Sample Run Time Research Articles

Rapid Intact Mass Analysis and Evaluation of the Separation Potential of Microfluidic Capillary Electrophoresis Mass Spectrometry for Oligonucleotides.

Oligonucleotide characterization is a rapidly advancing field in the biopharmaceutical industry. Understanding critical quality attributes, such as intact mass and impurities, requires a toolbox of analytical techniques, which commonly includes liquid chromatography-mass spectrometry (LC-MS). Oligonucleotide LC-MS analysis frequently requires sample run times upward of 15 min to achieve separation of multiple oligonucleotide species. Additionally, LC methods frequently employ mobile phase additives such as triethylamine and 1,1,1,3,3,3-hexafluoro-2-propanol that are not always desired for use in MS instrumentation. Here, microfluidic capillary electrophoresis mass spectrometry (CE-MS) via ZipChip technology was employed to enable rapid intact mass analysis of oligonucleotide single strands. Baseline separation of equal length oligonucleotides was achieved in less than 4 min. Additionally, the potential of the ZipChip platform for separation of oligonucleotide full-length products (FLPs) and their impurities was evaluated.

Structural Analysis and Identity Confirmation of Anthocyanins in Brassica oleracea Extracts by Direct Injection Ion Mobility-Mass Spectrometry.

Anthocyanins are a subclass of plant-derived flavonoids that demonstrate immense structural heterogeneity which is challenging to capture in complex extracts by traditional liquid chromatography-mass spectrometry (MS)-based approaches. Here, we investigate direct injection ion mobility-MS as a rapid analytical tool to characterize anthocyanin structural features in red cabbage (Brassica oleracea) extracts. Within a 1.5 min sample run time, we observe localization of structurally similar anthocyanins and their isobars into discrete drift time regions based upon their degree of chemical modifications. Furthermore, drift time-aligned fragmentation enables simultaneous collection of MS, MS/MS, and collisional cross-section data for individual anthocyanin species down to a low picomole scale to generate structural identifiers for rapid identity confirmation. We finally identify anthocyanins in three other Brassica oleracea extracts based on red cabbage anthocyanin identifiers to demonstrate our high-throughput approach. Direct injection ion mobility-MS therefore provides wholistic structural information on structurally similar, and even isobaric, anthocyanins in complex plant extracts, which can inform the nutritional value of a plant and bolster drug discovery pipelines.

Simple and high sample throughput LC/ESI-MS/MS method for bioequivalence study of prazosin, a drug with risk of orthostatic hypotension

Objective The study aimed to develop a rapid, simple and sensitive LC/ESI-MS/MS method to measure prazosin concentration in human plasma and apply bedside sampling in bioequivalence study of two prazosin tablets to resolve the adverse effect of orthostatic hypotension. Significance The LC/ESI-MS/MS prazosin method was highly sensitive and selective. Bedside sampling reduced the orthostatic hypotension incidence and subject dropout rate. Methods After sample preparation, prazosin and terazosin (IS) were detected on mass spectrometer operating in multiple reaction monitoring mode using positive ionization. Mobile phase flow rate was set at 0.40 mL/min with sample run time of 1.75 min. The bioanalytical method was validated as per EMEA and FDA guidelines. Bedside sampling was performed in bioequivalence study for the first 4 h after dosing. The three primary pharmacokinetic parameters, C max, AUC0- t and AUC0-∞ and 90% confidence interval were determined. Results The small injection volume of 1 μL minimized instrumentation contamination and prolonged the analytical column lifespan. Linearity was obtained between 0.5 and 30.0 ng/mL, with coefficient of determination, r 2 ≥ 0.99. The mean extraction recovery of prazosin and IS was >92%, with precision value (CV, %) ≤ 10.3%. Only two orthostatic hypotension adverse events were reported. The two prazosin formulations were found to be bioequivalent. Conclusion The LC/ESI-MS/MS method has shown robustness and reliability exemplified by the incurred sample re-analysis result. Bedside sampling should be proposed for bioequivalence or pharmacokinetic studies of drugs demonstrating adverse event of orthostatic hypotension.

