Operation steps of high performance liquid chromatograph

Operation steps of high performance liquid chromatograph:

1). Filter the mobile phase and select different filters according to your needs.

2). Ultrasonic degas the mobile phase after suction filtration for 10-20 minutes.

3). Turn on the HPLC workstation (including computer software and chromatograph), connect the mobile phase pipeline, and connect the detection system.

4). Enter the main menu of the HPLC control interface, click manual to enter the manual menu.

5). It was useless for a while, or changed to a new mobile phase, you need to flush the pump and injection valve first. Rinse the pump directly at the water outlet of the pump with a needle. To flush the injection valve, under the manual menu, first click purge, then click start, and the speed should not exceed 10 ml / min when flushing.

6). To adjust the flow rate and use a new mobile phase for the first time, you can try the pressure first. The greater the flow rate, the greater the pressure, generally not more than 2000. Click injure, select the appropriate flow rate, click on, walk the baseline, and observe the situation of the baseline.

7). Design aliasing methods. Click file, select select users and methods, you can select the existing various aliasing methods. To create a new method, click new method. Select the required accessories, including injection valve, pump, detector, etc., depending on the needs. After selecting, click protocol. A complete sampling method needs to include: a. Steady flow before injection, generally 2-5 minutes; b. Baseline zero; c. Loading-inject conversion of the injection valve; d. Sampling time, which varies with different samples different.

8). Inject and operate after injection. Select the aliasing method and click start. For injection, all samples need to be filtered. After the method is finished, click postrun to record data and mark. After all the samples have been walked, use the above method to walk the baseline for a while to wash off the residue.

9). When shutting down, turn off the computer first, then turn off the liquid chromatography.

10). Fill in the register and sign it by the person in charge.

Precautions:

1). The mobile phase requires chromatographic purity, and 20M deionized water is used for water. The mobile phase after degassing should be vibrated carefully so as not to cause bubbles.

2). The column is very fragile, the first time you do it, don't let liquid pass through the column.

3). All liquid passing through the column must be strictly filtered.

4). The pressure cannot be too great, it is best not to exceed 2000 psi

Maintenance of liquid chromatograph (hplc)

1. The daily operating conditions of HPLC: temperature: 10-30 ℃; relative humidity "80%; preferably constant temperature and constant humidity, away from high electrical interference and high vibration equipment

2. Pump maintenance:

1) Use mobile phase as clean as possible;

2) The sand core filter at the liquid inlet should be cleaned frequently;

3) Prevent precipitation during mobile phase exchange;

4) Avoid blockage or bubbles in the pump;

3. Sampler maintenance:

1) After each analysis, the sample inlet should be washed repeatedly to prevent cross-contamination of the sample;

4. Column maintenance:

1) The column must not collide, bend or vibrate strongly under any circumstances;

2) When the column is connected with the chromatograph, the valve parts or pipelines must be cleaned;

3) Pay attention to the degassing of the mobile phase;

4) Avoid using high viscosity solvents as mobile phase;

5) Purify the injected sample;

6) Strictly control the injection volume;

7) After the end of daily analysis work, the residual sample in the injection valve should be cleaned;

8) After the daily analysis and determination, the column must be cleaned with an appropriate solvent;

9) If the analytical column will not be used for a long time, it should be stored and closed with an appropriate organic solvent;

5. Maintenance of detector (UV):

1) Maintenance of the UV lamp: Before the analysis, when the column is almost balanced, turn on the detector; after the analysis is completed, turn off the detector immediately.

2) The sample cell should be maintained.