Paired-Fraction Electrochemical Programming of Enzymatic Dipeptide Synthesis in Water

The dipeptide Ac-Phe-Ala-NH2 is proposed as the smallest analytically complete test of paired-fraction electrochemical programming (PFEP) for peptide-bond formation. The reaction uses an activated acyl donor, Ac-Phe-OEt, an amine acceptor, Ala-NH2, and alpha-chymotrypsin or a qualified engineered ligase. Anodic and cathodic liquids are generated outside the reaction vessel, characterized for acid-base capacity, ionic composition, reactive impurities, gases, and state age, and then added as small doses to an electrode-free enzymatic reactor. PFEP is not proposed to make peptide bonds from unactivated free amino acids. Its testable role is to control protonation, enzyme activity, substrate partitioning, and the time-dependent competition between aminolysis and hydrolysis. The central hypothesis is that feedback-controlled fraction dosing can increase integrated dipeptide productivity or shorten exposure to hydrolytic conditions relative to a chemically reconstructed pH and composition trajectory. The hypothesis fails if matched conventional controls reproduce the result or if apparent gain reflects altered extraction, detector response, or enzyme loss. No new experimental yield is reported. Article type: Perspective, mechanistic hypothesis, and experimental framework. No experimental peptide yield, potency, clinical effect, or shelf-life evidence is reported.