Abstract
Laboratory Automation and High-Throughput Chemistry
Automated Flow-Through Synthesis of Heterocyclic Thioethers
Mark Ladlow et al. (GlaxoSmithKline Cambridge Technology Centre, University Chemical Laboratory, Cambridge, United Kingdom) describe the fully automated, sequential flow-through synthesis of a 44-member array of thioethers employing a resin capture-and-release reactor column. The array incorporates four different heterocyclic scaffolds, and the synthesis was performed with a custom-built robotic synthesizer that is able to load and regenerate the reactor column and to array each product into a single vial by using ultraviolet threshold detection. All the compounds were obtained in high yield (>75%) and excellent purity (>95%), without the need for further purification (J. Comb. Chem.
The Chemical Hunt for the Identification of Drugable Targets
Chemical biology has emerged as a new scientific discipline to change the way that scientists approach and study the interface among chemistry, biology, and physics. By integrating the knowledge base of the human genome with the power of diverse and flexible chemical technology platforms, the ultimate goal is to define the rules of engagement for small molecules as well as their use in basic biology and in drug discovery. Manfred Auer and colleagues (Novartis Institute for Biomedical Research, Vienna, Austria) highlight the current counterpoles of the chemical biology philosophy in the framework between conformational diversity and informational complexity. Lead finding with CONA (confocal nanoscanning-bead picking-AIDA technology), an integrated synthesis-screening and analysis methodology, is described. Three PICKOScreen instruments (jointly developed with and built by Evotec Technologies, Hamburg, Germany) consisting of a con-focal microscope, a scanning and picking stage, and a proprietary bead evaluation software are currently used for on-bead screening, and fluorescence fluctuation analysis at single-molecule resolution (Curr. Opin. Chem. Biol.
Kerstin Thurow, Ph.D.
Hilmar Weinmann, Ph.D.
Identification of Peptides that Promote the Rapid Precipitation of Germania Nanoparticle Networks via Use of a Peptide Display Library
Peptides that promote the rapid, room-temperature precipitation of amorphous germania nanoparticle networks from an alkoxide solution have been identified according to Matthew B. Dickerson et al. (Chem. Comm. 15(40),
Hydrophobic Polymer-Supported Scandium Catalyst for Carbon-Carbon Bond-Forming Reactions in Water
It has been revealed that a hydrophobic polymer-supported scandium(III) catalyst prepared from sulfonated polystyrene resin is an effective catalyst for carbon-carbon bond-forming reactions such as Mukaiyama aldol reactions in water. (Shinya Limura et al. in Tetrahedron 35(60),
Simple and Efficient Protocol for Catalyst Recycling and Product Recovery in the Pd-Catalyzed Homogeneous Suzuki Reaction
A simple and efficient protocol for catalyst recycling and product recovery in the Pd-catalyzed homogeneous Suzuki reaction is described by Sabrina M. Nobre et al. (Tetrahedron Lett. 34(45),
Poly(ethylene oxide)/methanol can be used as a solvent medium for the Pd-catalyzed Suzuki cross-coupling reaction under mild conditions. After the end of the reaction, the product is extracted with heptane, and the polar phase can be reused several times without any change in their activity. Pure biaryl products were obtained from the nonpolar phase in excellent isolated yields (>90%). The same catalyst-containing polar phase was also used for coupling different aryl halides furnishing the biaryl products in good to high isolated yields.
High-Throughput Purification of Combinatorial Libraries: Automated Separation of Single Diastereomers from a 4-Amido-Pyrrolidone Library Containing Intentional Diastereomer Pairs
A 4-amido-pyrrolidone library that was intentionally synthesized as pairs of diastereomers was produced by B. Yan and colleagues (Discovery Partners International, South San Francisco, CA, and Pfizer Global Research and Development, Ann Arbor, MI) via solution-phase parallel syntheses and purified by an automated high-throughput purification system. A total of 2592 4-amido-pyrrolidinones were ultimately isolated as single diastereomers from a matrix of 1920 syntheses. After the four-step synthesis and high-performance liquid chromatography purification, the average yield of a single diastereomer was 36.6%. The average chemical purity was > 90%, and the average diastereomeric purity was >87%. The choice of chiral amines used to make amides with heterocyclic acid chlorides had a dramatic effect on success. Analysis of the relationship between amines used for synthesis and the diastereomeric separation showed that amides made from chiral 1, 2-amino alcohols gave superior separation to amides from chiral morpholines. The presence of a hydrogen bond donor on the amide side chain seems to be required for a better diastereomeric separation (J. Comb. Chem.
