Direct coupling of capillary supercritical fluid chromatography with double-focusing high resolution mass spectrometry

An integral restrictor interface with jet separator for coupling capillary column supercritical fluid extraction – supercritical fluid chromatography with high resolution mass spectrometry (SFE-SFC-MS) has been built and used for the analysis of a fatty acid ester, and of polymer additives with a wide range of masses. The mobile phase used was supercritical carbon dioxide; a flame ionization detector (FID) was used in parallel with the mass spectrometer. Different SFC-MS interface operating conditions, e.g. temperature, restrictor position, flow rate, and sample transfer conditions were optimized to obtain good sensitivity and separation for these applications. In addition, the sensitivity of measurements performed with the direct insertion probe and by SFC-MS interface have been compared.     

Optimization of capillary SFC-MS for the determination of additives in polymers

A standard direct introduction capillary interface is used for the SFC-MS analysis of polymer additives. The system is optimized with respect to the position of the restrictor, probe tip temperature, and ion source temperature. El-like charge-exchange spectra are obtained. Cl using ammonia as the reagent gas is used for the quantitative analysis of a real world sample. The experimental capillary SFC-MS spectra obtained show a good similarity with those recorded using the direct insertion probe. The influence of the experimental conditions on the mass spectra obtained is evaluated statistically.     

Capillary supercritical fluid chromatography-mass spectrometry

Capillary supercritical fluid chromatography and its combination with mass spectrometry (SFC-MS) is an important analytical methodology for the analysis of thermally labile and high molecular weight compounds. The mass spectrometer provides sensitive and highly selective detection for the chromatographic effluent. The same physical-chemical properties of supercritical fluids that provide important chromatographic advantages are also important for the transport and gas phase introduction of analyte molecules into the mass spectrometer. The low mobile phase flow rates of small diameter (< 100 μm i.d.) capillary columns allow the total chromatographic effluent to be introduced with conventional, but often slightly modified, ion source configurations for quadrupole mass spectrometers with either electron impact (EI) or chemical ionization (CI) detection. The full range of CI reagents can also be used to obtain additional selectivity and complementary structural data. The instrumentation and operating parameters for capillary SFC-MS methods are described. The applicability and usefulness of various SFC-MS analysis methods are demonstrated using a variety of samples.     

Capillary supercritical fluid chromatography with simultaneous flame ionization and mass spectrometric detection

A simple interface between a capillary supercritical fluid chromatograph and an Extranuclear Simulscan mass spectrometer is described. The SFC column is directly inserted into the ion source through the existing GC-interface. The system is equipped with a splitting device which allows simultaneous EI/MS and flame ionization detection when CO₂ is used as the supercritical phase. The effect of source temperature and pressure on CO₂ clustering was studied for optimization of source conditions. The performance of the system was evaluated with a series of model compounds and standard mixtures.     

A simple approach for coupling liquid chromatography and electron ionization mass spectrometry

A miniaturized, aerosol based interface for directly coupling a liquid chromatograph with a mass spectrometer is presented. The interface is entirely within the electron ionization (EI) source of the mass spectrometer and no additional, external devices are needed. This simple and effective approach exploits micro-flow nebulization technology providing a new interface suitable for a variety of applications of environmental and biological interest. The new interface provides necessary linearity, ruggedness, sensitivity, and reproducibility of response for trace level analysis, and readily interpretable mass spectra for unambiguous identification of unknown compounds of small to medium molecular weight.     

