The role of free electrons in MALDI: electron capture by molecules of alpha-cyano-4-hydroxycinnamic acid

Low-energy (0-12 eV) electron attachment to molecules of a typical matrix substance used for matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS), namely alpha-cyano-4-hydroxicynnamic acid, has been investigated in the gas phase at different temperatures ranging from 140 degrees C to 260 degrees C by means of electron capture negative-ion mass spectrometry (ECNI MS). The yield of negative ions, formed by electron capture, was measured as a function of incident electron energy for four different temperatures. The long-lived parent molecular anion, [M]- (m/z 189), was observed in the negative-ion mass spectra of the substance under investigation. Its autodetachment lifetime was estimated to be approximately 600 micros. It was found that at 140 degrees C the main decay channel of the long-lived temporary molecular anion of alpha-cyano-4-hydroxicynnamic acid is a formation of the [M-COOH]-; fragment negative ion (m/z 144) with an intensity of 37.2% in percentage terms in respect of the total anion current. There are also [M-H]-, [M-CO2]- and [CN]- fragments in the spectra with intensities of about 7.7%, 21.6% and 3.1% at 140 degrees C. It was shown that the escape of the CO2 molecule from the parent molecular anion is a slow process. It takes [M]- about 10 micros to decay on carbon dioxide molecules and [M-CO2]- fragment anions. Increasing the temperature of the target molecule alters the negative-ion mass spectra of alpha-cyano-4-hydroxicynnamic acid significantly. A possible role for the findings in typical MALDI MS experiments is discussed.     

Synthesis of poly(ether esters) from reaction of alpha-cyano-4-hydroxycinnamic acid and group IVB metallocenes

Poly(ether esters) are rapidly synthesized in moderate yield employing the interfacial polycondensation reaction system from the reaction of alpha-cyano-4-hydroxycinnamic acid and Group IVB metallocenes. The products are high polymers. Infrared spectroscopy shows the formation of new bands derived from the M-O and M-O(CO) linkages. It also shows that the products exist as alternating M-O and M-O(CO) linkages. The products show outstanding inhibition of a variety of cancer cell lines including two pancreatic cancer cell lines. EC₅₀ values for the polymers are in the nanogram/mL range. The ability to inhibit the cancer cell lines is generally Hf>Zr>Ti. Thus, future synthesis and testing might consider using compounds containing hafnocene and zirconocene in addition to the titanocene moiety.     

Improved sensitivity for insulin in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry by premixing alpha-cyano-4-hydroxycinnamic acid matrix with transferrin

This report describes an enhancement of the signal intensities of proteins and peptides in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). When alpha-cyano-4-hydroxycinnamic acid (CHCA) premixed with human transferrin (Tf) was used as a matrix, the signal intensity of insulin was amplified to more than ten times that of the respective control in CHCA without Tf. The detection limit of insulin was 0.39 fmol on-probe in the presence of Tf, while it was 6.3 fmol in the absence of Tf. The signal intensity of insulin was also enhanced when the CHCA matrix was premixed with proteins other than Tf (80 kDa), such as horse ferritin (20 kDa), bovine serum albumin (BSA, 66 kDa), or human immunoglobulin G (150 kDa). The optimum spectrum of insulin was obtained when the added amount of protein was in the range 0.26-0.62 pmol, regardless of the molecular weight of the added protein. Tf and BSA outperformed the other tested proteins, as determined by improvements in the resulting spectra. When the mass spectra of several peptides and proteins were recorded in the presence of Tf or BSA, the signal intensities of large peptides such as glucagon were enhanced, though those of smaller peptides were not enhanced. In addition, the signal enhancement achieved with Tf and BSA was more pronounced for the proteins, including cytochrome C, than for the large peptides. This enhancement effect could be applied to improve the sensitivity of MALDI-TOFMS to large peptides and proteins.     

Group IVB metallocene poly(ether ester) polymers containing alpha-cyano-4-hydroxycinnamic acid that act as self-matrix materials in MALDI MS

Polymers derived from reacting Group IVB metallocene dihalides and the matrix alpha-cyano-4-hydroxycinnamic acid act as their own matrix agent when performing MALDI MS. Ion fragments containing two repeat units and greater are formed. The results are similar to those obtained employing graphite as the matrix material. The advantage of employing graphite as a comparative standard is that non-interfering ion fragment clusters are not produced by graphite. This is the second report of the inclusion of alpha-cyano-4-hydroxycinnamic acid into a condensation polymer and the use of the polymer itself as a matrix material indicating that this approach can be successfully applied to other systems. The addition of graphite as a matrix material allows the mass range to increase for the reflective mode.     

