Low ion Abundances on samples, normal tune file

We cleaned the ion source, cut the column, changed the inlet from front to back (because the front inlet has some issue). After these maintenances, the tune file looks good, the ion abundance in tune file is similar as before. However, low TIC was observed with 1uL injection of our samples, with about 0.2 relative abundance compared to before. And we tried to inject 5 uL to increase the signal, however, the TIC didn't improve significantly. The results obtained can go forward but we want to know what is the reason here. Does anyone meet similar issue? Thanks.

  • 5 µL of what solvent at what inlet temperature and pressure with what liner installed?  What inlet mode - Split or Splitless?  What are the column dimensions? Configuration? Flow rate?  Check out the GC Calculators available here:   GC Calculators and Method Translator Software | Agilent    Use the Vapor Volume calculator to see if what you are injecting will fit in the liner.  Just injecting more sample does not necessarily mean you will see larger peaks. 

    if the tune is fine, then the MS is fine.  Do you run the samples in Scan or SIM?  What about the detector setting - Absolute, Relative, or Gain?  What is the EMV used in the run?  Are you sure you have pointed to the correct tune file in the method?

    In windows file explorer under the method subdirectory (e.g.  SampleRun.M) is a file   acqmeth.txt  .  That file contains the GC and MS parameters used in your method.   Please upload that file here.

  • Are you sure the inlet parameters were matched correctly from front to back? Has the back inlet ever been used before?

  • Hi Paul, 

    Thanks for the response. We are using Agilent 5183-4702 liner, HP-5ms Ultra Inert Columns (Part Number:19091S-433UI, 30 m, 0.25 mm, 0.25 µm, 7 inch cage).

    We are running in Scan, absolute. I always use autotune, I didn't find a place to choose a tune file in the method setting.

    Please find attached acqmeth, now and before the maintenance

    
                                   INSTRUMENT CONTROL PARAMETERS:    GCMS#1
                                   ----------------------------------------
    
       C:\MSDCHEM\1\METHODS\ZXH-STEROL-M6-MS-1ULIN-SPLITLESS.M
          Fri Jun 02 13:19:43 2023
    
    Control Information
    ------- -----------
    
    Sample Inlet      :  GC
    Injection Source  :  GC ALS
    Mass Spectrometer :  Enabled
    
    Oven
    Equilibration Time                           0.5 min
    Oven Program                                 On
        200 �C for 1 min
        then 4 �C/min to 276 �C for 1 min
        then 2 �C/min to 290 �C for 2 min
    Run Time                                     30 min
    
    
    
    
    
    Front Injector
    Syringe Size                                 10 �L
    Injection Volume                             1 �L
    Solvent A Washes (PreInj)                    5
    Solvent A Washes (PostInj)                   5
    Solvent A Volume                             8 �L
    Solvent B Washes (PreInj)                    0
    Solvent B Washes (PostInj)                   2
    Solvent B Volume                             8 �L
    Sample Washes                                0
    Sample Wash Volume                           8 �L
    Sample Pumps                                 6
    Dwell Time (PreInj)                          0 min
    Dwell Time (PostInj)                         0 min
    Solvent Wash Draw Speed                      300 �L/min
    Solvent Wash Dispense Speed                  6000 �L/min
    Sample Wash Draw Speed                       300 �L/min
    Sample Wash Dispense Speed                   6000 �L/min
    Injection Dispense Speed                     6000 �L/min
    Viscosity Delay                              0 sec
    Sample Depth                                 Disabled
    
    
    
    Front SS Inlet He
    Mode                                         Splitless
    Heater                                       On    300 �C
    Pressure                                     On    16.089 psi
    Total Flow                                   On    54 mL/min
    Septum Purge Flow                            On    3 mL/min
    Gas Saver                                    On    20 mL/min After 2 min
    Purge Flow to Split Vent                     50 mL/min at 2 min
    
    
    Back SS Inlet He
    Mode                                         Split
    Heater                                       On    125 �C
    Pressure                                     On    19.417 psi
    Total Flow                                   On    14 mL/min
    Septum Purge Flow                            On    3 mL/min
    Gas Saver                                    On    20 mL/min After 2 min
    Split Ratio                                  10 :1
    Split Flow                                   10 mL/min
    
    
    Thermal Aux 2 {MSD Transfer Line}
    Heater                                       On
    Temperature Program                          On
        280 �C for 0 min
    Run Time                                     30 min
    
    
    Column #1
    
    350 �C: 30 m x 250 �m x 0.25 �m
    In: Front SS Inlet He
    Out: Vacuum
    
    (Initial)                                    200 �C
    Pressure                                     16.089 psi
    Flow                                         1 mL/min
    Average Velocity                             38.737 cm/sec
    Holdup Time                                  1.2907 min
    Flow Program                                 On
        1 mL/min for 0 min
    Run Time                                     30 min
    
