Abstract:
Electrospray ionization sources interfaced to mass spectrometers have become widely used tools in analytical applications Processes occurring in Electrospray (ES) ionization generally include the addition or removal of a charged species such as II+ or other cation to effect ionization of a sample species. Electrospray includes ionization processes that occur in the liquid and gas phase and in both phases ionization processes require a source or sink for such charged species. Electrolyte species, that aid in oxidation or reduction reactions occurring in Electrospray ionization, are added to sample solutions in many analytical applications to increase the ES ion signal amplitude detected by a mass spectrometer (MS). Electrolyte species that may be required to enhance an upstream sample preparation or separation process may be less compatible with the downstream ES processes and cause reduction in MS signal. A new set of Electrolytes has been found that increases positive and negative polarity analyte ion signal measured in ESMS analysis when compared with analyte ESMS signal achieved using more conventional electrolytes. The new electrolyte species increase ES MS signal when added directly to a sample solution or when added to a second solution flow in an Electrospray membrane probe. The new electrolytes can also be added to a reagent ion source configured in a combination Atmospheric pressure ion source to improve ionization efficiency.

Description:
RELATED APPLICATIONS 
       [0001]    This application claims the benefit of Provisional Patent Application No. 60/932,644 filed Jun. 1, 2007 the contents of which are incorporated by reference herein. 
     
    
     FIELD OF INVENTION 
       [0002]    This invention relates to the field of Atmospheric Pressure Ion (API) sources interfaced to mass spectrometers. Such API sources include but are not limited to Electrospray, Atmospheric Pressure Chemical Ionization (APCI), Combination Ion Sources, Atmospheric Pressure Charge Injection Matrix Assisted Laser Desorption, DART and DESI. The invention comprises the use of new electrolyte species to enhance the analyte ion signal generated from these API sources interfaced to mass spectrometers. 
       BACKGROUND OF THE INVENTION 
       [0003]    Charged droplet production unassisted or pneumatic nebulization assisted Electrospray (ES) requires oxidation of species (positive ion polarity ES) or reduction of species (negative ion polarity) at conductive surfaces in the sample solution flow path. When a metal Electrospray needle tip is used that is electrically connected to a voltage or ground potential, such oxidation or reduction reactions (redox) reactions occur on the inside surface of the metal Electrospray needle during Electrospray ionization. If a dielectric Electrospray tip is used in Electrospray ionization, redox reactions occur on an electrically conductive metal surface contacting the sample solution along the sample solution flow path. This conductive surface typically may by a stainless steel union connected to a fused silica Electrospray tip. The Electrospray sample solution flow path forms one half cell of an Electrochemical or voltaic cell. The second half of the Electrochemical cell formed in Electrospray operates in the gas phase. Consequently, operating rules that can be used to explain or predict the behavior of liquid to liquid Electrochemical cells may be applied to explain a portion of the processes occurring in Electrospray ionization. The electrolyte aids in promoting redox reactions occurring at electrode surfaces immersed in liquid in electrochemical cells. The electrolyte not only plays a role in the initial redox reactions required to form single polarity charged liquid droplets but also fundamentally affects the production of sample related ions from rapidly evaporating liquid droplets and their subsequent transport through the gas phase into vacuum. Additional charge exchange reactions can occur with sample species in the gas phase. The mechanism by which the electrolyte affects liquid and gas phase ionization of analyte species is not clear 
         [0004]    The type and concentration of electrolyte species effects ES ionization efficiency. The electrolyte type and concentration and sample solution composition will affect the dielectric constant, conductivity and pH of the sample solution. The relative voltage applied between the Electrospray tip and counter electrodes, the effective radius of curvature of the Electrospray tip and shape of the emerging fluid surface determine the effective electric field strength at the Electrospray needle tip. The strength of the applied electric field is generally set just below the onset of gas phase breakdown or corona discharge in Electrospray ionization. With an effective upper bound on the electric field that is applied at the Electrospray tip during Electrospray operation, the Electrospray total ion current is determined by the solution properties as well as the placement of the conductive surface along the sample solution flow path. The effective conductivity of the sample solution between the nearest electrically conductive surface in contact with the sample solution and the Electrospray tip plays a large role in determining the Electrospray total ion current. It has been found with studies using Electrospray Membrane probes that the ESMS analyte signal can vary significantly with Electrospray total ion current. A description of the Electrospray Membrane probe is given in U.S. patent application Ser. Nos. 11/132,953 and 60/840/095 and incorporated herein by reference. 
