Dear CADET Community,
I am currently working on a project that involves performing and analyzing organocatalytic reactions in an autonomous manner. However, I am struggling with optimizing the HPLC analysis, which I would like to be completely independent of human intervention. The HPLC module I am using is rather classical, with a column size of 250 mm x 4.6 mm, and the eluents I am using are a mixture of hexane and isopropanol. Unfortunately, I cannot change the eluents since the other column with chiral filling is integrated into the module. The analytes are small (100-400 u) and, in most cases, non-polar (with an average amount of isopropanol used in isocratic experiments being 2%). My goal is to find the gradient that will allow for the separation of 2-4 analytes. The separation does not have to be very precise; rather, I care about its reliability.
I found CADET to be a promising tool that can help me, but as I am not a specialist in analytical and physical chemistry (I mainly work in the fields of organic chemistry and computer science), I have some questions that I hope you can answer:
- Is it possible to build a model that can predict the retention time of a single analyte under a given gradient?
- If yes, could you please advise me on how many preliminary experiments I would need to conduct to find the parameters for the absorption model? Also, should the data be from isocratic or gradient experiments?
- What absorption model is suitable for such experiments? I have read that the Langmuir isotherm, for example, cannot be used in gradient conditions.
If these questions seem trivial or pointless, I would appreciate it if you could recommend some literature that could be helpful in this task, especially for experiments with classical normal phase HPLC. Most of the literature I have come across has been about ion-exchange chromatography or RP-LC.