Sample preparation and quantification of polar drug, allopurinol, in human plasma using LCMSMS.

A fast, selective and reproducible LC-MS/MS method with simple sample preparation was developed and validated for a polar compound, allopurinol in human plasma, using acyclovir as internal standard (IS). Chromatographic separation was achieved using Agilent Poroshell 120 EC-C18 (100 × 2.1 mmID, 2.7 µm) analytical column. The mobile phase was comprised of 0.1%v/v formic acid-methanol (95:05; v/v), at a flow rate of 0.45 mL/min. The effect of different protein precipitation agents used in sample preparation such as methanol, acetonitrile, a mixture of acetonitrile-methanol and a mixture of acetonitrile-acetone were evaluated to optimize the extraction efficiency of allopurinol and IS. The use of acetone-acetonitrile (50:50, v/v) as protein precipitating agent shortened the sample preparation time and improved the recovery of allopurinol to above 93%. The IS-normalised matrix factors at two concentration levels were 1.0, with CV of 5.1% and 4.2%. Allopurinol in plasma was stable at benchtop for 24 h, in autosampler tray for 48 h, in instrumentation room for 48 h, in freezer after 7 freeze-thaw cycles and in freezer for 140 days. Allopurinol stock standard solutions were stable for 140 days at room temperature and in the chiller. The short sample run time of the validated bioanalytical method allowed high throughput analysis of plasma samples in pharmacokinetic study of an allopurinol formulation. The robustness and reproducibility of the bioanalytical method was reaffirmed through incurred sample reanalysis (ISR).

2-μm fused-core column ultra-high-performance liquid chromatography/tandem mass spectrometric determination of donepezil in human plasma: Application to a bioequivalence study

Ultra-high-performance liquid chromatography (UHPLC) hyphenated to tandem mass spectrometric methods using the dimethylpentafluorophenylpropyl stationary phase with polar and nonpolar characteristics of a 2-μm fused-core silica particle packing were evaluated to perform efficient separation for the analysis of donepezil in human plasma. The fused-core silica particle design providing the shorter diffusional mass transfer path for the analytes is less affected in resolving power by increasing mobile-phase velocity for faster chromatographic resolution between the administered compound and multiple interfering peaks from the extracted quality control, calibration standard and study samples following simple protein precipitation extraction procedures. In this work, two UHPLC-MS/MS approaches requiring 1.2 min per sample run time and neither expensive ultra-high-pressure instrumentation nor new laboratory protocols were applied and compared for determination of donepezil in human plasma at sub-nanograms per milliliter region to support a randomized, crossover bioequivalence (BE) study in which healthy volunteers each received a single oral dose of the test and reference formulations of 10 mg donepezil hydrochloride. The column performance in terms of chromatographic separation efficiency, peak asymmetry and resolution and retention time reproducibility was found to be sustainable over a thousand extracted plasma injections. The linear dynamic range was detected over a concentration range of 0.2–50 ng/mL. The intra- and inter-day assay accuracy and precision were within 15% for the analyte in individual biological fluids. Positive correlation coefficients (r) greater than 0.98 and 0.99 for donepezil concentrations in study plasma samplers measured by the proposed UHPLC-MS/MS approaches and another validated HPLC-MS/MS method were observed.

