 # How is theoretical net peptide content calculated?

Theoretical net peptide content (calculated assuming that counterions are the only non-peptide components present in your peptide sample) can be estimated by dividing molecular weight (MW) of the peptide by a sum of this molecular weight and a number of trifluoroacetate (TFA) or acetate (AcO-) counterions that are required to neutralize the peptide multiplied by the molecular weight of the TFA counterion (MW= 114) and the AcO counterion is (MW= 59).

For example, a synthetic peptide with a TFA salt and a MW= 1000 with a free N-terminal amino group and one Lys has theoretical net peptide content of 1000 / (1000 + (2 x 114)) = 1000/1228 =0.81 or 81%. This example peptide has 2 positions for the TFA salt to bind, hence the 2×114.

Theoretical net peptide content formula: (peptide MW)/(peptide MW + (#bound salts x salt MW))

Counterions are not the only potential non-peptide components in the peptide sample. It can also contain residual water, adsorbed solvents and traces of other substances. As a result, the actual net peptide content is usually determined by either elemental analysis (N2 content) or quantitative amino acid analysis.