Equilibrium in the Catalytic Condensation of Carboxylic Acids with Methyl Ketones to 1,3-Diketones and the Origin of the Reketonization Effect

Acetone is the expected ketone product of an acetic acid decarboxylative ketonization reaction with metal oxide catalysts used in the industrial production of ketones and for biofuel upgrade. Decarboxylative cross-ketonization of a mixture of acetic and isobutyric acids yields highly valued unsymmetrical methyl isopropyl ketone (MIPK) along with two less valuable symmetrical ketones, acetone and diisopropyl ketone (DIPK). We describe a side reaction of isobutyric acid with acetone yielding the cross-ketone MIPK with monoclinic zirconia and anatase titania catalysts in the absence of acetic acid. We call it a reketonization reaction because acetone is deconstructed and used for the construction of MIPK. Isotopic labeling of the isobutyric acid’s carboxyl group shows that it is the exclusive supplier of the carbonyl group of MIPK, while acetone provides only methyl group for MIPK construction. More branched ketones, MIPK or DIPK, are less reactive in their reketonization with carboxylic acids. The proposed mechanism of reketonization supported by density functional theory (DFT) computations starts with acetone enolization and proceeds via its condensation with surface isobutyrate to a β-diketone similar to β-keto acid formation in the decarboxylative ketonization of acids. Decomposition of unsymmetrical β-diketones with water (or methanol) by the retrocondensation reaction under the same conditions over metal oxides yields two pairs of ketones and acids (or esters in the case of methanol) and proceeds much faster compared to their formation. The major direction yields thermodynamically more stable products—more substituted ketones. DFT calculations predict even a larger fraction of the thermodynamically preferred pair of products. The difference is explained by some degree of a kinetic control in the opposite direction. Reketonization has lower reaction rates compared to regular ketonization. Still, a high extent of reketonization occurs unnoticeably during the decarboxylative ketonization of acetic acid as the result of the acetone reaction with acetic acid. This degenerate reaction is the major cause of the inhibition by acetone of its own rate of formation from acetic acid at high conversions

Catalytic Condensation of Ketones with Carboxylic Acids

It is found that use of acetone in place of acetic acid in the reaction with isobutyric acid is effective for the synthesis of the cross-ketonization product, methyl isopropyl ketone (MIPK). Rate of MIPK formation is of the same order of magnitude, but slightly lower for acetone compared to acetic acid under similar conditions (Fig. 1). The most active catalyst is KOH-treated titania. The second product of this reaction, DIPK, results from the ketonic decarboxylative condensation of isobutyric acid with itself. The 13C labeled carbonyl group from isobutyric acid is almost exclusively (within the detection error) transferred to the MIPK product (Scheme 2). In the reaction of acetic acid with a more branched ketone, DIPK, only a negligible amount of MIPK is produced with all studied catalysts. Based on the experimental data, the proposed mechanism most likely includes enolization of acetone, followed by its condensation as the nucleophile with isobutyric acid as the electrophile, and completed by the retro-condensation to MIPK (Scheme 2). The order of the enolic components activity, acetic acid ≥ isobutyric acid \u3e acetone \u3e\u3e DIPK, is generally consistent with the order of their adsorption energies on metal oxides. Low or non-branched ketones could be efficiently used in place of one of the acids in the cross-ketonization reaction. Because of the relatively high reaction rate, this process needs to be accounted for in the kinetic scheme of the decarboxylative ketonization

DEVELOPMENT OF IN VITRO LIVER CULTURE TECHNIQUES TO INTEGRATE A PHYSIOLOGICALLY RELEVANT MICROENVIROMENT RECAPITULATING LIVER LOBULE ZONATION

During drug-discovery, cell monolayers fail to accurately predict drug-induced liver injury– a common reason for discontinuation during clinical trials. One reason is that monolayer cultures lack many aspects of the tissue microenvironment. The liver is composed of subunits known as lobules. In the lobule, a sinusoid connects a portal triad and central vein and is surrounded by hepatocytes to maximize nutrient exchange. A unidirectional flow leads to extracellular gradients of oxygen, nutrients, hormones, and morphogens. These gradients in the lobule impact hepatocyte function through the differential expression of hepatocytic pathways in the periportal (PP) and perivenous (PV) regions. To improve in vitro hepatocyte cultures, a platform that incorporates structural and microenvironmental factors is needed to recapitulate the liver. Factors include: coculturing liver-specific cells; inclusion of liver-specific extracellular gradients, specifically nutrients and morphogens; incorporating dynamic nutrient flow and distribution; and inclusion of appropriate three-dimensional hepatocyte organization, and a relevant extracellular matrix (ECM). In this work, I integrated physiological aspects– oxygen, morphogens, extracellular matrix stiffness, and a 3D paper scaffold culture– into in vitro cultures of different cell types with increasing hepatocyte-like functions: HepG2, HepaRG, and primary human hepatocytes (PHHs). I evaluated cell health, liver-specific function, metabolic enzyme activity, and transcriptional regulation as each physiological condition was introduced individually, and in concert. This work displayed that oxygen improves in vitro zonal patterning of SULT and UGT transcript, but oxygen alone could not achieve the drug metabolizing enzyme’s zonal distribution. The inclusion of Wnt alone increases CYP, UGT and SULT activity, as compared to a control, while the magnitude of enzymatic activity increase is modulated by oxygen. Overall, the HepaRG cell’s basal and inducible drug metabolizing enzyme activity is similar across a wide range of ECM stiffness suggesting that the HepaRG cells are adaptable to various ECMs, unlike PHHs. Lastly, direct comparisons of monolayers and paper-based cultured in PP and PV microenvironments, indicated zonation-like trends between the PP and PV conditions. The paper-based PV conditioned cells exhibited the highest drug metabolizing enzyme activity. This work developed a more representative in vitro liver models by incorporating tissue-specific features.Doctor of Philosoph

