Binding Motifs of Carboplatin and Oxaliplatin with Guanine: A Combined MS/MS, IRMPD, and Theoretical Study.

Complexes generated in the gas phase involving the purine nucleobase guanine bound to second and third generation platinum drugs, namely, carboplatin (CarboPt) and oxaliplatin (OxaliPt), were investigated by combining tandem mass spectrometry, collision-induced dissociation (CID), infrared multiple photon dissociation spectroscopy (IRMPD), and density functional theory (DFT) calculations. As the first step, a spectroscopic characterization of the protonated platinum drugs was accomplished. Protonation of both CarboPt and OxaliPt in the gas phase occurs on one of the two carbonyl groups of the cyclobutanedicarboxylate and oxalate ligand, respectively. Such protonation has been postulated by several theoretical studies as a key preliminary step in the hydrolysis of Pt drugs under acidic conditions. Subsequently, the protonated drugs react with guanine in solution to generate a complex of general formula [Pt drug + H + guanine]+, which was then mass-selected. CID experiments provided evidence of the presence of strong binding between guanine and platinum-based drugs within the complexes. The structures of the two complexes have also been examined by comparing the experimental IRMPD spectra recorded in two spectral regions with DFT-computed IR spectra. For each system, the IRMPD spectra agree with the vibrational spectra calculated for the global minimum structures, which present a monodentate complexation of Pt at the N7 position of canonical guanine. This binding scheme is therefore akin to that observed for cisplatin, while other coordination sites yield substantially less stable species. Interestingly, in the case of oxaliplatin, the IRMPD spectra are consistent with the presence of two isomeric forms very close in energy.

Chloride Binding Modulated by Anion Receptors Bearing Tetrazine and Urea.

Modulation and fine-tuning of the strength of weak interactions to bind anions are described in a series of synthetic receptors. The general design of the receptors includes both a urea motif and a tetrazine motif. The synthetic sequence towards three receptors is detailed. Impacts of H-bond strength and linker length between urea and tetrazine on chloride complexation are studied. Binding properties of the chloride anion are examined in both the ground and excited states using a panel of analytical methods (NMR spectroscopy, mass spectrometry, UV/Visible spectroscopies, and fluorescence). A ranking of the receptors by complexation strength has been determined, allowing a better understanding of the structure-properties relationship on these compounds.

Open questions on toxic heavy metals Cd, Hg and Pb binding small components of DNA and nucleobases. Are there any predictable trends?

In this perspective article, we provide a bibliographic compilation of experimental and theoretical work on Cd, Hg, and Pb, and analyze in detail the bonding of M2+ and CH3M+ (M = Zn, Cd, Hg, Pb) with urea and thiourea as suitable models for larger biochemical bases. Through the use of DFT calculations, we have found that although in principle binding energies decrease according to ionic size (Zn2+ > Cd2+ > Pb2+), Hg2+ largely breaks the trend. Through the use of EDA (Energy Decomposition Analysis) it is possible to explain this behavior, which is essentially due to the strong contribution of polarization to the binding. This conclusion is ratified by the NEDA (Natural Energy Decomposition Analysis) formalism, showing that the charge transfer term is very large in all cases, but particularly in the case of the mercury-thiourea system. The general trends observed for the interactions with CH3M+ monocations show however CH3Hg+ binding energies systematically smaller than the CH3Zn+ ones, likely because the relativistic contraction of the Hg orbitals is very much attenuated by the attachment to the methyl group. Finally, we have investigated the gas-phase reactivity between EtHg+ and uracil to compare it with that exhibited by CH3Hg+ and n-ButHg+ previously described in the literature. This comparison gathers new information that highlights the importance of the length of the alkyl chain attached to the metal on the mechanisms of these reactions. For methyl mercury, only the alkyl transfer process is allowed; for butyl mercury, protonation is clearly favored, and for ethyl mercury, both paths are competitive experimentally.

Dereplication of Acetogenins from Annona muricata by Combining Tandem Mass Spectrometry after Lithium and Copper Postcolumn Cationization and Molecular Networks.