Implementation of the chemical reaction interface mass spectrometry technique on a Hewlett-Packard mass-selective detector

A microwave-powered chemical reaction interface has been installed in a Hewlett-Packard gas chromatograph-mass spectrometer (GC-MS) system (5890 IT gas chromatograph-S971 mass-selective detector). The technical details and optimization strategies are discussed. The evaluation of this new setup is presented, showing detection limits of 1 ng of ¹³C-, ¹⁵N-, and Cl-containing compounds with signal-to-noise ratios greater than or egual to 3. Selective detection was evaluated with a urine sample from a dog dosed with ¹⁵N₃-midazolam that had been previously analyzed by using a differentially pumped research-level quadrupole mass spectrometer. The results show that the detection of ¹⁵N and Cl remains highly selective and the mass-selective detector gives comparable sensitivity to the larger instrument when the latter is operating over a conventional mass range. The capability for chemical reaction interface mass spectrometry can be easily accomplished with an inexpensive GC-MS system.     

Simultaneous extraction and analysis by high performance liquid chromatography coupled to diode array and mass spectrometric detectors of bixin and phenolic compounds from annatto seeds

This study was designed to identify and quantify the carotenoids and phenolic compounds from annatto seeds using high performance liquid chromatography coupled to diode array and mass spectrometer detectors (HPLC-DAD-MS/MS). Furthermore, using response surface methodology, an optimized procedure for simultaneous extraction of these compounds was established. In addition to bixin, known to be the main carotenoid in annatto seeds, hypolaetin and a caffeoyl acid derivative were identified as the main phenolic compounds. The optimized procedure involved 15 extractions using acetone:methanol:water (50:40:10, v/v/v) as solvent, a solid-liquid ratio of 1:9 (m/v) and an extraction time of 5 min. Validation data indicated that the HPLC method proposed provided good linearity, sensitivity, procedure accuracy, system precision and suggested its suitability for the simultaneous analysis of phenolic compounds and carotenoids in annatto seeds.     

Evaluation of the atmospheric pressure photoionization source for the determination of benzidines and chloroanilines in water and industrial effluents by high performance liquid chromatography-tandem mass spectrometry

A solid phase extraction-high performance liquid chromatography-tandem mass spectrometry based analytical method suitable for simultaneous analysis of benzidine, 3,3′-dichlorobenzidine, mono-, di-, and tri-chloroanilines has been developed.Normal phase separation by liquid chromatography was performed using a cyano propyl methyl silica column, and atmospheric pressure photoionization was employed as interface with mass spectrometer.The developed method was evaluated in terms of limit of detection, accuracy, and precision. The quantification limit for all the compounds ranged between 7 and 112 ng L⁻¹, while recovery for all the compounds was higher than 94%. The method was tested by analyzing different industrial wastes, showing residual contamination by most of the analytes.     

Application of high‐resolution ESI and MALDI mass spectrometry to metabolite profiling of small interfering RNA duplex

We investigated the application of a high‐resolution Orbitrap mass spectrometer equipped with an electrospray ionization (ESI) source and a matrix‐assisted laser desorption/ionization‐time‐of‐flight (MALDI‐TOF) mass spectrometer to the metabolite profiling of a model small interfering RNA (siRNA) duplex TSR#34 and compared their functions and capabilities. TSR#34 duplex was incubated in human serum in vitro, and the duplex and its metabolites were then purified by ion exchange chromatography in order to remove the biological matrices. The fraction containing the siRNA duplex and its metabolites was collected and desalted and then subjected to high‐performance liquid chromatography (HPLC) equipped with a reversed phase column. The siRNA and its metabolites were separated into single strands by elevated chromatographic temperature and analyzed using the ESI‐Orbitrap or the MALDI‐TOF mass spectrometer. Using this method, the 5' and/or 3' truncated metabolites of each strand were detected in the human serum samples. The ESI‐Orbitrap mass spectrometer enabled differentiation between two possible RNA‐based sequences, a monoisotopic molecular mass difference which was less than 2 Da, with an intrinsic mass resolving power. In‐source decay (ISD) analysis using a MALDI‐TOF mass spectrometer allowed the sequencing of the RNA metabolite with characteristic fragment ions, using 2,4‐dihydroxyacetophenone (2,4‐DHAP) as a matrix. The ESI‐Orbitrap mass spectrometer provided the highest mass accuracy and the benefit of on‐line coupling with HPLC for metabolite profiling. Meanwhile, the MALDI‐TOF mass spectrometer, in combination with 2,4‐DHAP, has the potential for the sequencing of RNA by ISD analysis. The combined use of these methods will be beneficial to characterize the metabolites of therapeutic siRNA compounds. Copyright © 2012 John Wiley & Sons, Ltd.     