Novel high throughput green UPLC-MS/MS methods for determination of desmopressin with special sample handling

Objectives of present investigation were to develop novel high throughput green UPLC-MS/MS methods for quality control of desmopressin formulations. Two liquid chromatographic methods: Method-I based on isocratic elution and method-II based on gradient elution were developed and validated. Methods were green, since less amount of organic solvent (≤75 ​μL, per sample run time) was utilized in mobile phase. Mobile phase (0.1% formic acid in ACN, and in water), flow rate was 300 ​μL/min. Desmopressin retention time and sample run time for Method-I were 0.59 and 1.0 ​min, respectively. Retention time and sample run time for method-II were 2.03 and 2.5 ​min, respectively. Double charged ions [M ​+ ​2H]2+ of desmopressin (m/z 535.33 ​> ​120) were monitored in multi-reaction monitoring mode. Desmopressin was directly analyzed in water samples. Calibration curves were prepared between 50 and 600 ​ng/mL. The values of r2 were 0.999. Assay and dissolution study of desmopressin formulation was successfully performed using these methods. Desmopressin was found to adsorb or react with different type of materials while handling. The specific filters (Chromafil® Xtra PTFE-45/25 0.45 ​μm, syringe filter) and tubes (glass tubes or Neutral graduated micro test tubes) were used for handling desmopressin samples.

Separation of Branched and Isoprenoid Glycerol Dialkyl Glycerol Tetraether (GDGT) Isomers in Peat Soils and Marine Sediments Using Reverse Phase Chromatography

Glycerol dialkyl glycerol tetraethers (GDGTs) have become one of the most investigated lipid classes in marine and terrestrial organic geochemical research. GDGTs are microbial membrane core lipids biosynthesized as multiple homologue series of isoprenoid or methyl-branched isomers whose relative abundance depend on a range of environmental parameters, including temperature. This has led to the development of GDGT-based temperature proxies. Critical for GDGT analysis and the further development of their use as environmental proxies is a good chromatographic separation of the full range of structural and stereo-isomers with potential for discovery of novel GDGT variants. Several HPLC methods have been developed to this extent, but partial co-elution of GDGTs remains an issue despite long run times. In this study, we investigate the effects of different types of reverse phase (RP) chromatography on the separation of GDGT isomers. The RP method presented here gives good separation of isoGDGT isomers in comparison to the recently developed double column HILIC analysis operated in normal phase (NP) and has a shorter run time. In marine samples, the regularly reported isoprenoid GDGTs separated in a similar way as in NP, however an earlier eluting group was observed to elute with the crenarchaeol isomer used in the TEX86 proxy. In a Swedish peat bog sample, a large range of isoGDGT isomers were observed. and for branched GDGTs, near baseline separation of the 5- and 6-methyl branched GDGT isomers was achieved, as well as asymmetric isomers like the 5/6 isomer. The C18-XB method is rapid and versatile and can be set up on either low-pressure HPLC systems (max 400 bar) with a sample run time of 25 minutes for brGDGTs and 45 minutes to include isoGDGTs. On UHPLC-MS systems (> 600 bar) the sample run time is reduced to 15 minutes. Both methods meet the demands of high-resolution sampling in shorter time and for low running costs. The C18-XB RP method presented here gives unusual separation of branched GDGTs and could be a useful tool for the further elucidation of the biological sources and environmental factors that play a role in the production of different GDGT isomers.

Quantum-classical reinforcement learning for decoding noisy classical parity information

Learning a hidden parity function from noisy data, known as learning parity with noise (LPN), is an example of intelligent behavior that aims to generalize a concept based on noisy examples. The solution to LPN immediately leads to decoding a random binary linear code in the presence of classification noise. This problem is thought to be intractable classically, but can be solved efficiently if a quantum oracle can be queried. However, in practice, a learner is likely to obtain classical data. In this work, we show that a naive application of the quantum LPN algorithm to classical data encoded in an equal superposition state requires an exponential sample complexity. We then propose a quantum-classical reinforcement learning algorithm to solve the LPN problem for classical data and demonstrate a significant reduction in the sample complexity compared with the naive approach. Simulations with a hidden bit string of length up to 12 show that the quantum-classical reinforcement learning performs better than known classical algorithms when the sample complexity, run time, and robustness to classical noise are collectively considered. Our algorithm is robust to any noise in the quantum circuit that effectively appears as Pauli errors on the final state.