Annonaceous acetogenins are natural products held responsible for atypical Parkinsonism due to chronic consumption in traditional medicine or as food, leading to the development of analytical strategies for their complete chemical characterization in complex mixtures. Characterization by tandem mass spectrometry (MS/MS) of acetogenins using collision-induced dissociation from lithium adducts provides additional structural information compared to protonated or sodiated species such as ketone location on the acetogenin backbone. However, very low intensity diagnostic ions together with the lack of extensive structural information regarding position of OH and THF substituents limit this approach. Copper adducts led to diagnostic fragment ions that allow us to identify the position of oxygen rings and hydroxyl substituents. Fragmentation rules were established on the basis of acetogenin standards allowing the identification of 45 over the 77 analogues observed in an extract of Annona muricata by LC-MS/MS using postcolumn infusion of copper sulfate (CuSO4) solution. Molecular networks that were generated thanks to specific fragmentations obtained with copper led to the distinction of THF ring position or to the identification of hydroxylated lactone, for instance.

Discrimination of sulfated isomers of chondroitin sulfate disaccharides by HILIC-MS.

Chondroitin sulfate (CS) glycosaminoglycans are biologically active sulfated polysaccharides that pose an analytical challenge for their structural analysis and functional evaluation. In this study, we developed a hydrophilic interaction liquid chromatography separation method and its on-line coupling to mass spectrometry (MS) allowing efficient differentiation and sensitive detection of mono-, di-, and trisulfated CS disaccharides and their positional isomers, without requiring prior derivatization. The composition of the mobile phase in terms of pH and concentration showed great influence on the chromatographic separation and was varied to allow the distinction of each CS without signal overlap for a total analysis time of 25 min. This methodology was applied to determine the disaccharide composition of biological reaction media resulting from various enzymatic transformations of CS, such as enzymatic desulfation of CS disaccharides by a CS 4-O-endosulfatase, and depolymerization of the CS endocan by chondroitinase lyase ABC.

Design and property investigation on a five-interaction-based fluorescent anion receptor clip.

A molecular clip combining a doubly substituted fluorescent anion-π donor probe and two flexible arms bearing H-bond motifs constitutes a new generation of anion receptors. Five simultaneous non-covalent interactions are highlighted by theoretical complexation studies with five different anions. A large range of analytical techniques (electrospray-tandem mass spectrometry, NMR, UV-visible, steady-state and time-resolved fluorescence) were deployed to evaluate the stoichiometry and association constants with the selected anions. The photophysical and anion-π donor properties of the tetrazine ring allowed fine characterization of the binding properties of the ligand. Based on previously published results, an anti-cooperativity effect in non-covalent interactions was demonstrated.

Intertwined Detection and Recognition Roles of Tetrazine in Synergistic Anion-π and H-bond Based Anion Receptor.

The intrinsic properties of tetrazine as a π-anion receptor and as an on/off recognition probe merged with H-bond ability of an urea motif into a single architecture constitutes a new generation of well-defined anion receptors. Complexation properties directly benefit from the dual and synergistic contribution of tetrazine and urea. In this study, we report on the synthesis and assessment of binding properties to anions of diverse geometries. Association constants have been predicted by theoretical calculations and evaluated by multiple and complementary experimental techniques including electrospray-mass tandem spectroscopy, NMR, UV-visible, steady state fluorescence spectroscopies and time resolved fluorescence. These results provide the basis for a better understanding of both the complexation and the anion-dependent quenching mechanism.

Kinetic study of azobenzene E/Z isomerization using ion mobility-mass spectrometry and liquid chromatography-UV detection.

