Pitfalls in Using Isotopic Distributions for Structural Interpretation by Mass Spectrometry.

This brief cautionary note reports a failure in a common and useful assumption, namely, that the isotopes of the elements occur in their natural abundance ratios in commercially sourced organic compounds. Some commercial sources of tris(pentafluorophenyl)borane, B(C6F5)3, show severely depleted 10B, while materials from other suppliers display natural isotopic abundances. The depletion varies from lot-to-lot, and it was confirmed by inductively coupled plasma (ICP) mass spectrometry. The isotope 10B is used in the nuclear power industry, as a neutron absorber in the power control rods. It is speculated that the residual 11B generated when preparing 10B-enriched boron carbide for control rod use, provides 11B-rich raw material that is then used for commercial B(C6F5)3 synthesis.

Spontaneous Oxidation in Aqueous Microdroplets: Water Radical Cation as Primary Oxidizing Agent.

Exploration of the unique chemical properties of interfaces can unlock new understanding. A striking example is the finding of accelerated reactions, particularly spontaneous oxidation reactions, that occur without assistance of catalysts or external oxidants at the air interface of both aqueous and organic solutions (provided they contain some water). This finding opened a new area of interfacial chemistry but also caused heated debate regarding the primary chemical species responsible for the observed oxidation. An overview of the literature covering oxidation in microdroplets with air interfaces is provided, together with a critical examination of previous findings and hypotheses. The water radical cation/radical anion pair, formed spontaneously and responsible for the electric field at or near the droplet/air interface, is suggested to constitute the primary redox species. Mechanisms of accelerated microdroplet reactions are critically discussed and it is shown that hydroxyl radical/hydrogen peroxide formation in microdroplets does not require that these species be the primary oxidant. Instead, we suggest that hydroxyl radical and hydrogen peroxide are the products of water radical cation decay in water. The importance of microdroplet chemistry in the prebiotic environment is sketched briefly and the role of partial solvation in reaction acceleration is noted.

Levitating Droplet Electroanalysis.

Chemical reactions that occur in droplets proceed much differently compared to bulk phases. For instance, many groups have studied droplets during levitation by mass spectrometry and fluorescence to gain more detailed mechanistic insight. Such droplets maximize the probability of solution species interacting with the solution-air interface, an interface that is inherently difficult to probe electrochemically. In this Technical Note, we overcome this limitation by developing a laser-pulled dual-barrel electrode. Having two microwires sealed within the same glass capillary allows one to make two-electrode measurements. We show that the electrode can be positioned within a levitating water droplet and that the voltammetry of a redox indicator (hexacyanoferrate (II/III)) can be observed in real-time. Such foundational measurement tools are important to probe a variety of chemical reactions at complex interfaces.

Oxazolone mediated peptide chain extension and homochirality in aqueous microdroplets.

Peptide formation from amino acids is thermodynamically unfavorable but a recent study provided evidence that the reaction occurs at the air/solution interfaces of aqueous microdroplets. Here, we show that i) the suggested amino acid complex in microdroplets undergoes dehydration to form oxazolone; ii) addition of water to oxazolone forms the dipeptide; and iii) reaction of oxazolone with other amino acids forms tripeptides. Furthermore, the chirality of the reacting amino acids is preserved in the oxazolone product, and strong chiral selectivity is observed when converting the oxazolone to tripeptide. This last fact ensures that optically impure amino acids will undergo chain extension to generate pure homochiral peptides. Peptide formation in bulk by wet-dry cycling shares a common pathway with the microdroplet reaction, both involving the oxazolone intermediate.

Re-Imagining Drug Discovery using Mass Spectrometry.

It is argued that each of the three key steps in drug discovery, (i) reaction screening to find successful routes to desired drug candidates, (ii) scale up of the synthesis to produce amounts adequate for testing, and (iii) bioactivity assessment of the candidate compounds, can all be performed using mass spectrometry (MS) in a sequential fashion. The particular ionization method of choice, desorption electrospray ionization (DESI), is both an analytical technique and a procedure for small-scale synthesis. It is also highly compatible with automation, providing for high throughput in both synthesis and analysis. Moreover, because accelerated reactions take place in the secondary DESI microdroplets generated from individual reaction mixtures, this allows either online analysis by MS or collection of the synthetic products by droplet deposition. DESI also has the unique advantage, amongst spray-based MS ionization methods, that complex buffered biological solutions can be analyzed directly, without concern for capillary blockage. Here, all these capabilities are illustrated, the unique chemistry at droplet interfaces is presented, and the possible future implementation of DESI-MS based drug discovery is discussed.

Quantitative Determination of Water in Organic Liquids by Ambient Mass Spectrometry.

This study uses a rapid tandem mass-spectrometry method to determine water content in complex organic solutions. Emphasis is placed on trace-water analysis by a fast and accurate alternative to the Karl-Fischer method. In this new method, water is captured by a charge-labeled molecular probe. Water binds strongly with high specificity to the strongly electrophilic aldehyde site in a charge-labelled molecule (N-methylpyridinium); competitive binding by other analytes is effectively discriminated against in the mass-measurement step. Quantitative determinations are made over a wide concentration range, 0.001 % (10 ppm) to 99 %, with better than 10 % relative standard deviation, along with short (1 min) analysis times using small sample volumes (several µL). Applications include water measurement in simple organic solvents, for example, deuterated solvents, as well as in complex mixtures, for example, organic reaction mixtures. Additionally, this method allows for water monitoring in levitated droplets. Mechanistic investigations into the impact of water on important chemical processes in organic synthesis and environmental science are reported.

