2015 Technical Posters

CoSMoS 2015 Technical Posters

August 17 – 19, 2014

Sheraton San Diego Hotel & Marina, San Diego, CA

 

POSTER RECEPTION:

Posters will be up at the start of the conference. We have dedicated time to view posters during session breaks and visit with our exhibitors each day and a reception Tuesday beginning at 5:00 PM. Poster authors will be available between 5:00 and 6:00 PM. Take advantage of this time to speak with the poster authors and our exhibitors while enjoying some food and drink.

 

Poster #1
Hanna S. Yuan
Academia Sinica

 

Inhibition of bacterial growth by the small molecule inhibitors of exonucleases
The 3′-to-5′ exonucleases that process nucleic acids by removing one nucleotide at a time from 3′ to 5′ end are essential for bacterial growth. These exonucleases bear various essential functions in prokaryotic or eukaryotic cells, including RNA maturation, RNA turnover, DNA replication and DNA repair. It has been shown that knockout of one or two exonucleases may generate a slow or no growth phenotype in E. coli, suggesting that inhibition of these exonucleases might be a way to reduce bacterial cell growth. Here we screened the small molecule compounds for the inhibition of exonucleases, and identified several potential inhibitors that can reduce the exonuclease activities of these enzymes. These compounds might be useful for the development of antibiotic or antivial agents.
Poster #2
John Jasper
Nature’s Fingerprint MIT LLC
Molecular Isotopic Engineering: Industrial Manufacture of Naproxen of Predetermined Stable-Isotopic Compositions for Reasons of Identity and Security
Molecular Isotope Technologies LLC has developed four patented or patent-pending generations of stable-isotopic methods and technologies: (i) product characterization (for both small molecules and biologics), (ii) process characterization (notably, process patent protection), (iii) in-process (continuous) analysis, and now (iv) molecular isotopic engineering. Early work in cooperation with the US FDA on the product characterization of naproxen revealed manufacturer-level isotopic provenance of this small analgesic molecule (Wokovich et al., 2004) which was referred to as “The Manufacturer’s Fingerprint.” This isotopic provenance represented the convergence of the effects of the stable-isotopic compositions of starting materials and isotopic effects of the synthetic process. Rather than merely accepting the random effects of variable sourcing and synthetic process on the stable-isotopic compositions of products, we take a proactive approach to purposefully determine the stable-isotopic composition of bio/pharmaceutical products. The main rationale for MIE is to predetermine the isotopic ranges of products for reasons of product identification and of product security, and also for intellectual property considerations. As an example of MIE, we analyzed the products of the isotopic-synthetic reactions for the last two steps of naproxen synthesis
Poster # 3
Joseph Simpkins
Virscidian
 
Feeding the NCE pipeline – Quality by design in high throughput analytical decision making
Software assisted results interpretation that supports quality by design strategies in the field of analytical results. Automated results interpretation strategies have been developed and are continually extended and refined to deliver up to five categories of end-points: Automated determination of suitability for review, automated QC of results, derivative result calculations, result interpretation strategies and automated messaging that reflects manual comments.
Poster # 4
Christine Andrews
Merck
 
Rank Ordering of Compounds in Complex, Microscale Mixtures via Relative Affinity Measurements Utilizing Two-dimensional Affinity Selection – Mass Spectrometry
ALIS (Automated Ligand Identification System) is an affinity selection-mass spectrometry based screening technique utilizing two dimensional (SEC-RPC) chromatography coupled to mass spectrometry to analyze protein: ligand complexes. ALIS is a label-free technique and each protein of interest can be incubated with complex mixtures of small molecule ligands, typically up to 2,500 compounds/ mix in screening mode. Given its flexible nature and low method development requirements, ALIS is an ideal technology to address multiple challenges in drug discovery.In the pharmaceutical industry, there is a continuous search for tools to expedite the selection of a preclinical candidate.  In addition to screening, ALIS has proven to be an effective tool to rank order analogs in mixtures at sub-microgram quantities.   This capability can be capitalized on to prioritize chemistry resources for potent compounds, and minimize work on low probability of success library components. Using the ALIS relative affinity ranking approach, we investigated the binding and relative ranking of compounds from crude library mixtures optimized for binding to the target of interest. At the same time, these compounds were purified and assayed using the standard biochemical assay for the program.   Results from the affinity ranking correlate well with activity data obtained on the purified compounds.   These experiments represent a unique capability to assay complex mixtures and unpurified samples, leading to more thorough and efficient SAR exploration by increasing return on investment, as well as potentially allowing the exploration of more challenging or costly synthetic avenues at a reasonable scale for evaluation. 
Poster # 5
Amber Guillen
Genentech
 