         [0005]    ES signal is enhanced when specific organic acid species such as acetic and formic acids are added to organic and aqueous solvents. Conversely, ES signal is reduced when inorganic acids such as hydrochloric or trifluoroacetic acid are added to Electrospray sample solutions. Although mechanisms underlying variation in Electrospray ionization efficiency due to different electrolyte counter ion species have been proposed, explanations of these root modulators underlying Electrospray ionization processes remain speculative. Conventional electrolytes added to sample solutions in Electrospray ionization are generally selected to maximize Electrospray MS analyte ion signal Alternatively, electrolyte species and concentrations are selected to serve as a reasonable compromise to optimize upstream sample preparation or separation system performance and downstream Electrospray performance. Trifluoroacetic acid may be added to a sample solution to improve a reverse phase gradient liquid chromatography sample separation but its presence will reduce the Electrospray MS signal significantly compared with Electrospraying with art organic electrolyte such as Formic or Acetic acid added to the sample solution Generally for polar analyte species, the highest Electrospray MS signal will be achieved using a polar organic solvent such as methanol in water with acetic or formic acid added as the electrolyte. Typically, a 30:70 to 50:50 methanol to water ratio is run with acetic or formic acid concentrations ranging from 0.1% to over 1%. Running non polar solvents, such as acetonitrile, with water will reduce the ESMS signal for polar compounds and adding inorganic acid will reduce ESMS signal compared to the signal achieved using a polar organic solvent in water with acetic or formic acid. Several species of acids bases and salts have been used at different concentrations and in different solvent compositions as electrolyte species in Electrospray ionization to maximize ESMS analyte species. For some less polar analyte samples that do not dissolve in aqueous solutions, higher ESMS signal is achieved running the sample in pure acetonitrile with an electrolyte. For compounds such as carbohydrates with low or no proton affinity, adding a salt electrolyte may product higher ESMS signal. 
         [0006]    The invention comprises using a new set of electrolyte species in Electrospray ionization to improve the Electrospray ionization efficiency of analyte species compared with ES ionization efficiency achieved with conventional electrolyte species used and reported for Electrospray ionization. Electrospraying with the new electrolyte species increases ESMS analyte signal amplitude by a factor of two to ten compared to the highest ESMS signal achieved using acetic or formic acids. ESMS signal enhancements have been achieved whether the new electrolytes are added directly to the sample solution or added to the second solution of an Electrospray membrane probe. When convention acid or salt electrolytes added to the sample solution are Electrosprayed in positive polarity mode, the anion from these electrolytes does not readily appear in the positive ion spectrum. As expected, the anion of these electrolytes does appear in the negative ion polarity ESMS spectrum. One distinguishing characteristic of the new electrolytes comprising the invention is that a characteristic protonated or deprotonated parent related ion from the electrolyte species appears in both positive and negative polarity spectrum acquired using Electrospray ionization. The positive polarity electrolyte ion appearing in the positive polarity Electrospray mass spectrum is the (M+H) +  species with the (M−H) −  species appearing in the negative polarity Electrospray mass spectrum. 
       SUMMARY OF THE INVENTION 
       [0007]    One embodiment of the invention comprises conducting Electrospray ionization of an analyte species with MS analysis where at least one of a new set of electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic is added directly to the sample solution. The electrolyte may be included in the sample solution from its fluid delivery system or added to the sample solution near the Electrospray tip through a tee fluid flow connection. 
         [0008]    Another embodiment of the invention is running at least one of a set (Anew electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic in the second solution flow of an Electrospray membrane probe during Electrospray of the sample solution. The concentration of the new electrolyte can be varied or scanned by running step functions or gradients through the second solution flow path. The second solution flow is separated from the sample solution flow by a semipermeable membrane that allows reduced concentration transfer of the new electrolyte into the sample solution flow during Electrospray ionization with MS analysis. 
         [0009]    Another embodiment of the invention is running at least one of a set of new electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic in the second solution of an Electrospray membrane probe during Electrospray of the sample solution that contains a second electrolyte species. The addition of the new electrolyte to the second solution flow increases the Electrospray MS signal even if the second electrolyte species, when used alone, reduces the ESMS analyte signal. The concentration of the new electrolyte in the second solution flow can be step or ramped to maximize analyte ESMS signal 
         [0010]    Another embodiment of the invention comprises running at least one of a set of new electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic in the downstream membrane section second solution flow of a multiple membrane section Electrospray membrane probe during Electrospray ionization with MS analysis. One or more membrane sections can be configured upstream in the sample solution flow path from the downstream Electrospray membrane probe. Electrocapture and release of samples species using upstream membrane sections can be run with electrolyte species that optimize the Electrocapture processes independently while a new electrolyte species is run through the downstream membrane section second solution flow to optimize Electrospray ionization efficiency of the analyte species. 
