**Liquid Chromatography Operating Procedures**
This document provides a detailed guide for the operation of a liquid chromatography system. It outlines the step-by-step process from startup to shutdown, including data analysis and method setup.
**1. Startup Sequence**
Before beginning, ensure all components are properly connected and powered on in the following order:
- Pump
- Detector
- Computer host
- Monitor
- Printer
Once all devices are powered on, open the Chromatography Workstation software by double-clicking the icon. Enter the password “SAGE†and click OK. Then, select the “Alltech-426†icon to proceed.
**2. Setting Chromatographic Conditions**
- Set the flow rate of the pump using the arrow keys. Once set, press RUN/STOP to start the pump.
- Adjust the detector settings, run a self-test, and set the required wavelength. Press ZER0 to reset the baseline and prepare for analysis.
**3. Preparing the Mobile Phase**
Mix the mobile phase according to the desired volume ratio, filter it, and sonicate for 20 minutes in an ultrasonic cleaner. Place a filter into the mobile phase, open the bypass valve, and press the PURGE button. Use a syringe to draw and flush the mobile phase until no air bubbles remain. Stop the pump after this step.
**4. Injection and Analysis**
On the computer, select the blue “RUN/SINGLE†icon. Rename the file (e.g., SAM01.001) and click OK. Load the sample into the injection valve, press the “ZERO†key on the detector, then switch the valve to INJECT and press the spacebar on the keyboard to start the analysis.
After the chromatogram is collected, the system will stop automatically. Select the “Analyze†icon to calculate the results, then go to “Reports†and choose “Area%†to view the data. To print, select the printer icon, choose “Print Report P†and “Custom C,†then initiate printing.
Repeat the process for each sample, with at least 5 minutes between runs.
**5. Shutdown Procedure**
Before turning off the system, flush the column with mobile phase for 20–30 minutes. If the column will not be used for an extended period, rinse it with acetonitrile for the same duration. Then, turn off the devices in the following order:
- Computer
- Monitor
- Pump
- Detector
Ensure the software is closed before powering down the computer.
**Notes:**
- The flow rate should not exceed 1.5 mL/min under normal conditions.
- The pump will not operate if the mobile phase is depleted.
- Always power off the system only after closing the operating software.
**Method Modification Guide**
If you need to adjust the analysis parameters, follow these steps:
**1. Modify Running Time**
Go to the “Setup†icon, set the sampling frequency to 2, and adjust the running time (e.g., 10 or 20 minutes). Ensure the channel status is ON and the trigger is set to the second option. Save the method by selecting “File†> “Save Method.â€
**2. Adjust Integral Parameters**
If the results are inaccurate, modify the integral settings:
- Adjust Minimun Area to exclude small impurities.
- Set Peak Width to around 2.2 for flat peaks or 0.5 for sharp ones.
- Set Threshold to 150.
Recalculate the results and save the method.
**3. Customize Report Format**
Access “Custom Report†and link the file name, user name, and date. Set scaling to “Autoscale to Largest Peak†and add retention times and peak areas. Define the format for each column (e.g., 2 or 3 decimal places).
**4. Access Previous Methods and Data**
Use the “Method†icon to select a saved method, and the “Data†icon to load previous chromatograms.
**Internal Standard Method**
- Select the method and click OK.
- Enter the standard sample and calculate the spectrum.
- Set up the internal standard peak, concentration, and calibration parameters.
- Recalibrate, record the RF value, and verify the accuracy.
- Run the sample and print the result.
**External Standard Method**
Follow similar steps as the internal standard method, but use external standards instead. Ensure the sample volume is correctly calculated and entered. Print the final result.
**Company Information**
For more information, visit [Shanghai Qite Analytical Instrument Co., Ltd.](http://) or contact via phone/fax.
For news and updates, visit [ChinaWJ.com.cn](http://news.chinawj.com.cn).
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