Preparation of Stereo-Divergent Compounds from the Natural Product Drupacine Based on Complexity-to-Diversity Strategy

The Complexity-to-Diversity (CtD) strategy was applied to synthesize a 23-member compound collection from the natural product drupacine, including 21 novel compounds. An unusual benzo [d] cyclopenta [b] azepin skeleton was constructed by Von Braun reaction to cleave C−N bond of drupacine. Moreover, compound 10 has potential cytotoxicity to human colon cancer cells with low toxicity to the normal human colon mucosal epithelial cell lines.     

Temperature manifold for a stopped-flow machine to allow measurements from −10 to +40 °C

Conducting enzymatic stopped-flow experiments at temperatures far removed from ambient can be very problematic because extremes in temperature (<10 °C or >30 °C) can damage the machine or the enzyme. We have devised a simple manifold that can be attached to most commercial stopped-flow systems that is independently heated or cooled separate from the main stopped-flow system. Careful calibration of the flow circuit allows the sample to be heated or cooled to the measurement temperature (−8 to +40 °C) 1 to 2 s before mixing in the reaction chamber. This approach allows measurements at temperatures where the stopped flow or the protein is normally unstable. To validate the manifold, we investigated the well-defined ATP-induced dissociation of rabbit muscle myosin subfragment 1 (S1) from its complex with pyrene-labeled actin. This process has both temperature-dependent and -independent components. Use of ethylene glycol allowed us to measure the reaction below 0 °C and up to 42 °C, and as expected the second-order rate constant (K₁k₊₂) and the maximum rate of dissociation (k₊₂) both increased with temperature, whereas 1/K₁ is unaffected by the change in temperature.     

Influence of chain length on mono- versus di-alkylation in the reactivity of [Pt2(μ-S)2(PPh3)4] towards α,ω-dihalo-n-alkanes; a synthetic route to platinum(II) ω-haloalkylthiolate complexes

The reactions of [Pt₂(μ-S)₂(PPh₃)₄] with α,ω-dibromoalkanes Br(CH₂)_nBr (n = 4, 5, 6, 8, 12) gave mono-alkylated [Pt₂(μ-S){μ-S(CH₂)_nBr}(PPh₃)₄]⁺ and/or di-alkylated [Pt₂(μ-S(CH₂)_nS}(PPh₃)₄]²⁺ products, depending on the alkyl chain length and the reaction conditions. With longer chains (n = 8, 12), intramolecular di-alkylation does not proceed in refluxing methanol, with the mono-alkylated products [Pt₂(μ-S){μ-S(CH₂)_nBr}(PPh₃)₄]⁺ being the dominant products when excess alkylating agent is used. The bridged complex [{Pt₂(μ-S)₂(PPh₃)₄}₂{μ-(CH₂)₁₂}]²⁺ was accessible from the reaction of [Pt₂(μ-S)₂(PPh₃)₄] with 0.5 mol equivalents of Br(CH₂)₁₂Br. [Pt₂(μ-S){μ-S(CH₂)₄Br}(PPh₃)₄]⁺ can be cleanly isolated as its BPh₄⁻ salt, but undergoes facile intramolecular di-alkylation at −18 °C, giving the known species [Pt₂(μ-S(CH₂)₄S}(PPh₃)₄]²⁺. The reaction of I(CH₂)₆I with [Pt₂(μ-S)₂(PPh₃)₄] similarly gives [Pt₂(μ-S){μ-S(CH₂)₆I}(PPh₃)₄]⁺, which is fairly stable towards intramolecular di-alkylation once isolated. These reactions provide a facile route to ω-haloalkylthiolate complexes which are poorly defined in the literature. X-ray crystal structures of [Pt₂(μ-S){μ-S(CH₂)₅Br}(PPh₃)₄]BPh₄ and [Pt₂(μ-S(CH₂)₅S}(PPh₃)₄](BPh₄)₂ are reported, together with a study of these complexes by electrospray ionisation mass spectrometry. All complexes fragment by dissociation of PPh₃ ligands, and the bromoalkylthiolate complexes show additional fragment ions [Pt₂(μ-S){μ-S(CH₂)_n−2CHCH₂}(PPh₃)_m]⁺ (m = 2 or 3; m ≠ 4), most significant for n = 4, formed by elimination of HBr.     

The mechanism of Rubisco‐catalysed oxygenation

Ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco) is the cornerstone of photosynthetic carbon assimilation because it catalyses the fixation of CO₂ onto ribulose‐1,5‐bisphosphate (RuBP). The enzyme also catalyses RuBP oxygenation, thereby evolving phosphoglycolate which is recycled along the photorespiratory pathway. Oxygenation is quantitatively important, because under ordinary gaseous conditions, more than one third of RuBP molecules are oxygenated rather than carboxylated. However, contrary to carboxylation, the chemical mechanism of oxygenation is not well known, and little progress has been made since the early 80s. Here, I review recent experimental data that provide some new insights into the reaction mechanism, and carry out simple calculations of kinetic parameters. Isotope effects suggest that oxygenation is less likely initiated by a redox phenomenon (such as superoxide production) and more likely involves concerted chemical events that imply interactions with protons. A possible energy profile of the reaction is drawn which suggests that the generation of the oxygenated reaction intermediate (peroxide) is irreversible. Possible changes in oxygenation‐associated rate constants between Rubisco forms are discussed.     