Z and E azobenzene isomers are molecular switches which can interconvert both photochemically and thermally. Presently, we studied a ketal-substituted bridged azobenzene in which two stable diastereomeric conformers (Z1 and Z2) photochemically interconvert through the transient E isomer. UV-VIS absorption spectroscopy is commonly used to study the relaxation kinetics of azobenzenes, but it does not allow direct quantitation of the process in this case. In the present paper, liquid chromatography coupled to UV detection (LC-UV) and ion mobility-mass spectrometry (IMS-MS) were combined to study the thermal back relaxation kinetics of the E isomer. LC separation of the three isomers was achieved in less than 10 minutes, allowing the characterization of the relatively slow thermal back relaxation kinetics at low temperature through UV detection. In addition, the faster E→Z thermal back relaxation at higher temperature was studied using IMS-MS, which allows shorter timescale separation than LC. Baseline separation of the two Z isomers was achieved in IMS-MS for [Z + Ag]+ ions, and their gas-phase conformations were determined by IRMPD experiments. Both IMS-MS and LC-UV methodologies succeeded to study the E→Z thermal back relaxation kinetics, and appeared to be complementary techniques. We show that the combination of the two techniques allows the characterization of the isomerization processes over a broad temperature range, and the determination of the associated thermodynamic observables.

Insights into Cisplatin Binding to Uracil and Thiouracils from IRMPD Spectroscopy and Tandem Mass Spectrometry.

The monofunctional primary complexes cis-[PtCl(NH3)2(L)]+, formed by the reaction of cisplatin, a major chemotherapeutic agent, with four nucleobases L, i.e., uracil (U), 2-thiouracil (2SU), 4-thiouracil (4SU), and 2,4-dithiouracil (24dSU), have been studied by a combination of infrared multiple photon dissociation (IRMPD) action spectroscopy in both the fingerprint (900-1900 cm-1) and the N-H/O-H stretching (3000-3800 cm-1) ranges, energy-resolved collision-induced dissociation (CID) mass spectrometry, and density functional calculations at the B3LYP/LACVP/6-311G** level. On the basis of the comparison across the experimental features and the linear IR spectra of conceivable structures, the cisplatin residue is found to promote a monodentate interaction preferentially with the O4(S4) atoms of the canonical forms of U, 4SU, and 24dSU and to the S2 atom of 2SU, yielding the most stable structures. Additional absorptions reveal the presence of minor, alternative tautomers in the sampled ion populations of 2SU and 24dSU, underlying the ability of cisplatin to increase the prospect of (therapeutically beneficial) nucleic acid strand disorder. Implication of these evidence may provide insights into drug mechanism and design.

Helically shaped cation receptor: design, synthesis, characterisation and first application to ion transport.

The methyl ester of 8-oxo-8H-indeno[2',1':7,8]naphtho[1,2-b]thiophene-2-carboxylic acid (1) and its corresponding PEGylated ester were synthesised and fully characterised. X-ray diffraction studies on (1) confirmed the helical structure of the receptor and that it is self-assembled into layers by π-π interactions. An in-depth study by DFT calculations and MS experiments (ESI-MS, MS/MS, IMRPD and ESI-IMS-MS) was carried out between (1) and the physiological cation K+. The formation of supramolecular complexes between (1) and K+ with different stoichiometries was demonstrated and the cation K+ preferentially interacts with the oxygen atoms of the carbonyl bond of the ketone and ester groups and the sulphur atom of the heterocycle. The ability of the two synthesized aromatic architectures to transport ions across a model lipid membrane has been studied by electrophysiology experiments. The formation of pores was observed, even at nanomolar concentrations. Since the PEGylated molecule showed more regular pore definitions than the hydrophobic molecule, the introduction of a polar hydrophilic chain made it possible to control the orientation of the aromatic architectures within the membrane.

Combined Experimental and Theoretical Survey of the Gas-Phase Reactions of Serine-Ca2+ Adducts.

