2-((1,1-Dioxidotetrahydrothiophen-3-yl)(methyl)amino)-2-oxoethyl 3-(3-nitrophenyl)acrylate

Profiling of hydroxycinnamoylquinic acids in plant extracts using in-source CID fragmentation.[Pubmed:27591562]

J Mass Spectrom. 2016 Dec;51(12):1130-1145.

Hydroxycinnamoylquinic acids (HCQAs) are a major class of phenolic plant secondary metabolites, belonging to the chlorogenic acid family. Various health-beneficial properties of HCQAs have been shown, which has drawn interest for HCQA profiling in plants of human consumption. However, this task remains challenging, because several isomeric HCQAs can be present in the sample with identical molecular formulae and the limited availability of reference standards poses additional challenges to their identification. In the present work, a high performance liquid chromatography-electrospray ionization-quadrupole time-of-flight-mass spectrometry (HPLC-ESI-Q/TOF-MS) method accompanied with an effective data filtering protocol is presented, which is shown to be suitable for the identification of HCQAs in plant materials in a non-targeted manner. Both collision-induced dissociation (CID) fragmentation performed in a collision cell and in-source (CID) fragmentation were used to produce accurate mass fragments. It was shown that fragmentation characteristics required for identification of regio-isomers of HCQAs can be achieved with in-source CID fragmentation, enabling the use of a single-stage MS system with in-source fragmentation for convincing identification of HCQAs. Based on a thorough validation of identified HCQA compounds using coffee bean extracts as reference samples, comprehensive profiling of HCQAs in two apricot (Prunus armeniaca L.) genotypes ('Preventa' and 'Gonci magyarkajszi') was carried out for the first time and the following 10 HCQAs were shown to be present in apricot fruit: 3-caffeoylquinic acid (CQA), cis-3-CQA, 4-CQA, 5-CQA, cis-5-CQA, 3,5-diCQA, 3-p-coumaroylquinic acid (pCoQA), 4-pCoQA, 3-feruloylquinic acid (FQA) and cis-3-FQA. Copyright (c) 2016 John Wiley & Sons, Ltd. HIGHLIGHTS: An HPLC-ESI-Q/TOF-MS method suitable for the identification of hydroxycinnamoyilquinic acids (HCQAs) in plant material in a non-targeted manner was developed. Single-stage, high-resolution MS system with in-source fragmentation was shown to be suitable for convincing identification of HCQAs. Comprehensive profiling of HCQAs in two apricot (Prunus armeniaca L.) genotypes was carried out for the first time. Copyright (c) 2016 John Wiley & Sons, Ltd.

Analysis of matrix-assisted laser desorption/ionization quadrupole time-of-flight collision-induced dissociation spectra of simple precursor ions and isobaric oligosaccharide ion mixtures based on product ion intensities and pattern recognition.[Pubmed:28299859]

Rapid Commun Mass Spectrom. 2017 May 30;31(10):873-885.

RATIONALE: Qualitative analysis of glycomic tandem mass spectrometry (MS/MS) data based on m/z values of product ions alone is widely used, and often sufficient for analysis of single analytes. However, most complex glycomic mixtures contain multiple isobaric oligosaccharides, in which case this approach is often limited. Here we show how ion intensity information can be used in order to enhance MS/MS data analysis, and extract both qualitative and semiquantitative information from complex glycomic MS/MS datasets. METHODS: A matrix-assisted laser desorption/ionization quadrupole time-of-flight (MALDI QTOF) mass spectrometer was used in this study. We compared the intensities of product ions within a single product ion series, determined by their length, across the whole glycomic MS/MS dataset. In order to detect discernable patterns, the intensity data was normalized to the intensity of each product ion within the series. In most cases, normalized data yielded easily discernable patterns, relevant either for analysis of specific glycomic structure types, or mechanistic MS studies. RESULTS: We used our approach on a glycomic sample consisting of human milk oligosaccharides. The approach yielded useful results for both qualitative and semiquantitative analyses. All normalizations performed were not equally rich in information and the information content of generated tables was not possible to predict. These analyses were shown to be independent of instrument manufacturer. CONCLUSIONS: Our approach enabled more detailed qualitative analysis of MS/MS spectra of precursor ions containing isobaric oligosaccharide structures. While limited semiquantitative information could be extracted from the raw data as well, the accuracy of this method should be significantly enhanced when standard calibration mixtures can be prepared. Copyright (c) 2017 John Wiley & Sons, Ltd.

Gas-Phase Fragmentation Behavior of Oxidized Prenyl Peptides by CID and ETD Tandem Mass Spectrometry.[Pubmed:27785692]

J Am Soc Mass Spectrom. 2017 Apr;28(4):704-707.

