Harnessing Triticale (× Triticosecale Wittmack) Malting for Amylases Enzymes Synthesis and Partial Purification

Abstract

Triticale malt is an underutilized cereal matrix with high amylolytic potential, which may be exploited for the recovery of plant- derived enzymes for food and biotechnological applications. This study aimed to obtain α-amylase- and β-amylase-enriched aqueous extracts from Bicentenario and Siglo XXI triticale malts and to partially purify these enzymes using ethanol/trisodium citrate aqueous two-phase systems (ATPS). Triticale grains were malted by germination for 5days and dried at 50°C. Aqueous extraction was performed using ground malt and deionized water at a 1:10 ratio under diferent extraction times (30, 120, and 270min) and temperatures (30°C and 40°C). The selected extracts were lyophilized and subjected to ATPS composed of ethanol and trisodium citrate. Enzyme activity was determined using the BETAMYL-3 method for β-amylase and the AOAC 2002.01, AACC 22-02.01, and ICC Standard No. 303 methods for α-amylase. Mild extraction conditions favored enzyme recovery, with the highestα-amylase activityobservedin SigloXXIextractat 30min/30°C,increasing from 99.10CU/gin malt to 507.42CU/gin the aqueous extract. β-Amylase activity was also higher in Siglo XXI than in Bicentenario extracts. The 30% ethanol/18% trisodium citrate ATPS was the most efective system, promoting diferential partitioning with α-amylase concentrated in the bottom phase (432.89CU/mL)andβ-amylaseinthetopphase(5.78BU/mL).Theseresultshighlightthepotentialoftriticalemaltasarenewable source of amylolytic enzymes and support ethanol/trisodium citrate ATPS as a mild, sustainable strategy for obtaining amylase- enriched fractions, although further optimization is needed to improve β-amylase recovery and scalability.