The association of Ca2+ to serine and the subsequent gas-phase unimolecular reactivity of the [Ca(Ser)]2+ (Ser = Serine) adduct was investigated throughout the use of tandem mass spectrometry techniques and B3LYP/6-311+G(3df,2p)//B3LYP/6-311+G(d,p) density functional theory calculations. In a first step, the structure and relative stability of all possible conformers of serine were obtained and analyzed, as well as the most stable [serine-Ca]2+ adducts. For the analysis of the different potential energy surfaces associated with the gas-phase unimolecular reactivity of these adducts, only those that differ by less than 100 kJ·mol-1 from the global minimum were taken into account. In agreement with previous studies, the serine-Ca2+ global minimum corresponds to a charge-solvated structure in which Ca is tricoordinated to neutral serine. The major peaks observed in the nanoelectrospray-MS/MS spectrum of [Ca(Ser)]2+ adduct correspond to both Coulomb explosions, yielding either CaOH+ + [C3,H6,N,O2]+ or [C2,H4,O,N]+ + [Ca(C,H3,O2)]+, and to the loss of neutrals, namely, CH2O and H2O. Our theoretical survey of the energy profile allow us to conclude that, although all the aforementioned fragmentation processes can have their origin at the global minimum, similar fragmentations involving low-lying conformers, both zwitterionic and nonzwitterionic, compete and should be considered to account for the observed reactivity. We have also found that in some specific cases post-transition state dynamics similar to the ones described before in the literature for formamide-Ca2+ reactions, may also play a role.

Topology and Electronic Density Driven Generation of Alkali Cation Complexes.

The formation and characterization of K+ and Cs+ complexes originating from the cooperativity of three non-covalent interactions is explored. The tridimensional preorganization of the naphthothiophene platform displays a favorable well-defined bay region combining a π fragment and a carbonyl moiety flanking a central sulfur atom. A joint theoretical and experimental infrared multiple photon dissociation (IRMPD) study allowed deciphering the key contribution of the orthogonal phenyl fragment to the elaboration of alkali metal complexes. In combination with S and CO interactions, the π-cation interaction significantly enhances the binding energies of naphthothiophene derivatives.

Interactions of Dimethyltin(IV) with Uracil As Studied in the Gas Phase.

The gas-phase interactions of uracil (Ura) with dimethyltin(IV) were studied by a combined experimental and theoretical approach. Positive-ion electrospray spectra show that the interaction of dimethyltin(IV) with Ura results in the formation of the [(CH3)2Sn(Ura-H)]+ ion. The tandem mass spectrometry spectrum of this complex is characterized by numerous fragmentation processes, notably associated with elimination of H,N,C,O and C3,H3,N,O moieties, as well as the unusual loss of C2H6 leading to the [Sn(Ura-H)]+ complex. In turn, the [Sn(Ura-H)]+ complex fragments according to pathways already observed for the [Pb(Ura-H)]+ analogue. Sequential losses of ·CH3 radicals are also observed from the [(CH3)2Sn(N,C,O)]+ species (m/z 192). Comparison between density functional theory-computed vibrational spectra and the infrared multiple photon dissociation spectrum recorded between 1000 and 1900 cm-1 shows a good agreement as far as the global minimum is concerned. This comparison points to a bidentate interaction with a deprotonated canonical diketo form of uracil, involving both the N3 and O4 electronegative centers. This binding scheme has been already reported for the Pb/uracil system. The bidentate form characterized by the interaction between dimethyltin with N3 and O2 centers is slightly less stable. Interconversion between the two structures is associated with a small activation barrier (56 kJ/mol). The potential energy surfaces were explored to account for the main fragmentations observed upon collision-induced dissociation.

Isomer separation and effect of the degree of polymerization on the gas-phase structure of chondroitin sulfate oligosaccharides analyzed by ion mobility and tandem mass spectrometry.