UHPLC Evaluation of Kinetex EVO C18 Columns 
It has been shown that pH can have profound effects on the selectivity [1] and loadability [2] of analytes in analytical and preparative reversed-phase HPLC separations. In today’s market, many column manufacturers produce fully-porous particles which are able to tolerate low and high pH conditions allowing analysts to take advantages of manipulating pH. Other developments within HPLC include the use of core-shell particles [3] to gain improvements in efficiency and pressure when compared to fully-porous of the same size on both HPLC and UHPLC instrumentation [4]. Until recently, core-shell technology did not tolerate high pH conditions. The introduction of Phenomenex’s Kinetex EVO C18 aims to combine the selectivity of varying pH and the efficiency of core-shell particles. To evaluate the performance of these new columns, the Van Deemter plots of two commercial small molecules were obtained on multiple columns of varying particle size, particle type and manufacturer. From this study, the Kinetex EVO stationary phase did show improved performance but was not the best column tested. 
Poster  #6
Yande Huang
Bristol-Myers Squibb Co
 
Structure Identification and Mechanistic Understanding of Two Color Degradants during the Early Development of BMS-911543
During the early development of BMS-911543 as a JAK2 inhibitor candidate for the treatment of myeloproliferative neoplasms, tablets were discolored when stored under room temperature/room light for a few days. Additionally, a batch of BMS-911543 drug substance (API) with a pale brown color was isolated from the mother liquor with a deep red color. This was unusual as the typical process produced off-white API from golden-brown mother liquor. Furthermore, when stressed under high intensity light/UV light for three days, the solution of the off-white API in ACN/H2O (1:1 v/v) changed from colorless to yellow, while the other off-white API solution in ACN became bright red. LC-UV/MS analysis and preparative HPLC isolation were used to investigate the root causes of the color changes in the mother liquor and the photodegradation solutions. A red degradant from the mother liquor and a pale yellow degradant from the photodegradation solution were isolated and characterized using 1D and 2D NMR and HRMS. These two degradants were also observed in the discolored tablets at trace level. Mechanistic analysis suggested the formation of these color degradants via singlet oxygen oxidation. This hypothesis was supported by the oxidative degradation of the API with Rose Bengal, a typical singlet oxygen sensitizer. Rationale of structure identification and proposed mechanism of formation will be presented.
Poster #7
Carol Preisig
ACD Labs
 
A Unified Software Platform for Laboratory Informatics
Wise implementation of informatics technology for creating knowledge management infrastructure to cope with the variety and volume of scientific workflows and analytical information being generated from modern instrumentation is the aim of R&D organizations. Simultaneously with simplifying IT infrastructure for data management are the need and opportunity to leverage advanced analytical science to generate knowledge from those data and support evolving user requirements and preferences. Amalgamation is one key data management theme for simplification of disparate chemistry-related software technologies such as CDS, ELN, LES, LIMS, SDMS, and data warehouses, which have proliferated throughout organizations. Simple amalgamation isn’t satisfactory, hence technologies that are designed for effectively integrating, automating and communicating in Laboratory Informatics Environments (LIE) are of growing interest. Reported here are capabilities in automated workflows involving analytical data with chemical structures using the ACD/Spectrus informatics platform. Specifically described herein is automated homogenization of data from a set of instruments, including NMR structure verification, as one solution and automated metabolite analysis as another. The automation of the LC/UV-MS purity check prior to compound registration is another solution widely deployed on ACD/Spectrus. The same platform forms the basis of a prototype integration with a commercial ELN. These ACD/Spectrus solutions feature a variety and volume of analytical data in their workflows and illustrate how unified laboratory intelligence technologies can address knowledge management needs in chemistry R&D subdisciplines. Furthermore, they showcase how a platform with an architecture that is amenable to integration and to cloud-based deployment can enable workflows with automated data analysis, interpretation, and knowledge capture along with visualization via thick or web or mobile client interfaces.
Poster #8
Ole Gron
Vertex Pharmaceuticals
 