Device for Preparing Combinatorial Libraries in Powder Metallurgy
Shoufeng Yang and Julian R. G. Evans (University of London) describe a powder-metering, -mixing, and -dispensing mechanism that can be used as a method for producing large numbers of samples for metallurgical evaluation or electrical or mechanical testing from multicomponent metal and cermet powder systems. It is designed to make use of the same commercial powders that are used in powder metallurgy and, therefore, to produce samples that are faithful to the microstructure of finished products. The particle assemblies produced by the device could be consolidated by die pressing, isostatic pressing, laser sintering, or direct melting. The powder metering valve provides both on-off and flow rate control of dry powders in open capillaries by using acoustic vibration. The valve is simple and involves no relative movement, avoiding seizure from fine powders. An orchestra of such valves can be arranged on a building platform to prepare multicomponent combinatorial libraries. (J. Comb. Chem.
Microfluidic Chip Technology and MicroReactor Technology
Combinatorial Synthesis in Micro Reactors
P. Watts and S. J. Haswell (University of Hull, Hull, UK) review the current and future applications of microreactors in the field of combinatorial chemistry and drug discovery. Liquid-phase reactions have been used to illustrate the advantages of performing chemical reactions in micro-reactors to illustrate that reactions can be performed very rapidly in high yield to enable the preparation of combinatorial libraries of structurally related compounds (Comb. Chem. High Throughput Screening
A New Synthetic Method for Controlled Polymerization by Using a Microfluidic System
K. L. Beers et al. (National Institute of Standards and Technology, Gaithersburg, MD) successfully designed a microchannel reactor for controlled polymerization. The molecular mass of the polymer produced is governed by the flow rate or polymerization time. The monomer conversion agrees well with the bulk reaction kinetics reported in the literature. The reactor is convenient and inexpensive to build, with a versatile design that can be reconfigured and prototyped in less than a day. This continuous microreactor can be used to produce libraries of materials for direct integration with high-throughput processing and characterization methods (J. Am. Chem. Soc.
Continuous Synthesis of CdSe-ZnS Composite Nanoparticles in a Microfluidic Reactor
An article in Chem. Commun. 2004, 48–49, by Hongzhi Wang et al., demonstrates that the continuous synthesis in a microreactor is a simple and efficient way to prepare composite particles with different structures and determine optimized experimental conditions.
Continuous synthesis is believed to be one advantage of microreactor technology, which can mean running up to 24 hours per day and doing analyses online. A multistep continuous synthesis in a microreactor is expected to provide better quality functional products with improved economics for complicated reactions. Now, the authors work with composite nanoparticles, such as core-shell structure nanoparticles. Here, different experimental conditions for the core and shell are usually needed, and in some cases, a new reactant has to be added during the reaction.
The experiments show that CdSe particles with different ZnS monolayers can be easily obtained by changing the flow rate or capillary lengths, which demonstrated that the multistep continuous synthesis in the microreactor was a simple and efficient way to control the composite particle structure and find the optimized experimental conditions. In summary, ZnS-coated CdSe particles have been obtained by using a total continuous method in a microfluidic reactor. The results not only displayed the same tendency in the ultraviolet and luminescence spectra as the conventional method in a flask, but also exhibited the obvious advantage of the microreactor technology in conveniently controlling the structure and properties of the nanoparticles. Through this system, CdSe particles and a ZnS coating can be produced in sequence, and particle size and layer thickness can be directly adjusted by the flow rate. At the same time, this experiment reveals the importance of the micromixer for the prefigured reaction. A more efficient mixer is expected to optimize and miniaturize this system in the future. In addition, the results show the possibility of operating a more complicated reaction in the microreactor, which means a greater potential for its application.
The Design of a Continuous Reactor for Fluorous Biphasic Reactions under Pressure and its Use in Alkene Hydroformylation
An article in J. Chem. Soc., Dalton Trans.,
The separation of the product from the catalyst and solvent can be a major barrier to the commercialization of homogeneous catalytic systems, which would otherwise be highly desirable because of their high activity, long lifetime, and tunable selectivity. This is particularly the case for high-boiling products, for which product distribution can cause decomposition of thermally sensitive catalysts. A variety of new approaches to this problem are being investigated.
The authors report a reactor in which reactions can be carried out under moderate pressures of gaseous reagents and illustrate its use in the rhodium-catalyzed hydroformylation of 1-octene. The gases are fed from gas cylinders through mass flow controllers into a stirred tank reactor fitted with a thermocouple and bursting disc. The fluorous solvent containing the catalyst and the liquid substrate (dissolved in a suitable solvent, if required) are fed through separate high-performance liquid chromatography (HPLC) pumps and a heat exchanger into the reactor. The reaction mixture can be removed from the bottom of the reactor through a dip tube and passed through the heat exchanger, a capillary, and a flow meter into a gravity separator, where the dissolved gases are released. At the opposite end from the entry port, the separator is fitted with an overflow tube through which the organic phase passes and is collected.