         [0011]    In yet another embodiment of the invention, at least one of the new electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic are added to the sample solution in a single APCI inlet probe or sprayed from a second solution in a dual APCI inlet probe to enhance the ion signal generated in Atmospheric Pressure Corona Discharge Ionization. 
         [0012]    In another embodiment of the invention, at least one of the new electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic are added to the solution Electrosprayed from a reagent ion source comprising an Electrospray ion generating source configured in a combination ion source including Electrospray ionization and/or Atmospheric Pressure Chemical Ionization. 
         [0013]    In yet another embodiment of the invention, at least one of the new electrolytes including but not limited to Benzoic acid, Cyclohexanecarboxylic Acid or Trimethyl Acetic are added to the solution that is nebulized followed by corona discharge ionization forming a reagent ion source configured in a combination ion source including Electrospray ionization and/or Atmospheric Pressure Chemical Ionization. 
     
    
     
       BRIEF DESCRIPTION OF THE INVENTION 
         [0014]      FIG. 1  is a schematic of an Electrospray Ion Source interfaced to a mass spectrometer. 
           [0015]      FIG. 2  is a cross section diagram of an Electrospray Membrane probe. 
           [0016]      FIG. 3  is a zoomed in view of the sample solution flow channel, the second solution flow channel and the semipermeable membrane in an Electrospray Membrane Probe 
           [0017]      FIG. 4  shows a single section Electrospray Membrane probe integrated with pneumatic nebulization sprayer mounted on an Electrospray ion source probe mounting plate. 
           [0018]      FIG. 5  is a schematic of a three section Electrospray Membrane probe 
           [0019]      FIG. 6  is a diagram of a combination atmospheric pressure ion source comprising a sample solution Electrospray inlet probe and an Electrospray reagent ion source. 
           [0020]      FIG. 7  shows the ESMS ion signal curves for a 1 μM Hexatyrosine in a 1:1 methanol:water solution Electrosprayed at a flow rate of 10 μl/min while running electrolyte concentration gradients in the Electrospray Membrane probe second solution flow using conventional electrolyte species and a new electrolyte species. 
           [0021]      FIG. 8  shows the ESMS signal curves for a 1 μM Hexatyrosine in a 1:1 methanol:water solution Electrosprayed at a flow rate of 10 μl/min while running conventional and new electrolyte species concentration gradients in the Electrospray Membrane probe second solution flow and with benzoic acid added directly to the sample solution at different concentrations. 
           [0022]      FIG. 9  shows a set of ESMS signal curves comparing ESMS ion signal of a 1 μM Hexatyrosine in a 1:1 methanol: water solution Electrosprayed at a flow rate of 10 μl/min for different concentrations of acetic acid and cyclohexanecarboxylic acid added directly to the sample solution 
           [0023]      FIG. 10  shows a set of ESMS signal curves comparing positive polarity ESMS ion signal of a 1 μM Hexatyrosine in a 1:1 methanol:water solution Electrosprayed at a flow rate of 10 μl/min while running acetic acid and benzoic acid electrolyte concentration gradients in the Electrospray Membrane probe second solution flow with pure solvent sample solutions and with 0.001% trifluoroacetic acid added to the sample solution. 
           [0024]      FIG. 11  shows a set of ESMS signal curves comparing negative polarity ESMS ion signal of a 1 μM Hexatyrosine in a 1:1 methanol:water solution Electrosprayed at a flow rate of 10 μl/min while running acetic acid and benzoic acid electrolyte concentration gradients in the Electrospray Membrane probe second solution flow with pure solvent sample solutions. 
           [0025]      FIG. 12  shows a set of ESMS signal curves comparing positive polarity ESMS ion signal of a 1 μM reserpine in 1:1 methanol:water solution running at a flow rate of 10 μl/min for acetic acid, benzoic acid and trimethyl acetic acids added individually to the sample solution at different concentrations. 
           [0026]      FIG. 13  shows a set of ESMS signal curves comparing positive polarity ESMS ion signal of a 1 μM leucine enkephalin in a 1:1 methanol:water solution running at a flow rate of 10 μl/min for acetic acid, benzoic acid, cyclohexanecaboxylic acid and trimethyl acetic acids added individually to the sample solution at different concentrations. 
           [0027]      FIG. 14A  is a positive polarity Electrospray mass spectrum of benzoic Acid and  FIG. 14B  is a negative polarity mass spectrum of benzoic acid. 