Evaluation of different approaches to multiple heart cut sampling in 2D-LC
In small molecule analysis the chromatographic complexity is in most instances less than it is in natural product and peptide analysis. Nonetheless, there are often unresolved or overlapping peaks which require improved separation. 2D-LC can be divided into two main groups: comprehensive and heart cut. Comprehensive 2D-LC is a way of increasing the peak capacity for a LC run with hundreds of peaks without any additional separation time. This requires a very short 2nd dimension run with some degree of orthogonality to the first dimension. Multiple Heart-Cut (m-HC) 2D-LC, on the other hand, is mainly focused on the analysis of fewer but more difficult to separate peaks. In mHC-2D-LC, the 2nd dimension gradient can be longer than in comprehensive 2D-LC because the loop fill time does not have to equal the 2nd dimension run time. This provides a greater opportunity to successfully resolve structurally similar co-eluting components from the 1st dimension.   This presentation will highlight work that has been performed in small molecule pharmaceutical development using m-HC 2D-LC and discuss the core principles and setup of such a system. It will highlight the importance of many of the parameters of which one needs to be aware in order to successfully couple the two dimensions. For example, injections of large plugs of solvents from the first dimension onto the 2nd dimension can be difficult and thus the selection of columns, flow rate, loop size and heart cut volume is important. Additionally, this presentation will discuss the importance of orthogonality of the two dimensions and show examples of small molecule separations illustrating 2D-LCs utility in detection and quantification of co-eluting impurities.
Poster #9
Charles McEwen
University of the Sciences
 
Results from a new prototype source for inlet ionization mass spectrometry
Inlet ionization technologies capable of converting small, large, volatile, and nonvolatile compounds into gas-phase ions for analysis by mass spectrometry are astonishingly simple and sensitive. No high-voltage, laser, or nebulizing gas is required, and clogged capillaries are eliminated. Ionization is assisted by the vacuum of the mass spectrometer and a matrix. The matrix can be either a solid small molecule similar to MALDI or a solution as in ESI. Sample is introduced into the atmospheric pressure to vacuum inlet of the mass spectrometer either continuously from a liquid chromatograph or discretely using a solid matrix. Matrix-assisted ionization (MAI) is similar to MALDI without the laser desorption, and electrospray ionization inlet (ESII) is ESI occurring within the inlet and thus not requiring nebulizing gases. MAI and ESII show reproducibility and quantification comparable to ESI. The commercial ESI/APCI ion source is replaced by a mechanical prototype source that allows a syringe needle tip, or fused silica capillary, to be guided into the inlet aperture of the mass spectrometer. For MAI, a 1 uL syringe pulls in 0.5 uL of sample, expels it back into the container and then pulls in 0.5 uL of a concentrated matrix solution. The matrix is expelled to dry on the syringe needle tip before it is inserted into the inlet using the prototype source. For ESII, a metal union with voltage applied is connected on one end to the LC column and on the other end to fused silica capillary, the exit of which is placed into the inlet. With MAI, full acquisition mass spectra are obtained in seconds consuming as little as 70 nL of a 100 ppb solution. With ESII, ion abundances 3 – 10X greater than ESI are obtained Examples include analysis of mixtures such as protein tryptic digests and drugs in biological fluids.
Poster #10
Sarah Trimpin
Wayne State University
 
