Hey everyone!

I'm doing peptide synthesis on Rink amide resin and sometimes I need to add amino acids with secondary amines or beta amino acids. Normally I synthesize 35 to 40 mer peptides. As a simple test (recommended by people who do SPPS), I add a few Fmoc deprotected beads to ONLY the ethanolic solution of ninhydrin (and not the complete ninhydrin reagent kit which includes KCN and phenol and pyridine and so on) and heat to approximately 95 degree C. Usually, after 20% piperidine treatment, an intense blue coloration ON the beads appear and we consider that as a positive result (i.e. free amine is present and ready for the next coupling). (Since we take very few beads, a purple solution is usually not seen, at least not in the first 10 or 20 minutes). For some secondary amino acids and beta amino acids, I see orange or brown coloration ON the beads.

Sometimes, we do not see any color at all and this mostly happens after we reach 20+ amino acids and this puts us in the blind. After that, we're unsure whether amino acids are coupled or not and whether the SPPS is proceeding.

Is there a way to know for certain that ninhydrin can be made reliable?

(I know maldi analysis is the go-to technique but its not cost effective)

If you guys could give some suggestions, that'd be great!