Evaluation of a differential mobility spectrometer/miniature mass spectrometer system

A planar differential mobility spectrometer (DMS) was coupled to a Mini 10 handheld rectilinear ion trap (RIT) mass spectrometer (MS) (total weight 10 kg), and the performance of the instrument was evaluated using illicit drug analysis. Coupling of DMS (which requires a continuous flow of drift gas) with a miniature MS (which operates best using sample introduction via a discontinuous atmospheric pressure interface, DAPI), was achieved with auxiliary pumping using a 5 L/min miniature diaphragm sample pump placed between the two devices. On-line ion mobility filtering showed to be advantageous in reducing the background chemical noise in the analysis of the psychotropic drug diazepam in urine using nanoelectrospray ionization. The combination of a miniature mass spectrometer with simple and rapid gas-phase ion separation by DMS allowed the characteristic fragmentation pattern of diazepam to be distinguished in a simple urine extract at lower limits of detection (50 ng/mL) than that achieved without DMS (200 ng/mL). The additional separation power of DMS facilitated the identification of two drugs of similar molecular weight, morphine (average MW = 285.34) and diazepam (average MW = 284.70), using a miniature mass spectrometer capable of unit resolution. The similarity in the proton affinities of these two compounds resulted in some cross-interference in the MS data due to facile ionization of the neutral form of the compound even when the ionic form had been separated by DMS.     

Recent developments of miniature ion trap mass spectrometers

The recent development of miniature ion trap mass spectrometer systems in the last ten years is reviewed in this paper. These instruments adopt different atmospheric pressure interfaces (APIs), which are membrane inlets (MIs), discontinuous atmospheric pressure interface (DAPI) and continuous atmospheric pressure interface (CAPI).     

Improving pulsed radiofrequency glow discharge for time-of-flight mass spectrometry simultaneous elemental and molecular analysis

The performance of radiofrequency (rf) millisecond pulsed glow discharge (PGD) coupled to a fast orthogonal time-of-flight mass spectrometer (TOFMS) for chemical characterization and quantification of organic volatile compounds was investigated by using two different GD chamber designs. The designs investigated had substantial differences in the way that the volatile organic compound is introduced into the GD and the distance between the cathode and the sampling cone of the mass spectrometer. Bromochloromethane was selected as the model analyte because of the practical interest of determining trihalomethanes at low concentrations, and also because of both its low boiling point (to avoid problems associated with condensations in the interface) and the fact that it has two different heteroatoms, making the fragmentation patterns easier to follow. Pulse shapes of element, fragment, and molecular parent ions obtained by using the two GD chambers under investigation were critically compared. Results revealed the critical effect of the GD chamber geometry in obtaining the three types of chemical information, temporally discriminated. The spectra of the gaseous samples and of a polymer containing TBBPA (solid sample) were also compared. Detection limits for bromochloromethane in the order of low ng L^?1, and the required high tolerance of the plasmas to the introduction of organic vapours, were achieved using one of the proposed GD designs. The capability of the designed system for the analysis of other volatile compounds, for example dimethyl disulfide and dimethyl selenide, was also successfully evaluated, making use of the analytical potential of the information obtained from the different pulse time regions. Figure

?     

