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Papers by Joseph Dalluge

Clinical Chemistry, 2001

Background: Cardiac troponin I (cTnI) results vary 100-fold among assays. As a step toward standa... more Background: Cardiac troponin I (cTnI) results vary 100-fold among assays. As a step toward standardization, we examined the performance of 10 candidate reference materials (cRMs) in dilution studies with 13 cTnI measurement systems.Methods: Solutions of 10 cTnI cRMs, each characterized by NIST, were shipped to the manufacturers of 13 cTnI measurement systems. Manufacturers used their respective diluents to prepare each cRM in cTnI concentrations of 1, 10, 25, and 50 μg/L. For the purpose of ranking the cRMs, the deviation of each cTnI measurement from the expected response was assessed after normalization with the 10 μg/L cTnI solution. Normalized deviations were examined in five formats. Parameters from linear regression analysis of the measured cTnI vs expected values were also used to rank performance of the cRMs.Results: The three cRMs demonstrating the best overall rankings were complexes of troponins C, I, and T. The matrices for these three cRMs values differed; one was recon...

BioTechniques, 2000

A rapid method for the identification and characterization of proteins in bacterial cell-free ext... more A rapid method for the identification and characterization of proteins in bacterial cell-free extracts has been developed using directly combined liquid chromatography-electrospray mass spectrometry. The usefulness of this technique is demonstrated for monitoring the expression and chemical modification of phosphoenolpyruvate-sugar phosphotransferase system (PTS) proteins from E. coli with molecular masses ranging from 9-65 kDa. The technique is characterized by minimal sample preparation, remarkable mass accuracy and resolution, reproducibility and the ability, unlike gel electrophoresis, to directly identify posttranslational modifications. The advantages of this technique over analogous matrix-assisted laser desorption ionization mass spectrometry approaches and its potential as a standard tool in the biomolecular research laboratory are discussed.

Anal. Methods, 2017

Checkpoints in processing of metabolomics data are essential to reliably define preliminary ident... more Checkpoints in processing of metabolomics data are essential to reliably define preliminary identifications of metabolomic studies.

Mucosal immunology, Nov 24, 2016

Morphine and its pharmacological derivatives are the most prescribed analgesics for moderate to s... more Morphine and its pharmacological derivatives are the most prescribed analgesics for moderate to severe pain management. However, chronic use of morphine reduces pathogen clearance and induces bacterial translocation across the gut barrier. The enteric microbiome has been shown to have a critical role in the preservation of the mucosal barrier function and metabolic homeostasis. Here, we show for the first time, using bacterial 16s rDNA sequencing, that chronic morphine treatment significantly alters the gut microbial composition and induces preferential expansion of Gram-positive pathogenic and reduction in bile-deconjugating bacterial strains. A significant reduction in both primary and secondary bile acid levels was seen in the gut, but not in the liver with morphine treatment. Morphine-induced microbial dysbiosis and gut barrier disruption was rescued by transplanting placebo-treated microbiota into morphine-treated animals, indicating that microbiome modulation could be exploite...

Fresenius' Journal of Analytical Chemistry, 2000

A number of different procedures have been developed in recent years that utilize mass spectromet... more A number of different procedures have been developed in recent years that utilize mass spectrometry for the direct determination of proteins in complex mixtures of biological origin. Specific examples of these include the use of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) or directly combined liquid chromatography-electrospray ionization mass spectrometry (LC/ESI-MS) for rapid profiling of protein expression in bacterial and eucaryotic cells and cell-free extracts. Approaches to sample cleanup, contaminant removal, and initial separation of analytes on-line for the direct determination of proteins in cells using MALDI-and ESI-MS are discussed. Advantages of these techniques over traditional biochemical methods are highlighted, and a critical review of their utility and potential as standard tools in the biomolecular and microbiological research laboratory is presented.