Laserless Matrix-Assisted Ionization-Mass Spectrometry: A Novel Method Potentially Suitable for Field Portable Applications
Laserless matrix-assisted ionization (MAI) is rapid, requires minimal training, and reduces the energy necessary for ionization making it potentially ideal for use in clinical, forensic, and homeland security applications, as well as field-portable mass spectrometers allowing simultaneous detection and quantification of a wide array of chemical and biological agents. Waters SYNAPT G2 and QDa mass spectrometers with the ESI ion source open and interlocks overridden were used in this study. The ion source temperature was maintained at 50 °C for positive and 80 °C for negative mode measurements. 3-Nitrobenzonitrile and 1,2-dicyanobenzene where used as matrices dissolved in 70% acetonitrile:water. Analyte was dissolved in water or 1:1 acetonitrile:water and the matrix and analyte solutions were mixed 1:1 and allowed to dry before being introduced to the mass spectrometer inlet. Biological fluids, as obtained, were mixed 1:1 with matrix solution. The matrix:analyte solution (1 µL) was touched dry against the inlet aperture.   MAI is applicable to ionizing analytes with sensitivities in the low femtomole range for drugs and proteins. We show reproducibility of <10% for detection of illicit drugs such as morphine using direct analysis with internal standards. This performance is competitive with ESI but at much higher speed (~4 sec per sample). Surfaces can be readily investigated such as paper and TLC plates. We demonstrate the feasibility of identifying drugs and metabolites directly from complex biological fluids including saliva, whole blood and urine using MAI-MS. Contrary to ESI, MAI is robust towards salts, buffers, and detergents. Furthermore, we demonstrate its applicability to a low pumping capacity mass spectrometer as a mimic for a field portable instrument. MAI-MS requires minimal training, making it ideal for use in clinical, forensic and homeland security applications.
Poster #11
Giuseppe Astarita
Waters Corporation
 
Multidimensional analytical approaches: combining ion mobility and spectrophotometric detection with current MS-based metabolomics and lipidomics workflows
Here we propose a novel strategy that combines a typical ultra high performance liquid chromatography (UHPLC), data-independent mass spectrometry (MSE) workflow with travelling wave ion mobility (TWIM) and UV detection, to improve the characterization of carotenoids and chlorophylls in complex biological matrices. UV detection selectively highlighted pigments absorbing at specific wavelengths, while TWIM coupled to MS was used to maximize the peak capacity. We applied this approach for the analysis of pigments in different microalgae samples, including Chlorella vulgaris, Dunaliella salina and Phaeodactylum tricornutum. Using UHPLC-UV-MSE information (retention time, absorbance at 450 nm and accurate masses of precursors and product ions), we tentatively identified 26 different pigments (carotenes, chlorophylls and xanthophylls). By adding TWIM information (collision cross sections), we further resolved 5 isobaric pigments, not resolved by UHPLC-UV-MSE alone. The characterization of the molecular phenotypes allowed us to differentiate the microalgae species. Our results demonstrate that a combination of TWIM and UV detection with traditional analytical approaches increases the selectivity and specificity of analysis, providing a new tool to characterize pigments in biological samples. We anticipate that such an analytical approach will be extended to other lipidomics and metabolomics applications.
Poster #12
Giorgis Isaac
Waters Corporation
 
Screening and Identification of Undeclared Synthetic Compounds as Adulterants using UPLC-Qtof-MS Coupled to a Novel Informatics Platform
Two commercial herbal product samples to relief pain and inflammations were obtained from India and Taiwan. 1g of powdered sample was extracted using 10 mL of methanol by ultrasonication for 20 minutes followed by centrifugation at RCF of 4472 g for 5 minutes. The supernatant was diluted 50 times with methanol and injected into an UPLC system using a sub-1.8 µm UPLC HSS T3 column (100 mm×2.1 mm) column coupled with a QTOF mass spectrometer in data independent acquisition mode which allowed simultaneous collection of both low (intact molecule) and high energy (fragment ions) data from a single injection. The instrument control, method development, data acquisition, processing and analysis were conducted using the informatics software platform provided from the instrument vendor. A natural product analytical workflow was used to analyze the mass spectra of the herbal products which utilize tools such as 3D peak detection, an adulterant database library containing accurate mass fragment ions for confirmation of analytes of interest, structure elucidation module and automatic reports to effectively provide the visualization of the results.For the commercial sample obtained from India, phenylbytazone, an anti-inflammatory agent was detected as an adulterant using an internal database library. In the US and UK, phenylbytazone is no longer approved for human use, as it can cause severe adverse effects such as suppression of white blood cell production and aplastic anemia. This drug was one of the main components in the 2013 meat adulteration scandal. In addition to phenylbytazone, an unknown metabolite was detected as well. Using the structure elucidation module which utilizes elemental composition determination, online library searches and fragment ion matching, oxyphenbutazone, a metabolite/derivative of phenylbutazone was also identified as an adulterant in the same sample. No major adulterants were found from the sample purchased from Taiwan. 
Poster #13
David Stranz
Sierra Analytics, Inc.
 
