In this study, isolation of compounds of low concentration from large volumes of weak solvent was effectively accomplished with simple HPLC method and plumbing modifications. Focusing the gradient improved the quality of the target compound isolation by increasing the resolution between contaminant peaks and permitting increased loading on the column. Whether using a larger loop and syringe or adding a tee and a loading pump to the HPLC configuration for sample introduction, the chromatography was remarkably consistent and isolated products had comparable purities.
Detection plays an important role in the success of product isolation. While large sample volumes very rapidly distort UV chromatograms and make product peak identification ambiguous, mass-directed purification clearly reveals where the target compound elutes and makes fraction collection easier. As the amount of sample volume increases, the efficiency of the overall compound isolation improves by reducing the solvent usage and time for completingthe purification process.