The catalyst phase is recycled to the reactor through the fluorous HPLC pump and the heat exchanger. The organic phase is replenished with new substrate from the organic reservoir through the organic HPLC pump and the heat exchanger. All flows can be varied independently, and the overall flow rate through the system is determined by the gas pressure and the resistance of the capillary (length and diameter). Four capillaries can be switched in or out of the flow by using multiport valves at both ends to allow different flow rates and, hence, residence times within the reactor. The authors describe the construction of a reactor that is capable of carrying out reactions using liquid and gaseous reagents under continuous flow for many hours. The use of the reactor is demonstrated with the hydroformylation of 1-octene, and the effects of phosphine leaching are described. In the reaction discussed, which was run for 1 hour in batch mode and a further 19 hours continuously, the catalyst underwent > 15,500 turnovers at an average rate of 750 per hour. This compares favorably with commercial rhodium-catalyzed processes for propene hydroformylation, which have rates in the region of 500 to 700 per hour.
High-Throughput Analytics
Prediction of MS/MS Data. 1. A Focus on Pharmaceuticals Containing Carboxylic Acids
The identification of metabolites is an important chapter in the development of safe and effective new drugs and is typically associated with HPLC-MS methods. In a report by H. Desaire and coworkers, recent efforts focused on the development of predictive rules for structural characterization of carboxylic acids in pharmaceuticals via MS/MS spectra are presented (Anal. Chem.
As the basis of the studies, the loss of CO2, an abundant fragmentation pathway, was analyzed for about 100 carboxylic acids under specified low-energy collision-induced dissociation conditions on a quadrupole ion trap mass spectrometer. Afterward, the tested substrates were systematically classified into four separate subclasses, based on functional groups in proximity to the carboxylic acid. Additional information was apparent, and general rules about CO2 dissociation could be revealed. To verify the predictability and completeness of the rules, 20 different pharmaceuticals that contained a carboxylic acid functional group were analyzed and compared with the assignments regarding the rules. A 90% success rate for this first test shows that MS/MS data acquired under specified conditions will be predictable, based on a compound's structure.
First Screening of Low Molecular Weight Compounds by Thin-Layer Chromatography and On-Spot MALDI-TOF Mass Spectrometry
M. N. Eberlin and coworkers (State University of Campinas, Sao Paulo, Brazil) developed a new implementation for the combination of thin-layer chromatography (TLC) and direct “one-spot” positive ion MALDI-TOF mass spectrometry (TMT-MS) for low molecular weight compounds (Anal. Chem.
Because of the widespread usage of TLC as a fast and easy separation technique, a combination with MALDI-TOF mass spectrometry has already been tested with high molecular weight analytes such as proteins, polymers, and peptides, whereas the detection of low molecular weight analytes suffered from interferences with background ions. The newly introduced TMT-MS method uses triethylamine-α-cyano-4-hydroxycinnamic acid (Et3N·α-CHCA), an ultraviolet-absorbing proton donor ionic liquid, as the matrix. Preparation of TLC plates can easily be achieved by doping each spot with a few microliters of a 0.1% (w/v) acetonitrile solution of the ionic liquid with subsequent solvent evaporation via exposure to high vacuum. Several alkaloids, anesthetics, and antibiotics were tested and showed a significant reduction of matrix background and an enhanced sensitivity compared with precedent on-spot TMT-MS methods. This seems to be an ideal method for analyzing on-the-bench TLC experiments as well as for quality monitoring in product manufacturing.
High-Throughput Characterization and Quality Control of Small-Molecule Combinatorial Libraries
To fully realize the potential of combinatorial synthesis and high-throughput screening for increasing the efficiency of the drug discovery and development process, issues related to compound purity must be addressed. Impurities, often present after synthesis, can lead to ambiguous screening results and inhibit the development of quality structure-activity relationships. The demand for high-throughput analytical characterization of combinatorial libraries has prompted the development of more rapid methods to keep pace with compound production. Recent progress has focused upon the development of parallel separation methods, multiplexed detector interfaces, and synergistic combinations of different detectors possessing complementary selectivities. J. R. Kenseth and S. J. Coldiron (CombiSep, Inc., Ames, IA) summarize recent trends in high-throughput characterization and quality control (Curr. Opin. Chem. Biol.
MS-Express: Data-Extracting and -Processing Software for High-Throughput Experimentation with Mass Spectrometry
High-throughput experiments (HTE) result in large amounts of raw data that have to be evaluated for sample classification. Mass spectrometry (MS) in particular, which is a widely used detection method in catalytic HTE applications, produces enormous amounts of data. In a typical HTE, the catalysts are tested sequentially. The recording of the scans, however, occurs continuously. For this reason, the scans of interest must be extracted from the raw data, and scans belonging to the same sample must be averaged in a tedious procedure before further processing. W. F. Maier and coworkers (University of Saarbruecken, Saarbruecken, Germany) present their custom-designed software, MS-Express (mass spectrometry data-extracting and -processing software), an efficient tool for HTE MS data evaluation. Besides sorting the data, the program also establishes statistical significance with the help of reference and blank data and provides concise information about abundance and intensity distributions of expected peaks. A special feature is that the program also reports unexpected MS signals, which potentially lead to unexpected discoveries (J. Comb. Chem.
