Synthesis of 7-Aminocoumarins from 7-Hydroxycoumarins via Amide Smiles Rearrangement.

N-Substituted 7-aminocoumarins can be synthesized from readily available 7-hydroxycoumarins via alkylation with α-bromoacetamides and subsequent tandem O → N Smiles rearrangement-amide hydrolysis. The key rearrangement sequence proceeds under mild conditions to provide convenient access to various N-alkyl and N-aryl products in moderate to high yields. The process is operationally simple, inexpensive, transition-metal-free, and can be telescoped into a one-pot process.

Ligand-Directed Approach to Activity-Based Sensing: Developing Palladacycle Fluorescent Probes That Enable Endogenous Carbon Monoxide Detection.

Carbon monoxide (CO) is an emerging gasotransmitter and reactive carbon species with broad anti-inflammatory, cytoprotective, and neurotransmitter functions along with therapeutic potential for the treatment of cardiovascular diseases. The study of CO chemistry in biology and medicine relative to other prominent gasotransmitters such as NO and H2S remains challenging, in large part due to limitations in available tools for the direct visualization of this transient and freely diffusing small molecule in complex living systems. Here we report a ligand-directed activity-based sensing (ABS) approach to CO detection through palladium-mediated carbonylation chemistry. Specifically, the design and synthesis of a series of ABS probes with systematic alterations in the palladium-ligand environment (e.g., sp3-S, sp3-N, sp2-N) establish structure-activity relationships for palladacycles to confer selective reactivity with CO under physiological conditions. These fundamental studies led to the development of an optimized probe, termed Carbon Monoxide Probe-3 Ester Pyridine (COP-3E-Py), which enables imaging of CO release in live cell and brain settings, including monitoring of endogenous CO production that triggers presynaptic dopamine release in fly brains. This work provides a unique tool for studying CO in living systems and establishes the utility of a synthetic methods approach to activity-based sensing using principles of organometallic chemistry.

Development of a General Aza-Cope Reaction Trigger Applied to Fluorescence Imaging of Formaldehyde in Living Cells.

Formaldehyde (FA) is a reactive signaling molecule that is continuously produced through a number of central biological pathways spanning epigenetics to one-carbon metabolism. On the other hand, aberrant, elevated levels of FA are implicated in disease states ranging from asthma to neurodegenerative disorders. In this context, fluorescence-based probes for FA imaging are emerging as potentially powerful chemical tools to help disentangle the complexities of FA homeostasis and its physiological and pathological contributions. Currently available FA indicators require direct modification of the fluorophore backbone through complex synthetic considerations to enable FA detection, often limiting the generalization of designs to other fluorophore classes. To address this challenge, we now present the rational, iterative development of a general reaction-based trigger utilizing 2-aza-Cope reactivity for selective and sensitive detection of FA in living systems. Specifically, we developed a homoallylamine functionality that can undergo a subsequent self-immolative ß-elimination, creating a FA-responsive trigger that is capable of masking a phenol on a fluorophore or any other potential chemical scaffold for related imaging and/or therapeutic applications. We demonstrate the utility of this trigger by creating a series of fluorescent probes for FA with excitation and emission wavelengths that span the UV to visible spectral regions through caging of a variety of dye units. In particular, Formaldehyde Probe 573 (FAP573), based on a resorufin scaffold, is the most red-shifted and FA sensitive in this series in terms of signal-to-noise responses and enables identification of alcohol dehydrogenase 5 (ADH5) as an enzyme that regulates FA metabolism in living cells. The results provide a starting point for the broader use of 2-aza-Cope reactivity for probing and manipulating FA biology.

Cinchonidinium acetate as a convenient catalyst for the asymmetric synthesis of cis-stilbenediamines.

Inexpensive and readily available cinchonidinium acetate is an effective catalyst for the syn-selective aza-Henry reaction of arylnitromethanes and aryl imines. The resulting masked cis-stilbenediamine products are produced in excellent diastereoselectivity and good enantioselectivity, and enantiopure material can be achieved via recrystallization. The features of the cinchona catalyst needed for selectivity are discussed, with specific emphasis on formation of a kinetically controlled syn-product without epimerization of the highly acidic α-nitro stereocenter.

Quantification of electrophilic activation by hydrogen-bonding organocatalysts.

