Survival Yield Method Research Articles

Oxygen Atom from Carbonyl Group as an Important Binding Agent to the G-Quadruplex - Study Case of Flavonoids.

In the field of anticancer therapy study it is of great interest to find effective G-quadruplex ligands which may be of potential use in medical treatment or cancer prevention. Since among the compounds of natural origin, flavonoids have attracted notable attention because of their unique properties and promising therapeutic applications, an interesting question was to identify the flavonoid structural features that could provide effective binding properties toward G-quadruplex. By using electrospray ionization mass spectrometry, followed by the survival yield method, it has been shown that the flavonoid molecules which contain an available C4=O carbonyl group form more stable adducts with G-tetrads than the other ones. Molecular docking has shown that C4=O carbonyl group can be a source of hydrogen bonds and/or π-stacking interactions. Therefore, the flavonoid molecules which contain an available C4=O carbonyl group can be regarded as good binders of G-quadruplexes.

Study of protonated dimers of cytosine, cytidine, and deoxycytidine using survival yield method and quantum mechanics calculations

RationaleCytosine and its conjugates are prone to form protonated, triply‐bonded dimers. Therefore, the nucleic‐acid cytosine‐rich sequence forms the four‐stranded noncanonical secondary structure known as the intercalated motif (i‐motif). This process has resulted in studies on cytosine protonated dimers. This communication focuses on the protonated dimers of cytosine and its nucleoside using the survival yield (SY) method and quantum mechanics calculations.MethodsTo obtain the precursor ion fragmentation curve, the plot of SY against Ecomδ, the product ion spectra of the protonated dimers were obtained using a Waters/Micromass Q‐TOF Premier mass spectrometer. Quantum mechanics calculations were performed using GAUSSIAN 16, and full geometry optimizations and energy calculations were performed within the density functional theory framework at B3LYP/6‐31G(d,p).ResultsThe precursor ion fragmentation curve allowed the rating of the gas‐phase stabilities of the analyzed protonated dimers. Substitution of sugar moiety at N1 cytosine atom decreased the gas‐phase stabilities of the protonated dimers. The deoxycytidine dimer was found to be more stable than the cytidine dimer and cytidine–deoxycytidine dimer. Quantum chemical calculations indicated that cytosine aminohydroxy tautomer may be involved in the formation of protonated cytosine–cytosine nucleoside dimers but not in the formation of cytosine dimers.ConclusionsThe results obtained for nucleoside dimers indicated that the SY method may reflect the i‐motif stabilities observed under physiological conditions. Therefore, the analysis of other protonated dimers of variously substituted cytosine–cytosine nucleoside using the SY method may be important to study the effect of cytosine substitution on the i‐motif stabilities. Cytosine tautomer containing C2‐OH… N(2H)‐C4 moiety may be involved in the formation of protonated cytosine–cytosine nucleoside dimers.

Binding of Quercetin Derivatives toward G-Tetrads as Studied by the Survival Yield Method.

Recently, much interest has been devoted to finding effective G-quadruplex ligands, both of synthetic or natural origins, which may be of potential use in the field of cancer therapy. Among compounds of natural origin, a common flavonol quercetin has attracted notable attention. Yet, only a modest number of papers have been concerned with a comparison of quercetin conjugates binding to G-quadruplexes. In this study, we applied the survival yield (SY) method in order to compare the stability of G-tetrad complexes with quercetin and its conjugates, namely, 3-O-glycosides and O-methylated conjugates. According to the determined values of Ecomδ50, flavonol glycosides bind most effectively with G-tetrads, whereas, among flavonols, 3-O-methylquercetin makes the most effective bonds. Because the aglycone structure is of crucial importance for biological processes, 3-O-methylquercetin seems to be a suitable candidate for anticancer therapeutics, and the extracts from the plants, which contain high amounts of 3-O-methylquercetin or its glycosides, should be considered as interesting materials for preparation of pharmaceuticals or dietary supplements.

Exploring Glycosylated Soy Isoflavones Affinities toward G-tetrads as Studied by Survival Yield Method.