           [0028]      FIG. 15A  is a positive polarity Electrospray mass spectrum of trimethyl acetic acid and  FIG. 15B  is a negative polarity mass spectrum of trimethyl acetic acid. 
           [0029]      FIG. 16A  is a positive polarity Electrospray mass spectrum of cyclohexanecarboxylic acid and  FIG. 16B  is a negative polarity mass spectrum of cyclohexanecarboxylic acid. 
       
    
    
     DESCRIPTION OF THE INVENTION 
       [0030]    Electrospray total ion current, for a given applied electric field, is a function of the sample solution conductivity between the Electrospray tip and the first electrically conductive surface in the sample solution flow path. The primary charge carrier in positive ion Electrospray is generally the H+ ion which is produced from redox reactions occurring at electrode surfaces in contact with the sample solution in conventional Electrospray or a second solution in Electrospray Membrane probe. The electrolyte added to the sample or second solution plays a direct or indirect role in adding or removing H+ ions in solution during Electrospray ionization. The indirect role in producing H+ ions is the case where the electrolyte aids in the electrolysis of water at the electrode surface to produce H+ ions. The direct role an electrolyte can play is to supply the H+ ion directly from dissociation of an acid and loss of an electron at the electrode surface. The type and concentration of the electrolyte anion or neutral molecule in positive ion polarity and even negative ion polarity significantly affects the Electrospray ionization efficiency for most analyte species. The mechanism or mechanisms through which the electrolyte operates to affect ion production in Electrospray ionization is not well understood. Even the role an electrolyte plays in the redox reactions that occur during Electrospray charged droplet formation is not well characterized. Consequently, the type and concentration of the electrolyte species used in Electrospray ionization is determined largely through trial and error with decisions based on empirical evidence for a given Electrospray MS analytical application. To this end, a number of electrolyte species were screened using an Electrospray membrane probe to determine if electrolyte species different from those used conventionally or historically provided improved Electrospray performance. A set of such new electrolytes was found which demonstrated improved analyte ESMS signal in both positive and negative positive modes. The set of new electrolytes comprises but may not be limited benzoic acid, trimethylacetic acid and cyclohexanecaboxylic acid. 
         [0031]    As noted above, unlike electrolytes conventionally or historically used in Electrospray ionization, when Electrospraying with a new electrolyte, a characteristic electrolyte ion peak is generated in both positive and negative ion polarity mode. The (M+H) +  ion is generated for benzoic acid, trimethyl acetic acid and cyclohexanecarboxylic acid in positive polarity Electrospray ionization. Conversely, the (M−H) −  ion, as expected, is generated when Electrospraying benzoic acid, trimethyl acetic acid and cyclohexanecarboxylic acid in negative polarity as shown in  FIGS. 14 ,  15  and  16 . The mechanism or mechanisms by which the new electrolyte enhances the Electrospray signal may occur in the liquid phase, gas phase or both. Benzoic acid has a low gas phase proton affinity so protonated benzoic acid ion may readily donate an H+ to gas phase neutral analyte species or may reduce the neutralization of the Electrospray produced analyte ion by transferring protons to competing higher proton affinity contamination species in the gas phase. 
         [0032]    A cross section schematic of Electrospray ion source  1  is shown in  FIG. 1 . Electrospray sample solution inlet probe  2  comprises sample solution flow channel or tube  3 , Electrospray tip  4  and annulus  5  through which pneumatic nebulization gas  7  flows exiting concentrically  6  around Electrospray tip  4 . Different voltages are applied to endplate and nosepiece electrode  11 , capillary entrance electrode  12  and cylindrical lens  13  to generate single polarity charged droplets in Electrospray plume  10 . Typically, in positive polarity Electrospray ionization, Electrospray tip  4  would be operated at ground potential with −3 KV, −5 KV and −6 KV applied to cylindrical lens  13 , nosepiece and endplate electrode  11  and capillary entrance electrode  12  respectively. Gas heater  15  heats countercurrent drying gas flow  17 . Charged droplets comprising charged droplet plume  10  produced by unassisted Electrospray or Electrospray with pneumatic nebulization assist evaporate as they pass through Electrospray source chamber  18 . Heated countercurrent drying gas  14  exiting through the orifice in nosepiece electrode  11  aids in the drying of charged liquid droplets comprising Electrospray plume  10 . A portion of the ions generated from the rapidly evaporating charged liquid droplets are directed by electric fields to pass into and through orifice  20  of dielectric capillary  21  into vacuum. Ions exiting capillary orifice  20  are directed through skimmer orifice  27  by the expanding neutral gas flow and the relative voltages applied to capillary exit lens  23  and skimmer electrode  24 . Ions exiting skimmer orifice  27  pass through ion guide  25  and into mass to charge analyzer  28  where they are mass to charge analyzed and detected as is known in the art 
         [0033]    The analyte ion signal measured in the mass spectrometer is due in large part to efficiency of Electrospray ionization for a given analyte species. The Electrospray ionization efficiency includes the processes that convert neutral molecules to ions in the atmospheric pressure ion source and the efficiency by which the ions generated at atmospheric pressure are transferred into vacuum. The new electrolyte species may play a role in both mechanisms that affect Electrospray ionization efficiency. In one embodiment of the invention, at least one of the new electrolytes including, benzoic acid, trimethyl acetic acid and cyclohexanecarboxylic acid is added to sample solution  8  delivered through sample solution flow channel  3  to Electrospray tip  4  where the sample solution is Electrosprayed into Electrospray ion source chamber  18 . 