PP165—Evaluation of JNJ-26489112, a Novel Antiepileptic Drug: A Placebo-Controlled, Exploratory Study

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Assessing Function and Endurance in Adults with Spinal and Bulbar Muscular Atrophy: Validity of the Adult Myopathy Assessment Tool

Purpose. The adult myopathy assessment tool (AMAT) is a performance-based battery comprised of functional and endurance subscales that can be completed in approximately 30 minutes without the use of specialized equipment. The purpose of this study was to determine the construct validity and internal consistency of the AMAT with a sample of adults with spinal and bulbar muscular atrophy (SBMA). Methods. AMAT validity was assessed in 56-male participants with genetically confirmed SBMA (mean age, 53 ± 10 years). The participants completed the AMAT and assessments for disease status, strength, and functional status. Results. Lower AMAT scores were associated with longer disease duration (r = -0.29; P \u3c 0.03) and lower serum androgen levels (r = 0.49-0.59; P \u3c 0.001). The AMAT was significantly correlated with strength and functional status (r = 0.82-0.88; P \u3c 0.001). The domains of the AMAT exhibited good internal consistency (Cronbach\u27s α  = 0.77-0.89; P \u3c 0.001). Conclusions. The AMAT is a standardized, performance-based tool that may be used to assess functional limitations and muscle endurance. The AMAT has good internal consistency, and the construct validity of the AMAT is supported by its significant associations with hormonal, strength, and functional characteristics of adults with SBMA. This trial is registered with Clinicaltrials.gov identifier NCT00303446

Tau-Mediated Nuclear Depletion and Cytoplasmic Accumulation of SFPQ in Alzheimer's and Pick's Disease

Tau dysfunction characterizes neurodegenerative diseases such as Alzheimer's disease (AD) and frontotemporal lobar degeneration (FTLD). Here, we performed an unbiased SAGE (serial analysis of gene expression) of differentially expressed mRNAs in the amygdala of transgenic pR5 mice that express human tau carrying the P301L mutation previously identified in familial cases of FTLD. SAGE identified 29 deregulated transcripts including Sfpq that encodes a nuclear factor implicated in the splicing and regulation of gene expression. To assess the relevance for human disease we analyzed brains from AD, Pick's disease (PiD, a form of FTLD), and control cases. Strikingly, in AD and PiD, both dementias with a tau pathology, affected brain areas showed a virtually complete nuclear depletion of SFPQ in both neurons and astrocytes, along with cytoplasmic accumulation. Accordingly, neurons harboring either AD tangles or Pick bodies were also depleted of SFPQ. Immunoblot analysis of human entorhinal cortex samples revealed reduced SFPQ levels with advanced Braak stages suggesting that the SFPQ pathology may progress together with the tau pathology in AD. To determine a causal role for tau, we stably expressed both wild-type and P301L human tau in human SH-SY5Y neuroblastoma cells, an established cell culture model of tau pathology. The cells were differentiated by two independent methods, mitomycin C-mediated cell cycle arrest or neuronal differentiation with retinoic acid. Confocal microscopy revealed that SFPQ was confined to nuclei in non-transfected wild-type cells, whereas in wild-type and P301L tau over-expressing cells, irrespective of the differentiation method, it formed aggregates in the cytoplasm, suggesting that pathogenic tau drives SFPQ pathology in post-mitotic cells. Our findings add SFPQ to a growing list of transcription factors with an altered nucleo-cytoplasmic distribution under neurodegenerative conditions

Keeping Our Brains in Shape: The Exploration of the Relationship Between Aerobic Exercise, Cognitive Training, and Cognitive Decline

Mentor: Mark McDaniel From the Washington University Undergraduate Research Digest: WUURD, Volume 8, Issue 1, Fall 2012. Published by the Office of Undergraduate Research, Joy Zalis Kiefer Director of Undergraduate Research and Assistant Dean in the College of Arts & Sciences