Taking soy-based food supplements for menopausal symptoms by women may reduce the risk of cancer. Therefore, the interaction between nucleic acids (or their constituents) and ingredients of the supplements, e. g., isoflavone glucosides, on the molecular level, has been of interest with respect to cancer therapy. In this work, the interaction between isoflavone glucosides and G-tetrads, namely [4G+Na]+ ions (G stands for guanosine or deoxyguanosine), were analyzed by using electrospray ionization-collision induced dissociation-mass spectrometry (ESI-CID-MS) and survival yields method. The strength of isoflavone glucosides-[4G+Na]+ interaction in the gas phase was determined from Ecom50 - the energy required to fragment 50 % of selected precursor ions. Glycitin-[4G+Na]+ interaction was found to be the strongest, and the interaction between isoflavone glucosides and guanosine tetrad was established to be stronger than that between isoflavone glucosides and deoxyguanosine tetrad.

Differences in the internal energies of ions in electrospray ionization mass spectrometers equipped with capillary-skimmer and capillary-RF lens interfaces.

Small metabolites are commonly analyzed using electrospray ionization mass spectrometry (ESI-MS). Although the protonated form of a compound of interest is typically the target ion in ESI-MS, the protonated forms of small metabolites occasionally undergo fragmentation during ion transmission from ambient conditions to vacuum conditions, hindering the unambiguous identification of analyte molecules. To estimate the fragmentation efficiency during ESI processes, the internal energy distribution of the ions (P(E)) must be evaluated. The common approach for the P(E) evaluation is the survival yield method, which uses thermometer ions. In this study, the P(E) of ions produced by an ESI source in a commercial triple quadrupole mass spectrometer equipped with a capillary-skimmer and capillary-RF lens interfaces was evaluated using benzyl ammonium thermometer ions. Furthermore, this study proposes the use of 3-(aminomethyl)indole and related compounds, which have the lowest Eapp values among the reported thermometer ions, to obtain P(E) values of the ions more accurately. Results showed that P(E) strongly depends on whether a capillary-skimmer interface or capillary-RF lens interface was used for ion transport to the vacuum. ESI-MS with a capillary-skimmer interface provided a considerably lower and narrower P(E) of ions than that with a capillary-RF lens interface, thereby producing intact protonated molecules without significant fragmentation of most small metabolites. However, ESI-MS equipped with capillary-RF lens interfaces provided a higher efficiency of ion transmission than ESI-MS equipped with a capillary-skimmer interface, allowing for highly sensitive analysis of metabolites.

Fragmentation of Protonated Histamine and Histidine by Electrospray Ionization In-Source Collision-Induced Dissociation

Electrospray ionization (ESI) generally produces intact gas-phase ions without extensive fragmentation; however, for histamine and histidine, ESI provides fragment ions through in-source collision-induced dissociation (CID). In this study, we investigated the fragmentation of these compounds both experimentally and using density functional theory calculations. We found that histamine undergoes protonation with subsequent NH3 loss by ESI in-source CID. The corresponding fragmentation mainly produces bicyclo and spiro compounds. In contrast, the ESI in-source CID of protonated histidine preferentially results in H2O loss rather than NH3 loss. However, the corresponding fragment ion is not observed in the ESI mass spectrum of histidine, because it undergoes further CO loss within 100 ps. Consequently, protonated histidine produces a fragment ion arising from a 46 Da loss, which corresponds to the masses of H2O and CO, by ESI in-source CID. The fragment ion yields of histamine and histidine produced by ESI in-source CID are then estimated from the dissociation rate constant and internal energy of the analyte ion, respectively. The dissociation rate constant and internal energy of the analyte ion were determined by double-hybrid density functional theory calculations and the survival yield method using benzylpyridinium thermometer ions, respectively. Because intense fragment ion signals are present in the ESI mass spectrum, the analysis of the fragment ions produced by ESI in-source CID facilitates the identification of metabolites originating from aromatic amino acids, such as histamine.

Study of Substituted Phenethylamine Fragmentation Induced by Electrospray Ionization Mass Spectrometry and Its Application for Highly Sensitive Analysis of Neurotransmitters in Biological Samples.