         [0034]      FIG. 2  shows the cross section diagram of an Electrospray Membrane Probe  30  that is used in an alternative embodiment of the invention. Electrospray Membrane probe  30 , more fully described in U.S. patent application Ser. No. 11/132,953 and incorporated herein by reference, comprises sample solution flow channel  31 A through which sample solution flow  31  flows exiting at Electrospray tip  4 . Common elements with  FIG. 1  retain the element numbers. A second solution  32 , in contact with electrode  33 , passes through second solution flow path  32 A. Voltage is applied to electrode  33  from power supply  35 . Sample solution  31  and second solution  32  are separated by semipermeable membrane  34 . Semipermeable membrane  34  may comprise a cation or anion exchange membrane. A typical cation exchange membrane is Nafion™ that may be configured with different thicknesses and/or conductivity characteristics in Electrospray Membrane probe assembly  30 . Second solution  32  flow is delivered into second solution flow channel  32 A from an isocratic or gradient fluid delivery system  37  through flow channel  36  and exits through channel  38 . Sample solution  31  flow is delivered to sample solution flow channel  31 A from isocratic or gradient fluid delivery system  40  through flow channel  41 . Dielectric probe body sections  42  and  43  comprise chemically inert materials that do not chemically react with sample solution  31  and second solution  32 . Sample solution  31  passing through flow channel  31 A is Electrosprayed from Electrospray tip  4  with or without pneumatic nebulization assist forming Electrospray plume  10 . Electrospray with pneumatic nebulization assist is achieved by flowing nebulization gas  7  through annulus  5  exiting at  6  concentrically around Electrospray tip  4 . To effect the Electrospray generation of single polarity charged liquid droplets, relative voltages are applied to second solution electrode  33 , nosepiece and endplate electrode  11  and capillary entrance electrode  12  using power supplies  35 ,  49  and  50  respectively. Heated counter current drying gas  14  aids in drying charge liquid droplets in spray plume  10  as they move towards capillary orifice  20  driven by the applied electric fields. A portion of the ions produced from the rapidly evaporating droplets in Electrospray plume  10  pass through capillary orifice  20  and into mass to charge analyzer  28  where they are mass to charge analyzed and detected. 
         [0035]      FIG. 3  is a diagram of one Electrospray Membrane probe  30  operating mode for positive polarity Electrospray ionization employing an alternative embodiment of the invention At least one new electrolyte species comprising benzoic acid, trimethyl acetic acid and cyclohexanecarboxylic acid is added in higher concentration to the solution contained in Syringe  54  of fluid delivery system  37 . Syringe  55  is filled with the same solvent composition as loaded into Syringe  54  but without a new electrolyte species added. A specific isocratic new electrolyte concentration or a new electrolyte concentration gradient for second solution  32  can be delivered to second solution flow channel  32 A by setting the appropriate ratios of pumping speeds on syringes  54  and  55  in fluid delivery system  37 . During positive ion polarity Electrospray ionization, H+ is produced at the surface of second solution electrode  33  and passes through semipermeable membrane  34 , most likely as H 3 O + , into sample solution  31 , driven by the electric field. A portion of the new electrolyte species flowing through second solution flow channel  32 A also passes through semipermeable membrane  34  entering sample solution  31  and forming a net concentration of new electrolyte in sample solution  31 . The new electrolyte concentration in solution  31  during Electrospray operation is well below the new electrolyte concentration in second solution  32 . The Electrospray total ion current and consequently the local sample solution pH at Electrospray tip  4 , the new electrolyte concentration in sample solution  31  and the sample ion Electrospray MS signal response can be controlled by adjusting the new electrolyte concentration in second solution  32  flowing through second solution flow channel  32 A. The solvent composition of second solution  32  can be configured to be different from the solvent composition of the sample solution to optimize solubility and performance of a new electrolyte species. 