Journal of Chromatography B: Biomedical Sciences and Applications, 2001

Liquid chromatography (LC) in direct combination with mass spectrometry (MS) has been shown to be... more Liquid chromatography (LC) in direct combination with mass spectrometry (MS) has been shown to be a good analytical technique for the selective separation and detection of labile folate monoglutamates. Reversed-phase LC and electrosprayionization MS conditions were developed and optimized for the separation and detection of 5-methyltetrahydrofolic acid, 5-formyl tetrahydrofolic acid, tetrahydrofolic acid, dihydrofolic acid and folic acid in aqueous samples. Representative and reproducible positive ion mass spectra were generated for each folate under mild MS conditions. The selective MS detection and identification of endogenous 5-methyltetrahydrofolic acid in human plasma was accomplished through the development of a straightforward C-based solid-phase extraction procedure. This procedure allows for the qualitative assessment of 18 5-methyltetrahydrofolic acid in plasma. Based upon an isotope-dilution internal standard calibration study with standards, the LC-MS limit of quantitation for 5M-THF was estimated to be 0.39 ng / ml.

Biointerphases, Jan 6, 2015

The mechanism of interaction of cold nonequilibrium plasma jets with mammalian cells in physiolog... more The mechanism of interaction of cold nonequilibrium plasma jets with mammalian cells in physiologic liquid is reported. The major biological active species produced by an argon RF plasma jet responsible for cell viability reduction are analyzed by experimental results obtained through physical, biological, and chemical diagnostics. This is complemented with chemical kinetics modeling of the plasma source to assess the dominant reactive gas phase species. Different plasma chemistries are obtained by changing the feed gas composition of the cold argon based RF plasma jet from argon, humidified argon (0.27%), to argon/oxygen (1%) and argon/air (1%) at constant power. A minimal consensus physiologic liquid was used, providing isotonic and isohydric conditions and nutrients but is devoid of scavengers or serum constituents. While argon and humidified argon plasma led to the creation of hydrogen peroxide dominated action on the mammalian cells, argon-oxygen and argon-air plasma created a ...

Analytical Chemistry, 2014

The cellular phospholipid membrane plays an important role in cell function and cell−cell communi... more The cellular phospholipid membrane plays an important role in cell function and cell−cell communication, but its biocomplexity and dynamic nature presents a challenge for examining cellular uptake of phospholipids and the resultant effects on cell function. Platelets, small anuclear circulating cell bodies that influence a wide variety of physiological functions through their dynamic secretory and adhesion behavior, present an ideal platform for exploring the effects of exogenous phospholipids on membrane phospholipid content and cell function. In this work, a broad range of platelet functions are quantitatively assessed by leveraging a variety of analytical chemistry techniques, including ultraperformance liquid chromatography−tandem electrospray ionization mass spectrometry (UPLC−MS/MS), vasculature-mimicking microfluidic analysis, and single cell carbon-fiber microelectrode amperometry (CFMA). The relative enrichments of phosphatidylserine (PS) and phosphatidylethanolamine (PE) were characterized with UPLC−MS/MS, and the effects of the enrichment of these two phospholipids on both platelet secretory behavior and adhesion were examined. Results show that, in fact, both PS and PE influence platelet adhesion and secretion. PS was enriched dramatically and decreased platelet adhesion as well as secretion from δ-, α-, and lysosomal granules. PE enrichment was moderate and increased secretion from platelet lysosomes. These insights illuminate the critical connection between membrane phospholipid character and platelet behavior, and both the methods and results presented herein are likely translatable to other mammalian cell systems.