A Validated Ultra-Performance Liquid Chromatography Tandem Triple Quadrupole Mass Spectrometric Method for Fast Determination of Losartan in Rabbit Plasma.

A rapid UPLC-MS-MS method was developed and validated for determination of losartan in rabbit plasma. Protonated adducts of losartan and eprosartan (IS) were monitored in multiple reaction monitoring mode. Molecular masses of daughter species of losartan were m/z 423.19 > 207 and m/z 423.19 > 180; and of eprosartan were m/z 425.11 > 135 and m/z 425.11 > 107. Losartan from plasma samples was extracted by protein precipitation method. The mobile phase comprising water (0.1% formic acid) (A) and acetonitrile (0.1% formic acid) (B) was used in gradient mode. Analytes were eluted on Acquity UPLC®BEH C18 1.7 μm, 2.1 × 50 mm column. Sample run time was 3.0 min. The validation parameters: accuracy, precision and recovery were within recommended limits. Losartan as well as internal standard remains stable in benchtop stability study as well as in post-preparative stability study. Pharmacokinetic parameters such as Cmax (182.79 ± 23.80 ng/mL), Tmax (1.16 ± 0.28 h), AUC0-t (1188.57 ± 404.60 ng h/mL) and Kel (0.0954 ± 0.0140 h-1) of losartan were measured. Method was successfully applied for pharmacokinetic investigation in rabbits and can be used for losartan determination in plasma sample obtained from other animals.

Quantification of Cefepime, Meropenem, Piperacillin, and Tazobactam in Human Plasma Using a Sensitive and Robust Liquid Chromatography-Tandem Mass Spectrometry Method, Part 1: Assay Development and Validation.

The highly variable pharmacokinetics of β-lactam antibiotics and β-lactamase inhibitors poses a significant challenge to clinicians in ensuring appropriate antibiotic doses in critically ill patients. Therefore, routine monitoring of plasma concentrations is important for individualization of antimicrobial therapy. Accordingly, a simple and robust analytical method for the simultaneous measurement of multiple β-lactam antibiotics and β-lactamase inhibitors is highly desirable to ensure quick decisions on dose adjustments. In this study, a sensitive, simple, and robust method for the simultaneous quantification of cefepime, meropenem, piperacillin, and tazobactam in human plasma was developed and rigorously validated according to FDA guidance. Sample extraction was accomplished by simple protein precipitation. Chromatographic separation of analytes was achieved using stepwise gradient elution. Analytes were monitored using tandem mass spectrometry (MS/MS) with a turbo ion spray source in positive multiple-reaction-monitoring mode. The calibration curve ranged from 0.5 to 150 μg/ml for cefepime, 0.1 to 150 μg/ml for meropenem and piperacillin, and 0.25 to 150 μg/ml for tazobactam. Inter- and intraday precision and accuracy, sensitivity, selectivity, dilution integrity, matrix effect, extraction recovery, and hemolysis effect were investigated for all four analytes, and the results met the acceptance criteria. Compared to other reported methods, our method is more robust because of the combination of the following features: (i) a simple sample extraction procedure, (ii) a short sample run time, (iii) a wide dynamic range, and (iv) the small plasma sample volume needed. Since our method already covers β-lactams and a β-lactamase inhibitor with highly heterogeneous physicochemical properties, further antibiotic candidates may easily be incorporated into this multianalyte method.

