The primary goal of short-term shutdown (e.g., weekends) is to prevent the crystallization of residues and blockage of tubing, while ensuring system stability for a quick restart after the holiday.
Remove harmful mobile phases: If the system contains buffer salts, flush thoroughly with pure water (or 5% methanol in water) to prevent salt crystallization. If it contains solvents prone to decomposition, such as chloroform, also flush them out completely.
Tubing replacement: After cleaning, flush the entire system (including pump, column, and detector cell) with 60/40 methanol/water for approximately 30 minutes to ensure the tubing is stored in a solvent compatible with the column.
After flushing, turn off the power to the pump, detector, autosampler, and column oven sequentially. Close the chromatography workstation and computer. Cover with a dust cover, and turn off the voltage regulator and the main laboratory power supply.
Long-term shutdown (e.g., Spring Festival or National Day holidays) requires more thorough maintenance to prevent microbial growth in tubing, component damage, and system performance degradation.
This is the most critical step in the entire process. The cleaning protocol depends on the type of mobile phase last used.
|
Mobile Phase Type |
Cleaning Procedure |
|
Pure organic phase (e.g., methanol, acetonitrile) |
Flush the system directly with 100% methanol for 40–60 minutes. |
|
Buffer salts (e.g., phosphates, ammonium acetate) |
Step 1: Flush with 95% water + 5% methanol for 80–100 minutes to completely remove salts. Step 2: Flush with 100% methanol for 40 minutes. |
Important Notes:
When cleaning the column, it is not recommended to use 100% pure water; adding 5% methanol prevents the stationary phase from "collapsing."
The same principle applies to manual injection valves. Ensure cleaning solution passes through both the "Load" and "Inject" positions, and never leave the valve in the intermediate position.
Final Principle: Ensure all tubing is filled with and stored in 100% methanol. For quaternary pumps, all channel tubing should be stored in organic solvent to prevent microbial growth and blockage.
After cleaning, stop the pump in the workstation. Turn off the detector, pump, and column oven power sequentially. Close the workstation and computer, and cover with a dust cover. Turn off the voltage regulator and unplug the instrument power cord to ensure complete power disconnection and prevent fans in some modules from continuing to run. If the column needs to be removed, tightly seal both ends with plugs to prevent the bed from drying out. Back up important data in the workstation in advance. Check and empty the waste bottle. Close laboratory doors, windows, water, electricity, and gas supplies.
Proper post-holiday startup directly affects HPLC analytical precision and is a key concern for analysts returning to work.
1) Pre-Startup Inspection: Eliminate Appearance and Connection Issues
First, check that the instrument appears intact and all tubing connections are secure. If the column was stored long-term, reinstall it, ensuring the fittings are sealed without leaks. Verify that laboratory temperature and humidity meet instrument operating requirements to avoid startup instability caused by substandard environmental conditions.
2) Gradual Startup and System Flushing: Prevent Component Damage
Power on in the following order: power supply → column oven → pump → detector. Gradually increase the pump flow rate from low to high (initial flow rate recommended at 0.2 mL/min), flushing tubing and the column with appropriate solvents to remove air bubbles. If using buffer salts in the mobile phase, first flush with 5%–10% methanol/acetonitrile in water before transitioning to the target mobile phase to prevent salt precipitation.
3) Performance Verification: Ensure Data Accuracy
After the system has stabilized, inject a standard sample and verify that retention time, peak area, resolution, and other parameters are consistent with pre-holiday values. Only after confirming no performance anomalies should formal experiments begin. If issues such as peak distortion or abnormal pressure appear, promptly troubleshoot for blocked tubing or column contamination.
Environmental Control: For instruments not used over extended periods, power them on regularly (e.g., 1–2 times per week) to prevent moisture buildup and maintain operational readiness.
Personnel Management: If instrument use is required during holidays, prior application and approval by management are necessary.
Safety Awareness: Before leaving the laboratory, confirm that all water, electricity, and gas supplies are turned off, and doors and windows are securely locked.
Q1: Can the HPLC be left on standby during holidays?
A: Long-term standby is not recommended. Extended standby not only consumes electricity but may also cause tubing blockages or component damage due to pressure fluctuations or mobile phase degradation. For short holidays (up to 3 days), the pump can be kept at a low flow rate to maintain low pressure, provided the mobile phase is stable. For holidays exceeding 3 days, a full shutdown with proper maintenance is recommended.
Q2: What if we forgot to flush the column before shutdown? How should we handle it after the holiday?
A: Upon post‑holiday startup, first flush the column at low flow rate with a compatible solvent (e.g., methanol for reversed‑phase columns) while monitoring pressure changes. If pressure is too high, gradually increase the flow rate or switch to a different flushing solvent. If pressure does not recover, column regeneration or replacement may be necessary.
Q3: What if there is a power outage during the holiday?
A: Prepare UPS units in advance to provide emergency power for critical equipment. After a power outage, once power is restored, follow the post‑holiday startup procedure. Focus on checking tubing for leaks caused by sudden pressure changes and ensure the column has not been affected by moisture.
Q4: After installing a long‑stored column post‑holiday, the pressure is abnormally high. How can this be resolved?
A: First, check for air bubbles or blockages in the tubing connections. Then flush the column at low flow rate with the storage solvent for 30 minutes. If pressure remains high, the column may be back‑flushed (if supported by the column manufacturer) or the guard column replaced.
