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Liquid Chromatography-Mass Spectrometry (LC-MS)
Standard Operating Procedure
INSTRUMENT:
Perkin Elmer Series 200 Liquid Chromatograph
Perkin Elmer Series 200 Pump
Perkin Elmer Series 200 Column Oven
Perkin Elmer API 150EX Mass Spectrometer
INSTRUMENT START-UP:
A. Degassing Solvents
1.
2.
3.
4.
5.
Ensure there are enough degassed solvents before running a sample
Get a 500 mL or 1000 mL Erlenmeyer filter flask
Place your solvent and a stir bar into the flask and sit it on a hot plate
Hook up a vacuum line and stopper the top of the flask
Turn on the vacuum and the stir plate
a. DO NOT HEAT
6. The solvent is degassed when bubbles no longer appear
7. Place parafilm over the filtering arm and parafilm the top of the flask after
you put the tubing in.
A. The instrument should always be on, if not see alternative start up procedure:
B. Open the nitrogen valve on the bench top
C. Open the nitrogen valve in the gas storage room
D. Open the nitrogen tank
1. Make sure the regulator pressure is set to 40 psi
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E. Prime the pump
1. Open the panel on the LC-MS Pump
2. Open the black pump valve (turn counter-clockwise)
Valve
3. On the LC-MS Pump screen press Purge
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4.
5.
6.
7.
8.
9.
Press F3 and enter a flow rate of 1.00
Press Enter
Allow the system to purge for 30 seconds
Press Stop
Close the black pump valve (turn clock-wise)
Close the panel door
A. Open Analyst Software 1.3.2
B. Click on Hardware Configuration
C. Click LCMS then Activate Profile and close the dialogue box
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D. On the lower right hand corner, double click on the first icon
1. This will bring up a dialogue box giving you a detailed status of the
instrument
10. When the instrument is ready to use the icons will turn green. If the
icons remain yellow, you may run the experiment.
11. The instrument will need to pump down to a vacuum. Approximately less
than 5.0x10-5 Torr (This will take some time)
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--------------------------------------------------------------------------------------------------------------------SAMPLE PREPARATION
A.
B.
C.
D.
Place each sample into an LC-MS Sample vial located in cabinet #
Include a sample vial filled with DI water
Label the vials
Load the samples into the autosampler
--------------------------------------------------------------------------------------------------------------------BUILDING A BATCH:
A. Select Build Acquisition Batch from the menu located on the left side of the
screen
B. Sample Tab
1. Type in a Set Name
2. Click Add Set
3. Choose a method from the acquisition list drop down menu
a. For Instrumental Analytical Chemistry use the method caffeine DInj
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4. Click method editor to create or edit a method
C. Adding Samples
1. Click add samples
2. Sample Name
a. Type a prefix in the prefix box and it will be attached to all sample
names
b. If the sample number box is checked, the program will add numbers
at the end of your sample name
c. The number of digits will be the number of digits at the end of your
sample name
3. Data File
a. Type a prefix in the prefix box and it will be attached to all data file
names
b. Clicking Set Name will add the set name to the data file name
c. Sub Folder can be added to store your data
4. New Samples
a. Type the number of samples you want to add
5. Click OK
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6. Under vial position, enter the numerical positions of your samples
D. Submit Tab
1. Make sure the information is correct
2. Click Submit
--------------------------------------------------------------------------------------------------------------------RUNNING AN EXPERIMENT:
A. From the View menu select Sample Queue
B. From the Acquire menu select Ready
C. From the Acquire menu select Start Sample
--------------------------------------------------------------------------------------------------------------------DATA COLLECTION:
A. From the File menu select Open Data File, select your Set Name and click OK
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B. Right click on the screen and select List Data
C. Select the Peak List Tab on lower window
D. Right Click in the lower window and select TurboChrom Parameters
E. Select Bunching Factor
1. Bunching Factor – The number of data points that are grouped to form
one bunch point.
2. Ideally this value should be set so there are approximately 20 data
points for each peak from start to end
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--------------------------------------------------------------------------------------------------------------------INSTRUMENT SHUT-DOWN:
A.
B.
C.
D.
E.
F.
G.
H.
I.
J.
Remove vials from autosampler and dispose of solutions
Select Hardware Configuration → LCMS→ Deactivate Profile
Close the Analyst Software Program
Close the nitrogen valve on the bench top
Close the nitrogen valve in the gas storage room
Close the nitrogen cylinder in the gas storage room
Check the waste; if full, change the waste container. As a lab assistant if
you need help
DO NOT TURN OFF THE ROUGHING PUMPS
DO NOT TURN OFF THE INSTRUMENT
DO NOT TURN OFF THE COMPUTER
--------------------------------------------------------------------------------------------------------------------WASTE CONSIDERATIONS FOR INSTRUMENTAL CHEMISTRY COURSE:
A. Samples and Standards may be disposed of in the sink
--------------------------------------------------------------------------------------------------------------------ALTERNATE INSTRUMENT START-UP:
A.
B.
C.
D.
E.
Open the nitrogen cylinder in the gas storage room
Open the nitrogen valve in the gas storage room
Open the nitrogen valve on the bench top
Turn on the Roughing Pumps
Turn on the Mass Spectrometer
1. A large black switch on the back right hand side of the instrument
F. The instrument will need to pump down to a vacuum
1. Approximately less than 5.0x10-5 Torr
2. This will take time
G. Follow steps starting on page 2
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