Comparative esterase kinetics and hypoxia-induced, 4-hydroxynonenal-mediated regulation of carboxylesterase in three Neotropical Characiform fish.

The serine-hydrolase esterases-acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and carboxylesterase (CarbE)-mediate xenobiotic detoxification and lipid metabolism, yet their comparative kinetics and regulatory mechanisms remain largely unknown in Neotropical fish. Here, we provide the first integrated kinetic and mechanistic characterization of AChE, BChE, and CarbE in the serum and liver of three ecologically and commercially important Characiform fish: curimbatá (Prochilodus lineatus), pacu (Piaractus mesopotamicus), and piavussu (Leporinus macrocephalus). The dominant serum esterase was species-specific: CarbE predominated in curimbatá (Vmax = 29.85 U·mL-1) and pacu (4.69 U·mL-1), whereas piavussu serum was dominated by BChE (17.87 U·mL-1). Selective-inhibitor profiling confirmed the B-esterase identity of serum CarbE, which was inhibited by the organophosphate methyl-paraoxon with approximately 10-fold higher potency in curimbatá than in pacu (IC50 = 74 vs. 691 nM), indicating interspecific differences in organophosphate-scavenging capacity. Semi-purified CarbE fractions from the two species were biochemically distinct (49 vs. 56 kDa; Vmax = 3426 vs. 330 U·mg-1 protein). Pacu CarbE hydrolyzed p-nitrophenyl palmitate (p-NPP) in serum and liver microsomes, demonstrating long-chain ester hydrolase activity, and the microsomal activity decreased after 42 h of hypoxia (0.5 mg O2·L-1), with a more pronounced effect during spring/summer. The lipid aldehyde 4-hydroxynonenal (4-HNE) inhibited serum CarbE more potently than the microsomal form (approximate IC50 ≈ 2 vs. ≈ 4 mM), supporting a potential link between hypoxia-associated lipid peroxidation and reduced esterase activity. These findings establish a biochemical framework for esterase function in Neotropical Characiform fish and identify 4-HNE-mediated CarbE inhibition as a candidate regulatory mechanism associated with oxygen limitation.