Two dimensional preparation of liquid phase

Preparation of liquid phaseThe purpose is to extract and purify the monomers of the product. Compared with traditional purification methods such as distillation and extraction, it is a more effective separation method and is therefore widely used in the extraction and purification of samples and products. With the increasing demand for high-purity components in fields such as synthesis, plant chemistry, biochemistry, and pharmaceuticals, the application of preparative liquid chromatography is also rapidly expanding and developing.

Preparation of liquid phaseSteps for using a chromatograph

1、 Power on

1. Connect the computer and instrument power (before connecting the power, you can first take the suction head into the mobile phase bottle to be used);

2. Turn on the power switch of the instrument, usually from top to bottom, with the detector switch at the bottom;

3. Check if the mobile phase is sufficient and if the solvent amount recorded on the instrument is correct;

4. Turn on the computer;

5. Open the instrument online software to enter the interface, click start to set the pump flow rate to 10, and there should be liquid flowing out of the instrument at this time;

6. Click on balance and then click on each wavelength and adjust;

After a period of time, the sample can be injected once the pressure stabilizes.

2、 Sample injection

1. Twist the sampler to load up and extract a certain amount of the liquid to be separated

Quick injection, what was done was to aspirate 0.2 milliliters;

2. Click the "Run Control" button in the menu bar of the control interface to set the sample information storage path (to be done every time);

3. Twist the sampler to enter the unload state and start timing the retention time;

4. Peak connection;

5. After all peaks have appeared, click on the interrupt button in the menu bar to prepare for repeated injection.

3、 Shut down

1. After clicking the interrupt button, rinse for a few minutes and set the flow rate to 0. After the pressure reaches 0, transfer the suction head to methanol and set the flow rate to 10;

2. It usually takes half an hour to rush until there is nothing left to rush down. At this point, the pressure is usually already stable;

3. Set the flow rate to 0 and click 'Close' on the interface (next to 'Start');

4. Close the window, and click 'OK' on all pop-up windows, such as' OK '.

Daily simple maintenance and upkeep

1. The filtration requirements for mobile phase: chromatography pure reagents, secondary distilled water, and buffer salts must be filtered.

2. Preparation type liquid chromatography suction filter head: In special circumstances, the filter head can be removed to extract and determine whether it is clogged; Alternatively, a syringe can be used to draw in the flowing fluid and filter it out to determine if it is blocked. If there is any blockage, isopropanol ultrasonic cleaning can be used; If the cleaning is unsuccessful, it needs to be replaced.

3. Unidirectional valve: If the pump pressure is unstable or the mobile phase cannot be extracted, it may be a problem with the one-way valve. Remove it and use isopropanol ultrasonic cleaning. When cleaning, place it in a small beaker in reverse according to its installation direction. Remember not to wash it upside down from the installation direction!

4. Pump head: If there is a leakage or other malfunction of the pump head, it is generally necessary to apply for maintenance (such as leakage, or replacement of the plunger rod and its sealing gasket, etc.).

5. Filter cartridge: When abnormal chromatographic peaks occur, it is possible that this component has been contaminated. Remove and use isopropanol to super wash or replace the filter gasket of the preparative liquid chromatograph

6. Drain valve: When there is no sealing or leakage in this area, it is usually due to contamination or wear of the gasket. The gasket can be removed and cleaned with isopropanol ultrasonic or replaced.

7. Manual sampler: Pay attention to cleaning with secondary distilled water and methanol during loading and injection analysis; If there is a leakage phenomenon, the cause is likely to be wear or contamination of the rotor sealing gasket, and generally requires repair or replacement of the preparation type liquid chromatograph accessories.

8. Preparation type liquid chromatography flow cell: When the chromatographic peak is abnormal, it may be due to contamination of this component. After disassembly, remove the gasket and rinse it with isopropanol (which can be operated or repaired according to the instructions).

9. Workstation: In the event of a system crash, the computer can be restarted; When it is not running normally, the first thing to do is to replace the computer and test its hardware failure; Or re plug and unplug the interface or reinstall the software on the local machine.

When conducting analysis, the retention time sometimes drifts and sometimes changes rapidly due to the following reasons:

1. The solution to poor temperature control is to use a constant temperature device to maintain a constant column temperature.

2. The solution to prevent changes in the mobile phase is to prevent evaporation, reactions, etc.

3. The column is not balanced properly and needs to be balanced for a longer period of time.

4. The solution to the change in flow rate is to reset it to maintain stability.

5. There are bubbles in the pump, which can be expelled through exhaust and other operations.

6. If the mobile phase is not suitable, the solution is to change the mobile phase or mix it appropriately in the control room.

Preparation of liquid phaseChromatography is mainly used in the following fields:

1. Natural medicinal chemistry and traditional Chinese medicine chemistry. This is currently a widely used method in China. In order to clarify the trace active ingredients in plants, manual chromatography columns are no longer sufficient. Almost every laboratory engaged in natural medicine has at least one liquid phase, and wealthy laboratories have one liquid phase per person.

2. Protein purification. Compared to the former, protein purification started slightly later, but has developed rapidly. Meanwhile, due to the relatively small advantages of foreign instrument manufacturers in this field, China is also accelerating its development in this area and seizing territory.

3. Purification of drug impurities. Due to the fact that the synthesis process cannot achieve 100% yield and often involves by-products, obtaining trace impurities and determining their structure and content relies on liquid chromatography.

4. Preparation of chiral compounds. This is mainly used in organic synthesis and methodology.