RATIONALE: Chondroitin sulfate (CS) glycosaminoglycans are bioactive sulfated polysaccharides comprising repeating units of uronic acid and N-acetyl galactose sulfated at various positions. The optimal length and sulfation pattern of the CS bioactive sequences remain elusive so that structure-activity relationships cannot be easily established. Development of efficient analytical methods allowing the differentiation of the various sulfation patterns of CS sequences is therefore of particular importance to correlate their biological functions to the sulfation pattern. METHODS: Discrimination of different oligomers (dp2 to dp6) of synthetic chondroitin sulfate isomers was evaluated by electrospray ionization tandem mass spectrometry (ESI-MS/MS) in the negative-ion mode from deprotonated and alkali adduct species. In addition, ion mobility mass spectrometry (IMS-MS) was used to study the influence of both the degree of polymerization and sulfate group location on the gas-phase conformation of CS oligomers. RESULTS: ESI-MS/MS spectra of chondroitin sulfate isomers show characteristic product ions exclusively from alkali adduct species (Li, Na, K and Cs). Whatever the alkali adducts studied, MS/MS of chondroitin oligosaccharides sulfated at position 6 yields a specific product ion at m/z 139 while CS oligosaccharides sulfated at position 4 show a specific product ion at m/z 154. Being observed for the different CS oligomers di-, tetra- and hexasaccharides, these fragment ions are considered as diagnostic ions for chondroitin 6-O-sulfate and chondroitin 4-O-sulfate, respectively. IMS-MS experiments reveal that collision cross-sections (CCS) of CS oligomers with low charge states evolved linearly with degrees of polymerization indicating a similar gas-phase conformation. CONCLUSIONS: This study allows the fast and unambiguous differentiation of CS isomers sulfated at position 6 or 4 for both saturated and unsaturated analogues from MS/MS experiments. In addition, the CCS linear evolution of CS oligomers in function of the degree of polymerization indicates that no folding occurs even for hexasaccharides.

Undervalued N3 Coordination Revealed in the Cisplatin Complex with 2'-Deoxyadenosine-5'-monophosphate by a Combined IRMPD and Theoretical Study.

The complex obtained by the reaction of cisplatin and 2'-deoxyadenosine-5'-monophosphate (5'-dAMP) in water has been isolated and detected by electrospray ionization mass spectrometry. The so-formed cis-[PtCl(NH3)2(5'-dAMP)]+ complex has been studied in detail by infrared multiple photon dissociation (IRMPD) spectroscopy in two spectral ranges, namely, 700-1900 and 2800-3800 cm-1, backed by quantum-chemical calculations at the B3LYP/LACV3P/6-311G** level of theory. In agreement with the computational results, the vibrational spectroscopic characterization of cis-[PtCl(NH3)2(5'-dAMP)]+ shows that the sampled ionic population comprises two major isomers, differentiated in the X-H stretching region by their distinct fragmentation patterns. One of these species presents coordination of the platinum moiety at the N3 position of adenine, whereas in the second one, platinum is bound at the N1 position of adenine. IRMPD kinetics have allowed an estimation of their relative proportions. Surprisingly, the most abundant component of cis-[PtCl(NH3)2(5'-dAMP)]+ is the N3 isomer, although it is slightly less stable than the other potential isomers in the gas phase. In contrast, the lowest-energy species, namely, the one showing cisplatin binding to the N7 position of adenine, seems to be the one less represented in the sampled ionic population. These findings suggest that the reaction of cisplatin with 5'-dAMP is governed by the kinetics of the process occurring in solution rather than by the thermodynamic factors.

Characterization of Protonated Model Disaccharides from Tandem Mass Spectrometry and Chemical Dynamics Simulations.

The fragmentation mechanisms of prototypical disaccharides have been studied herein by coupling tandem mass spectrometry (MS) with collisional chemical dynamics simulations. These calculations were performed by explicitly considering the collisions between the protonated sugar and the neutral target gas, which led to an ensemble of trajectories for each system, from which it was possible to obtain reaction products and mechanisms without pre-imposing them. The ß-aminoethyl and aminopropyl derivatives of cellobiose, maltose, and gentiobiose were studied to observe differences in both the stereochemistry and the location of the glycosidic linkage. Chemical dynamics simulations of MS/MS and MS/MS/MS were used to suggest some primary and secondary fragmentation mechanisms for some experimentally observed product ions. These simulations provided some new insights into the fundamentals of the unimolecular dissociation of protonated sugars under collisional induced dissociation conditions.

Identification of acylation products in SHAPE chemistry.