Simultaneous and Spontaneous Oxidation and Reduction in Microdroplets by the Water Radical Cation/Anion Pair.

Microdroplets show unique chemistry, especially in their intrinsic redox properties, and to this we here add a case of simultaneous and spontaneous oxidation and reduction. We report the concurrent conversions of several phosphonates to phosphonic acids by reduction (R-P → H-P) and to pentavalent phosphoric acids by oxidation. The experimental results suggest that the active reagent is the water radical cation/anion pair. The water radical cation is observed directly as the ionized water dimer while the water radical anion is only seen indirectly though the spontaneous reduction of carbon dioxide to formate. The coexistence of oxidative and reductive species in turn supports the proposal of a double-layer structure at the microdroplet surface, where the water radical cation and radical anion are separated and accumulated.

Spontaneous Water Radical Cation Oxidation at Double Bonds in Microdroplets.

Spontaneous oxidation of compounds containing diverse X=Y moieties (e.g., sulfonamides, ketones, esters, sulfones) occurs readily in organic-solvent microdroplets. This surprising phenomenon is proposed to be driven by the generation of an intermediate species [M+H2O]+·: a covalent adduct of water radical cation (H2O +· ) with the reactant molecule (M). The adduct is observed in the positive ion mass spectrum while its formation in the interfacial region of the microdroplet (i.e., at the air-droplet interface) is indicated by the strong dependence of the oxidation product formation on the spray distance (which reflects the droplet size and consequently the surface-to-volume ratio) and the solvent composition. Importantly, based on the screening of a ca. 21,000-compound library and the detailed consideration of six functional groups, the formation of a molecular adduct with the water radical cation is a significant route to ionization in positive ion mode electrospray, where it is favored in those compounds with X=Y moieties which lack basic groups. A set of model monofunctional systems was studied and in one case, benzyl benzoate, evidence was found for oxidation driven by hydroxyl radical adduct formation followed by protonation in addition to the dominant water radical cation addition process. Significant implications of molecular ionization by water radical cations for oxidation processes in atmospheric aerosols, analytical mass spectrometry and small-scale synthesis are noted.

Spontaneous Oxidation of Aromatic Sulfones to Sulfonic Acids in Microdroplets.

Reactions in microdroplets can be accelerated and can present unique chemistry compared to reactions in bulk solution. Here, we report the accelerated oxidation of aromatic sulfones to sulfonic acids in microdroplets under ambient conditions without the addition of acid, base, or catalyst. The experimental data suggest that the water radical cation, (H2O)+•, derived from traces of water in the solvent, is the oxidant. The substrate scope of the reaction indicates the need for a strong electron-donating group (e.g., p-hydroxyl) in the aromatic ring. An analogous oxidation is observed in an aromatic ketone with benzoic acid production. The shared mechanism is suggested to involve field-assisted ionization of water at the droplet/air interface, its reaction with the sulfone (M) to form the radical cation adduct, (M + H2O)+•, followed by 1,2-aryl migration and C-O cleavage. A remarkably high reaction rate acceleration (∼103) and regioselectivity (∼100-fold) characterize the reaction.

Reaction Acceleration Promoted by Partial Solvation at the Gas/Solution Interface.

The kinetics of organic reactions of different types in microvolumes (droplets, thin films, and sealed tubes) show effects of gas/solution interfacial area, reaction molecularity and solvent polarity. Partial solvation at the gas/solution interface is a major contributor to the 104 -fold reaction acceleration seen in bimolecular but not unimolecular reactions in microdroplets. Reaction acceleration can be used to manipulate selectivity by solvent choice.

High-yield gram-scale organic synthesis using accelerated microdroplet/thin film reactions with solvent recycling.

A closed system has been designed to perform microdroplet/thin film reactions with solvent recycling capabilities for gram-scale chemical synthesis. Claisen-Schmidt, Schiff base, Katritzky and Suzuki coupling reactions show acceleration factors relative to bulk of 15 to 7700 times in this droplet spray system. These values are much larger than those reported previously for the same reactions in microdroplet/thin film reaction systems. The solvent recycling mode of the new system significantly improves the reaction yield, especially for reactions with smaller reaction acceleration factors. The microdroplet/thin film reaction yield improved on recycling from 33% to 86% and from 32% to 72% for the Katritzky and Suzuki coupling reactions, respectively. The Claisen-Schmidt reaction was chosen to test the capability of this system in gram scale syntheses and rates of 3.18 g per h and an isolated yield of 87% were achieved.

Internal Activation of Peptidyl Prolyl Thioesters in Native Chemical Ligation.

Prolyl thioesters have shown significantly lower reactivities in native chemical ligation (NCL) in comparison to that of the alanyl thioester. This report describes a mild and efficient internal activation protocol of peptidyl prolyl thioesters in NCL without using any thiol-based additives, where the introduction of a 4-mercaptan substituent on the C-terminal proline significantly improves the reactivity of prolyl thioesters via the formation of a bicyclic thiolactone intermediate. The kinetic data indicate that the reaction rate is comparable to that of the reported data of alanyl thioesters, and the mechanistic studies suggest that the ligation of two peptide segments proceeds through an NCL-like pathway instead of a direct aminolysis, which ensures the chemoselectivity and compatibility of various amino acid side chains. This 4-mercaptoprolyl thioester-based protocol also allows an efficient one-pot ligation-desulfurization procedure. The utility of this method has been further demonstrated in the synthesis of a proline-rich region of Wilms tumor protein 1.