The thermodynamic properties of 4-pentenoic acid

The enthalpy of formation of liquid 4-pentenoic acid was determined by combustion calorimetry. The vapor pressure and enthalpy of vaporization of the compound were measured by the transfer method over the temperature range 289–324 K. Conformational analysis was performed. The equilibrium structure, fundamental vibrations, moments of inertia, and total energy of the stablest acid conformers were calculated by the B3LYP/6-311G(d, p) and G3MP2 quantum-chemical methods. The experimental IR spectrum and calculated vibrational frequencies were used to assign IR bands. The thermodynamic properties of monomeric 4-pentenoic acid in the ideal gas state were calculated over the temperature range 0–1500 K. Additive and quantum-chemical methods were used to estimate the Δ_f H°(g) and Δ_vap H° values. Close agreement between the calculation results and experimental data was obtained. It was shown that additive and quantum-chemical methods could be used for estimating the enthalpies of formation and vaporization of nonconjugated alkenoic acids.     

Equilibrium sublimation and thermodynamic properties of SnS

Knudsen effusion studies of the sublimation of polycrystalline SnS, prepared by annealing and chemical vapor transport, have been performed employing vacuum micro-balance techniques in the temperature range 733–944 K and at pressures ranging from about 6 × 10^?3 to 11 Pa.The third-law heats of sublimation and second-law entropy of reaction SnS(s) = SnS(g) were determined to be ΔH⁰₂₉₈ = 220.4 ± 3.0 kJ mole^? and ΔS⁰₂₉₈ = 162.4 ± 4.5 J K^?1 mole^?1. From these data the standard heat of formation and absolute entropy of SnS(s) were calculated to be ?102.9 ± 4.0 kJ mole^?1 and 79.9 ± 6.0 J K^?1, respectively.     

Disappearance of interfering alkali-metal adducted peaks from matrix-assisted laser desorption/ionization mass spectra of peptides with serine addition to alpha-cyano-4-hydroxycinnamic acid matrix

It has been described that ion yield in both positive- and negative-ion matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) of peptides is often inhibited by trace amounts of alkali metals and that the MALDI mass spectra are contaminated by the interfering peaks originating from traces of alkali metals, even when sample preparation is carefully performed. Addition of serine to the commonly used MALDI matrix alpha-cyano-4-hydroxycinnamic acid (CHCA) significantly improved and enhanced the signals of both protonated and deprotonated peptides, [M+H](+) and [M-H](-). The addition of serine to CHCA matrix eliminated the alkali-metal ion adducts, [M+Na](+) and [M+K](+), and the CHCA cluster ions from the mass spectra. Serine and serinephosphate as additives to CHCA enhanced and improved the formation of molecular-related ions of phosphopeptides in negative-ion MALDI mass spectra.     

Some thermodynamic parameters for hydroxy amino acids: Bicine and tricine

pK values of N,N-dihydroxyethylglycine (bicine) and N-[tris(hydroxymethyl)methyl]-glycine (tricine) have been determined by the Irving-Rossotti method in an aqueous medium at 25, 30, 35, 40, 45, and 50°C and at different ionic strengths (I = 0.1, 0.5, and 1.0). Plots between pK_a(NH) and 1/T for various ionic strengths have been obtained and the values of slopes have been used to calculate the ΔH, ΔS, and ΔG for the dissociation reactions of bicine and tricine. The ΔH, ΔS, and ΔG values for bicine were found to be 10.6 ± 0.6 kcal mol^?1, ?1.9 ± 1.8 e.u., and 11.1 ± 0.06 kcal mol^?1, respectively, and for tricine 11.2 ± 0.6 kcal mol^?1, 1.6 ± 1.6 e.u., and 10.7 ± 0.06 kcal mol^?1, respectively. The pK_a(NH) values decrease with rise in temperature but the influence of ionic strength is not significant.     

phi-Values beyond the ribosomally encoded amino acids: kinetic and thermodynamic consequences of incorporating trifluoromethyl amino acids in a globular protein

The consequences of the substitution of 4,4,4-trifluorovaline for valine on the folding kinetics and thermodynamics of a globular protein are presented. Variants of the N-terminal domain of L9, a small alpha-beta protein, were prepared in which V3 or V21 was replaced by trifluorovaline. CD and NMR demonstrate that the structure is not perturbed. Both are more stable, the V3 variant by 0.8 kcal mol-1 and the V21 variant by 1.4 kcal mol-1. The increase of stability is significantly larger than that observed in coiled-coils on a per trifluoromethyl group basis. Folding is two-state, and the variants both fold faster than the wild type. The Phi-values are 0.16 and 0.11, respectively.     

