In contrast to that of starch, the rate of digestion of glycogen by the midgut homogenate was nearly constant over time ( Fig. 7(b). This result is likely a consequence of the higher number of branches composed of α-1,6 glucose residues in the glycogen molecule. An effective digestion of glycogen probably requires the action of a debranching enzyme to hydrolyze the α-1,6-glycosidic linkage at the branch point and release of a linear α-1,4 glucose polymer that could then be hydrolyzed by the α-amylase. A glycogenolytic

system like this was proposed for the bacteria Bacillus subtilis ( Shim et al., 2009). Lumacaftor To search for an enzyme capable of hydrolyzing the α-1,6 linkages present in glycogen, we performed an assay using isomaltose (Glu-α-1,6-Glu) as a substrate at pH 6.5. According to our results, the L. longipalpis larvae were ineffective at hydrolyzing this disaccharide or dextran molecules, a glucose polymer formed by glycosidic α-1,6 linkages with ramifications of the α-1,3-type linkages.

An efficient debranching activity could be detectable only using substrates containing α-1,6-glycosidic residues bound to linear α-1,4 glucose polymers. In nature, this debranching activity may be performed by debranching enzymes such as those produced by some bacteria or plants ( Zhu et al., 1998, Delatte et al., 2006, EPZ015666 supplier Shim et al., 2009 and Bijttebier et al., 2010). How L. longipalpis larvae address branched substrates is a problem to be solved in the future. The final digestion of the oligosaccharides generated by the hydrolysis of starch and glycogen molecules can be attributed to an α-glucosidase. This enzyme predominates in the posterior midgut and is associated with the midgut wall (Fig. 8). More specifically, this enzyme is bound to the microvilli of the enterocytes. From this site, the α-glucosidase can digest products of starch hydrolysis such as maltose, maltotriose and other oligosaccharides with high molecular masses. Adhesion to the midgut wall maintains the enzyme in the appropriate anatomical site despite the counter-flow mechanism, presumably present in most insects, which could be responsible for the Telomerase reutilization of the soluble digestive enzymes such as the α-amylase

and others (Terra and Ferreira, 1994 and Fazito do Vale et al., 2007). The posterior midgut is the correct site for α-glucolytic activity because this enzyme requires a neutral or acidic environment (Fig. 9). In addition, starch and other polysaccharides must first be pre-digested in the anterior midgut to generate the substrates to be digested by the α-glucosidase in the posterior midgut. Recently, Moraes et al. (2012) reported the presence of two peaks of α-glucosidase activity in L. longipalpis larval midgut by using gel filtration chromatography. One of these peaks was eluted as an enzyme of 66 kDa, a molecular mass similar of that found in the present work ( Fig. 4(b). The authors also reported the presence of a peak corresponding to a high molecular mass (>200 kDa).