Although liquid chromatography-tandem mass spectrometry (LC-MS/MS) equipped with electrospray ionization (ESI) is widely employed for metabolite analysis, substituted phenethylamines commonly undergo fragmentation during ESI in-source collision-induced dissociation (CID). Unexpected fragmentation hampers not only unambiguous identification but also accurate metabolite quantification. ESI in-source CID induces N-Cα bond dissociation in substituted phenethylamines lacking a β-hydroxy group to produce fragment ions with a spiro[2.5]octadienylium motif. In contrast, phenethylamines with a β-hydroxy group generate substituted 2-phenylaziridium through ESI in-source CID-induced H2O loss. The fragment ion yield produced by ESI in-source CID can be estimated by the dissociation rate constant and internal energy of the analyte ion, determined by employing density functional theory calculations and the survival yield method using a thermometer ion, respectively. Fragmentation is strongly enhanced by the presence of an β-hydroxy group, whereas N-methylation suppresses fragmentation. In particular, octopamine and noradrenaline, which contain an β-hydroxy and primary amine groups, produce more intense fragment ion signals than protonated molecules. Regarding the quantitative analysis of phenethylamines present in the mouse brain, the noradrenaline fragment ion used as the precursor in multiple reaction monitoring (MRM) provided a higher signal-to-noise ratio in the resulting spectra than protonated noradrenaline. The present method allows for the quantitative analysis of substituted phenethylamines with high sensitivity.

Comparison of internal energy distributions generated by supercritical fluid chromatography versus liquid chromatography hyphenated with electrospray high resolution mass spectrometry

The internal energy distributions of thermometer ions dissolved in supercritical and liquid solvents and produced in the gas phase by electrospray (ESI) were measured and compared by the survival yield method. The influence of different chromatographic conditions such as the nature of solvents, the composition of the mobile phase, the pressure of the back pressure regulator was studied for supercritical fluid chromatography (SFC) whereas the influence of the composition and of the flow rate of the mobile phase was investigated for liquid chromatography (LC). The MS instrumental parameters were studied in parallel for SFC and LC showing that the drying gas temperature and the fragmentor voltage affected the internal energy distribution, whereas the capillary voltage did not modify the internal energy distribution. A comparison of the internal energy distributions generated by SFC and LC was carried out leading to conclude that SFC led to higher internal energy, i.e. more in-source fragment ions, than LC.

Internal Energy Deposition in Infrared Matrix-Assisted Laser Desorption Electrospray Ionization With and Without the Use of Ice as a Matrix.

The internal energy deposited into analytes during the ionization process largely influences the extent of fragmentation, thus the appearance of the resulting mass spectrum. The internal energy distributions of a series of para-substituted benzyl pyridinium cations in liquid and solid-state generated by infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) were measured using the survival yield method, of which results were subsequently compared with conventional electrospray ionization (ESI). The comparable mean internal energy values (e.g., 1.8-1.9eV at a collision energy of 15eV) and peak widths obtained with IR-MALDESI and ESI support that IR-MALDESI are essentially a soft ionization technique where analytes do not gain considerable internal energy during the laser-induced desorption process and/or lose energy during uptake into charged electrospray droplets. An unusual fragment ion, protonated pyridine, was only found for solid IR-MALDESI at relatively high collision energies, which is presumably resulted from direct ionization of the pre-charged analytes in form of salts. Analysis of tissue with an ice layer consistently yielded ion populations with higher internal energy than its counterpart without an ice layer, likely due to a substantially enhanced number of IR absorbers with ice. Further measurements with holo-myoglobin show that IR-MALDESI-MS retains the noncovalently bound heme-protein complexes under both native-like and denaturing conditions, while complete loss of the heme group occurred in denaturing ESI-MS, showing that the softness of IR-MALDESI is equivalent or superior to ESI for biomolecules.

Establishment of a Charge Reversal Derivatization Strategy to Improve the Ionization Efficiency of Limaprost and Investigation of the Fragmentation Patterns of Limaprost Derivatives Via Exclusive Neutral Loss and Survival Yield Method.

