product. The biological activity assessment of samples with increased levels of these species quantified by the method exhibited a linear decline in potency with increasing levels of mispaired species in the desired product. We also demonstrated the utility of the technique for testing in-process intermediate materials to determine and assess downstream purification process capability in removing diverse mispaired IgG-like species and LMWS to a certain level during the downstream purification process. Our investigation demonstrates that adopting this method was vital in developing asymmetric bispecific mAb from the initial stage of cell line development to manufacturing process development. Therefore, this tool could be used in the control strategy to monitor and control mispaired species during manufacturing, thus improving the quality control of the final product.

Extraction of naturally occurring peptides versus the tryptic digestion of proteins from fetal versus adult bovine serum for LC-ESI-MS/MS.

The naturally occurring peptides and digested proteins of fetal versus adult bovine serum were compared by LC-ESI-MS/MS after correction against noise from blank injections and random MS/MS spectra as statistical controls. Serum peptides were extracted by differential precipitation with mixtures of acetonitrile and water. Serum proteins were separated by partition chromatography over quaternary amine resin followed by tryptic digestion. The rigorous X!TANDEM goodness of fit algorithm that has a low error rate as demonstrated by low FDR q-values (q ≤ 0.01) showed qualitative and quantitative agreement with the SEQUEST cross correlation algorithm on 12,052 protein gene symbols. Tryptic digestion provided a quantitative identification of the serum proteins where observation frequency reflected known high abundance. In contrast, the naturally occurring peptides reflected the cleavage of common serum proteins such as C4A, C3, FGB, HPX, A2M but also proteins in lower concentration such as F13A1, IK, collagens and protocadherins. Proteins associated with cellular growth and development such as actins (ACT), ribosomal proteins like Ribosomal protein S6 (RPS6), synthetic enzymes and extracellular matrix factors were enriched in fetal calf serum. In contrast to the large literature from cord blood, IgG light chains were absent from fetal serum as observed by LC-ESI-MS/MS and confirmed by ELISA.

Evaluation of the Antiproliferative Potential of Yellow Jaboticaba (Myrciaria glazioviana) Extracts Against Human Cervical Cancer (HeLa cells line) and the Analysis of Their Chemical Composition by HPLC-HRESIMS.

Cervical cancer is a specific type of cancer that affects women around the world, with an incidence of 604 thousand new cases per year and 341 thousand deaths. There is a high demand for new effective antineoplastic drugs with few side effects. In this sense, recent research highlights the potential of compounds of natural origin in treating and preventing different types of cancer. Myrciaria glazioviana is a Brazilian native species belonging to the Myrtaceae family, which has previously described biological activities such as antimicrobial, anti-inflammatory, and antioxidant properties. This study aims to evaluate the anticancer activity of the dichloromethane extract (MGD) and ethyl acetate extract (MGA) of M. glazioviana leaves against human cervical cancer cell line (HeLa), as well as to identify their bioactive compounds. Using HPLC-HRESIMS technique, ten compounds were characterized in both samples: quinic acid, ellagic acid, Tri-O-methyl ellagic acid, two derivatives of Tetra-O-methyl flavellagic acid, quercetrin, Di-O-methyl ellagic acid, and three derivatives of pentamethyl coruleoellagic acid. Through MTT assays using HeLa cells and NIH/3T3 cells, it was observed that MGD and MGA were selective against tumor cells, with IC50 values of 24.31 and 12.62 µg/mL, respectively. The samples induced the tumor cell death by apoptosis, as evidenced by the activation of caspases 3/7, cell shrinkage, and pyknotic nuclei. Both samples were also able to inhibit the migration of HeLa cells after 24 hours of treatment, indicating a potential antimetastatic effect. Therefore, the present research highlights the antiproliferative and antimigratory potential of this species against HeLa cells.

Concentric Hybrid Nanoelectrospray Ionization-Atmospheric Pressure Chemical Ionization Source for High-Coverage Mass Spectrometry Analysis of Single-Cell Metabolomics.