         [0036]      FIG. 4  shows one embodiment of Electrospray Membrane probe  57  comprising single membrane section assembly  58  connected to pneumatic nebulization Electrospray inlet probe assembly  59  mounted on Electrospray ion source probe plate  61 . Common elements diagrammed in  FIGS. 1 ,  2  and  3  retain the same element numbers. 
         [0037]      FIG. 5  is a diagram of three membrane section Electrospray Membrane probe assembly  64  comprising Electrocapture dual membrane section  67  and single Electrospray Membrane section  68 . Each membrane section operates in a manner similar to the single section Electrospray membrane probe described in  FIGS. 2 and 3 . Electrocapture Dual membrane section  67  comprises second solution flow channel  70  with electrode  71  and semipermeable membrane section  76  and second solution flow channel  72  with electrode  73  and semipermeable membrane section  77 . Single membrane section  68  comprises second solution flow channel  74  and electrode  75  with semipermeable membrane  78 . The electrolyte type and concentration and solution composition can be controlled in second solutions  80 ,  81  and  82  as described previously. Common elements described in  FIGS. 1 through 4  retain their element numbers in  FIG. 5 . Electrical potential curve  84  is a diagram of one example of relative electrical potentials set along the sample solution flow path for positive polarity Electrospray ionization and positive ion Electrocapture. Dual membrane Electrocapture section  67  can be operated to trap and release positive or negative polarity sample ions in the sample solution as described in pending PCT Patent Application Number PCT/SE2005/001844 incorporated herein by reference. In an alternative embodiment of the invention, at least one new electrolyte including benzoic acid, trimethyl acetic acid or cyclohexanecarboxylic acid species is added to second solution  82  with the concentration controlled to maximize Electrospray sample ion signal as described above. Second solution  82  composition and flow rate can be varied and controlled independently from second solutions  80  and  81  compositions and flow rates to independently optimize Electrocapture and on line Electrospray performance. 
         [0038]      FIG. 6  is a diagram of atmospheric pressure combination ion source  88  comprising Electrospray inlet probe assemblies  90  and  91  with pneumatic nebulization assist. Electrospray inlet probe  90  comprises Electrospray tip  4  and auxiliary gas heater  92  heating gas flow  93  to aid in the drying of charged liquid droplets comprising Electrospray plume  10 . Voltage applied to ring electrodes  94  and  95  allow control of the production of net neutral or single polarity charged liquid droplets from Electrospray inlet probes  90  and  91  respectively while minimizing undesired electric fields in spray mixing region  96 . Electrospray inlet probe  91  provides a source of reagent ions that when drawn through spray plume  10  by electric fields  97  effect atmospheric chemical ionization of a portion of the vaporized neutral sample molecules produced from evaporating charged droplets in spray plume  10 . Combination ion source  88  can be operated in Electrospray only mode, APCI only mode or a combination of Electrospray and APCI modes as described in pending U.S. patent application Ser. No. 11/396,968 incorporated herein by reference. In an alternative embodiment of the invention, at least one new electrolyte, including benzoic acid, trimethyl acetic acid or cyclohexanecarboxylic acid, can be added to the sample flow solution of Electrospray inlet probe  90  and/or to the reagent solution of Electrospray inlet probe  91  which produces reagent ions to promote gas phase atmospheric pressure chemical ionization in mixing region  96  New electrolyte species run in sample solutions can increase the sample ESMS ion single as described above. In addition, new electrolytes in the reagent solution Electrosprayed from Electrospray probe  91  form low proton affinity protonated ions in positive ion polarity Electrospray which serve as reagent ions for charge exchange in atmospheric pressure chemical ionization or combination ES and APCI operation New electrolyte species may also be added to sample solution in corona discharge reagent ion sources or APCI sources to improve APCI source performance. 