    
    Column #2
    
    350 �C: 30 m x 250 �m x 0.25 �m
    In: Back SS Inlet He
    Out: Front Detector FID
    
    (Initial)                                    200 �C
    Pressure                                     19.417 psi
    Flow                                         1 mL/min
    Average Velocity                             30.944 cm/sec
    Holdup Time                                  1.6158 min
    Flow Program                                 On
        1 mL/min for 0 min
    Run Time                                     30 min
    
    
    
    
    
    Front Detector FID
    Heater                                       Off
    H2 Flow                                      Off
    Air Flow                                     Off
    Makeup Flow                                  On    5 mL/min
    Const Col + Makeup                           Off
    Flame                                        Off
    Electrometer                                 On
    
    
    
    
    
    
    
    
    
    
    
    Signals
     Test Plot                                   Save Off
                                                 50 Hz
     Test Plot                                   Save Off
                                                 50 Hz
     Test Plot                                   Save Off
                                                 50 Hz
     Test Plot                                   Save Off
                                                 50 Hz
    
    
                                    MS ACQUISITION PARAMETERS
    
    
    General Information
    ------- -----------
    
    Tune File                : atune.u
    Acquistion Mode          : Scan
    
    
    MS Information
    -- -----------
    
    Solvent Delay            : 10.00 min
    
    EMV Mode                 : Gain Factor
    Gain Factor              : 1.00
    Resulting EM Voltage     : 1753
    
    [Scan Parameters]
    
    Low Mass                 : 50.0
    High Mass                : 550.0
    Threshold                : 150
    Sample #                 : 2       A/D Samples    4
    Plot 2 low mass          : 50.0
    Plot 2 high mass         : 550.0
    
    [MSZones]
    
    MS Source                : 230 C   maximum 250 C
    MS Quad                  : 150 C   maximum 200 C
    
    
                                 END OF MS ACQUISITION PARAMETERS
    
    
                                  TUNE PARAMETERS for SN: US83140737
                            -----------------------------
    
     Trace Ion Detection is OFF.
    
     EMISSION    :      34.610
     ENERGY      :      69.922
     REPELLER    :      21.914
     IONFOCUS    :      90.157
     ENTRANCE_LE :      16.000
     EMVOLTS     :    1752.941
                                   Actual EMV  :    1752.94
                                   GAIN FACTOR :       1.00
     AMUGAIN     :    1586.000
     AMUOFFSET   :     113.438
     FILAMENT    :       1.000
     DCPOLARITY  :       0.000
     ENTLENSOFFS :      19.325
     MASSGAIN    :    -775.000   
     MASSOFFSET  :     -42.000   
    
                               END OF TUNE PARAMETERS
                          ------------------------------------
    
    
    
                                     END OF INSTRUMENT CONTROL PARAMETERS
                                     ------------------------------------
    
    
    .

    
                                   INSTRUMENT CONTROL PARAMETERS:    GCMS#1
                                   ----------------------------------------
    
       C:\MSDCHEM\1\METHODS\ZXH-STEROL-M10-40MIN-MS-1ULIN-SPLITLESS-BACK.M
          Thu Aug 31 13:15:54 2023
    
    Control Information
    ------- -----------
    
    Sample Inlet      :  GC
    Injection Source  :  GC ALS
    Mass Spectrometer :  Enabled
    
    Oven
    Equilibration Time                           0.5 min
    Oven Program                                 On
        200 �C for 1 min
        then 4 �C/min to 270 �C for 1 min
        then 1 �C/min to 290 �C for 0.5 min
    Run Time                                     40 min
    
    
    
    
    
    Back Injector
    Syringe Size                                 10 �L
    Injection Volume                             1 �L
    Solvent A Washes (PreInj)                    2
    Solvent A Washes (PostInj)                   0
    Solvent A Volume                             5 �L
    Solvent B Washes (PreInj)                    0
    Solvent B Washes (PostInj)                   2
    Solvent B Volume                             5 �L
    Sample Washes                                0
    Sample Wash Volume                           1 �L
    Sample Pumps                                 6
    Dwell Time (PreInj)                          0 min
    Dwell Time (PostInj)                         0 min
    Solvent Wash Draw Speed                      300 �L/min
    Solvent Wash Dispense Speed                  6000 �L/min
    Sample Wash Draw Speed                       300 �L/min
    Sample Wash Dispense Speed                   6000 �L/min
    Injection Dispense Speed                     6000 �L/min
    Viscosity Delay                              0 sec
    Sample Depth                                 Disabled
    
