Enzymatic modification of amadumbe flour for gluten-free applications
Authors:   , Manhivi Vimbainashe Edina 
Published: July 2021

The production of gluten-free bread from gluten-free flours remains a technological challenge. Different strategies have been employed to improve the dough rheological properties. Enzymatic modification of the proteins in dough may result in polymers that mimic gluten. In this study gluten-free amadumbe flour was modified using single and optimised multiple crosslinking enzyme systems for the improvement of rheological properties and bread quality. Specifically, compositional, rheological and thermal properties of amadumbe and cactus mucilages were investigated as potential hydrocolloids and as substrates for crosslinking enzymes in gluten-free bread production. The effects of laccase, tyrosinase and transglutaminase on amadumbe dough rheology were also investigated. Model reactions were used to demonstrate the different enzymatic reactions occurring in amadumbe dough treated with the crosslinking enzymes. The combination of enzymes was then optimised using response surface methodology (RSM), to produce dough with improved G’ and G”. Xanthan gum, amadumbe mucilage or cactus mucilage were then added to the dough with an optimum enzyme combination. The effect of these enzymes and hydrocolloids were studied on the bread properties. The mucilages had a similar composition of monosaccharides and amino acids, except for the absence of rhamnose in amadumbe mucilage. Cactus mucilage showed a pseudoplastic flow behaviour whilst amadumbe mucilage showed a Newtonian flow behaviour at up to 5% (w/v) concentrations. The mucilages contained phenolics and amino acids such as lysine, tyrosine and glutamine, which are potential enzyme substrates. Trametes versicolor laccase catalysed the crosslinking of phenolics and thiols producing a wide range of crosslinking products which included homo- and hetero-conjugates, as demonstrated by mass spectroscopy. Thiol and total phenolic contents of dough decreased by up to 28% and 93%, respectively, as laccase activity was increased (0-3 U/g flour), confirming crosslinking reactions. Laccase-catalysed modification of amadumbe dough increased dough viscoelasticity, as shown by the increase in G’ and G”. Tan δ decreased with increase in laccase activity indicating an increase in the elastic character of the dough. Tyrosinase oxidation resulted in a 7.7 – 39.4% decrease in dough free amine and a 16.8 – 46.3% decrease in the dough thiol content as activity was increased (0-80 U/g flour). Transglutaminase treatment decreased the dough free amino groups by 10 – 38.1% as activity was increased from 0 to 2 U/g flour. An increase in dough G’ and G”, showed that both transglutaminase and tyrosinase improved dough viscoelasticity. Evidence of transglutaminase and tyrosinase crosslinking was provided by relevant model reactions monitored by mass spectrometry. Reaction model data showed the formation of the glutamyl-lysine bond due to transglutaminase crosslinking, whilst tyrosinase crosslinking resulted in disulphide and dityrosine bond formation. The viscosity and elasticity of amadumbe dough containing soy protein were optimised using a central composite design and the enzyme combination resulting in maximum G’ and G”, and minimum Tan δ was selected and verified. The predicted optimal enzyme activities (LAC, 1.78 U/g flour), (TYR, 79 U/g flour) and TG, 1.97 U/g flour) resulted in amadumbe dough that had a higher G’ and G”, as well as bread with a higher specific volume and lower crumb hardness compared to the dough without enzymes or with a single enzyme system. Addition of cactus and or amadumbe mucilage to the dough containing the optimum enzyme combination further improved dough viscoelasticity, improved bread specific volume, and significantly (p < 0.05) reduced bake loss and crumb moisture loss. The better bread was produced from dough with an optimum enzyme combination and 2% cactus mucilage. Sensory evaluation revealed that enzymes and cactus mucilage improved bread texture, appearance and overall acceptability but did not significantly affect bread aroma and taste. Overall, the combined effect of multiple enzyme-catalysed modification of gluten-free amadumbe flour and amendment with hydrocolloid resulted in more acceptable bread quality than single enzyme systems or unmodified flour. Therefore the optimised combination of enzymes have potential for application in gluten-free bread production.