1. Introduction

2. Maintenance Objectives and Scope

2.1 Maintenance Objectives

• Ensure the precision and accuracy of HPLC analytical results, with relative standard deviation (RSD) of peak area and retention time meeting analytical requirements (usually ≤ 2%).
• Maintain stable operation of the instrument, avoid abnormal phenomena such as pressure fluctuations, baseline drift, peak distortion, and leakage.
• Extend the service life of key components, including the pump head, check valve, injector rotor, chromatographic column, detector lamp, and flow cell.
• Minimize the risk of instrument failures, reduce maintenance costs and the impact of unplanned downtime on experimental work.
• Keep the instrument in compliance with relevant industry standards and laboratory quality management requirements (such as ISO 17025).

2.2 Maintenance Scope

3. Maintenance Cycle and Classification

3.1 Daily Maintenance (Per Operation Day)

3.2 Weekly Maintenance (Every 7 Days)

3.3 Monthly Maintenance (Every 30 Days)

3.4 Quarterly Maintenance (Every 90 Days)

4. Practical Maintenance Tips for Key Components

4.1 Mobile Phase System Maintenance

1. Mobile Phase Preparation: Use high-purity solvents (HPLC grade) and ultrapure water (resistance ≥ 18.2 MΩ·cm) to prepare the mobile phase. Avoid using expired or contaminated solvents. For buffer solutions (such as phosphate buffer), prepare them fresh daily to prevent microbial growth and precipitation. When preparing mixed mobile phases, mix them in the correct proportion and stir evenly to avoid concentration gradients.
2. Degassing Treatment: Dissolved air in the mobile phase will cause baseline noise, peak splitting, and pressure fluctuations. Before use, degas the mobile phase by ultrasonic degassing (15-20 minutes), helium sparging, or online degasser. Ensure the degassing is sufficient; for easily volatile solvents, reduce the ultrasonic time to avoid solvent loss.
3. Storage and Replacement: Store the prepared mobile phase in a clean, sealed container, and label it with the preparation date, composition, and concentration. Avoid storing buffer solutions in direct sunlight or high-temperature environments. Replace the mobile phase every 24-48 hours; if precipitation or turbidity is found, replace it immediately. After the experiment, flush the solvent lines with pure solvent (such as methanol or acetonitrile) to prevent buffer crystallization and blockage.
4. Solvent Bottle Maintenance: Clean the solvent bottle and cap weekly with ultrapure water and methanol alternately, and dry them thoroughly. Replace the solvent filter (inlet filter) every month or when blockage is found (indicated by increased pump pressure). Soak the filter in dilute nitric acid (5%) for 30 minutes and rinse with ultrapure water if it is slightly clogged.

4.2 Solvent Delivery System (Pump) Maintenance

1. Daily Inspection and Flushing: Before starting the pump, check the mobile phase level, ensure the solvent lines are free of air bubbles, and prime the pump to remove air in the pump head. After the experiment, flush the pump with pure solvent (methanol or acetonitrile) at a low flow rate (0.5-1 mL/min) for 15-20 minutes to remove residual buffer or sample components, preventing corrosion and blockage of the pump head and check valve.
2. Pressure Monitoring: Monitor the pump pressure during operation. Record the normal operating pressure; if the pressure increases by more than 10% or fluctuates significantly, check for blockage in the solvent lines, filter, or column. If the pressure is too low, check for leaks in the solvent lines or pump head.
3. Check Valve Maintenance: The check valve is prone to contamination by residual samples or buffer precipitates, leading to unstable flow rate or pressure. Clean the check valve monthly: disassemble it, soak it in methanol or acetonitrile for 30 minutes, rinse with ultrapure water, and reassemble. If the check valve is worn or damaged (indicated by inability to maintain pressure), replace it in time.
4. Pump Head Maintenance: Inspect the pump head seal every quarter. If there is solvent leakage around the pump head, replace the seal gasket. Lubricate the pump piston with the specified lubricant every 6 months to reduce wear. Avoid using corrosive solvents for a long time to prevent damage to the pump head.

4.3 Sample Introduction System (Injector) Maintenance

1. Sample Preparation: Ensure the sample is fully dissolved and filtered through a 0.22 μm or 0.45 μm filter (consistent with the column particle size) to remove insoluble impurities, preventing blockage of the injector and column. Avoid introducing samples with high viscosity or suspended solids.
2. Injector Flushing: After each sample injection, flush the injector with the mobile phase or a suitable solvent (compatible with the sample and mobile phase) 3-5 times to remove residual sample, preventing cross-contamination. After the experiment, flush the injector with pure methanol or acetonitrile for 10-15 times to clean thoroughly.
3. Rotor and Needle Maintenance: Check the injector rotor monthly for wear or contamination. If the injection precision decreases or there is leakage, replace the rotor. Clean the injection needle regularly with a needle cleaner or ultrasonic cleaner; if the needle is bent or blocked, replace it immediately. Avoid touching the needle tip with hard objects to prevent damage.
4. Seal Maintenance: Inspect the injector seal every month. If there is solvent leakage during injection, replace the seal. Store the seals in a clean, dry place to avoid contamination and aging.