SHAPE chemistry (selective 2'-hydroxyl acylation analyzed by primer extension) has been developed to specifically target flexible nucleotides (often unpaired nucleotides) independently to their purine or pyrimidine nature for RNA secondary structure determination. However, to the best of our knowledge, the structure of 2'-O-acylation products has never been confirmed by NMR or X-ray data. We have realized the acylation reactions between cNMP and NMIA under SHAPE chemistry conditions and identified the acylation products using standard NMR spectroscopy and LC-MS/MS experiments. For cAMP and cGMP, the major acylation product is the 2'-O-acylated compound (>99%). A trace amount of N-acylated cAMP has also been identified by LC-UV-MS2. While for cCMP, the isolated acylation products are composed of 96% of 2'-O-acylated, 4% of N,O-diacylated, and trace amount of N-acylated compounds. In addition, the characterization of the major 2'-O-acylated compound by NMR showed slight differences in the conformation of the acylated sugar between the three cyclic nucleotides. This interesting result should be useful to explain some unexpected reactivity of the SHAPE chemistry.

Conformational Dynamics in Ion Mobility Data.

The shape of the spectral features in arrival time distributions (ATDs) recorded by ion mobility spectrometry (IMS) can often be interpreted in terms of the coexistence of different isomeric species. Interconversion between such species is also acknowledged to influence the shape of the ATD, even if no general quantitative description of this effect is available. We present an analytical model that allows simulating ATDs resulting from interconverting species. This model is used to reproduce experimental data obtained on a bistable system and to interpret discrepancies between measurements on different types of instruments. We show that the proposed model can be further exploited to extract kinetic and thermodynamic data from tandem-IMS measurements.

Gas-phase interactions of organotin compounds with cysteine.

The gas-phase interactions of cysteine with di-organotin and tri-organotin compounds have been studied by mass spectrometry experiments and quantum calculations. Positive-ion electrospray spectra show that the interaction of di- and tri-organotins with cysteine results in the formation of [(R)2 Sn(Cys-H)]+ and [(R)3 Sn(Cys)]+ ions, respectively. MS/MS spectra of [(R)2 Sn(Cys-H)]+ complexes are characterized by numerous fragmentation processes, notably associated with elimination of NH3 and (C,H2 ,O2 ). Several dissociation routes are characteristic of each given organic species. Upon collision, both the [(R)3 Sn(Gly)]+ and [(R)3 Sn(Cys)]+ complexes are associated with elimination of the intact amino acid, leading to the formation of [(R)3 Sn]+ cation. But for the latter complex, two additional fragmentation processes are observed, associated with the elimination of NH3 and C3 H4 O2 S. Calculations indicate that the interaction between organotins and cysteine is predominantly electrostatic but also exhibits a considerable covalent character, which is slightly more pronounced in tri-organotin complexes. A preferred bidentate interaction of the type -η2 -S-NH2 , with sulfur and the amino group, is observed. As for the [(R)3 Sn(Cys)]+ complexes, their stability is due to the combination of the hydrogen bond taking place between the amino group and the sulfur lone pair and the interaction between the carboxylic oxygen atom and the metal. Copyright © 2016 John Wiley & Sons, Ltd.

On the gas phase fragmentation of protonated uracil: a statistical perspective.

The potential energy surface of protonated uracil has been explored by an automated transition state search procedure, resulting in the finding of 1398 stationary points and 751 reactive channels, which can be categorized into isomerizations between pairs of isomers, unimolecular fragmentations and bimolecular reactions. The use of statistical Rice-Ramsperger-Kassel-Marcus (RRKM) theory and Kinetic Monte Carlo (KMC) simulations allowed us to determine the relative abundances of each fragmentation channel as a function of the ion's internal energy. The KMC/RRKM product abundances are compared with novel mass spectrometry (MS) experiments in the collision energy range 1-6 eV. To facilitate the comparison between theory and experiments, further dynamics simulations are carried out to determine the fraction of collision energy converted into the ion's internal energy. The KMC simulations show that the major fragmentation channels are isocyanic acid and ammonia losses, in good agreement with experiments. The third predominant channel is water loss according to both theory and experiments, although the abundance obtained in the KMC simulations is very low, suggesting that non-statistical dynamics might play an important role in this channel. Isocyanic acid (HNCOH(+)) is also an important product in the KMC simulations, although its abundance is only significant at internal energies not accessible in the MS experiments.