Comprehensive Chemical Formula Assignment to Mass Spectra of Complex Sample Mixtures
Most methods for assignment of chemical formulas to mass spectral features do so by considering the peaks on a one-by-one basis. In spectra of complex mixtures, there is typically a large number of relationships among spectral features, such as peaks belonging to the same isotope cluster, peaks related by charge state or adduct difference, and peaks related by m/z differences that can be attributed to chemical formula differences. These reinforcing relationships are either inherent to the substance itself (the isotope cluster), induced through the sampling and ionization process (charge states and adducts), or arise through coincidental or chemical family relationships among sample components.
Poster #14
Ryan Cohen
Merck
 
Comparison of quantitative NMR using CRAFT versus chromatography for complex mixture analysis
Quantitative NMR is a widely used method in the pharmaceutical industry for determination of salt ratios, residual solvents, and major component assays. Several recognized advantages are a broad linear dynamic range, uniform response, and often simple method development. However, a few major disadvantages have hindered its usage versus other techniques (e.g., chromatography). These include the expense of purchasing and maintaining high field spectrometers, analyst-to-analyst variability, poor sensitivity, and inadequate spectral resolution for complex mixtures. Many of these limitations have recently been or are currently being addressed by advances in NMR technology and pulse sequence development.
Here we applied CRAFT (Complete Reduction to Amplitude Frequency Tables) for analysis of several complex small molecule mixtures. CRAFT enables quantitation in the time domain and therefore does not require phase correction, baseline manipulation, and manual integration.   Thus, the method is inherently less prone to analyst variability. In addition, the technique allows for a novel form of spectral
deconvolution, such that nearly overlapping signals may be accurately quantified. Furthermore, results obtained by CRAFT were directly compared versus traditional quantitative NMR, as well as, chromatographic data. Several cases were observed where accuracy was much better using CRAFT, such as overlapping signals or signals on top of rising baselines (e.g., from broad resonances).
Poster #15
Lucas Westling
Novartis
Throughput Comparison of Fast Inlet Instruments for ADME Assays 
ADME (absorption, distribution, metabolism, elimination) assays provide information which medicinal chemists use to optimize the properties of their compounds. Fast turnaround time of assay screening data is crucial to efficient drug discovery. These rapid screening assays create large numbers of samples for LC/MS/MS analysis and it is important that the analytical instrumentation used is fast enough to keep up with the demand. Using traditional LC/MS and LC/MS/MS instrumentation and methods, analysis time is typically limited by the autosampler cycle time and instrument-PC communications. This additional overhead can add 30-45 seconds to each injection beyond the LC gradient time, reducing the impact of rapid LC methods. The advent of fast serial sampling systems such as the Agilent RapidFire and Apricot Designs Dual Autosampler (ADDA) have enabled significantly shorter analysis times. The RapidFire and ADDA coupled with Agilent binary pumps and AB Sciex API4000 mass spectrometers allow sensitive trap & elute methods with cycle times of less than 15 seconds and less than 30 seconds, respectively, per injection while still deliveringdata comparable to traditional instrumentation. After switching from ballistic gradient methods using a traditional autosampler to trap & elute methods using the RapidFire and ADDA instruments, our assay runtimes decreased from approximately 10 hours for a full 384-well plate to just under 2.5 hours.
Poster #16
Kate Comstock
Thermo Fisher Scientific
Effective Workflow for Pharmaceutical Impurity Analysis using HR- LCMS and Compound Discoverer 
Pharmaceutical impurity analysis is crucial for drug R&D, production, and post-marketing surveillance. LCMS is routinely used for impurity analysis because of its speed and sensitivity. For rapid, accurate, and confident impurity ID, very high resolution mass spectrometer and effective data processing software are essential.This study demonstrates a workflow for Fexofenadine API impurity profiling. The HRAM data acquired from a Thermo Scientific™ Orbitrap Elite™ was processed using Thermo Scientific™ Compound Discoverer™ (CD) software. From component extraction to structure characterization, the advanced algorithms and architecture design featured in Compound Discoverer improved the overall speed and confidence of impurity ID.The HRAM data acquired on the Orbitrap Elite mass spectrometer provided a wealth of information for Fexofenadine API impurity identification. The accurate mass full scan data with high fidelity isotopic pattern allowed confident impurity identification and elemental composition assignment. The information-rich HCD MS/MS fragments proved to be very valuable for structure elucidation.
 