4.4 Separation System (Chromatographic Column) Maintenance

1. Column Activation and Conditioning: A new chromatographic column should be activated before use. For reversed-phase columns, flush with methanol or acetonitrile at a low flow rate (0.5 mL/min) for 30-60 minutes, then equilibrate with the mobile phase for 20-30 minutes until the baseline is stable. For normal-phase columns, use n-hexane or isopropanol for activation.
2. Sample and Mobile Phase Protection: As mentioned earlier, ensure the sample is filtered and the mobile phase is pure and degassed to avoid contamination and blockage of the column. Avoid using mobile phases with extreme pH values (usually reversed-phase columns: pH 2-8; normal-phase columns: pH 3-7) to prevent damage to the stationary phase.
3. Column Flushing: After each experiment, flush the column with a solvent compatible with the mobile phase to remove residual sample and mobile phase components. For reversed-phase columns, if the mobile phase contains buffer, first flush with 50% methanol-water (or acetonitrile-water) for 20-30 minutes, then flush with pure methanol or acetonitrile for 30 minutes. For normal-phase columns, flush with n-hexane or isopropanol.
4. Storage: When the column is not in use for a long time, store it in a suitable storage solvent. Reversed-phase columns are stored in pure methanol or acetonitrile; normal-phase columns are stored in n-hexane. Seal both ends of the column to prevent the stationary phase from drying out. Store the column in a cool, dry place, avoiding direct sunlight and high temperature.
5. Regular Inspection: Check the column pressure and separation effect monthly. If the column pressure increases significantly, the peak shape becomes distorted, or the resolution decreases, the column may be clogged or the stationary phase may be damaged. For slightly clogged columns, flush with a suitable solvent (such as THF for reversed-phase columns) at a low flow rate to remove impurities. If the performance cannot be restored, replace the column.

4.5 Detection System Maintenance

1. UV-Vis Detector: Check the lamp intensity daily; if the intensity is lower than 50% of the initial value, replace the lamp. Clean the flow cell weekly: flush with pure methanol or acetonitrile, then with ultrapure water. If the flow cell is contaminated, soak it in dilute nitric acid (5%) for 30 minutes, rinse with ultrapure water, and dry. Calibrate the wavelength accuracy monthly using a standard reference material (such as holmium oxide). Keep the detector cell compartment clean and dry to avoid mold growth.
2. Fluorescence Detector: Replace the lamp every 1000 hours of use or when the signal intensity decreases significantly. Clean the flow cell and excitation/emission slits monthly to remove contamination. Avoid exposing the detector to strong light when not in use to protect the photomultiplier tube. Calibrate the sensitivity regularly using a standard fluorescent substance (such as quinine sulfate).
3. Refractive Index Detector: Keep the detector at a constant temperature (usually 30℃) to ensure stable refractive index measurement. Clean the flow cell weekly with pure solvent and ultrapure water. Avoid sudden changes in mobile phase composition to prevent damage to the detector.

4.6 Data Acquisition and Processing System Maintenance

1. Software Maintenance: Update the data processing software regularly to fix bugs and improve performance. Back up the analytical methods, data files, and system parameters daily to prevent data loss. Avoid installing unrelated software on the data system computer to prevent system crashes or conflicts.
2. Hardware Maintenance: Check the connection between the data system and the detector/pump monthly to ensure stable signal transmission. Clean the computer fan and interface regularly to prevent dust accumulation and poor heat dissipation. Replace the computer hard drive every 2-3 years to avoid data loss due to hard drive failure.

5. Maintenance Notes and Safety Precautions

5.1 Maintenance Notes

• All maintenance work must be carried out in accordance with the maintenance cycle and tips specified in this document. Record the maintenance time, content, replaced components, and instrument performance after maintenance in detail to form a maintenance record for future inspection and tracking.
• Use original or compatible spare parts (such as seals, filters, lamps, and columns) to ensure the performance and compatibility of the instrument. Do not use inferior spare parts to avoid instrument damage or performance degradation.
• When disassembling and assembling components (such as check valves, injectors, and flow cells), use professional tools and operate gently to avoid damaging precision components. Follow the instrument manual to ensure correct disassembly and assembly.
• After completing maintenance, conduct a performance test (such as measuring the baseline stability, peak reproducibility, and column efficiency) to ensure the instrument operates normally before conducting analytical experiments.
• Regularly review and update the maintenance plan according to the instrument operation status, usage frequency, and experimental environment to improve the pertinence and effectiveness of maintenance.

5.2 Safety Precautions

• Before carrying out any maintenance work, shut down the instrument, cut off the power supply and mobile phase supply, and release the pressure in the system to prevent solvent leakage, pressure shock, or electrical shock accidents.
• When using organic solvents (such as methanol, acetonitrile, and THF) during maintenance, operate in a well-ventilated fume hood. Wear personal protective equipment (PPE) such as safety gloves, safety glasses, and anti-static clothing to avoid skin contact and inhalation of harmful vapors. Keep the solvents away from open flames and high temperatures to prevent fire and explosion.
• When handling corrosive substances (such as dilute nitric acid and buffer solutions with extreme pH values), wear acid-base resistant gloves and goggles. Avoid splashing the substances on the instrument or skin; if splashed, rinse immediately with plenty of water.
• Do not disassemble the instrument components randomly without professional training to avoid damage to the instrument or personal injury. If a major fault occurs, contact professional maintenance personnel for handling.
• Dispose of waste solvents, filters, and other maintenance wastes in accordance with laboratory waste disposal regulations, and do not discharge them randomly to avoid environmental pollution.

6. Maintenance Record and Filing

7. Conclusion