A spectrophotometric sensor is described that provides a useful assessment of the LUMO-lowering provided by catalysts in Diels-Alder and Friedel-Crafts reactions. A broad range of 33 hydrogen-bonding catalysts was assessed with the sensor, and the relative rates in the above reactions spanned 5 orders of magnitude as determined via (1)H- and (2)H NMR spectroscopic measurements, respectively. The differences between the maximum wavelength shift of the sensor with and without catalyst (Δλ(max)(-1)) were found to correlate linearly with ln(k(rel)) values for both reactions, even though the substrate feature that interacts with the catalyst differs significantly (ketone vs nitro). The sensor provides an assessment of both the inherent reactivity of a catalyst architecture as well as the sensitivity of the reaction to changes within an architecture. In contrast, catalyst pK(a) values are a poor measure of reactivity, although correlations have been identified within catalyst classes.

Amino acid-dependent stability of the acyl linkage in aminoacyl-tRNA.

Aminoacyl-tRNAs are the biologically active substrates for peptide bond formation in protein synthesis. The stability of the acyl linkage in each aminoacyl-tRNA, formed through an ester bond that connects the amino acid carboxyl group with the tRNA terminal 3'-OH group, is thus important. While the ester linkage is the same for all aminoacyl-tRNAs, the stability of each is not well characterized, thus limiting insight into the fundamental process of peptide bond formation. Here, we show, by analysis of the half-lives of 12 of the 22 natural aminoacyl-tRNAs used in peptide bond formation, that the stability of the acyl linkage is effectively determined only by the chemical nature of the amino acid side chain. Even the chirality of the side chain exhibits little influence. Proline confers the lowest stability to the linkage, while isoleucine and valine confer the highest, whereas the nucleotide sequence in the tRNA provides negligible contribution to the stability. We find that, among the variables tested, the protein translation factor EF-Tu is the only one that can protect a weak acyl linkage from hydrolysis. These results suggest that each amino acid plays an active role in determining its own stability in the acyl linkage to tRNA, but that EF-Tu overrides this individuality and protects the acyl linkage stability for protein synthesis on the ribosome.

Minimizing the amount of nitromethane in palladium-catalyzed cross-coupling with aryl halides.

A method for the formation of arylnitromethanes is described that employs readily available aryl halides or triflates and small amounts of nitromethane in a dioxane solvent, thereby reducing the hazards associated with this reagent. Specifically, 2-10 equiv (1-5% v/v) of nitromethane can be employed in comparison to prior work that used nitromethane as solvent (185 equiv). The present transformation provides high yields at relatively low temperatures and tolerates an array of functionality, including heterocycles and substantial steric encumbrance.

Nitroethylation of vinyl triflates and bromides.

A two-carbon homologation of vinyl triflates and bromides for the synthesis of homoallylic nitro products is described. This palladium-catalyzed double coupling of nitromethane exploits the anion stabilizing and leaving group properties of nitromethane, generating the homo allyl nitro products via a tandem cross-coupling/π-allylation sequence. The resultant process provides a mild and convenient entry to nitroethylated products, which are versatile precursors to ß,γ-unsaturated carbonyls, homoallylic amines, and nitrile oxides.

Rapid quantification of the activating effects of hydrogen-bonding catalysts with a colorimetric sensor.

A sensor has been developed to quickly and simply assess the relative reactivity of different hydrogen-bonding catalysts. Specifically, blue-shifts seen upon treatment of H-bonding catalysts with the colorimetric compound 7-methyl-2-phenylimidazo[1,2-a]pyrazin-3(7H)-one correlate well to the K(eq) of binding to the sensor. The blue-shifts also show a high degree of correlation with relative rates in Diels-Alder reactions of methyl vinyl ketone and cyclopentadiene employing the H-bonding catalysts. The relevance of the sensor blue-shifts to the LUMO-lowering abilities of the H-bonding catalysts is discussed.

Palladium-catalyzed nitromethylation of aryl halides: an orthogonal formylation equivalent.

An efficient cross-coupling reaction of aryl halides and nitromethane was developed with the use of parallel microscale experimentation. The arylnitromethane products are precursors for numerous useful synthetic products. An efficient method for their direct conversion to the corresponding oximes and aldehydes in a one-pot operation has been discovered. The process exploits inexpensive nitromethane as a carbonyl equivalent, providing a mild and convenient formylation method that is compatible with many functional groups.