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The separation of eight antibiotics belonging to 5-nitroimidazole family was carried out by means of CEC coupled with MS. Preliminary experiments were carried out with ultraviolet detection in order to select the proper stationary and mobile phase. Among the different stationary phases studied (namely Lichrospher C18, 5 mu m particle size; Cogent (TM) Bidentate C18, 4.2 mu m; Pinnacle II (TM) Phenyl, 3 mu m; Pinnacle II (TM) Cyano, 3 mu m), Cogent (TM) Bidentate C18 (4.2 mu m) gave the best performance. For CEC-MS coupling, a laboratory assembled liquid-junction-nano-spray interface was used. In order to achieve a good sensitivity, special attention was paid to both optimization of the sheath liquid composition as well as selection of the injection mode. Under optimized CEC-ESI-MS conditions, the separation was accomplished within 22 min by using a column packed with a mixture of Bidentate C18:Lichrospher Silica-60 (5 mu m) 3:1 w/w, an inlet pressure of 11 bar, a voltage of 15 kV, and a mobile phase composed by 45:10:45 v/v/v ACN/MeOH/water containing ammonium acetate (5 mM pH 5). A combined hydrodynamic and electrokinetic injection of 8 bar, 15 kV, and 96 s was adopted. The method was validated in terms of repeatability and intermediate precision of retention times and peak areas, linearity, and LODs and LOQs. RSDs values were

In this work we present a new polyimide-based liquid junction nanospray interface, combining the advantages of sheath flow and sheathless designs. The liquid junction interface was prepared as a hybrid microfabricated plastic liquid junction sprayer allowing to perform analysis in standard (30-100 micro m ID) separation capillaries. The interface incorporates a self-aligning port for coupling of the separation capillary as well as ports for automated flushing of both the liquid junction and the separation capillary. The interface permits performing CE both with and without electroosmotic flow. The use of large bore capillaries allows injection and preconcentration of larger sample volumes, favoring detection sensitivity. Different compositions of the spray liquid and BGE can be used when needed.

The optimization of working parameters controlling the transfer of an analyte from the separation into the spray capillary at the liquid junction interface is a complex problem. The numerical models of hydrodynamic flow, electric field strength and the consequential mass transfer provide a valuable insight into the function of the miniaturized device. The results revealed that the most important parameter is the electric field strength inside the gap between the separation and spray capillaries. In a strong electric field, the analyte leaving the separation capillary is immediately transferred into the spray capillary at its maximum concentration. Although the losses of analyte outside the interface are dominant in this case, the sprayed concentration determines the detection sensitivity. Since only a small amount of the sprayed material enters mass spectrometer, the losses at the interface do not influence the sensitivity. At low electric field strength the total amount of analytes is transferred into the spray capillary. In this case, however, the analytes enter the capillary slowly and, as a result, are significantly diluted. The electric field strength, pressure and dimensions determine the mass transfer in the interface and must be considered when the optimum conditions of an analysis are chosen. Several fold improvement in sensitivity and efficiency of the method can be expected when working under the optimum conditions.

A rapid, high-resolution and high-efficiency novel method for the simultaneous determination of mesoporphyrin (MP), coproporphyrin (CP) and uroporphyrin (UP) in human urine by capillary electrophoresis coupled with electrospray ionization mass spectrometry (CE-ESI-MS) was developed. The simple laboratory-made interface was developed by using a micro-tee connection and a liquid junction. The porphyrins were well separated by CE with a positive 12 kV voltage in a 20 mM ammonium acetate buffer (pH 10.5) containing 40% v/v acetonitrile. The sheath liquid was composed of acetonitrile-water (50:50 v/v) containing 5 mM ammonium acetate and 0.1% v/v formic acid (pH 3.0) with a flow rate of 5 microL/min. The porphyrins were identified with the criteria of migration time and m/z values, and quantified with the measurement of peak area of [M+H]+ ion. Limits of detection for MP, CP and UP were 85 nM, 140 nM and 300 nM, respectively. The correlation coefficients of calibration curves were between 0.9956 and 0.9862. The intra-day precisions in terms of relative standard deviations (n = 12) of the method were 1.1-3.9% for migration times and 2.3-14.1% for peak areas. The porphyrin recoveries of spiked urine samples were between 73% and 96%. The liquid junction interface for CE-ESI-MS was demonstrated to be applicable for the potential quantitative analysis of clinical samples.