Journal of Microcolumn Separations, 1998

A micellar electrokinetic chromatographic MEKC separation system was developed for the efficient ... more A micellar electrokinetic chromatographic MEKC separation system was developed for the efficient separation of biologically and clinically important Ž. green tea catechins. Six structurally isomeric tea catechins, e.g., y epigallo-Ž. Ž. Ž. Ž. catechin-3-gallate EGCG , y epicatechin gallate ECG , y gallocatechin gal-Ž. Ž. Ž. Ž. Ž. Ž. late GCG , y epigallocatechin EGC , y epicatechin EC , and q catechin Ž. C , were effectively separated using a tetraboratersodium dodecyl sulfate Ž. SDS r␤-cyclodextrin micellar buffer system. The effects of several critical separa-Ž tion parameters micelle charge type, surfactant type, organic solvent buffer. modifier, micelle concentration, buffer pH, and cyclodextrin buffer modifier were evaluated for enhanced isomer separation and resolution. The pH was shown to be a strong factor influencing the separation of the isomers, but baseline resolution of all six structural isomers was only effected when a micellar buffer system contained ␤-cyclodextrin buffer modifier. The proficiency of the developed method was tested Ž. by determining the analytical levels mgrg of crude catechins in aqueous extrac-Ž. tions of three varieties of green tea Camellia sinesis .

PloS one, 2014

The arginine decarboxylase pathway, which converts arginine to agmatine, is present in both human... more The arginine decarboxylase pathway, which converts arginine to agmatine, is present in both humans and most bacterial pathogens. In humans agmatine is a neurotransmitter with affinities towards α2-adrenoreceptors, serotonin receptors, and may inhibit nitric oxide synthase. In bacteria agmatine serves as a precursor to polyamine synthesis and was recently shown to enhance biofilm development in some strains of the respiratory pathogen Pseudomonas aeruginosa. We determined agmatine is at the center of a competing metabolism in the human lung during airways infections and is influenced by the metabolic phenotypes of the infecting pathogens. Ultra performance liquid chromatography with mass spectrometry detection was used to measure agmatine in human sputum samples from patients with cystic fibrosis, spent supernatant from clinical sputum isolates, and from bronchoalvelolar lavage fluid from mice infected with P. aeruginosa agmatine mutants. Agmatine in human sputum peaks during illness...

The Analyst, 2013

Secreted bioactive lipids play critical roles in cell-to-cell communication and have been implica... more Secreted bioactive lipids play critical roles in cell-to-cell communication and have been implicated in inflammatory immune responses such as anaphylaxis, vasodilation, and bronchoconstriction. Analysis of secreted bioactive lipids can be challenging due to their relatively short lifetimes and structural diversity. Herein, a method has been developed using ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to quantify five cell-secreted, structurally and functionally diverse bioactive lipids (PGD2, LTC4, LTD4, LTE4, PAF) that play roles in inflammation. Sample analysis time is 5 min, and isotopically labeled internal standards are used for quantification. This method was applied to an immortal secretory cell line (RBL-2H3), a heterogeneous primary cell culture containing peritoneal mast cells, and murine platelets. In RBL cell supernatant samples, intrasample precisions ranged from 7.32-21.6%, averaging 17.0%, and spike recoveries in cell supernatant matrices ranged from 88.0-107%, averaging 97.0%. Calibration curves were linear from 10 ng mL(-1) to 250 ng mL(-1), and limits of detection ranged from 0.0348 ng mL(-1) to 0.803 ng mL(-1). This method was applied to the determination of lipid secretion from mast cells and platelets, demonstrating broad applicability for lipid measurement in primary culture biological systems.

