What should I do if there is bacterial contamination when preparing the mobile phase in the liquid phase?

The symptom of "blocking" is an abnormal increase in column pressure, and the direct cause is poor flow path. The main location of clogging is at the front end of the chromatographic column. The main reason is that there are impurities in the mobile phase, and the main source of impurities is bacteria.

First of all, we must realize that general domestic methanol does not require additional filtration, and there is no problem in using it directly. Even some solid particulate impurities can be removed on the filter head at the front of the liquid flow path system. What is really likely to cause problems are bacteria in the water.

Newly prepared pure water will grow bacteria if left indoors for a few days. Although these bacteria are invisible to the naked eye, they are enough to clog the gaps in the column packing particles, causing the column to be scrapped quickly. This is the cause of bacterial contamination when preparing mobile phases, and the solution to it is simply to ensure the reliability of the water.

Here are two recommended ways: ??

(1) The most ideal way is of course to purchase a laboratory-specific pure water machine, which is convenient and reliable, and the quality is assured. The only drawback is that it's expensive.

(2) Purchase commercially available brands of purified water in boxes. The quality of this water is sufficient to meet the requirements of liquid chromatography. First randomly select one to do a bacterial plate test, and use it only after the number of colonies is qualified.

Here is a detail: in most books, when it comes to preparing mobile phase, there is a filtration step at the end. But from the actual effect of our long-term use, as long as the quality of the water can be guaranteed, this step can and should be eliminated. There are three reasons:

(1) Mobile phase filtration is beneficial in theory, but in actual operation, since it is impossible to use a dedicated bottle, it is easy to cause cross-contamination, which has a greater impact on mobile phases with complex ratios.

(2) Mobile phase filtration is not cost-effective in terms of economic cost. It costs more than 6,000 yuan to buy a filtering device, and filters are recognized as relatively easily damaged equipment. The most important thing is that the cost of the filter is too high, one piece costs tens of yuan. Calculated based on the normal service life of a general liquid phase column, the cost of the filter will be much higher than the cost of the chromatographic column.

(3) Mobile phase filtration is not cost-effective for work efficiency. Using a solvent filter requires a process of pre-cleaning, equipment, use, cleaning after use, and drying, which should take at least an hour. This cost cannot be ignored.

In addition to bacterial contamination of the mobile phase caused during preparation, there are also contamination caused by improper use and operation, which in turn leads to blockage. The next article will introduce the detection and treatment measures.

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