This work describes the application of the Agave salmiana cuticle as a new protonic exchange biological membrane (0.080 ± 0.001 mm thickness). Different chemical, electrochemical and mechanical treatments were evaluated to stimulate the ionic exchange properties of the cuticle. Thermal treatment was adequate for its application in a two-chamber electrolyzer. Under optimal conditions an ionic conductivity value of 10 ± 3 mS cm−1 was obtained; this value is similar to the value achieved using a Nafion membrane. The thermally-activated bio-membrane was also evaluated in a fuel cell, where the highest potential was obtained using methanol and hydrogen (0.46 ± 0.01 V). This result makes the Agave salmiana cuticle a competitive choice to replace the commercial membrane. Its surface morphology and their functional groups were evaluated through scanning electron microscopy (SEM), infrared spectroscopy and impedance spectroscopy. This thermally-treated Agave salmiana cuticle is an ecofriendly alternative to replace Nafion membranes in electrolyzer and fuel cells.