    
    
    Front SS Inlet He
    Mode                                         Splitless
    Heater                                       Off
    Pressure                                     Off
    Total Flow                                   Off
    Septum Purge Flow                            Off
    Gas Saver                                    On    20 mL/min After 2 min
    Purge Flow to Split Vent                     50 mL/min at 2 min
    
    
    Back SS Inlet He
    Mode                                         Splitless
    Heater                                       On    300 �C
    Pressure                                     On    19.417 psi
    Total Flow                                   On    19 mL/min
    Septum Purge Flow                            On    3 mL/min
    Gas Saver                                    On    20 mL/min After 2 min
    Purge Flow to Split Vent                     15 mL/min at 0.75 min
    
    
    Thermal Aux 2 {MSD Transfer Line}
    Heater                                       On
    Temperature Program                          On
        230 �C for 0 min
    Run Time                                     40 min
    
    
    Column #1
    
    350 �C: 30 m x 250 �m x 0.25 �m
    In: Back SS Inlet He
    Out: Vacuum
    
    (Initial)                                    200 �C
    Pressure                                     16.089 psi
    Flow                                         1 mL/min
    Average Velocity                             38.737 cm/sec
    Holdup Time                                  1.2907 min
    Flow Program                                 Off
        1 mL/min for 0 min
    Run Time                                     40 min
    
    
    Column #2
    
    350 �C: 30 m x 250 �m x 0.25 �m
    In: Back SS Inlet He
    Out: Front Detector FID
    
    (Initial)                                    200 �C
    Pressure                                     19.417 psi
    Flow                                         1 mL/min
    Average Velocity                             30.944 cm/sec
    Holdup Time                                  1.6158 min
    Flow Program                                 Off
        1 mL/min for 0 min
    Run Time                                     40 min
    
    
    
    
    
    Front Detector FID
    Heater                                       Off
    H2 Flow                                      Off
    Air Flow                                     Off
    Makeup Flow                                  On    3 mL/min
    Const Col + Makeup                           Off
    Flame                                        Off
    Electrometer                                 On
    
    
    
    
    
    
    
    
    
    
    
    Signals
     Test Plot                                   Save Off
                                                 50 Hz
     Test Plot                                   Save Off
                                                 50 Hz
     Test Plot                                   Save Off
                                                 50 Hz
     Test Plot                                   Save Off
                                                 50 Hz
    
    
                                    MS ACQUISITION PARAMETERS
    
    
    General Information
    ------- -----------
    
    Tune File                : atune.u
    Acquistion Mode          : Scan
    
    
    MS Information
    -- -----------
    
    Solvent Delay            : 10.00 min
    
    EMV Mode                 : Gain Factor
    Gain Factor              : 1.00
    Resulting EM Voltage     : 2165
    
    [Scan Parameters]
    
    Low Mass                 : 50.0
    High Mass                : 550.0
    Threshold                : 150
    Sample #                 : 2       A/D Samples    4
    Plot 2 low mass          : 50.0
    Plot 2 high mass         : 550.0
    
    [MSZones]
    
    MS Source                : 230 C   maximum 250 C
    MS Quad                  : 150 C   maximum 200 C
    
    
                                 END OF MS ACQUISITION PARAMETERS
    
    
                                  TUNE PARAMETERS for SN: US83140737
                            -----------------------------
    
     Trace Ion Detection is OFF.
    
     EMISSION    :      34.610
     ENERGY      :      69.922
     REPELLER    :      33.641
     IONFOCUS    :      90.157
     ENTRANCE_LE :       3.000
     EMVOLTS     :    2188.235
                                   Actual EMV  :    2164.71
                                   GAIN FACTOR :       0.99
     AMUGAIN     :    1604.000
     AMUOFFSET   :     104.938
     FILAMENT    :       1.000
     DCPOLARITY  :       0.000
     ENTLENSOFFS :      19.827
     MASSGAIN    :    -767.000   
     MASSOFFSET  :     -47.000   
    
                               END OF TUNE PARAMETERS
                          ------------------------------------
    
    
    
                                     END OF INSTRUMENT CONTROL PARAMETERS
                                     ------------------------------------
    
    

  • Hey thank you. Yes, both the two inlets are using. We changed the parameters accordingly. Please find the attached files above.

  • Looking at this gave me a headache knowing you swapped inlets but I'm not sure every parameter matches. I'll let Paul take this one, but I noticed your MSD transfer line temps are different. The first one is at 280 (typical) and the 2nd one is 230.

  • I see your EMV went up from1753 to 2165 but you say you lost 80% response, yes something is wrong. Your flows from col #1 don't match the new col #2. In the first one your flow programs are on and now they are off and pressures don't match either.

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