Generation of mass tags by the inherent electrochemistry of electrospray for protein mass spectrometry

We present herein a review of our work on the on-line electrochemical generation of mass tags toward cysteine residues in peptides and proteins. Taking advantage of the inherent electrochemical nature of electrospray generated from a microfabricated microspray emitter, selective probes for cysteine were developed and tested for on-line nonquantitative mass tagging of peptides and proteins. The nonquantitative aspect of the covalent tagging thus allows direct counting of free cysteines in the mass spectrum of a biomolecule through additional adduct peaks. Several substituted hydroquinones were investigated in terms of electrochemical properties, and their usefulness for on-line mass tagging during microspray experiments were assessed with L-cysteine, peptides, and intact proteins. Complementarily, numerical simulations were performed to properly understand the respective roles of mass transport, kinetics of electrochemical-chemical reactions, and design of the microspray emitter in the mass tagging overall efficiency. Finally, the on-line electrochemical tagging of cysteine residues was applied to the analysis of tryptic peptides of purified model proteins for protein identification through peptide mass fingerprinting.     

HS-SPME-GC-MS analysis of antioxidant degradation products migrating to drinking water from PE materials and PEX pipes

Polyethylene (PE) and cross-linked polyethylene (PEX) pipes are frequently used in water supply systems. Such pipes contain added antioxidants with phenolic structures, e.g. Irgafos 168, Irganox 1010 and 1076, in order to improve durability. However, phenol, ketone and quinone antioxidant degradation products may leach and enter drinking water. The aim of this investigation was to develop a method for measuring these degradation products with a performance meeting the drinking water quality criteria of 20?µg?L^?1. Using headspace solid phase microextraction coupled to a gas chromatograph with a mass spectrometer, a method was established revealing limits of detection and quantification less than 0.4 and 1?µg?L^?1 respectively. The method was applied to migration experiments for two PEX pipes and one PE material, quantifying the release of two degradation products. Highest concentrations were observed for 2,6-di-tert-butyl-p-benzoquinone which in one of the two pipes was found in concentrations of 18–57?µg?L^?1 in each of eight consecutive release experiments.     

Total synthesis of natural and non-natural Δ(5,6)Δ(12,13)-jatrophane diterpenes and their evaluation as MDR modulators

We report the details of the total synthesis of natural and non-natural jatropha-5,12-dienes. The successful tactic for the assembly of the strained trans-bicyclo[10.3.0]pentadecane scaffold employed a B-alkyl Suzuki-Miyaura cross-coupling for the formation of the C5/C6 double bond and a ring-closing metathesis for the construction of the C12/C13 double bond. The key step of the synthesis of the cyclopentane fragment, an uncatalyzed intramolecular carbonyl-ene reaction, was studied computationally by DFT calculations. The members of the ensemble of synthetic natural and non-natural jatrophanes were subsequently examined as modulators for the ABCB1, ABCG2, and ABCC1 efflux proteins, which are associated with multidrug resistance in cancer chemotherapy.     

Ionic‐Liquid‐Based Polymer Electrolytes for Battery Applications

The advent of solid‐state polymer electrolytes for application in lithium batteries took place more than four decades ago when the ability of polyethylene oxide (PEO) to dissolve suitable lithium salts was demonstrated. Since then, many modifications of this basic system have been proposed and tested, involving the addition of conventional, carbonate‐based electrolytes, low molecular weight polymers, ceramic fillers, and others. This Review focuses on ternary polymer electrolytes, that is, ion‐conducting systems consisting of a polymer incorporating two salts, one bearing the lithium cation and the other introducing additional anions capable of plasticizing the polymer chains. Assessing the state of the research field of solid‐state, ternary polymer electrolytes, while giving background on the whole field of polymer electrolytes, this Review is expected to stimulate new thoughts and ideas on the challenges and opportunities of lithium‐metal batteries.     

Difference between Extra‐ and Intracellular T1 Values of Carboxylic Acids Affects the Quantitative Analysis of Cellular Kinetics by Hyperpolarized NMR

Incomplete knowledge of the longitudinal relaxation time constant (T₁) leads to incorrect assumptions in quantitative kinetic models of cellular systems, studied by hyperpolarized real‐time NMR. Using an assay that measures the intracellular signal of small carboxylic acids in living cells, the intracellular T₁ of the carboxylic acid moiety of acetate, keto‐isocaproate, pyruvate, and butyrate was determined. The intracellular T₁ is shown to be up to four‐fold shorter than the extracellular T₁. Such a large difference in T₁ values between the inside and the outside of the cell has significant influence on the quantification of intracellular metabolic activity. It is expected that the significantly shorter T₁ value of the carboxylic moieties inside cells is a result of macromolecular crowding. An artificial cytosol has been prepared and applied to predict the T₁ of other carboxylic acids. We demonstrate the value of this prediction tool.     

Direct coupling of a flow–flow electromembrane extraction probe to LC-MS

A fully integrated and automated electromembrane extraction LC-MS (EME-LC-MS) system has been developed and characterized. Hyphenation of a flow–flow EME probe to LC-MS was accomplished by using an in-built 10-port switching valve of the LC-MS system. The 10-port switching valve decoupled the high pressure of the UHPLC-system from the low pressure required for operation of the EME-probe by automated switching between a sample extraction/analysis and a sample load position. In the sample load position the extracted analytes were loaded into a HPLC sample loop. By switching the valve to the sample extraction/analysis position the setup allowed simultaneous analysis of previously loaded analytes while extracting a new sample. Performance of the system was characterized with respect to precision and linearity (RSD < 2.5%, R²: 0.998) and the setup was applied for studying the in-vitro metabolism of methadone by rat liver microsomes. As the metabolic reaction proceeded, methadone and its metabolites were extracted and analyzed in parallel by LC-MS using either isocratic or gradient elution. Compared to a conventional in-vitro metabolism analysis based on protein precipitation followed by LC-MS analysis the fully automated EME-LC-MS system offers a significant time saving and in addition demonstrates increased sensitivity as the analytes were automatically enriched during the extraction process. The experiment revealed 6 to 16 times higher S/N ratios of the EME-LC-MS method compared to protein precipitation followed by LC-MS and thus concomitantly lower LOD and LOQ. The setup integrates a complete analytical workflow of rapid extraction, enrichment, separation and detection of analytes in a fully automated manner. These attributes make the developed system a powerful alternative approach for a wide range of analytical applications.     

Plausible origins of homochirality in the amino acid catalyzed neogenesis of carbohydrates

The intrinsic ability of amino acids to catalyze the asymmetric formation of carbohydrates, which enzymes have mediated for millions of years, with significant amplification of enantiomeric excess suggests a plausible ancient catalytic process for the evolution of homochirality.