Optimization of Enzyme-Assisted Thermoultrasound for Protein Extraction and Hydrolysis of Mexican Caviar (Ahuautle)

Introduction: Mexican caviar (ahuautle), the eggs of aquatic insects traditionally consumed in central Mexico, represents an underutilized sustainable protein source. Combining enzymatic hydrolysis with thermoultrasound, a green emerging technology, offers a promising strategy to enhance protein extraction efficiency and functionality for functional food applications.

Methods: A Box-Behnken design with two replicates was employed to optimize enzyme-assisted thermoultrasound parameters (pH: 7–9, time: 10–20 min, enzyme concentration/E/S ratio: 1.5–2.5% w/v) for maximizing protein solubility and free amino group release from ahuautle protein concentrate. Response surface methodology (RSM) was applied using Statgraphics software.

Results and Discussion: Response surface analysis revealed that optimal conditions differed between responses. Maximum protein solubility was achieved at alkaline pH (9.0) with 20 min thermoultrasound treatment and enzyme concentration of 2.3% w/v. Maximum free amino group release occurred at pH 8.0, 16 min, and E/S ratio of 2.3% w/v. The model showed adequate fit with R² = 87.22%, confirming that the selected variables effectively explain the variability in protein extraction and hydrolysis responses. These divergent optima demonstrate that protein extraction and controlled hydrolysis require distinct processing parameters.

Conclusions: Enzyme-assisted thermoultrasound successfully optimized both protein extraction and hydrolysis of ahuautle, confirming its potential as a sustainable technology for producing functional protein ingredients or bioactive peptide precursors for novel food applications.