Formation of electronically excited states during the interaction of p-benzoquinone with hydrogen peroxide

The reaction between p-benzoquinone and H₂O₂ in slightly alkaline solutions yields three major quinoid products that accumulate in the reaction mixture: (a) 2,3-epoxy-p-benzoquinone, (b) 2-hydroxy-p-benzoquinone and (c) p-benzohydroquinone. The reaction is accompanied by photoemission, probably originating from excited triplet 2-hydroxy-p-benzoquinone. These products originate from hydrogen peroxide and hydroxide nucleophilic addition to the C₂?C₃ double bond, as well as secondary redox interactions. The hydroxy substituent and the epoxide ring exert a substantial influence on the electronic distribution in the p-benzoquinone molecule leading to a decrease in the half-wave potential, as compared to the parent p-benzoquinone. The generation of electronically excited states is the result of reactions secondary to the nucleophilic additions involving 2-hydroxy-p-benzosemiquinone, H₂O₂ and hydroxyl radical. The process involves the primary oxidation of 2-hydroxy-p-benzosemiquinone by hydrogen peroxide, followed by oxidation of the semiquinone by hydroxyl radical leading to the formation of the electronically excited quinone. The decay of the excited triplet to the ground state is accompanied by photoemission with maximal intensity at 485–530 nm. Thermodynamic calculations along with an observed increase of photoemission intensity in anaerobiosis point to the triplet (n, π*) multiplicity of the excited state. The efficiency of chemiluminescence could be calculated as 10^?8 photons/2-hydroxy-p-benzoquinone molecule formed. Photoemission arising from the p-benzoquinone/H₂O₂ reaction was inhibited efficiently by addition of GSH to the reaction mixture. This may be due to deactivation of the triplet quinone by a 2-glutathionyl-p-benzohydroquinone adduct, involving thioether α-hydrogen atom-transfer to the triplet ketone.     

Multicomponent chemistry in the synthesis of carbonic anhydrase inhibitors

Abstract

Carbonic anhydrase inhibitors (CAIs) are of growing interest since various isoforms of the enzyme are identified as promising drug targets for treatment of disease. The principal drawback of the clinically used CAIs is the lack of isoform selectivity, which may lead to observable side effects. Studies aiming at the design of isoform-selective CAIs entail generation and biological testing of arrays of compounds, which is a resource- and time-consuming process. Employment of multicomponent reactions is an efficient synthetic strategy in terms of gaining convenient and speedy access to a range of scaffolds with a high degree of molecular diversity. However, this powerful tool appears to be underutilized for the discovery of novel CAIs. A number of studies employing multicomponent reactions in CAI synthesis have been reported in literature. Some of these reports provide inspiring examples of successful use of multicomponent chemistry to construct novel potent and often isoform-selective inhibitors. On critical reading of several publications, however, it becomes apparent that for some chemical series designed as CAIs, the desired inhibitory properties are only assumed and never tested for. In these cases, the biological profile is reported based on the results of phenotypical cellular assays, with no correlation with the intended on-target activity. Present review aims at critically assessing the current literature on the multicomponent chemistry in the CAI design.     

Evaluation of monocyte subsets and markers of activation in leprosy reactions

Understanding host immune pathways associated with tissue damage during reactions are of upmost importance to the development of immune intervention strategies. The participation of monocytes in leprosy reactions was evaluated by determining the frequency of monocyte subsets and the degree of cellular activation through the expression of MHCII and the co-stimulatory molecules CD40, CD80, CD86. Leprosy subjects with or without reactions were included in this cross-sectional study. Peripheral blood mononuclear cell were isolated and stained ex vivo to determine monocyte subsets and the degree of cellular activation by flow cytometry. Intermediate monocytes were increased in leprosy patients with reactions when compared to patients without reactions. Although no difference was detected in the frequency of monocyte subsets between type 1 and 2 reactions, the expression of CD80 was increased in monocytes from patients with type 1 reactions and CD40 was higher in paucibacillary subjects presenting type 1 reactions. Moreover, CD86 and MHC II expression were higher in intermediate monocytes when compared to the other subsets in leprosy reaction types 1 and 2. Intermediate monocyte activation with CD86 and MHCII expression is involved with both type 1 and 2 reactions, whereas CD80 and CD40 expression is related to type 1 reactions.     

Application of immunoproteomics to rapid cytokine detection

Cytokines are pivotal to a balanced innate or cell-mediated immune response, and can be indicative of disease progression and/or resolution. Methods to measure key cytokines rapidly with accuracy, precision, and sensitivity are consequently important. The current assay technologies, which are based on RT-PCR, immunoassays, or bioassays, are limited in their use in the clinic, in particular because of the long time (1-3 h) required to carry out the assays. An alternative, semi-quantitative approach described here, uses an immunological capture step and a mass spectral readout. The goal of the assay is speed rather than sensitivity or precision.