Equilibrium sublimation and thermodynamic properties of SnSe and SnSe2

Knudsen effusion studies of the sublimation of polycrystalline SnSe and SnSe₂, prepared by annealing and chemical vapor transport reactions, respectively, have been carried out using vacuum microbalance techniques in the temperature ranges 736–967 K and 608–760 K, respectively. From experimental mass-loss data for the sublimation reaction SnSe(s) = SnSe(g), the recommended values for the heat of formation and absolute entropy of SnSe(s) were calculated to be ΔH°_298,f = ?86.4 ± 9.9 kJ · mol^?1 and S°₂₉₈ = 89.0 ± 7.1 J · K^?1 · mol^?1. From mass-loss data for the decomposition reaction \documentclass{article}\pagestyle{empty}\begin{document}$ {\rm SnSe}_{\rm 2} ({\rm s)} = {\rm SnSe(s)} + \frac{1}{{\rm x}}{\rm Se}_{\rm x} ({\rm g) (x} = 2 - 8) $\end{document}, the recommended values for the heat of formation and absolute entropy of SnSe₂(s) were determined to be ΔH°_298,f = ?118.1 ± 15.1 kJ · mol^?1 and S°₂₉₈ = 111.8 ± 11.8 J · K^?1 mol^?1.     

Abc amino acids: design, synthesis, and properties of new photoelastic amino acids

Photoisomerizable amino acids provide a direct avenue to the experimental manipulation of bioactive polypeptides, potentially allowing real-time, remote control of biological systems and enabling useful applications in nanobiotechnology. Herein, we report a new class of photoisomerizable amino acids intended to cause pronounced expansion and contraction in the polypeptide backbone, i.e., to be photoelastic. These compounds, termed Abc amino acids, employ a photoisomerizable azobiphenyl chromophore to control the relative disposition of aminomethyl and carboxyl substituents. Molecular modeling of nine Abc isomers led to the identification of one with particularly attractive properties, including the ability to induce contractions up to 13 A in the backbone upon trans --> cis photoisomerization. This isomer, designated mpAbc, has substituents at meta and para positions on the inner (azo-linked) and outer rings, respectively. An efficient synthesis of Fmoc-protected mpAbc was executed in which the biaryl components were formed via Suzuki couplings and the azo linkage was formed via amine/nitroso condensation; protected forms of three other Abc isomers were prepared similarly. An undecapeptide incorporating mpAbc was synthesized by conventional solid-phase methods and displayed characteristic azobenzene photochemical behavior with optimal conversion to the cis isomer at 360 nm and a thermal cis --> trans half-life of 100 min at 80 degrees C.     

Hydroxamic acids as weak base indicators: protonation in strong acid media.

The protonation equilibria of N-phenylbenzohydroxamic, benzohydroxamic, salicylhydroxamic, and N-p-tolylcinnamohydroxamic acids have been studied at 25 degrees C in concentrated sulfuric, hydrochloric, and perchloric acid media; the UV-vis spectral measurements were analyzed using the Hammett equation and the Bunnett-Olsen and excess acidity methods. The medium effects observed in the UV spectral curves were corrected with the Cox-Yates and vector analysis methods. The H(A) acidity function based on benzamides provided the best results. The range of variation of the solvation coefficient m is similar to that of amides, this indicating similar solvation requirements for amides and hydroxamic acids. For the same substrate, the observed variations of pK(BH)(+) with the mineral acid used was justified by formation of solvent-separated ion pairs; for the same mineral acid, the observed changes in pK(BH)(+) can be explained by the solvation of BH(+). The change of the pK(BH)(+) values was in reasonably good agreement with the sequence of the catalytic efficiency of the mineral acids used, HCl > H(2)SO(4) > HClO(4).     

Hydration thermodynamic properties of amino acid analogues: a systematic comparison of biomolecular force fields and water models

We present an extensive study on hydration thermodynamic properties of analogues of 13 amino acid side chains at 298 K and 1 atm. The hydration free energies DeltaG, entropies DeltaS, enthalpies DeltaH, and heat capacities Deltac(P)() were determined for 10 combinations of force fields and water models. The statistical sampling was extended such that precisions of 0.3, 0.8, 0.8 kJ/mol and 25 J/(mol K) were reached for DeltaG, TDeltaS, DeltaH, and Deltac(P)(), respectively. The three force fields used in this study are AMBER99, GROMOS 53A6, and OPLS-AA; the five water models are SPC, SPC/E, TIP3P, TIP4P, and TIP4P-Ew. We found that the choice of water model strongly influences the accuracy of the calculated hydration entropies, enthalpies, and heat capacities, while differences in accuracy between the force fields are small. On the basis of an analysis of the hydrophobic analogues of the amino acid side chains, we discuss what properties of the water models are responsible for the observed discrepancies between computed and experimental values. The SPC/E water model performs best with all three biomolecular force fields.     