A Provable Approach for Double-Sparse Coding

Sparse coding is a crucial subroutine in algorithms for various signal processing, deep learning, and other machine learning applications. The central goal is to learn an overcomplete dictionary that can sparsely represent a given dataset. However, storage, transmission, and processing of the learned dictionary can be untenably high if the data dimension is high. In this paper, we consider the double-sparsity model introduced by Rubinstein, Zibulevsky, and Elad (2010) where the dictionary itself is the product of a fixed, known basis and a data-adaptive sparse component. First, we introduce a simple algorithm for double-sparse coding that can be amenable to efficient implementation via neural architectures. Second, we theoretically analyze its performance and demonstrate asymptotic sample complexity and running time benefits over existing (provable) approaches for sparse coding. To our knowledge, our work introduces the first computationally efficient algorithm for double-sparse coding that enjoys rigorous statistical guarantees. Finally, we support our analysis via several numerical experiments on simulated data, confirming that our method can indeed be useful in problem sizes encountered in practical applications.

Synthesis of high-performance thin film composite (TFC) membranes by controlling the preparation conditions: Technical notes

Reverse osmosis (RO) is one of the most globally competitive technologies for desalination and water purification, in which thin film composite (TFC) membrane is the common form of membranes used. In this paper, we examined changes in membrane performance by studying the effects of 1) m-phenylenediamine (MPD) and trimesoyl chloride (TMC) exposure times; 2) different phase inversion methods to prepare polysulfone (PSU) support sheets; 3) the support sheet thickness, and 4) the fabricated membrane’s drying temperature. Operational conditions of 300 psi, 25 °C, 2000 ppm NaCl, and sample run time of at least 8 h were applied. Optimal performance results were achieved when the interfacial polymerization time was short, and a vertical immersion during PSU layer preparation was applied. Increasing PSU sheet thickness increased the salt rejection but decreased the water flux, while rising the curing temperature led to an enhancement in salt rejection and water flux at short reaction times. Based on this study, a TFC membrane of NaCl rejection 98.8 ± 0.5% and water flux 76.1 ± 2.7 L/m2 h was fabricated using 25 s MPD contacting time, 5 s TMC reaction time, and 110 °C drying temperature. This result is considered high among reported outcomes in the literature for simply prepared TFC membranes.

LC-Ion Trap-MS Method for the Determination of Fluconazole in Plasma for Bioequivalence Studies of Pharmaceutical Formulations Using Semi-Automated Sample Handling

An LC–ion trap-MS method for the determination of fluconazole in human plasma has been developed to be applied for bioequivalence studies. The human plasma samples were extracted with methyl-tert-butyl ether. The extract was evaporated to dryness, reconstituted with mobile phase and injected into the LC–MS apparatus. Fluconazole d4 was used as the internal standard (IS). A C18 analytical column was used with a mobile phase consisting of 40% formic acid 0.1% and 60% acetonitrile. A robotic liquid handler was used to increase the method’s throughput. Fluconazole was detected by monitoring the mass transition of m/z 307.1 to m/z 238.0 (m/z 311.1 to m/z 242.0 for the IS). The method was validated according to the European Medicines Agency (EMEA) regulations and was found to be accurate, precise, and selective throughout the calibration range (0.05–10.0 µg/mL). Compared to the published methods for the determination of fluconazole in its usual therapeutic levels, the method provides the shortest sample runtime. For the first time, it was demonstrated that ion trap mass spectrometry can be used for the determination of fluconazole in plasma.

Development of An Online LC-LVI-GC × GC System: Design and Preliminary Applications

The identification of unknown compounds is of fundamental importance for a range of applications in chromatography including, but not limited to, environmental pollution, food/natural product analysis, metabolomics, sports testing, petrochemicals and biofuel analysis. Critical to the success of each application is the ability to separate the compounds of interest, both from each other and the sample matrix, which may be present at concentrations orders of magnitude higher than the analyte(s). Selectivity and sensitivity are key to such analyses and both may be increased by the use of multiple dimensions of separation. Here, we report on the construction of an online, liquid chromatography hyphenated large volume injection, cryogenically modulated, multi-dimensional gas chromatography (LC-LVI-GC × GC) system for the characterisation of complex matrices using existing instruments that are common in most analytical laboratories. We detail the design of the instrument and demonstrate its performance and potential on a range of sample types. The combination of the LC, and large volume injection cryogenic GC × GC, was found to lead to a high selectivity and peak capacity with little sample preparation needed, but with a trade of a large sample run time of ~40 (but up to ~60) minutes in each case. The system therefore has great potential for the targeted and untargeted analysis of very complex sample types.