Sensitivity is generally an issue in bioassays of prostaglandins and their synthetic analogs due to their extremely low concentration in vivo. To improve the ionization efficiency of limaprost, an oral prostaglandin E1 (PGE1) synthetic analog, we investigated a charge reversal derivatization strategy in electrospray ionization mass spectrometry (ESI-MS). We established that the cholamine derivative exhibits much greater signal intensity in the positive-ion mode compared with limaprost in the negative ion mode. Collision-induced dissociation (CID) involved exclusive neutral mass loss and positive charge migration to form stable cationic product ions with the positive charge on the limaprost residue rather than on the modifying group. This has the effect of maintaining the efficiency and specificity of multiple reaction monitoring (MRM) and avoiding cross talk. CID fragmentation patterns of other limaprost derivatives allowed us to relate the dissociation tendency of different neutral leaving groups to an internal energy distribution scale based on the survival yield method. Knowledge of the energy involved in the production of stabilized positive ions will potentially assist the selection of suitable derivatization reagents for the analysis of a wide variety of lipid acids. Graphical Abstract ᅟ.

How Hot are Your Ions Really? A Threshold Collision-Induced Dissociation Study of Substituted Benzylpyridinium "Thermometer" Ions.

The first absolute experimental bond dissociation energies (BDEs) for the main heterolytic bond cleavages of four benzylpyridinium "thermometer" ions are measured using threshold collision-induced dissociation in a guided ion beam tandem mass spectrometer. In this experiment, substituted benzylpyridinium ions are introduced into the apparatus using an electrospray ionization source, thermalized, and collided with Xe at varied kinetic energies to determine absolute cross-sections for these reactions. Various effects are accounted for, including kinetic shifts, multiple collisions, and internal and kinetic energy distributions. These experimentally measured 0K BDEs are compared with computationally predicted values at the B3LYP-GD3BJ, M06-GD3, and MP2(full) levels of theory with a 6-311+G(2d,2p) basis set using vibrational frequencies and geometries determined at the B3LYP/6-311+G(d,p) level. Additional dissociation pathways are observed for nitrobenzylpyridinium experimentally and investigated using these same levels of theory. Experimental BDEs are also compared against values in the literature at the AM1, HF, B3LYP, B3P86, and CCSD(T) levels of theory. Of the calculated values obtained in this work, the MP2(full) level of theory with counterpoise corrections best reproduces the experimental results, as do the similar literature CCSD(T) values. Lastly, the survival yield method is used to determine the characteristic temperature (Tchar) of the electrospray source prior to the thermalization region and to confirm efficient thermalization. Graphical Abstract ᅟ.

Internal energy deposition for low energy, femtosecond laser vaporization and nanospray post-ionization mass spectrometry using thermometer ions.

The internal energy of p-substituted benzylpyridinium ions after laser vaporization using low energy, femtosecond duration laser pulses of wavelengths 800 and 1042nm was determined using the survival yield method. Laser vaporization of dried benzylpyridinium ions from metal slides into a buffered nanospray with 75μJ, 800nm laser pulses resulted in a higher extent of fragmentation than conventional nanospray due to the presence of a two-photon resonance fragmentation pathway. Using higher energy 800nm laser pulses (280 and 505μJ) led to decreased survival yields for the four different dried benzylpyridinium ions. Analyzing dried thermometer ions with 46.5μJ, 1042nm pulse-bursts resulted in little fragmentation and mean internal energy distributions equivalent to nanospray, which is attributable to the absence of a two-photon resonance that occurs with higher energy, 800nm laser pulses. Vaporization of thermometer ions from solution with either 800nm or 1042nm laser pulses resulted in comparable internal energy distributions to nanospray ionization.

Detailed characterization of poly(2‐ethyl‐2oxazoline)s by energy variable collision‐induced dissociation study

Poly(2-oxazoline)s are important polymers and can be considered as pseudo-peptides which makes them important for biomedical and life science applications. This prompts the need for a detailed characterization of these polymers via different analytical tools such as mass spectrometry. Here, the energy-variable collision-induced dissociation (CID) of poly(2-ethyl-2-oxazoline)s was studied by electrospray ionization quadrupole time-of-flight tandem mass spectrometry (ESI-QTOFMS/MS) to gain further structural information about this polymer type. All polymers were analyzed using manual flow injection of samples into an ESI-QTOF mass spectrometer. Mass spectra (MS and MS/MS) were obtained in the positive ion mode over a mass-to-charge (m/z) range from 50 to 3000. The dependency of the fragmentation patterns as a function of collision energy was examined and the characteristic collision energy (CCE or CE50 ) values for various poly(2-ethyl-2-oxazoline)s with different end-groups were calculated. The effect of molar masses on the CCE values was investigated via the survival yield (SY) method and a linear relationship between CCE values and the degree of polymerization for the PEtOx polymers was observed. This study showed that ESI-MS/MS is very useful for differentiating poly(2-ethyl-2-oxazoline)s with different end-groups by varying the collision energy. The SY method has the potential to determine the importance of the end-groups on the fragmentation behavior of this polymer type.