Automated in-line gel filtration for native state mass spectrometry

Characterization of protein-ligand complexes by nondenaturing mass spectrometry provides direct evidence of drug-like molecules binding with potential therapeutic targets. Typically, protein-ligand complexes to be analyzed contain buffer salts, detergents, and other additives to enhance protein solubility, all of which make the sample unable to be analyzed directly by electrospray ionization mass spectrometry. This work describes an in-line gel-filtration method that has been automated and optimized. Automation was achieved using commercial HPLC equipment. Gel column parameters that were optimized include: column dimensions, flow rate, packing material type, particle size, and molecular weight cut-off. Under optimal conditions, desalted protein ions are detected 4 min after injection and the analysis is completed in 20 min. The gel column retains good performance even after >200 injections. A demonstration for using the in-line gel-filtration system is shown for monitoring the exchange of fatty acids from the pocket of a nuclear hormone receptor, peroxisome proliferator activator-delta (PPARdelta) with a tool compound. Additional utilities of in-line gel-filtration mass spectrometry system will also be discussed.     

Determination of butyl- and phenyltin compounds in human urine by HS-SPME after derivatization with tetraethylborate and subsequent determination by capillary GC with microwave-induced plasma atomic emission and mass spectrometric detection

A headspace solid-phase micro-extraction (HS-SPME) method was developed and optimized for gas chromatographic separation and determination of commonly found organotin compounds in human urine after potential exposure. Butyl- and phenyltin compounds were in situ derivatized to ethylated derivatives by sodium tetraethylborate (NaBEt₄) directly in the urine matrix. The relevant parameters affecting the yield of the SPME procedure were examined using tetrabutyltin as internal standard. The method was optimized for direct use in the analysis of undiluted human urine samples and mono-, di- and tri-substituted butyl- and phenyltin compounds could be determined after a 15-min headspace extraction time at room temperature. The selectivity of the microwave-induced plasma atomic emission detector (MIP-AED) as an element specific detector in combination with the relatively selective sample preparation technique of HS-SPME allowed the interference-free detection of the organotin compounds in all cases. A quadrupole mass spectrometer was used in parallel experiments as a detector for the confirmation of the identity molecular structure of the eluted compounds. The performance characteristics of the developed method are given for the determination of mixtures of these compounds. Finally the proposed method was applied to the analysis of several human urine samples.     

Increasing the mass resolution in a TOF aerosol mass spectrometer using a nonlinear post focusing method

The mass resolution for the time of flight aerosol mass spectrometer for aerosol component analysis is dependent on the initial direction and energy of the ions. We have found that the shape of the optimum post focusing electric field is nonlinear. The maximum electric potential should be applied to the ions whose initial direction is 90°. To determine the post focusing effects, a laser ablation mass spectrometer was installed. By using this LA-MS, it was found that the average energy distribution of the laser ablated ions was 8 eV. To establish an optimum mass resolution, a time delay and a high voltage are needed. The study results showed that 1500 ns and 3.7 kV, respectively, were the optimum parameters for time delay and voltage for this system. Using these optimized parameters, good resolution between the isotope mass signals of copper was achieved.     

Optimization of a quadrupole ion storage trap as a source for time‐of‐flight mass spectrometry

Designs of a quadrupole ion trap (QIT) as a source for time‐of‐flight (TOF) mass spectrometry are evaluated for mass resolution, ion trapping, and laser activation of trapped ions. Comparisons are made with the standard hyperbolic electrode ion trap geometry for TOF mass analysis in both linear and reflectron modes. A parallel‐plate design for the QIT is found to give significantly improved TOF mass spectrometer performance. Effects of ion temperature, trapped ion cloud size, mass, and extraction field on mass resolution are investigated in detail by simulation of the TOF peak profiles. Mass resolution (m/Δm) values of several thousand are predicted even at room temperature with moderate extraction fields for the optimized design. The optimized design also allows larger radial ion collection size compared with the hyperbolic ion trap, without compromising the mass resolution. The proposed design of the QIT also improves the ion–laser interaction volume and photon collection efficiency for fluorescence measurements on trapped ions. Copyright © 2011 John Wiley & Sons, Ltd.     