A Fast and Accurate UPLC Method for Analysis of Proteinogenic Amino Acids

A reliable and inexpensive method for the measurement of proteinogenic amino acids was developed. It allows the screening of large sample volumes that is important for current food and feed applications, but also for future biobased applications. The method is based on acid hydrolysis and automated pre-column derivatision in the injection needle of the autosampler using o-phthalaldehyde/ethanethiol reagent in combination with 9-fluorenylmethyl chloroformate. The calculated mean limit of detection and limit of quantification of pure amino acids were 2.3 and 4.6 μM, respectively. With this method, it was possible to accurately analyse the amino acid composition of bovine serum albumin, soy meal, Jatropha press cake, and cyanophycin. Mean reproducibility of all amino acids from bovine serum albumin was 6.8 %, with a mean recovery of 95 %. The sample run time is 16 min with a total cycle time from injection to injection of an acid hydrolyzed sample of 22 min, resulting in a greener method due to a reduction in solvent consumption.

Application of single immunoaffinity clean-up for simultaneous determination of regulated mycotoxins in cereals and nuts

A rapid and sensitive analytical strategy for the simultaneous determination of twelve mycotoxins (aflatoxins, fumonisins, zearalenon, deoxynivalenol, ochratoxin A, T-2 and HT-2 toxins) using ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) was developed and validated. The method was validated for peanuts, barley and maize-breakfast cereals; selected as they represent the matrices most often contaminated by mycotoxins. The method is designed for fast and reliable analyses of mycotoxins in regulatory, industrial and private laboratories. Multi-target immunoaffinity columns containing antibodies for all mycotoxins studied herein were used for sample clean-up. Method optimization was predominantly focused on the simplification of extraction and clean-up procedure recommended by column producers. This newly developed and simplified procedure decreased both the sample preparation time and the solvent volumes used for their processing. The analysis of all regulated mycotoxins was conducted by a newly developed UHPLC-MS/MS method with a sample run time of only ten minutes. The method trueness was tested with analytical spikes and certified reference materials, with recoveries ranging from 71% to 112% for all of the examined mycotoxins.

Online solid-phase extraction–liquid chromatography–electrospray–tandem mass spectrometry determination of multiple classes of antibiotics in environmental and treated waters

An online solid-phase extraction and liquid chromatography in combination with tandem mass spectrometry method was developed for the simultaneous determination of 31 antibiotics in drinking water, surface water and reclaimed waters. The developed methodology requires small sample volume (10 mL), very little sample preparation and total sample run time was 20 min. An Ion Max API heated electrospray ionization source operated in the positive mode with two selected reaction monitoring transitions was used per antibiotic for positive identity and quantification performed by the internal standard approach, to correct for matrix effects and any losses in the online extraction step. Method detection limits were in the range of 1.2-9.7, 2.2-15, 5.5-63 ng/L in drinking water, surface water and reclaimed waters, respectively. The method accuracy in matrix spiked samples ranged from 50-150% for the studied antibiotics. The applicability of the method was demonstrated using various environmental and reclaimed water matrices. Erythromycin was detected in more than 85% of the samples in all matrices (28-414, n.d.-199, n.d.-66 ng/L in reclaimed, river and drinking waters respectively). The other frequently detected antibiotics in reclaimed waters were nalidixic acid, clarithromycin, azithromycin, trimethoprim, and sulfamethoxazole.