Collision‐induced dissociation study of poly(2‐ethyl‐2‐oxazoline) using survival yields and breakdown curves

Poly(2-ethyl-2-oxazoline), a synthetic polymer was analysed by mass spectrometry using different ion sources. Two distributions could be identified in the mass spectra which related to two different polymer series (one with hydrogen and hydroxyl end-groups and the other with methyl and hydroxyl end-groups). The fragmentation behaviour of the protonated oligomers was studied in a quadrupole time-of-flight mass spectrometer (MS) with electrospray, atmospheric pressure chemical ionization and direct analysis in real time soft ionization techniques. Three product ion series were identified in the MS/MS spectra independently of the ion source used. Based on the results, a mechanism was proposed for the dissociation by means of the accurate mass of the product ions, pseudo MS(3) experiments and the energy dependence of the product ion intensity, i.e. breakdown curves. The survival yield method was used to highlight the correlation between the size of the oligomers and the laboratory frame collision energy.

Laser-Induced Thermal Desorption Facilitates Postsource Decay of Peptide Ions

We investigated the thermal mechanism involved in laser desorption/ionization (LDI) of thermally labile molecules from the flat surfaces of amorphous Si (a-Si) and crystalline Si (c-Si). a-Si was selected for this study because of its thermal property, such as low thermal conductivity; thus, it was predicted to be highly susceptible to laser-induced surface heating. By virtue of lack of surface nanostructures, the flat surfaces offer a simple model system to focus on the thermal mechanism, avoiding other effects, including possible non-thermal contributions that can arise from the physical existence of surface nanostructures. For the energetics study, the internal energies of substituted benzylpyridinium ions produced by LDI on the bare and coated surfaces of a-Si and c-Si were obtained using the survival yield method. The results, including LDI thresholds, ion yields, and internal energies all suggested that the LDI mechanism would be indeed thermal, which is most likely promoted by thermal desorption caused by laser-induced surface heating. In addition, the LDI process driven by laser-induced thermal desorption (LITD) was also found to be capable of depositing an excessive internal energy in resulting LDI ions, which underwent a dissociation. It exhibited the essentially same features as in postsource decay (PSD) in MALDI-TOF/TOF mass spectrometry. We report that the LDI process by LITD offers not only a way of intact ionization but also a facile means for PSD of peptide ions, which this work demonstrates is well suited to peptide sequencing using TOF/TOF mass spectrometry.

Internal energy deposition and ion fragmentation in atmospheric-pressure mid-infrared laser ablation electrospray ionization

Mid-infrared laser ablation of water-rich targets at the maximum of the 2.94 μm absorption band is a two-step process initiated by phase explosion followed by recoil pressure induced material ejection. Particulates and/or droplets ejected by this high temperature high pressure process can be ionized for mass spectrometry by charged droplets from an electrospray. In order to gauge the internal energy introduced in this laser ablation electrospray ionization (LAESI®) process, we apply the survival yield method and compare the results with electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI). The results indicate that LAESI yields ions with internal energies indistinguishable from those produced by ESI. This finding is consistent with the recoil pressure induced ejection of low micrometre droplets that does not significantly change the internal energy of solute molecules.

Non‐thermal internal energy distribution of ions observed in an electrospray source interfaced with a sector mass spectrometer

The internal energy distribution P(E(int)) of ions emitted in an electrospray (ESI) source interfaced with a sector mass spectrometer is evaluated by using the experimental survival yield (SY) method including the kinetic shift. This method is based on the relationship between the degree of fragmentation of an ion and its amount of internal energy and uses benzylpyridinium cations due to their simple fragmentation scheme. Quantum chemical calculations are performed, namely at G3(MP2)//B3LYP and QCISD/MP2 levels of theory. The results show that the internal energy distribution of the ions emitted in the ESI source interfaced with a sector analyzer is very narrow. The MassKinetics software is used to confirm these observations. The P(E(int)) is the parameter that allows to fit the experimental SY of each substituted benzylpyridinium cation with theoretical mass spectra generated by the MassKinetics software. The resulting internal energy distributions are similar to the ones obtained with the experimental SY method. This indicates that in the present experimental conditions, P(E(int)) cannot be compared with a 'thermal-like' Boltzmann distribution. In addition, it appears that with the sector analyzer, increasing the collision energy in the first pumping stage of the ESI source does not correspond to a warm-up of the produced ions.