         [0039]      FIG. 7  shows a set of ESMS ion signal curves for 1 μM Hexatyrosine sample in a 1:1 methanol:water sample solutions Electrosprayed using an Electrospray Membrane probe configuration  30  as diagrammed in  FIGS. 1 ,  2  and  3 . All sample solutions were run at a flow rate of 10 μl/min. Concentration gradients of different electrolyte species were run in the second solution flow channel while acquiring Electrospray mass spectrum. The second solution solvent composition was methanol:water for all electrolytes run with the exception of Naphthoxyacetic acid which was run in a methanol second solution. As the concentration of the added electrolyte increased in the second solution flow, the Electrospray total ion current increased. Each curve shown in  FIG. 7  is effectively a base ion chromatogram with the Hexatyrosine peak amplitude plotted over Electrospray total ion current. Signal response curves  100 ,  101 ,  102 ,  103  and  104  for Hexatyrosine versus Electrospray total ion current were acquired when running second solution concentration gradients of acetic acid (up to 10%), 2 napthoxyacetic acid (up to 37M), trimellitic acid (up to 244 M), 1,2,4,5 Benzene Carboxylic acid (up to 233 M) and terephthalic acid (saturated) respectively. Conventional electrolyte, acetic acid, provided the highest hexatyrosine ESMS signal amplitude for this set of electrolytes as shown in  FIG. 6 . Hexatyrosine signal response curve  108  was acquired while running a concentration gradient in the second solution of new electrolyte cyclohexanecarboxylic acid (up to 195 M). The maximum hexatyrosine signal achieved with new electrolyte run in the second solution of Electrospray Membrane probe  30  was two times the maximum amplitude achieved with acetic acid as an electrolyte. The limited cross section area of the semipermeable membrane in contact with the sample solution limited the Electrospray total ion current range with new electrolyte cyclohexanecarboxylic acid run in the second solution. As will be shown in later figures, higher analyte signal can be achieved by adding new electrolyte species directly to the sample solution. The difference in the shape and amplitude of curve  108  illustrates the clear difference in performance of the Electrospray ionization process when new electrolyte cyclohexanecarboxylic acid is used. 
         [0040]      FIG. 8  shows another set of ESMS ion signal curves for 1 μM hexatyrosine sample in a 1:1 methanol:water sample solutions Electrosprayed using an Electrospray Membrane probe configuration  30  as diagrammed in  FIGS. 1 ,  2  and  3 . Hexatyrosine Electrospray MS signal response curves  110  through  112  and  115  were acquired while running electrolyte concentration gradients in the second solution flow of Electrospray Membrane probe  30 . Hexatyrosine Electrospray MS signal response curve  118  was acquired by Electrospraying different sample solutions having different new electrolyte benzoic acid concentrations added directly to the sample solution. ESMS signal response curve  114  with end data point  113  for hexatyrosine was acquired by Electrospraying different sample solutions comprising different concentrations of citric acid added directly to the sample solutions. No Electrospray membrane probe was used to generate curves  114  or  118 . Signal response curves  110 ,  111 ,  112  and  115  for Hexatyrosine versus Electrospray total ion current were acquired when running second solution concentration gradients of conventional electrolytes, acetic acid (up to 10% in the second solution), formic acid (up to 5%) and nitric acid (up to 1%) and new electrolyte benzoic acid (up to 0.41 M in the second solution) respectively. Comparing the hexatyrosine ESMS signal response with new electrolyte benzoic acid added to the second solution of membrane probe  30  or directly to the sample solution during Electrospray ionization, similar ion signals are obtained for the same Electrospray ion current generated. Electrospray performance with the electrolyte added to the Electrospray Membrane probe second solution generally correlates well with the Electrospray performance with the same electrolyte added directly to the sample solution during Electrospray ionization for similar Electrospray total ion currents. As shown by curves  115  and  118 , increased hexatyrosine ESMS signal is achieved when new electrolyte benzoic acid is added to the second solution of Electrospray Membrane probe  30  or directly to the sample solution during Electrospray ionization. The maximum hexatyrosine ESMS signal shown by signal response curve  118  was over five times higher than that achieved with any of the conventional electrolytes acetic, formic or nitric acids or non conventional electrolyte citric acid. 
         [0041]    Electrospray MS signal response curves  120  and  121  for 1 μM hexatyrosine sample in a 1:1 methanol:water solutions are shown in  FIG. 9  Curve  121  was generated by Electrospraying different sample solutions containing different concentrations of conventional electrolyte acetic acid. Curve  120  was generated by Electrospraying different sample solutions containing different concentrations of new electrolyte cyclohexanecarboxylic acid. The maximum hexatyrosine ESMS signal achieved with new electrolyte cyclohexanecarboxylic acid was over two time higher than the maximum hexatyrosine signal achieved with conventional electrolyte acetic acid. 