Electrospray ionization (ESI) is the most widely used ionization method in on-line coupling of capillary electrophoresis-mass spectrometry (CE-MS). The conventional coaxial sheath flow electrospray interface is currently being replaced by the more sensitive nanoelectrospray technique. The usual limitation of nanoelectrospray CE-MS interface has been its short lifetime caused by deterioration of the metal coating on the CE capillary terminus. This article describes an easy way to construct a more durable and sensitive nanospray interface for nonaqueous CE-MS. In this approach a very thin glass spray capillary (ca. 30 microm outer diameter) is partly inserted inside the CE capillary, the junction being surrounded by the electrolyte medium, which is in contact with the platinum electrode. The interface was tested with five pharmaceuticals: methadone, pentazocine, levorphanol, dihydrocodeine, and morphine. Detection limits ranged from 12 to 540 fmol. Separation efficiency and reproducibility were also studied. The CE current was found to be stable and the migration times were highly reproducible. All the CE separations were carried out in a nonaqueous background electrolyte solution.

A new nano-liquid-junction interface for coupling both capillary electrochromatography (CEC) or nano-liquid chromatography (nano-LC) with mass spectrometry (MS) was studied. The interface was a small T piece of polymeric material where capillary column and tip capillary were positioned at 180° while the third exit (at 90°) was occupied by a capillary delivering a liquid-assisting spray ionization for CEC experiments or by the electrode for the high voltage spray for nano-LC. Experiments were carried out analyzing mixtures of some organophosphorus pesticides (OPPs) or anti-inflammatory and related acidic drugs with MS detection in positive or negative ion mode, respectively. Analyzed OPPs compounds were baseline resolved utilizing the novel interface in both nano-LC and CEC obtaining good sensitivity and repeatability. For CEC-MS, the limits of detection ranged between 0.03 and 6.80 μg/mL and the intra-day repeatability was RSD

A simple self-aligning liquid junction-electrospray interface for coupling a capillary electrophoresis (CE) system to an atmospheric pressure ionization (API) mass spectrometer (CE-MS) was developed. In contrast to previous liquid junction interfaces, the self-aligning liquid junction interface simplifies the precise alignment of the CE capillary and the sprayer needle and uses a positive make-up flow. Several capillary CE-MS applications were run using both the self-aligning liquid junction interface and the widely used sheath flow interface for comparison purposes. The electrospray stability of the self-aligning liquid junction interface is consistently better even when non-volatile electrolyte solutions are used. At first, some band broadening was obtained with the self-aligning liquid junction interface. Experiments with different CE buffer systems suggested that this band broadening was caused by the materials used in constructing the interface. By using a more inert material for the sprayer needle, the self-aligning liquid junction exhibits excellent electrophoretic resolution, comparable sensitivity, and higher signal-to-noise ratios when run under the same conditions as the sheath flow interface.

Nano-LC and CEC were studied for the separation of cytochrome c tryptic digest. The peptides mixture was analyzed using either a nano-LC commercial or a laboratory assem-bled instrumentation coupled with an IT-ESI-MS by using a nanospray interface. CEC experiments were carried out with a CE apparatus coupled with the IT-ESI-MS through a liquid junction interface. Analytes were separated utilizing C18 silica based stationary phases, of different properties and origin, silica derivatized with cyano groups and C18 monolithic material. The last column, just because the chemical composition (absence of charged/chargeable groups) was tested only using nano-LC. Best results mainly related to the highest number of peptides separated and column equilibration time were obtained by nano-LC employing the C18 stationary phase (detection of 20 peptides, coverage of 88%). Similar results were achieved using both commercial and laboratory assembled instru-mentation. The use of CEC revealed a higher separation efficiency and shorter analysis time. However, the number of separated peptides were lower than those observed in nano-LC. In CEC the use of capillaries packed with cyanosilica particles offered better results; however, less satisfactory than those observed in theminiaturized LC technique. Provided the use of the same stationary phase and taking into account the driving forces, the two techniques can be considered complementary, offering different information related to the retention times of the studied peptides.

This article describes a method for the separation and determination of nine drugs of abuse in human urine, including amphetamines, cocaine, codeine, heroin and morphine. This method was based on SPE on a strong cation exchange cartridge followed by CEC-MS. The CEC experiments were performed in fused silica capillaries (100 microm x 30 cm) packed with a 3 mum cyano derivatized silica stationary phase. A laboratory-made liquid junction interface was used for CEC-MS coupling. The outlet capillary column was connected with an emitter tip that was positioned in front of the MS orifice. A stable electrospray was produced at nanoliter per minute flow rates applying a hydrostatic pressure (few kPa) to the interface. The coupling of packed CEC columns with mass spectrometer as detector, using a liquid junction interface, provided several advantages such as better sensitivity, low dead volume and independent control of the conditions used for CEC separation and ESI analysis. For this purpose, preliminary experiments were carried out in CEC-UV to optimize the proper mobile phase for CEC analysis. Good separation efficiency was achieved for almost all compounds, using a mixture containing ACN and 25 mM ammonium formate buffer at pH 3 (30:70, v/v), as mobile phase and applying a voltage of 12 kV. ESI ion-trap MS detection was performed in the positive ionization mode. A spray liquid, composed by methanol-water (80:20, v/v) and 1% formic acid, was delivered at a nano-flow rate of approximately 200 nL/min. Under optimized CEC-ESI-MS conditions, separation of the investigated drugs was performed within 13 min. CEC-MS and CEC-MS(2) spectra were obtained by providing the unambiguous confirmation of these drugs in urine samples. Method precision was determined with RSDs valuesor=3.3% for retention times andor=16.3% for peak areas in both intra-day and day-to-day experiments. LODs were established between 0.78 and 3.12 ng/mL for all compounds. Linearity was satisfactory in the concentration range of interest for all compounds (r(2)or=0.995). The developed CEC-MS method was then applied to the analysis of drugs of abuse in spiked urine samples, obtaining recovery data in the range 80-95%.