Energy-Dependent Collision-Induced Dissociation of Lithiated Polytetrahydrofuran: Effect of the Size on the Fragmentation Properties

The fragmentation properties of singly and doubly lithiated polytetrahydrofuran (PTHF) were studied using energy-dependent collision-induced dissociation. The product ion spectrum of [PTHF + Li](+) showed the formation of three different series corresponding to product ions with hydroxyl, aldehyde and vinyl end-groups. Interestingly, besides these series, two additional, non-lithiated product ions C(4)H(9)O(+) and C(4)H(7)(+) were identified in the MS/MS spectra. The MS/MS of the doubly lithiated PTHF ([PTHF + 2Li](2+)) with a number of repeat units ranging from 8 to 27 showed the formation of product ions similar to those of the singly lithiated series, however, doubly lithiated product ions and product ions formed by the loss of one Li(+)-ion from the precursor ion also appeared with significant abundances. Analysis of the breakdown curves for the singly and doubly charged PTHF indicated that the series A ions are formed most probably together with the series B ions, while members of the series C ions appeared at significantly higher collision energies. The fragmentation properties of [PTHF + Li](+) and [PTHF + 2Li](2+) were also interpreted using the survival yield method. It was found that the collision energy/voltage necessary to obtain 50% fragmentation (CV(50)) was dependent linearly on the number of the repeat units, i.e., on the size, or the number of degrees of freedom (DOF).

Fragmentation of Benzylpyridinium “Thermometer” Ions and Its Effect on the Accuracy of Internal Energy Calibration

Electrospray ionization mass spectrometry (ESI-MS) is a powerful analytical method to study biomolecules and noncovalent complexes. The prerequisite for their intact observation is soft ionization. In ESI, the internal energy of ions is primarily influenced by collisional activation in the source. The survival yield method is frequently used to probe the energy deposition in ions during the electrospray process. In the present work, we investigate the fragmentation pathways of para-substituted benzylpyridinium ions, the most widely used "thermometer ions" in the survival yield method. In addition to the C-N bond cleavage, alternative fragmentation channels were found for the compounds studied. We consider these pathways to result from intramolecular rearrangements. The effect of these additional fragments on the accuracy of the internal energy calibration is estimated for both collision-cell and in-source collision-induced dissociation (CID). Altogether, results presented suggest that a correction of the energy scale is necessary for the method based on benzylpyridinium ions to precisely quantify ion internal energies.

Internal energy distribution of peptides in electrospray ionization : ESI and collision‐induced dissociation spectra calculation

The internal energy of ions and the timescale play fundamental roles in mass spectrometry. The main objective of this study is to estimate and compare the internal energy distributions of different ions (different nature, degree of freedom 'DOF' and fragmentations) produced in an electrospray source (ESI) of a triple-quadrupole instrument (Quattro I Micromass). These measurements were performed using both the Survival Yield method (as proposed by De Pauw) and the MassKinetics software (kinetic model introduced by Vékey). The internal energy calibration is the preliminary step for ESI and collision-induced dissociation (CID) spectra calculation. meta-Methyl-benzylpyridinium ion and four protonated peptides (YGGFL, LDIFSDF, LDIFSDFR and RLDIFSDF) were produced using an electrospray source. These ions were used as thermometer probe compounds. Cone voltages (V(c)) were linearly correlated with the mean internal energy values (<E(int) >) carried by desolvated ions. These mean internal energy values seem to be slightly dependent on the size of the studied ion. ESI mass spectra and CID spectra were then simulated using the MassKinetics software to propose an empirical equation for the mean internal energy (<E(int) >) versus cone voltage (V(c)) for different source temperatures (T): < E(int) > = [405 x 10(-6) - 480 x 10(-9) (DOF)] V(c)T + E(therm)(T). In this equation, the E(therm)(T) parameter is the mean internal energy due to the source temperature at 0 V(c).