Development of Optically Pure Pyrethroid-Like Fluorescent Substrates for Carboxylesterases

Abstract

Pyrethroids are now the world's most extensively used insecticides. One of the common metabolic routes of pyrethroid insecticides in living systems is hydrolysis by carboxylesterases, and this hydrolysis may be stereospecific since most pyrethroid insecticides have chiral centers. In previous studies, pyrethroid-like fluorescent substrates have been shown to be hydrolyzed in a fashion similar to actual pyrethroids. It is important to synthesize the stereoisomers of pyrethroid-like fluorescent substrates to study the stereointeraction between carboxylesterases and these substrates. In this study, an effective synthetic method for preparing optically enriched (R)- and (S)-α-2-hydroxy-2-(6-methoxy-2-naphthyl)acetonitrile was developed. With this alcohol, an efficient synthetic route for preparation of optically pure cypermethrin and fenvalerate analogues was provided. Identification of these stereoisomers was determined based on GC, HPLC, 1H NMR, and X-ray crystallography. In addition, stereointeraction between carboxylesterases and chiral fluorescent substrates indicated that (i) stereospecificity of recombinant mouse liver carboxylesterases (NCBI accession nos. BAC36707 and NM_133960) varied significantly (up to 300-fold difference) with different stereoisomers of cypermethrin and fenvalerate analogues; (ii) on the basis of V max, the sensitivity of this analytical method, using a single stereoisomer of cypermethrin analogues instead of a mixture of eight stereosiomers, could be enhanced by 4−6 times for detection of these carboxylesterases; and (iii) possible usage of these carboxylesterases for chiral synthesis is discussed.

Characterization of pyrethroid hydrolysis by the human liver carboxylesterases hCE-1 and hCE-2

“…The preferences of hCE-2 for the optical isomers of A3 and A4 were similar to those found for two murine pyrethroid-hydrolyzing carboxylesterases [11]. The (1S)-trans-isomers of A3 were hydrolyzed much faster by hCE-2 in a similar manner as the mouse pyrethroid-hydrolyzing carboxylesterases.…”

“…In this study, the mechanism of lower hydrolytic activity of hCE-1 for cis-A3 was caused by protrusion of the dichlorovinyl group of cis-A3 into oxyanion hole, resulting in the inability of the carbonyl carbon to access the active pocket of the enzyme. This mechanism may be applicable to other pyrethroidhydrolyzing carboxylesterases.The preferences of hCE-2 for the optical isomers of A3 and A4 were similar to those found for two murine pyrethroid-hydrolyzing carboxylesterases [11]. The (1S)-trans-isomers of A3 were hydrolyzed much faster by hCE-2 in a similar manner as the mouse pyrethroid-hydrolyzing carboxylesterases.…”

“…, cis-A7, trans-A7, A8, and four fenvalerate isomers (αR)(2R)-B5, (αR)(2S)-B5, (αS)(2R)-B5, and (αS)(2S)-B5) were all previously synthesized in this laboratory [6,[8][9][10][11]. The term α refers to the α-carbon of the alcohol moiety and 2 refers to the 2-position carbon of the acid moiety.…”

“…Thus, there are eight stereoisomers for cypermethrin and four stereoisomers for fenvalerate. To examine the hydrolytic activity of esterases for these optical isomers, eight and four fluorescent stereoisomer surrogates resembling cypermethrin and fenvalerate, respectively, were synthesized in our laboratory [11] and tested against hCE-1 and hCE-2 (Table 4). Both hCE-1 and hCE-2 hydrolyzed the trans-isomers of A3 faster than the corresponding cis-counterparts as was observed with the authentic pyrethroids.…”

“…On the other hand, hCE-2 showed a preference for (1S)-trans-A3, indicating that the cis-or transconfiguration at the C-1 and C-3 carbons are much more important than the C-α carbon. In mice, pyrethroidhydrolyzing carboxylesterases also prefer trans-isomers of A3 [11]; and trans-permethrin is over 78-fold less toxic than their cis-counterparts [29]. This suggests that hCE-1 and hCE-2 partly participate in the detoxification of pyrethroids with a preference for trans-isomers, possibly causing diffrent toxicity with certain optical isomers of pyrethroids in humans.…”