High-coverage mass spectrometry analysis of single-cell metabolomics remains challenging due to the extremely low abundance and wide polarity of metabolites and ultra-small volume in single cells. Herein, a novel concentric hybrid ionization source, nanoelectrospray ionization-atmospheric pressure chemical ionization (nanoESI-APCI), is ingeniously designed to detect polar and nonpolar metabolites simultaneously in single cells. The source is constructed by inserting a pulled glass capillary coaxially into a glass tube that acts as a dielectric barrier layer. Benefitting from the integrated advantages of nanoESI and APCI, its limit of detection is improved by one order of magnitude to 10 pg mL-1. After the operational parameter optimization, 254 metabolites detected in nanoESI-APCI are tentatively identified from a single cell, and 82 more than those in nanoESI. The developed nanoESI-APCI is successively applied to study the metabolic heterogeneity of human hepatocellular carcinoma tissue microenvironment united with laser capture microdissection (LCM), the discrimination of cancer cell types and subtypes, the metabolic perturbations to glucose starvation in MCF7 cells and the metabolic regulation of cancer stem cells. These results demonstrated that the nanoESI-APCI not only opens a new avenue for high-coverage and high-sensitivity metabolomics analysis of single cell, but also facilitates spatially resolved metabolomics study coupled with LCM.

Effects of Nano-Electrospray Ionization Emitter Position on Unintentional In-Source Activation of Peptide and Protein Ions.

Native ion mobility-mass spectrometry (IM-MS) typically introduces protein ions into the gas phase through nano-electrospray ionization (nESI). Many nESI setups have mobile stages for tuning the ion signal and extent of co-solute and salt adduction. However, tuning the position of the emitter capillary in nESI can have unintended downstream consequences for collision-induced unfolding or collision-induced dissociation (CIU/D) experiments. Here, we show that relatively small variations in the nESI emitter position can shift the midpoint (commonly called the "CID50" or "CIU50") potential of CID breakdown curves and CIU transitions by as much as 8 V on commercial instruments. A spatial "map" of the shift in CID50 for the loss of heme from holomyoglobin onto the emitter position on a Waters Synapt G2-Si mass spectrometer shows that emitter positions closer to the instrument inlet can result in significantly greater in-source activation, whereas different effects are found on an Agilent 6545XT instrument for the ions studied. A similar effect is observed for CID of the singly protonated leucine enkephalin peptide and Shiga toxin 1 subunit B homopentamer on the Waters Synapt G2-Si instrument. In-source activation effects on a Waters Synapt G2-Si are also investigated by examining the RMSD between CIU fingerprints acquired at different emitter positions and the shifts in CIU50 for structural transitions of bovine serum albumin and NIST monoclonal antibody.

LC/Q-TOF-MS-based structural characterization of enasidenib degradation products and establishment of a stability-indicating assay method: Insights into chemical stability.

RATIONALE: Enasidenib (EDB) is an orally active selective mutant isocitrate dehydrogenase-2 enzyme inhibitor approved by the U.S. Food and Drug Administration to treat acute myeloid leukemia. It lacks a reported forced degradation study and a stability-indicating assay method (SIAM). This study addresses this gap by establishing a degradation profile in accordance with the International Council for Harmonisation Q1A and Q1B (R2) guidelines and developing a validated SIAM for EDB. METHODS: EDB was exposed to forced degradation under various conditions (hydrolytic, photolytic, oxidative, and thermal stress). Degradation samples were analyzed using high-performance liquid chromatography on an Agilent ZORBAX Eclipse Plus C18 column with a mobile phase consisting of 0.1% formic acid in Milli-Q water and acetonitrile at a flow rate of 1 mL/min and detection at 270 nm. Liquid chromatography-quadrupole time-of-flight-high-resolution mass spectrometry (LC/Q-TOF HRMS) was used for the identification and characterization of degradation products. Nitrosamine risk assessment was conducted using a modified nitrosation assay procedure (NAP) test due to the presence of a secondary amine group in the drug, which is liable to forming nitrosamine drug substance-related impurities (NDSRI). RESULTS: The drug exhibited significant degradation under acidic, basic, photolytic, and oxidative conditions in the solution state. A total of nine degradation products (DP) were formed (DP-I, DP-III, and DP-IV: acidic conditions; DP-I and DP-III: basic conditions; DP-II, DP-V, DP-VI, and DP-VII: oxidative stress; and DP-VII, DP-VIII, and DP-IX: photolytic conditions), which were separated and identified using reversed-phase high-performance liquid chromatography and characterized using liquid chromatography-tandem mass spectrometry. The mechanism behind the formation of EDB degradation products has been discussed, and this study was the first to develop a degradation pathway for EDB. In addition, the possibilities of NDSRI formation for EDB were studied using a modified NAP test, which can contribute to the risk assessment of the drug. CONCLUSIONS: Forced degradation studies were conducted by establishing a SIAM for EDB. All the degradation products were characterized by mass spectral data obtained using LC/Q-TOF-HRMS.