Data processing was achieved by using the advanced processing workflow in Compound Discoverer. This workflow effectively searched for related expected and related unexpected impurities in one single process. Adducts of the components were grouped together, which simplifies data review. Multiple samples were grouped within the study design and processed together for comparison. Elemental compositions of putative impurities were automatically confirmed by isotopic pattern matching and spectral distance scores. FIShTM Scoring node provided automatic fragment ion matching score and structure annotation directly on the MS/MS spectra.
 
The results display the impurities that are structurally related to parent compound, the structure transformation, and structure annotation of MS/MS spectra. Also listed are the impurities which are structurally unrelated to the parent compound.
 
The reporting tool in Compound Discoverer allowed easy modification based on customizable templates with the desired information, including spectra, and structure annotation, etc
 
Poster #17
Patrick Wheeler
ACD Labs
Better, Faster Routes to Accurate Structures for Natural ProductsAccurate structures for natural products are attainable from smaller samples than ever before, thanks to the advances in NMR probes and pulse sequences. Still, the interpretation of this data can be arduous, requires great expertise, and is error-prone. Of course, other techniques are used to confirm structures as well: synthetic reproduction of natural products has a long tradition of success in the elucidation of molecules containing intricate elements, including multiple stereo centers. However, despite rigorous analysis by qualified chemists, these methods still sometimes arrive at erroneous results1-5. Astute application of modern technology can speed the rate at which structures are solved, while also crucially reducing errors that result either from synthetic methods or from unassisted analysis of instrumental data. Computer Assisted Structure Elucidation (CASE) has developed over the past decades to relieve the burden of work in ascertaining correct structures. In this presentation, we will discuss how CASE is used to objectively analyze complex sets of NMR data in order to test structural hypotheses, conduct de novo structure elucidation, and query large databases of known structures for matches to already identified natural products.
Poster #18
Kate Comstock
Thermo Fisher Scientific
Effective Workflow for Pharmaceutical Impurity Analysis using HR- LCMS and Compound Discoverer
Pharmaceutical impurity analysis is crucial for drug R&D, production, and post-marketing surveillance. LCMS is routinely used for impurity analysis because of its speed and sensitivity. For rapid, accurate, and confident impurity ID, very high resolution mass spectrometer and effective data processing software are essential.This study demonstrates a workflow for Fexofenadine API impurity profiling. The HRAM data acquired from a Thermo Scientific™ Orbitrap Elite™ was processed using Thermo Scientific™ Compound Discoverer™ (CD) software. From component extraction to structure characterization, the advanced algorithms and architecture design featured in Compound Discoverer improved the overall speed and confidence of impurity ID.The HRAM data acquired on the Orbitrap Elite mass spectrometer provided a wealth of information for Fexofenadine API impurity identification. The accurate mass full scan data with high fidelity isotopic pattern allowed confident impurity identification and elemental composition assignment. The information-rich HCD MS/MS fragments proved to be very valuable for structure elucidation.
 
Data processing was achieved by using the advanced processing workflow in Compound Discoverer. This workflow effectively searched for related expected and related unexpected impurities in one single process. Adducts of the components were grouped together, which simplifies data review. Multiple samples were grouped within the study design and processed together for comparison. Elemental compositions of putative impurities were automatically confirmed by isotopic pattern matching and spectral distance scores. FIShTM Scoring node provided automatic fragment ion matching score and structure annotation directly on the MS/MS spectra.
 
The results display the impurities that are structurally related to parent compound, the structure transformation, and structure annotation of MS/MS spectra. Also listed are the impurities which are structurally unrelated to the parent compound.
 
The reporting tool in Compound Discoverer allowed easy modification based on customizable templates with the desired information, including spectra, and structure annotation, etc
 

 

 

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