I am newly interested in LC, particularly, preparative reversed phase. Iíve called/written many universitiesí chemistry departments in my state (NC) for assistance, hoping I may locate a student interested in assisting. I know this would likely require compensation and Iím making arrangements for this. I would like to do an isolation of an alkaloid, specifically Mitragynine, from a crude extract of leaf material originating from Mitragyna speciosa. My interest in this substance has peaked due to a lack of knowledge of it and an increase in demand due to new possible uses being investigated worldwide. Iíve been attempting to find the easiest preparatory method for such an extraction through reading from various sources of information related to the subject online. Thus far, I believe a 2D method *(1st dimension being a mixed-mode sample prep) preparatory reversed phase liquid chromatography would be my best bet. Before proceeding to LC, Iím thinking a simple multi-stage acid-base separation inside of a standard separatory funnel to try and remove as many impurities (specifically more acidic compounds) as possible before moving on to the RPLC phase. I figure that I should be aiming somewhere between the pH of Mitragynine (what ever this is) and the pH of the final acid-base extraction. With that said, I have no earthly idea how one would formulate a specific method for isolating my intended compound (Mitragynine) via RPLC.
1) Concerning the mixed-mode sample prep (for pre-RPLC), how do I develop this method?
2) What packing material do I select (or how would I figure this out)?
3) How would I select the most appropriate mobile phase?
4) How exactly would I pack the column to a consistent density?
5) How do I quantify the specific affinity that the Mitragynine would have to the packing material and how do I know this would differ enough from the affinities of the other compounds present after the final acid-base extraction stage?
6) At the RPLC phase, how would I quantify the length of time necessary to see a peak in my target compound (Mitragynine) with/without a detector?
7) Would a detector be detrimental to the entire effort or is an occasional sample tested through TLC paper sufficient?
8) How would I figure the proper operating parameters?
9) Have I even decided on the proper methods?
10) Where could a non-chemist find help, preferably very cheap help, on this subject? Are there not directories/social networks for chemistry students/professionals interested in providing help related to this?