         [0042]    Three ESMS signal response curves using Electrospray membrane probe  30  for 1 μM hexatyrosine sample in 1:1 methanol:water solutions are shown in  FIG. 10 . Curve  122  was generated by running a concentration gradient of acetic acid in the Electrospray Membrane probe second solution flow. Over a factor of two increase in hexatyrosine signal was achieved by running a concentration gradient of benzoic acid in the second solution of the Electrospray Membrane probe as shown by signal response curve  123 . The addition of inorganic electrolytes to the sample solution generally reduces the analyte signal response for a given Electrospray total ion current. Hexatyrosine signal response curve  124  was acquired with 0.001% trifluoroacetic acid (TFA) added to the sample solution while running a concentration gradient of benzoic acid in the Electrospray Membrane probe second solution. The Electrospray total ion current of approximately 100 nA was measured at data point  125  on curve  124 . A data point  125 , the Electrospray signal of hexatyrosine was lower with 0.001% TFA added to the sample solution compared with the ESMS signal response with acetic acid added to the ES Membrane probe second solution. Very low concentration benzoic acid was added to the second solution when data point  125  was acquired. Increasing the concentration of benzoic acid in the second solution increased the hexatyrosine signal overcoming the ESMS signal reducing effect of TFA in the sample solution. Even with 0.001% TFA added to the sample solution, the addition of new electrolyte benzoic acid to the second solution of an ES Membrane probe increases the hexatyrosine ESMS signal to a maximum of over two times the maximum hexatyrosine ESMS signal achieved with acetic acid added to the second solution 
         [0043]      FIG. 11  shows negative ion polarity ESMS signal response curves for 1 μM hexatyrosine sample in 1:1 methanol:water solutions run using an Electrospray membrane probe. Curve  127  was acquired while running a concentration gradient of acetic acid in the second solution. Signal response curve  128  was acquired while running a concentration gradient of benzoic acid in the second solution of Electrospray Membrane probe  30 . The maximum negative ion polarity hexatyrosine ESMS signal acquired with new electrolyte benzoic acid was over two times the maximum ESMS signal achieved with conventional electrolyte acetic acid. 
         [0044]    1 μM reserpine sample in 1:1 methanol:water solutions were Electrosprayed to generate the ESMS signal response curves shown in  FIG. 12  New electrolytes benzoic acid and trimethyl acetic acid and conventional electrolyte acetic acid were added at different concentrations to different sample solutions to compare ESMS signal response. As shown by reserpine ESMS signal response curves  127 ,  128  and  129 , a two times signal increase can be achieve when new electrolyte species benzoic acid and trimethyl acetic acid are added to the sample solution compared to the ES MS signal achieved by Electrospraying with conventional electrolyte acetic acid added to the sample solution. 
         [0045]    A comparison of ESMS signal response for 1 μM leucine enkephalin sample in 1:1 methanol:water solutions using four electrolytes added to the sample solution is shown in  FIG. 13 . New electrolytes, benzoic acid, trimethyl acetic acid and cyclohexane carboxylic acid and conventional electrolyte acetic acid were added at different concentrations to different leucine enkephalin sample solutions to generate ESMS signal response curves  130 ,  131 ,  132  and  133  respectively. When running the new electrolytes, a maximum leucine enkephalin signal response increase of two times was achieved compared with the maximum signal response achieved with electrolyte acetic acid. Individually, a factor of three increase in leucine enkephalin ESMS maximum signal response was achieved by adding benzoic acid to the sample solution. 
         [0046]    A characteristic of the new electrolytes is the presence of an (M+H) +  electrolyte parent ion peak ion in the ESMS spectrum acquired in positive ion polarity Electrospray as shown in  FIGS. 14A ,  15 A and  16 A for benzoic acid, trimethyl acetic acid and cyclohexanecarboxylic acid respectively Such a parent positive ion is not generally observed when running conventional electrolytes in Electrospray ionization. As expected, the presence of an (M−H) −  electrolyte species peak was observed in the ESMS spectrum acquired in negative ion polarity mode as shown in  FIGS. 14B ,  15 B and  16 B The presence of gas phase electrolyte parent ions present in positive ion polarity Electrospray may play a role in increasing the ESMS analyte signal. 
         [0047]    The use of new electrolytes benzoic acid, trimethyl acetic acid and cyclohexanecarboxylic acid increases ESMS signal amplitude for samples run in positive or negative ion polarity Electrospray ionization. An increase in Electrospray MS analyte signal can be achieved by adding a new electrolyte directly to the sample solution or by running a new electrolyte in the second solution of an Electrospray Membrane probe during Electrospray ionization. Having described this invention with respect to specific embodiments, it is to be understood that the description is not meant as a limitation since further modifications and variations may be apparent or may suggest themselves. It is intended that the present application cover all such modifications and variations.