Improving mass spectrometer sensitivity using a high-pressure electrodynamic ion funnel interface

We report on a new electrodynamic ion funnel that operates at a pressure of 30 torr with no loss of ion transmission. The enhanced performance compared with previous ion funnel designs optimized for pressures of <5 torr was achieved by reducing the ion funnel capacitance and increasing the RF drive frequency (1.7 MHz) and amplitude (100-170 V peak-to-peak). No degradation of ion transmission was observed for pressures from 2 to 30 torr. The ability to operate at higher pressure enabled a new tandem ion funnel mass spectrometer interface design that can accommodate a greater gas load (e.g., from an ESI source). When combined with a multicapillary inlet, the interface provided more efficient introduction of ions, resulting in a significant enhancement in mass spectrometer sensitivity and detection limits.     

Analysis of recombinant and modified proteins by capillary zone electrophoresis coupled with electrospray ionization tandem mass spectrometry.

A method for rapid characterization of recombinant and modified proteins with known sequences is described. The analytical system consists of a capillary zone electrophoresis (CZE) instrument coupled to an electrospray ionization ion trap tandem mass spectrometer via a sheath-flow interface. Following the procedure consists of proteolytic fragmentation, CZE peptide separation, tandem mass spectrometry (MS-MS) analysis of separated peptides, sequence database search and monitoring of the specific peptides, C 125 S mutated interleukin 2 (S-125-IL2) and bovine beta-casein were characterized as a model of recombinant protein and naturally modified protein, respectively. A tryptic peptide mixture derived from the synthetic salmon calcitonin (s-CT) was also analyzed to test the performance of the system. Although a conventional sheath-flow interface with much higher flow-rate compared to the microspray interface and nanospray interface was used, the proteins were identified at the low picomole level.     

Phenolics in selected European hardwood species by liquid chromatography-electrospray ionisation mass spectrometry

The phenols in beech (Fagus sylvatica), birch (Betula pendula) and ash (Fraxinus excelsior) wood dusts were compared using a mass spectrometer fitted with an electrospray ionisation interface with liquid chromatographic separation. Hardwood dust is a carcinogen, and an analysis of the polyphenol profile is a useful method for identifying the dust source in workplace air. The mass spectrometer was operated in both the negative and positive ion modes. Phenolic compounds were identified by comparing mass spectra and retention times from liquid chromatography with those for standard compounds and data in the literature. The phenol contents of the studied wood species varied considerably, and only a few common compounds were found in them.     

Strong anion‐exchange liquid chromatography coupled with isotope ratio mass spectrometry using a Liquiface interface

The introduction of liquid chromatography coupled with isotope ratio mass spectrometry (LC/IRMS) as an analytical tool for the measurement of isotope ratios in non‐volatile analytes has somewhat simplified the analytical cycle from sample collection to analysis mainly due to the avoidance of the extensive sample processing and derivatisation that were necessary for gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). Here we test the performance of coupling strong anion exchange to IRMS using only the second commercially available interface; the Liquiface. The system was modified from installation specification to improve peak resolution in the interface and maintain peak separation from the column to the mass spectrometer. The system performance was assessed by the determination of sensitivity, accuracy and precision attained from carbohydrate separations. The system performed satisfactorily after modifications, resulting in maintenance of peak resolution from column to mass spectrometer. The sensitivity achieved suggested that ～150 ng carbon could be analysed with acceptable precision (<0.3‰). Accuracy was maintained in the interface as determined by correlation with offline techniques, resulting in regression coefficient of r² = 0.98 and a slope of 0.99. The average precision achieved for the separation of seven monosaccharides was 0.36‰. The integration of a carbonate removal device limited the effect of background carbon perturbations in the mass spectrometer associated with eluent gradients, and the coupling of strong anion‐exchange chromatography with IRMS was successfully achieved using the Liquiface. Copyright © 2010 John Wiley & Sons, Ltd.