Development of a Python-based electron ionization mass spectrometry amino acid and peptide fragment prediction model.

The increased fragmentation caused by harsher ionization methods used during mass spectrometry such as electron ionization can make interpreting the mass spectra of peptides difficult. Therefore, the development of tools to aid in this spectral analysis is important in utilizing these harsher ionization methods to study peptides, as these tools may be more accessible to some researchers. We have compiled fragmentation mechanisms described in the literature, confirmed them experimentally, and used them to create a Python-based fragment prediction model for peptides analyzed under direct exposure probe electron ionization mass spectrometry. This initial model has been tested using single amino acids as well as targeted libraries of short peptides. It was found that the model does well in predicting fragments of peptides composed of amino acids for which the model is well-defined, but several cases where additional mechanistic information needs to be incorporated have been identified.

M2 tumor-associated macrophages and CXCL2 induce lipid remodeling in hepatocellular carcinoma cell lines.

Primary hepatocellular carcinoma (HCC) is one of the most common malignant tumors, but its pathogenesis remains incompletely elucidated. Recently, many studies indicated that lipid remodeling plays an important role in the occurrence and development of HCC. Furthermore, lipids have been proven to be indispensable mediators in promoting communication between tumor cells and extracellular matrix in the tumor microenvironment. Thus, this study aims to comprehensively investigate the process of lipid remodeling during HCC metastasis based on the LC-electrospray ionization-MS (LC-ESI-MS) combined with multiple reaction monitoring technology. M2 tumor-associated macrophages and the recombinant human protein CXCL2 were used to simulate the tumor microenvironment. After co-incubating SMMC7721 and MHCC97-H cell lines with M2 tumor-associated macrophages or the recombinant human protein CXCL2 for 48 h, LC-ESI-MS was used to quantify the levels of two major classes of lipid molecules, namely, glycerophospholipids and sphingolipids. Our results suggest that lipid remodeling in the tumor microenvironment may promote the migration and invasion of HCC cell lines.

Determination of three C18-oxygenated steroids in adrenal lesion segments in primary aldosteronism by super-selective adrenal venous sampling and LC/ESI-MS/MS.

Super-selective adrenal venous sampling (ssAVS) can collect the adrenal tributary venous blood in the aldosterone (ALD)-hypersecreting segments in primary aldosteronism. The concentrations of the C18-oxygenated steroids, especially 18-oxocortisol (18-oxoF), in the lesion segments might be more useful indices than those in the peripheral or adrenal central veins (current candidate indexes) for the differential diagnosis of unilateral ALD-producing adenoma (APA) and bilateral adrenal hyperplasia (BAH). To verify this hypothesis, we developed a liquid chromatography/electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) method for simultaneously quantifying ALD, 18-oxoF and 18-hydroxycortisol in the adrenal tributary venous serum sample collected by ssAVS (ssAVS serum) and compared their concentrations between APA and BAH patients. Only deproteinization was required for a 10 µl sample prior to the LC/ESI-MS/MS analysis. Endogenous corticoids did not interfere with the quantifications, and the intra-assay and interassay precisions (≤ 8.3%) and accuracies (94.2-102.7%) were acceptable. The clinical study revealed that the 18-oxoF concentration was significantly higher in the ALD-producing tumor tissues (from APA patients) than in the hyperplastic tissues (from BAH patients). However, in conclusion, the 18-oxoF concentration in the ssAVS serum sample can be a rough indication but cannot be decisive for the differential diagnosis between APA and BAH owing to the significant individual difference.

UHPLC-ESI-QTOF-MS metabolite profiles of different extracts from Pelargonium endlicherianum parts and their biological properties based on network pharmacological approaches.