Nucleic Acids Research, 1996

A method has been developed for the microscale determination of 5,6-dihydrouridine, the most comm... more A method has been developed for the microscale determination of 5,6-dihydrouridine, the most common post-transcriptional modification in bacterial and eukaryotic tRNA. The method is based on stable isotope dilution liquid chromatography-mass spectrometry (LC/ MS) using [1,3-15 N 2 ]dihydrouridine and [1,3-15 N 2 ]uridine as internal standards. RNA samples were enzymatically digested to nucleosides before addition of the internal standards and subsequently analyzed by LC/MS with selected ion monitoring of protonated molecular ions of the labeled and unlabeled nucleosides. Sample quantities of ∼1 pmol tRNA and 5 pmol 23S rRNA were analyzed for mole% dihydrouridine. Dihydrouridine content of Escherichia coli tRNA Ser VGA and tRNA Thr GGU as controls were measured as 2.03 and 2.84 residues/ tRNA molecule, representing accuracies of 98 and 95%. Overall precision values for the analyses of E.coli tRNA Ser VGA and E.coli tRNA Thr GGU , unfractionated tRNA from E.coli and 23S rRNA from E.coli were within the range 0.43-2.4%. The mole% dihydrouridine in unfractionated tRNA and 23S rRNA from E.coli were determined as 1.79 and 0.0396%, corresponding to 1.4 and 1.1 residues/RNA molecule respectively.

Nucleic Acids Research, 1995

We have synthesized four different 5'-diphosphorylated oligoribonucleotides, varying in length fr... more We have synthesized four different 5'-diphosphorylated oligoribonucleotides, varying in length from 11 to 13 nucleotides by a new solid phase method. After deprotection and partial purification the 5'-diphosphorylated oligoribonucleotides could be converted to capped (m7Gppp)-oligoribonucleotides using guanylyl transferase. Radiolabelled capped oligoribonucleotides acted as primers for the influenza A virus RNA polymerase in vitro. The solid phase method described here should also allow the addition of 5'-diphosphates to synthetic oligodeoxyribonucleotides and be capable of automation.

Journal of Mass Spectrometry, 2000

Reversed-phase liquid chromatography with atmospheric pressure chemical ionization mass spectrome... more Reversed-phase liquid chromatography with atmospheric pressure chemical ionization mass spectrometry (LC/APCI-MS) in the positive-ion mode was utilized to analyze crude ether extracts from the root bark of Maclura pomifera, a tree known to have a high content of prenylated xanthones and flavanones. Identification of three xanthones and two flavanones was based on their unique mass spectra. Under optimum conditions peaks corresponding to the [MH](+) ion and characteristic fragments for each compound were observed. (1)H NMR data were used to confirm the identities of two xanthones that had the same molecular mass and similar fragmentation patterns. Fragmentation of the analytes was achieved by application of an electrostatic potential at the entrance of the single quadrupole mass spectrometer. The optimum voltage for fragmentation was found to be related to the class of compounds analyzed and, within each class, to be dependent on the structure of the prenyl moiety. Collision-induced pathways consistent with precedent literature describing the MS characterization of similar compounds and with the observed fragmentation patterns are tentatively proposed.

Journal of Chromatography A, 1998

A study of a variety of stationary phases and elution conditions for the liquid chromatographic (... more A study of a variety of stationary phases and elution conditions for the liquid chromatographic (LC) determination of six biologically active green tea catechins has resulted in the development of two well-defined, reproducible systems for such analyses which overcome limitations of previously described methods. Comparison of six reversed-phase columns indicates that deactivated stationary phases, which utilize ultrapure silica and maximize coverage of the silica support, provide significantly improved separation and chromatographic efficiencies for catechin analyses using LC, compared to conventional monomeric or polymeric C18 columns. Evaluation of elution conditions used for the separations reveals that the presence of acid in the mobile phase (0.05% trifluoroacetic acid) is essential for both the complete resolution of the catechins present in tea and the efficient chromatography of these compounds. The efficacy of one of the developed systems was demonstrated by the quantitative measurement of the six biologically active catechins in aqueous infusions of green tea (Camellia sinensis). Overall precision values for the analyses were within the range 0.3-1% (relative standard deviation).

