Amylase, maltase and sucrase activities in hepatopancreas of the euryhaline crab <i>Neohelice granulata</i> (Decapoda: Brachyura: Varunidae): partial characterization and response to low environmental salinity

Abstract

Studies on digestive adjustments at the biochemical level in relation to salinity in euryhaline crabs are lacking. Moreover, knowledge of biochemical digestive characteristics of euryhaline crabs (i.e. occurrence and characteristics of key digestive enzyme activities) is still scarce and fragmentary. We studied the occurrence, characteristics and response to low salinity of amylase, maltase and sucrase activities in the hepatopancreas of the euryhaline crab Neohelice (Chasmagnathus) granulata. Maximal amylase and maltase activities were found at pH 5.2. Sucrase activity was maximal within the pH range 3.6-5.2. Amylase, maltase and sucrase activities showed a Michaelis-Menten kinetics (km = 0.41±0.10 mg ml-1 ; 1.37±1.03 mM and 0.55±0.45 mM, respectively). In crabs acclimated to low salinity (10 psu; hyperregulating conditions), amylase activity (7263±980 µg maltose min-1 mg prot-1) was higher than in 35 psu (osmoconforming conditions) (3605±340 µg maltose min-1 mg prot-1). Maltase and sucrase activities (497±98 and 64±16 µg glucose min-1 mg prot-1 , respectively) were similar in both salinities. The response of amylase activity to low salinity suggests a role in digestive adjustments upon hyperregulation. This study contributes to a better understanding of the complexity of the biochemical adaptations to low salinity in euryhaline crabs.

Amylase in the hepatopancreas of a euryhaline burrowingcrab: characteristics and modulation

“…A sample of hemolymph was withdrawn for assaying osmolality as described below. The hepatopancreas was immediately excised, mixed with 50 mM Tris/HCl of pH 8.0 (4 mL g -1 of hepatopancreas tissue), and homogenized (CAT homogenizer × 120, tool T10) on ice (Asaro et al, 2011;Michiels et al, 2015a). The homogenate was centrifuged at 10,000 × g for 15 min at 4 °C.…”

“…2.6. Biochemical assays Amylolytic activity was measured as previously described (Asaro et al, 2011). In the standard assay, amylolytic activity was determined in a reaction mix containing 15 mg mL -1 starch in 50 mM phosphate buffer (pH 5.2) at 30 °C.…”

“…In the Mar Chiquita coastal lagoon (Argentina), this crab successfully occupies the whole intertidal area, where it is exposed to a broad range of environmental salinities (Spivak et al, 1994;Luppi et al, 2013). Previous work of our laboratory showed that complex and integrative responses occur upon acclimation to low and high salinity (López Mañanes et al, 2000;Schleich et al, 2001;Pinoni et al, 2005Pinoni et al, , 2013Pinoni et al, , 2015Pinoni and López Mañanes, 2009;Asaro et al, 2011;Michiels et al, 2015a). The aims of this work were to determine the occurrence of amylase forms in the hepatopancreas of the intertidal euryhaline crab N. granulata, to examine the effect of acclimation of crabs to low and high salinity on the pattern of amylase forms, and to determine biochemical characteristics of partially purified amylase.…”

“…In animals, α-amylases (α-1,4 glucan-4-gluconohydrolase, EC 3.2.1.1) have a central role in the initial steps of the digestion of key glycogenic substrates such as dietary starch and dietary and/or storage glycogen (Singh and Kayastha, 2014;Date et al, 2015). However, in spite of their inherent physiological importance, the current knowledge about forms, modulation by environmental factors, and biochemical features of amylase in the hepatopancreas (the main site of digestive enzyme synthesis and of digestion and absorption of nutrients and storage of energy substrates) of groups of ecological importance such as intertidal euryhaline crabs is still scarce and fragmentary (Blandamer and Beechey, 1966;Van Wormhoudt et al, 1995;Asaro et al, 2011;Zeng et al, 2010).…”

“…modulation of key digestive enzymes) in response to variations in environmental factors (del Valle et al, 2004(del Valle et al, , 2006Karasov et al, 2011;Karasov and Douglas, 2013). In some species of intertidal euryhaline crabs, biochemical adaptation to low and high salinity involves adjustments in various tissues and organs such as the hepatopancreas, although the mechanisms are still not fully understood from a metabolic perspective (Pinoni et al, 2005(Pinoni et al, , 2008Pinoni and López Mañanes, 2009;Asaro et al, 2011;Romano and Zeng, 2012;Romano et al, 2014;Michiels et al, , 2015aPinoni et al, 2013Pinoni et al, , 2015. The modulation of digestive enzyme activities in the hepatopancreas (e.g., amylase) in response to different environmental salinity could lead to adjustments in the digestive capacity for specific substrates (starch/glycogen) and therefore in the availability of key metabolites (i.e.…”