Farnesylation and geranylgeranylation are the two types of prenyl modification of proteins. Prenylated peptides are highly hydrophobic and their abundances in biological samples are low. In this report, we studied the oxidized prenylated peptides by electrospray ionization mass spectrometry and identified them by collision-induced dissociation (CID) and electron-transfer dissociation (ETD) tandem mass spectrometry. Modified prenyl peptides were generated utilizing strong and low strength oxidizing agents to selectively oxidize and epoxidize cysteine sulfur and prenyl side chain. We selected three peptides with prenyl motifs and synthesized their prenylated versions. The detailed characteristic fragmentations of oxidized and epoxidized farnesylated and geranylgeranylated peptides were studied side by side with two popular fragmentation techniques. CID and ETD mass spectrometry clearly distinguished the modified version of these peptides. ETD mass spectrometry provided sequence information of the highly labile modified prenyl peptides and showed different characteristic fragmentations compared with CID. A detailed fragmentation of modified geranylgeranylated peptides was compared by CID and ETD mass spectrometry for the first time. Graphical Abstract .

Modeling collision energy transfer in APCI/CID mass spectra of PAHs using thermal-like post-collision internal energy distributions.[Pubmed:27802636]

J Chem Phys. 2016 Oct 28;145(16):164311.

The internal energy transferred when projectile molecular ions of naphthalene collide with argon gas atoms was extracted from the APCI-CID (atmospheric-pressure chemical ionization collision-induced dissociation) mass spectra acquired as a function of collision energy. Ion abundances were calculated by microcanonical integration of the differential rate equations using the Rice-Ramsperger-Kassel-Marcus rate constants derived from a UB3LYP/6-311G+(3df,2p)//UB3LYP/6-31G(d) fragmentation mechanism and thermal-like vibrational energy distributions pME,Tchar. The mean vibrational energy excess of the ions was characterized by the parameter Tchar ("characteristic temperature"), determined by fitting the theoretical ion abundances to the experimental breakdown graph (a plot of relative abundances of the ions as a function of kinetic energy) of activated naphthalene ions. According to these results, the APCI ion source produces species below Tchar = 1457 K, corresponding to 3.26 eV above the vibrational ground state. Subsequent collisions heat the ions up further, giving rise to a sigmoid curve of Tchar as a function of Ecom (center-of-mass-frame kinetic energy). The differential internal energy absorption per kinetic energy unit (dEvib/dEcom) changes with Ecom according to a symmetric bell-shaped function with a maximum at 6.38 +/- 0.32 eV (corresponding to 6.51 +/- 0.27 eV of vibrational energy excess), and a half-height full width of 6.30 +/- 1.15 eV. This function imposes restrictions on the amount of energy that can be transferred by collisions, such that a maximum is reached as kinetic energy is increased. This behavior suggests that the collisional energy transfer exhibits a pronounced increase around some specific value of energy. Finally, the model is tested against the CID mass spectra of anthracene and pyrene ions and the corresponding results are discussed.

Identification of small-molecule inhibitors of the colorectal cancer oncogene Kruppel-like factor 5 expression by ultrahigh-throughput screening.[Pubmed:21885866]

Mol Cancer Ther. 2011 Nov;10(11):2043-51.

The transcription factor Kruppel-like factor 5 (KLF5) is primarily expressed in the proliferative zone of the mammalian intestinal epithelium, where it regulates cell proliferation. Studies showed that inhibition of KLF5 expression reduces proliferation rates in human colorectal cancer cells and intestinal tumor formation in mice. To identify chemical probes that decrease levels of KLF5, we used cell-based ultrahigh-throughput screening (uHTS) to test compounds in the public domain of NIH, the Molecular Libraries Probe Production Centers Network library. The primary screen involved luciferase assays in the DLD-1/pGL4.18hKLF5p cell line, which stably expressed a luciferase reporter driven by the human KLF5 promoter. A cytotoxicity counterscreen was done in the rat intestinal epithelial cell line, IEC-6. We identified 97 KLF5-selective compounds with EC(50) < 10 mumol/L for KLF5 inhibition and EC(50) > 10 mumol/L for IEC-6 cytotoxicity. The two most potent compounds, CIDs (PubChem Compound IDs) 439501 and 5951923, were further characterized on the basis of computational, Western blot, and cell viability analyses. Both of these compounds, and two newly synthesized structural analogs of CID 5951923, significantly reduced endogenous KLF5 protein levels and decreased viability of several colorectal cancer cell lines without any apparent impact on IEC-6 cells. Finally, when tested in the NCI-60 panel of human cancer cell lines, compound CID 5951923 was selectively active against colon cancer cells. Our results show the feasibility of uHTS in identifying novel compounds that inhibit colorectal cancer cell proliferation by targeting KLF5.