In the present study, we aimed to investigate the chemical profiles and biological activities of different extracts (ethyl acetate, dichloromethane, ethanol, and water) of Pelargonium endlicherianum parts (aerial parts and roots). Free radical scavenging, reducing power, phosphomolybdenum, and metal chelating were assayed for antioxidant properties. To detect enzyme inhibitory properties, cholinesterase, amylase, glucosidase, and tyrosinase were chosen as target enzymes. The ethanol extract of the aerial parts contained higher amounts of total bioactive compounds (120.53 mg GAE/g-24.46 mg RE/g). The ethanol and water extracts of these parts were tentatively characterized by UHPLC-ESI-QTOF-MS and 95 compounds were annotated. In addition, the highest acetylcholiesterase (3.74 mg GALAE/g) and butyrylcholinesterase (3.92 mg GALAE/g) abilities were observed by the ethanol extract of roots. The water extract from aerial parts exhibited the most pronounced inhibitory effects on multiple cancer cell lines, especially A549 (IC50: 23.2 µg/mL) and HT-29 (IC50: 27.43 µg/mL) cells. Using network pharmacology, P. endlicherianum compounds were studied against cancer, revealing well-connected targets such as epidermal growth factor receptor (EGFR), phosphoinositide-3-kinase (PI3K), AKT, receptor tyrosine-protein kinase erbB-2, and growth factor receptor bound protein 2 (GRB2) with significant impact on cancer-related pathways. The results could open a new path from natural treasure to functional applications with P. endlicherianum and highlight a new study on other uninvestigated Pelargonium species.

Phenolic diversity and antioxidant potential of different varieties of bamboo leaves using LC-ESI-QTOF-MS/MS and LC-ESI-QqQ-MS/MS.

Bamboo is a highly sustainable plant with a wide variety of leaves, yet little is known about its bioactive composition. Therefore, this study aims to characterize the phenolic profile and antioxidant capacity of 11 different varieties of bamboo leaves using liquid chromatography coupled with mass spectrometry. As a result, 81 phenolic compounds were tentatively identified, 29 of which were identified for the first time in the literature for bamboo leaves. The tentatively identified compounds fell into five classes (hydroxybenzoic and hydroxycinnamic acids, flavones, flavanones, and flavonols). The concentration of phenolic compounds ranged from 103 to 1291 mg/100 g. Among the provisionally identified compounds, there was a predominance of derivatives from the luteolin and apigenin group, with orientin and schaftoside being the majority in each group, respectively. The leaves also showed significant variation in antioxidant activity, highlighting the potential bioactive composition of bamboo leaves for future applications in the food industry.

Biomarker Identification in Liver Cancers Using Desorption Electrospray Ionization Mass Spectrometry (DESI-MS) Imaging: An Approach for Spatially Resolved Metabolomics.

Liver cancers are characterized by interindividual and intratumoral heterogeneity, which makes early diagnosis and the development of therapies challenging. Desorption electrospray ionization mass spectrometry (DESI-MS) imaging is a potent and sensitive MS ionization technique for direct, unaltered 2D and 3D imaging of metabolites in complex biological samples. Indeed, DESI gently desorbs and ionizes analyte molecules from the sample surface using an electrospray source of highly charged aqueous spray droplets in ambient conditions. DESI-MS imaging of biological samples allows untargeted analysis and characterization of metabolites in liver cancers to identify new biomarkers of malignancy. In this chapter, we described a detailed protocol using liver cancer samples collected and stored for histopathology examination, either as frozen or as formalin-fixed, paraffin-embedded specimens. Such hepatocellular carcinoma samples can be subjected to DESI-MS analyses, illustrating the capacity of spatially resolved metabolomics to distinguish malignant lesions from adjacent normal liver tissue.

MS3 Imaging Enables the Simultaneous Analysis of Phospholipid C═C and sn-Position Isomers in Tissues.

Mass spectrometry (MS) imaging of lipids in tissues with high structure specificity is challenging in the effective fragmentation of position-selective structures and the sensitive detection of multiple lipid isomers. Herein, we develop an MS3 imaging method for the simultaneous analysis of phospholipid C═C and sn-position isomers by on-tissue photochemical derivatization, nanospray desorption electrospray ionization (nano-DESI), and a dual-linear ion trap MS system. A novel laser-based sensing probe is developed for the real-time adjustment of the probe-to-surface distance for nano-DESI. This method is validated in mouse brain and kidney sections, showing its capability of sensitive resolving and imaging of the fatty acyl chain composition, the sn-position, and the C═C location of phospholipids in an MS3 scan. MS3 imaging of phospholipids has shown the capability of differentiation of cancerous, fibrosis, and adjacent normal regions in liver cancer tissues.