Delayed extraction experiments using a repulsing potential before ion extraction: evidence of non-covalent clusters as ion precursor in UV matrix-assisted laser desorption/ionization. Part II--Dynamic effects with alpha-cyano-4-hydroxycinnamic acid matrix

Delayed extraction experiments were undertaken to gain a better insight into the dynamic effects involved in the ion formation in UV matrix-assisted laser desorption/ionization. Part I1 was devoted to a 2,5-dihydroxybenzoic (2,5-DHB) matrix. The results clearly demonstrated the existence and the role of high-mass precursors corresponding to a non-covalent matrix-analyte association in ion formation. In this complementary study, ion flight time and abundance were studied as a function of the delay extraction time using the matrix alpha-cyano-4-hydroxycinnamic acid (HCCA). Under our instrumental conditions, where ejected ions experienced a low repulsing electric field before extraction, two main results were obtained: (i) two ion components are observed in the peak profiles depending on the repulsing field, a first, major component (I) similar to that observed for 2,5-DHB and a second, minor component (II) apparently triggered by the delayed extraction pulse, and (ii) ion time-of-flight variation vs delay time remained lower than that noted with 2,5-DHB matrix, indicating that the initial axial velocity is smaller. The initial kinetic energy of matrix and low molecular mass peptide ions for the component I is not high enough to overcome the repulsing potential in the delay time range (200-2200 ns) and we have to assume that ions have non-covalent clusters as precursors. Complete desolvation of these clusters-aggregates would be achieved through the extraction step. Simulations of the ion time-of-flight as a function of the delay time allow the determination of the average size of the precursors, typically 4500, 40000 and 50000 u for HCCA, ACTH 7-38 and bovine insulin quasi-molecular ion, respectively, assuming that the precursors are singly charged. The size of these ion precursors is greater than that of those generated for 2,5-DHB. For component II, ions are probably not solvated and they are directly desorbed from the target. Taking into account the results on HCCA and 2,5-DHB matrices and other results from the literature, a general model for ion formation based on clusters as ion precursors is proposed and discussed.     

Thermal degradation behavior of polyamides having 2- and 4-hydroxycinnamic acid dimer components

Two new types of polyamides having anti head-to-tail hydroxycinnamic acid dimer component in the main chain were synthesized. Their thermal degradation behavior was investigated by TG-DSC analysis in comparison with those of their model compounds and the polyamides derived from anti head-to-head coumarin dimer. The key reactions in the thermal degradation were clarified to be lactonization and cyclic imide formation. The polyamide, derived from 4-hydroxycinnamic acid dimer and hexamethylenediamine, showed good heat stability. © 1993 John Wiley & Sons, Inc.     

Dielectric and vibrational properties of amino acids

We calculate polarizability tensors and normal mode frequencies for the amino acids alanine, leucine, isoleucine, and valine using density functional perturbation theory implemented within the plane wave pseudopotential framework. It is found that the behavior of the electron density under external fields depends to a large extent on the geometrical structure of the molecule in question, rather than simply on the constituent functional groups. The normal modes are able to help distinguish between the different types of intramolecular hydrogen bonding present, and help to explain why leucine is found in the zwitterionic form for the gaseous phase. Calculated IR spectra show a marked difference between those obtained for zwitterionic and nonzwitterionic molecules. These differences can be attributed to the different chemical and hydrogen bonds present. Effective dynamical charges are calculated, and compared to atomic charges obtained from Mulliken population analysis. It is found that disagreement exists, largely due to the differing origins of these quantities.     

Zeolite 4A: heat capacity and thermodynamic properties

The heat capacity of zeolite 4A (also known as LTA, Linde Type A and sodium zeolite A), in the temperature range from 37 to 311 K, is reported. The heat capacity shows no anomalies in this temperature range. Thermodynamic parameters, H, S and G, relative to their values at T=0 K were derived. From these data, we find that zeolite 4A is stabilized by strong enthalpic interactions. Furthermore, its thermodynamic stability results from the strong Si---O and Al---O bonds in the primary building units, with bond strengths very close to those in other similar materials.