Lizard cholinesterases as biomarkers of pesticide exposure: Enzymological characterization

Abstract

Here we report the results of a study conducted to elucidate the enzymological characteristics of lizard cholinesterases (ChEs) in order to use them as potential biomarkers for pesticide exposure. Serum and brain tissue of the lizard Gallotia galloti were used as ChE sources and in vitro assays were performed to identify acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activities. The pH, substrate concentration, and specificity for ChE assays as well as the response of serum BChE to the reactivating agent pyridine-2-aldoxime methochloride (2-PAM) were also investigated in order to assess the possibilities of this methodology in biomonitoring programs. By the use of selective substrates and the inhibitor tetraisopropyl pyrophosphoramide, AChE and BChE activities were identified in lizard serum, while brain contained solely AChE. Likewise, butyrylthiocholine iodide was the optimum substrate for determining BChE activity and acetylthiocholine iodide for assaying both serum and brain AChE activities. The optimal ranges of pH and substrate concentrations were 7.5 to 8.0 and 5 to 10 mM, respectively. Serum was incubated with different doses of the organophosphorus (OP) compounds dichlorvos and paraoxon and subsequently incubated in the presence of two concentrations of 2-PAM (2 x 10(-2) or 2 x 10(-4) M). Reactivation rate of phosphorylated BChE was related to the degree of inhibition of BChE and the dose of 2-PAM. It was found that a 90-min incubation time with 2 x 10(-4) M of 2-PAM satisfactorily increased the OP-inhibited BChE activity. The enzymological properties of serum BChE activity and its in vitro reactivation in the presence of 2-PAM represent the initial justification for its use in monitoring OP contamination in the field.

A Comparison of Multiple Esterases as Biomarkers of Organophosphate Exposure and Effect in Two Earthworm Species

“…Our study supports earlier research performed in other species using both CbE and AChE as biomarkers of organophosphate exposure (Bonacci et al 2004;Sanchez-Hernandez and Moreno Sanchez 2002) by confirming that CbE, AChE, and BChE are present and quantifiable in both earthworm species. Additionally, all of the esterases were successfully inhibited by malathion exposure (Fig.…”

Characterization of Cholinesterases in Plasma of Three Portuguese Native Bird Species: Application to Biomonitoring

“…Plasma BChE of M. bassanus and C. ciconia showed a higher affinity to BuSCh (K m  = 19.4 and 4.8 µM, respectively) than other vertebrates such as the mink Mustela vison (K m  = 240 µM) and the lizard Gallotia galloti (K m  = 1×10 3 µM), but similar to the pigeon Columbia livia (K m  = 32 µM) and the rat Rattus novergicus (K m  = 13 µM) [25], [26]. The K m values of the grey heron ( A. cinerea ) obtained with BuSCh was higher than the ones reported for the other two studied species, but smaller than the ones obtained with M. vison , G. galloti and the fish Pioractus mesopotamicus (K m  = 1.2×10 3 µM) [25], [26].…”

Earthworm biomarkers of pesticide contamination: Current status and perspectives

“…31) Earthworms are among the organisms that show the slowest recovery rates, in terms of several months, of phosphorylated ChEs. 5,32) This extremely slow recovery makes ChE inhibition an excellent biomarker of pesticide contamination.…”

“…should be investigated when biomarker responses are used for field monitoring of pesticide contamination. This aspect of biomarker research has been extensively studied in vertebrates such as birds and fish, 10,31) and with certain biomarkers such as cytochrome P450-dependent monooxygenases (CYP) induction. 38) To date, only a few studies have examined the impact of confounding factors on earthworm biomarkers.…”