Bioautomation and Screening
A Novel Encoded Particle Technology that Enables Simultaneous Interrogation of Multiple Cell Types
The implementation of multiplexing technologies in biological and drug discovery research is recognized to have clear advantages: decreased reagent usage, smaller sample requirement, increased sample throughput, and the use of internal controls for comparisons.
Beske et al. (J. Biomol. Screen.
A Miniaturized Column Chromatography Method for Measuring Receptor-Mediated Inositol Phosphate Accumulation
The signaling of many neurotransmitters and hormones is critically dependent on G protein-coupled receptor (GPCR) activated cascades. The rapid and accurate assessment of GPCR-mediated phospholipase C (PLC) pathway activation, particularly accumulation of inositol phosphates (IPs), such as inositol-1,4,5-phosphate (IP3), is valuable for a number of pharmaceutical discovery and basic research needs. Classic methods for measuring intracellular accumulation of these molecules include time-consuming high-performance liquid chromatography separation or large-volume, gravity-fed anion-exchange column chromatography. More recent approaches (radio-receptor and AlphaScreen assays) offer higher throughput but rely on measurement of IP3 itself, rather than its accumulation with other downstream IPs.
Benjamin et al. (J. Biomol. Screen.
The miniaturized protocol described saves time and reagents. It is conducted in a standard 96-well compound screening format and is compatible with common HTS laboratory equipment.
Flow Cytometry for High-Throughput, High-Content Screening
Flow cytometry is a mature platform for quantitative multiparameter measurement of cell fluorescence. L. A. Sklar and colleagues (University of New Mexico, Albuquerque, NM) review recent innovations that allow up to 30-fold faster serial processing of bulk cell samples. Homogeneous discrimination of free and cellbound fluorescent probes eliminates wash steps to streamline sample processing. Compound screening throughput may be further enhanced by multiplexing of assays on color-coded bead or cell suspension arrays and by integrating computational techniques to create smaller, focused compound libraries. Novel bead-based assay systems allow studies of real-time interactions between solubilized receptors, ligands, and molecular signaling components that recapitulate and extend measurements in intact cells. These new developments, and their broad use, position flow cytometry as an attractive analysis platform for high-throughput, high-content biological testing, and drug discovery (Curr. Opin. Chem. Biol.
Theory and Applications of NMR-Based Screening in Pharmaceutical Research
Since the first report appeared in 1996 describing the use of NMR spectroscopy to screen for potential drug molecules, the field of NMR-based screening has evolved rapidly. Over the last several years, a variety of novel approaches have been introduced and have found widespread application in both pharmaceutical and academic research settings. These NMR-driven platforms have provided new pathways for inhibitor design against an increasing number of therapeutically relevant drug targets. In pharmaceutical research, NMR screening has become an important component in an integrated arsenal of biophysical, biochemical, and computational methods designed to discover and optimize drug leads. A comprehensive review of this technology platform is presented by Jonathan M. Moore et al. (Vertex Pharmaceuticals Inc. and University of Notre Dame, Notre Dame, IN) in Chem. Rev.
Reliable High-Throughput Functional Screening with 3-FABS
An NMR method called 3-FABS has extended the capabilities of NMR, enabling rapid, efficient, and reliable high-throughput functional screening for the identification of inhibitors and for measuring their 50% mean inhibition concentration with accuracy. The substrate is tagged with a CF3 moiety, and 19F NMR spectroscopy is used for the detection of the substrate and product components. Claudio Dalvit and colleagues (Chemistry and Biology Departments, Nerviano Medical Science, Nerviano, Italy) provide comprehensive insight into 3-FABS, including a discussion of its strengths and weaknesses when compared with other HTS techniques and a presentation of some of its applications to the screening of different enzymes and to multiple screening (Drug Discovery Today
Virtual Screening Methods that Complement HTS
Florence L. Stahura and Jürgen Bajorath (Albany Molecular Research, Bothell, WA, and University of Washington, Seattle, WA) discuss a number of computational methods that have been developed or adapted for molecule classification and virtual screening of compound databases. In particular, they focus on approaches that are complementary to high-throughput screening. They describe areas that greatly benefit from combining virtual and biological screening and discussed computational methods that are most suitable to contribute to the integration of screening technologies (Combinatorial Chem. High Throughput Screen.