If we want to isolate impurity, in case preparative HPLC is not available whether impurity can be isolated using analytical column ( 250 mm* 4.6, 5 micron, 20 microlitre injector valve)for characterisation by Mass, NMR and IR.
We use analytical scale HPLC columns to collect larger quantities of sample for analysis via other techniques (i.e. for identification).
Summary: To do so successfully, start by developing the BEST LC method you can which shows a lot of resolution between the peak of interest and everything else. This must be the focus of the method. The better the job you do in developing the method, the more sample you will be able to collect each run. Next, if enough sample is available, do a loading study to find out how much sample can be loaded on to your column. Maximize sample concentration over sample volume (large volumes compromise separations). Run the method with the large sample, collect the fraction(s), then analyze them for purity (to make sure you collected the desired pure peak). Run the method multiple times and collect the sample using a very short, narrow ID capillary exiting the detector.
Hi Charu, in your case you can use an analytical fraction collector (e.g: https://www.agilent.com/en/products/liquid-chromatography/infinitylab-lc-series-modules/lc-fraction-collectors/1260-infinity-analytical-scale-fraction-collector).
Hello Pandya
you can use the analytical column ( 250 mm* 4.6, 5 micron, 20 microlitre injector valve) but the thing is your developed method should be perfect. another source i want to suggest is by devloping a good method on HPTLC Metal plate where you can cut the desired band, collect in conical flask, by dissolving it to appropriate solvent, filter it and evaporate to dryness after that you can use it for characterisation by Mass, NMR and IR .
If impurities are at trace level they are difficult to isolate without prep HPLC. If impurity % is at higher, optional technique your can have is Flash chromatography or HPTLC as said by Mr Sharad in above comment.
- Badges
- Science topic
- Similar topics
- Analytical Chemistry
- Column