An Extraction Assay Analysis for Galanthamine in Guinea Pig Plasma and its Application to Nerve Agent Countermeasures

Galanthamine hydrobromide (GAL HBr), approved material for treatment of mild to moderate Alzheimer’s disease, is a centrally-acting reversible acetylcholinesterase inhibitor (AChEI) that is currently under evaluation as a therapeutic countermeasure against organophosphorus G- and V-Series nerve agents, which can induce rapid lethality in guinea pigs and humans. It has been shown that upon combination with atropine (ATR) and pyridine-2-aldoxime methochloride (2-PAM), a single dose of GAL administered before or soon after the acute exposure to a lethal dose of organophosphorus compounds can safely counteract toxicity in guinea pigs. To that end a new sample preparation extraction method analysis assay has been explored to enable future high-throughput, reproducible, and sensitive assays for the extraction of galanthamine in guinea pig plasma. Samples were prepared with diphenhydramine hydrochloride (DPH HCl) internal standard and recovered with 10 min liquid-liquid trichloromethane extractions. Samples were analyzed with a reversed phase liquid chromatographic column interfaced to a triple quadrupole mass spectrometer (LC/MS/MS) operating in the positive ion multiple reaction monitoring (MRM) Turbo Ionspray mode. Precursor to product ion (M+H)+ transitions of 288-to-213 m/z and 256-to-167 m/z for GAL and DPH were observed, respectively. Sample run times of 1.50 min were achieved.

Development of a Novel LC/MS/MS Extraction Assay for Galanthamine in Guinea Pig Plasma and its Application to Nerve Agent Countermeasures

Galanthamine Hydrobromide (GAL HBr), approved material for treatment of mild to moderate Alzheimer’s disease, is a centrally-acting reversible Acetylcholinesterase Inhibitor (AChEI) that is currently under evaluation as a therapeutic countermeasure against organophosphorus G- and V-Series nerve agents, which can induce rapid lethality in guinea pigs and humans. It has been shown that upon combination with Atropine (ATR) and pyridine-2Aldoxime Methochloride (2-PAM), a single dose of GAL administered before or soon after the acute exposure to a lethal dose of organophosphorus compounds can safely counteract toxicity in guinea pigs. To that end a new sample preparation extraction method analysis assay has been developed to enable future high-throughput, reproducible, and sensitive assays to quantitate galanthamine in guinea pig plasma. Samples were prepared with Diphenhydramine Hydrochloride (DPH HCl) internal standard and recovered with a 10 min liquid-liquid trichloromethane extraction. Samples were analyzed with a reversed phase liquid chromatographic column interfaced to a triple quadrupole mass spectrometer (LC/MS/MS) operating in the positive ion Multiple Reaction Monitoring (MRM) Turbo Ionspray mode. Precursor to product ion (M+H)+ transitions of 288-to-213 m/z and 256-to-167 m/z for GAL and DPH were observed, respectively. Sample run times of 1.50 min were achieved. Overall extraction method development proved to be acceptable and rugged.

Field guide to next‐generation DNA sequencers

The diversity of available 2(nd) and 3(rd) generation DNA sequencing platforms is increasing rapidly. Costs for these systems range from < $100,000 to more than $1,000,000, with instrument run times ranging from minutes to weeks. Extensive trade-offs exist among these platforms. I summarize the major characteristics of each commercially available platform to enable direct comparisons. In terms of cost per megabase (Mb) of sequence, the Illumina and SOLiD platforms are clearly superior (≤ $0.10/Mb vs. > $10/Mb for 454 and some Ion Torrent chips). In terms of cost per nonmultiplexed sample and instrument run time, the Pacific Biosciences and Ion Torrent platforms excel, with the 454 GS Junior and Illumina MiSeq also notable in this regard. All platforms allow multiplexing of samples, but details of library preparation, experimental design and data analysis can constrain the options. The wide range of characteristics among available platforms provides opportunities both to conduct groundbreaking studies and to waste money on scales that were previously infeasible. Thus, careful thought about the desired characteristics of these systems is warranted before purchasing or using any of them. Updated information from this guide will be maintained at: http://dna.uga.edu/ and http://tomato.biol.trinity.edu/blog/.