Carbamates enzyme inhibition

FIGURE 5.46 Interaction of the serine hydroxyl residue in the catalytically active site of acetylcholinesterase enzyme with esters of organophosphates or carbamates. The interaction leads to binding of the chemical with the enzyme, inhibition of the enzyme, inhibition of acetylcholine hydrolysis, and thus accumulation of acetylcholine in the synapses. 

Toxin (Enzyme Inhibition) Biosensors Enzyme affectors (inhibitors and activators) that influence the rate of biocatalytic reactions can also be measured. Sensing probes for organophosphate and carbamate pesticides, for the respiratory... 

Organophosphate and carbamate pesticides are potent inhibitors of the enzyme cholinesterase. The inhibition of cholinesterase activity by the pesticide leads to the formation of stable covalent intermediates such as phosphoryl-enzyme complexes, which makes the hydrolysis of the substrate very slow. Both organophosphorus and carbamate pesticides can react with AChE in the same manner because the acetylation of the serine residue at the catalytic center is analogous to phosphorylation and carbamylation. Carbamated enzyme can restore its catalytic activity more rapidly than phosphorylated enzyme [17,42], Kok and Hasirci [43] reported that the total anti-cholinesterase activity of binary pesticide mixtures was lower than the sum of the individual inhibition values. 

T. Noguer, B. Leca, G. Jeanty, and J.L. Marty, Biosensors based on enzyme inhibition detection of organophosphorus and carbamate insecticides and dithiocarbamate fungicides. Field Anal. Chem. Tech. 3, 171-178 (1999). 

S. Jin, Z. Xu, J. Chen, X. Liang, Y. Wu, and X. Qian, Determination of organophosphate and carbamate pesticides based on enzyme inhibition using a pH-sensitive fluorescence probe. Anal. Chim. Acta 523, 117-123 (2004). 

In our early work, we attempted the alkylation of the amine 81 with various carbohydrate triflates, namely 82-84 [57]. Somewhat disappointingly, the main products isolated were the alkenes, e.g. 85 and 86. Only a small amount of the desired N-linked carbasugar, e. g. 87 was ever obtained. At one stage we attempted an alkylation of the amine 81 with the triflate 82 and with the triflate 84 in the presence of potassium carbonate to our surprise, carbamates were formed, 88 and 89 These two carbamates have subsequently been deprotected to give the methyl -o-glucoside 90 and the free sugar 91, interesting candidates for enzyme inhibition studies [57]. 

Insecticidal carbamates also inhibit the enzyme acetylcholinesterase by transferring a carbamoyl group to the active hydroxyl. However, they differ from the phosphates in that they inhibit the enzyme reversibly and so a better fit at the active site is required for high activity. In consequence, a narrower range of structures is active. The chemistry, biochemistry, metabolism and toxicology of carbamate insecticides have been thoroughly reviewed (B-76MI10702). 

Enzymatic techniques have also been employed in the analysis of these compounds. The toxicity of carbamate insecticides is due to the inhibition of the enzyme acetylcholine esterase, so the determination of these compounds can be achieved by enzyme inhibition (2,83,119), bioassay (118,167), or enzyme-linked immunosorbent assay (ELISA) (168-171). In the detection of carbamates by fluorimetric enzyme inhibition, the effluent from a reversed-phase chromatographic column was incubated with cholinesterase, which was introduced via a postcolumn reagent delivery pump. Then, the resulting partially inhibited cholinesterase was reacted with N-methyl indoyl acetate to produce a fluorophore and a reduction in the baseline fluorescence (172). 

The action spectrum for DDT and its structural analogs is known to be quite broad and cannot be attributed to simple enzyme inhibition, as in the case of the carbamates or the organophosphates. The presence of chloroaryl moieties, as well as steric effects at receptor sites, both appear to be factors affecting insecticidal activity. In addition to DDT itself, its metabolites DDE and DDA and DDD have some activity. Efforts to overcome insect resistance and to produce more biodegradable analogs led to the introduction of substituents other than chlorine for... 

These devices are now being tested in demilitarization plants as monitors of stack exhaust atmospheres and work areas. We have not had the opportunity to determine the sensitivity of the devices to the numerous enzyme-inhibiting compounds which would be of interest to others. Generally, however, it is reasonable to expect, for instance, less sensitivity to P=S and P—S linkages and conversely better sensitivity to compounds where the sulfur is replaced by oxygen due to greater toxicity of these compounds. Such compounds as the carbamates, due to their inhibiting qualities, would also be expected to be detected with these systems. 

Derivatives of aminoformic acid react with ChE similarly to OPC, i.e. in two stages with establishing a covalent bond. A degree of the anti-cholinesterase action of carbamates depends upon strength of the complexes formed. Evidently, the phosphorylated ChE becomes disabled for a more prolonged period, as compared with the carbamylated ChE. Rapidity of the initial activity restoration of the enzyme inhibited by carbamates is determined, respectively, by the carbamylated enzyme hydrolysis speed, which depends upon the inhibitor structure. Carbamy-... 

Early studies of the relation of aryl N-methylcarbamate structure with activity and enzyme inhibition were first reported in 1966 and showed rather simple dependence on substituent effects for limited sets of compounds (4, 5) The apparent simplicity was partly due to the fact that QSAR techniques were still in the first decade of development. The true complexity of carbamate inhibition is revealed in later studies by Fujita and co-workers on brown plant-hopper AChE (6, 7), and by Hansch and co-workers on housefly head AChE (8). The latter study shows that 12 significant factors are required to correlate the pI50 data for a set of 269 carbamates. 

Ortho Substituted Phenyl N-Methylcarbamates. Table 1 shows the enzyme Inhibition data for all of the tested carbamates. With the single exception of the 2-phenyl compound, all of the carbamates were stronger Inhibitors (higher pI50) of AChE from S-straln of the leafhopper. This clearly confirms the supposition that resistance to carbamates Is due mainly to changes In enzyme Inhibition. It Is also consistent with a study by Hama and Iwata showing a correlation between the degree of resistance and both Insecticidal activity and AChE Inhibition for several carbamate Insecticides (22). 

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