Capillary electrochromatography (CEC) was coupled with mass spectrometry (MS) for the separation ofsome selected pesticides and drug enantiomers. CEC separations were carried out in fused silica capillaries packed with either 5 micron RP18 silica or 5 micron silica modified vancomycin particles. The capillary column was connected with the MS utilizing a laboratory-made liquid-junction interface equipped with a 50 micron I.D. capillary-tip positioned at a few mm from the orifice of the MS. The CEC–MS set-up was operated without external pressure assistance during the electrochromatographic run commonly used to avoid bubble formation. However a hydrostatic pressure of a few kPa was applied only to the liquid-junction interface to optimize the ion-spray due to the low I.D. of the capillary-tip. In order to optimize the CEC–MS method, several experimental parameters were studied, namely the inlet pressure, the hydrostatic pressure into the liquid-junction interface, the type of sheath-liquid and the mobile phase. The application of an inlet pressure influenced only analyte retention times that were shortened by increasing the pressure. On the contrary the hydrostatic pressure applied to the interface increased the flow rate into the tip also increasing the ion-signal recorded in the mass spectrometry. The ion-signal raised almost linearly by increasing the outlet pressure till 3.5 kPa and then decreased. The separation of the selected pesticides was not influenced at all changing the hydrostatic pressure on the interface. Some basic enantiomeric compounds of pharmaceutical interest were successfully separated by CEC achieving good resolution. They were detected by MS with limit of detection in a range of 0.24–0.60 microg/mL.

A pressurized liquid junction nanoelectrospray interface was designed and optimized for reliable on-line CE-MS coupling. The system was constructed as an integrated device for highly sensitive and selective analyses of proteins and peptides with the separation and spray capillaries fixed in a pressurized spray liquid reservoir equipped with the electrode for connection of the electrospray potential. The electrode chamber on the injection side of the separation capillary and the spray liquid reservoir were pneumatically connected by a Teflon tube filled with pressurized nitrogen. This arrangement provided precisely counterbalanced pressures at the inlet and outlet of the separation capillary. The pressure control system was driven by an electrically operated valve and maintained the optimum flow rate for the electrospray stability. All parts of the interface being in contact with the CEBGE, spray liquid and/or sample were made of glass or Teflon. The use of these materials minimized the electrospray chemical noise often caused by plastic softeners or material degradation. During optimization, the transfer of the separated zones between the separation and electrospray capillaries was monitored by UV absorbance and contactless conductivity detectors placed at the outlet of the separation capillary and inlet of the electrospray tip, respectively. This arrangement allowed independent monitoring of the effects of pressure, CE voltage and geometry of the liquid junction on the spreading and dilution of the separated zones after passage through the interface.

A simple and cost-effective laboratory-made liquid junction interface was used for coupling of CE with MS. In this device the capillary column and the spray tip were positioned in the electrode vessel containing appropriate spray liquid. The electrospray potential was applied on the electrode inside the liquid junction. A stable electrospray was produced at nanoliter per minute flow rates generated in the emitter tip without using an external pump. This arrangement provided high durability of the spray tip and independent optimization of the CE separation (use of coated capillaries) and ESI conditions. CE-MS analysis of mixtures of drugs, peptides, tryptic digests of proteins and biological fluids was optimized with respect to the effects of the distance between the separation capillary and electrospray tip and pressure applied on the liquid junction. The sensitivity of the system, in terms of the LOD (base peak monitoring) was below 10 ng/mL for the beta-blocker drugs and below 200 ng/mL for peptide analysis.

The on-line coupling of capillary electrophoresis and mass spectrometry using a continuous-flow fast-atom bombardment system in combination with a liquid-junction interface is described. The influence of the liquid-junction coupling on the efficiency and the resolution is investigated. Qualitative analysis of small peptides and quantitative analysis of dextromethorphan is demonstrated with detection limits in the low ng/mL range, indicating the feasibility of the method in bioanalysis. Copyright © 1989 Heyden & Son Ltd