Journal of Chromatography A, 2000

An overview of analytical methods for the measurement of biologically important tea catechins is ... more An overview of analytical methods for the measurement of biologically important tea catechins is presented. Liquid chromatography and capillary electrophoresis are the most cited techniques for catechin separation, identification and quantitation. Liquid chromatography with ultraviolet detection is frequently used; however, mass spectrometry, electrochemical, fluorescence and chemiluminescence detection are also utilized in cases where more sensitive or selective detection is needed. Two modes of capillary electrophoresis, capillary zone electrophoresis and micellar electrokinetic capillary chromatography, have been employed for the determination of catechins. Both modes of capillary electrophoresis are based on ultraviolet detection. Additional analytical techniques, such as gas chromatography, thin-layer chromatography, paper chromatography, spectrophotometry, biosensing, chemiluminescence and nuclear magnetic resonance spectroscopy have also been utilized for the determination of catechins and are reviewed herein.

Journal of Chromatography A, 2004

Monitoring amino acid metabolism during fermentation has significant potential from the standpoin... more Monitoring amino acid metabolism during fermentation has significant potential from the standpoint of strain selection, optimizing growth and production in host strains, and profiling microbial metabolism and growth state. A method has been developed based on rapid quantification of underivatized amino acids using liquid chromatography-electrospray tandem mass spectrometry (LC-MS-MS) to monitor the metabolism of 20 amino acids during microbial fermentation. The use of a teicoplanin-based chiral stationary phase coupled with electrospray tandem mass spectrometry allows complete amino acid analyses in less than 4 min. Quantification is accomplished using five isotopically labeled amino acids as internal standards. Because comprehensive chromatographic separation and derivatization are not required, analysis time is significantly less than traditional reversed-or normal-phase LC-based amino acid assays. Intra-sample precisions for amino acid measurements in fermentation supernatants using this method average 4.9% (R.S.D.). Inter-day (inter-fermentation) precisions for individual amino acid measurements range from 4.2 to 129% (R.S.D.). Calibration curves are linear over the range 0-300 g/ml, and detection limits are estimated at 50-450 ng/ml. Data visualization techniques for constructing semi-quantitative fermentation profiles of nitrogen source utilization have also been developed and implemented, and demonstrate that amino acid profiles generally correlate with observed growth profiles. Further, cellular growth events, such as lag-time and cell lysis can be detected using this methodology. Correlation coefficients for the time profiles of each amino acid measured illustrate that while several amino acids are differentially metabolized in similar fermentations, a select group of amino acids display strong correlations in these samples, indicating a sub-population of analytes that may be most useful for fermentation profiling.

Journal of Agricultural and Food Chemistry, 2003

A method has been developed for simultaneous identification of soyasaponins and soy isoflavones i... more A method has been developed for simultaneous identification of soyasaponins and soy isoflavones in soy products, based on liquid chromatography/electrospray ionization-mass spectrometry (LC/ ESI-MS). Soy-based nutraceutical products were analyzed by LC/ESI-MS with detection of protonated and sodiated molecular ions, as well as characteristic fragment ions for these compounds. Soy isoflavones were characterized by a strong protonated molecular ion in addition to corresponding [aglycone + H] + ions. Monitoring the soyasaponin-specific protonated aglycone and dehydrated aglycone ions throughout the chromatogram provided a unique fingerprint for soyasaponin content in the samples. This mass spectrometric fingerprint also allowed immediate classification of the soyasaponin analytes as group A or B soyasaponins, based on the unique masses of aglycone ions observed for each class. Quantification of soyasaponin B b in soy-derived materials, based on the use of a purified soyasaponin B b standard and a glycyrrhizin internal standard, has been accomplished.

Current Protein & Peptide Science, 2002

Novel approaches to protein measurement based on mass spectrometry are being developed that chall... more Novel approaches to protein measurement based on mass spectrometry are being developed that challenge more traditional methods. This review summarizes the emergence of mass spectrometry as a tool for clinical protein, peptide, and amino acid determination. Specific applications of mass spectrometry to the measurement of transferrin, transthyretin, glycated hemoglobin, and homocysteine will be discussed, as will the limitations of the technology, and future directions for clinical protein measurement.

Chemical Biology & Drug Design, 2014