High performance liquid chromatography (HPLC) is a commonly used separation and analysis technology, which has the advantages of high separation efficiency, high sensitivity and good selectivity. It uses a high-pressure pump to pump the liquid mobile phase into the system, and realizes the separation of the mixture through the interaction between the stationary phase and the mobile phase. High performance liquid chromatography is widely used in chemistry, biology, medicine and other fields, which can be used to analyze components in complex mixtures, detect trace substances, and prepare and separate compounds.
The separation principle of HPLC is based on the distribution balance of different components between stationary phase and mobile phase. By adjusting the composition and flow rate of mobile phase, as well as the type and particle size of stationary phase, different components can be effectively separated. HPLC is usually equipped with highly sensitive detectors, such as ultraviolet-visible light detectors and fluorescence detectors, to detect the concentration of target components.
Attention should be paid to the following points when using HPLC:
1. mobile phase: select a suitable mobile phase to ensure compatibility with the substance to be tested and avoid bubbles or precipitation.
2. Sample treatment: ensure the purity of samples and avoid pollution and cross-contamination. Filter or centrifuge if necessary.
3. Temperature control: ensure the temperature stability of chromatographic column and detector to obtain the best separation effect.
4. Pressure monitoring: Pay close attention to the system pressure, and check it in time if it is abnormal.
5. Regular maintenance: Conduct regular maintenance on key components such as chromatographic columns and detectors to prolong their service life.
6. Data interpretation: accurately identify the peak value in combination with the standard to avoid misjudgment.
7. Safe operation: Understand the nature of chemicals used, follow safety guidelines, and prevent leakage and accidents.
8. Training and experience: Operators need to receive training and accumulate experience to deal with possible problems.
9. Instrument calibration: Ensure the accuracy and stability of the instrument and follow the manufacturer's calibration recommendations.