Eenantiomer Separation And Stereoselective Metabolism In Vitro Of Chiral Pesticides Containing S/P

Nowadays,the chiral phenomenon of pesticides exists generally all over the world.Enantiomers of chiral pesticides always show different or even opposite effects in biological activity,toxicity,distribution and metabolic degradation when spraying to the environment,although they are quite similar in physical and chemical properties.Therefore,it is significant for the rational use of chiral pesticides to study the chiral separation and enantioselective degradation of them.Six chiral pesticides containing S/P?isofenphos-methyl,isofenphos,isocarbophos,flufiprole,fipronil and ethiprole?were separated by supercritical fluid chromatography?SFC?with supercritical CO₂ as main mobile phase.The effects of chiral stationary phase?CSP;Chiralcel OD-H,Chiralpak AS-H,Chiralpak AD-3,Chiralpak IB,Lux 3u Cellulose-1 and Sino-Chiral OJ?,different type and concentration of modifiers?methanol,ethanol and2-propanol?,column temperature?29⁴1??and back pressure?130¹70bar?on the separation were investigated to obtain the optimal separation condition.Meanwhile,the degradation extracts of flufiprole in rat and human liver microsomes were determined on HPLC-MS/MS to explore the stereoselective differences of enantiomers and measure their metabolic kinetic parameters.Five chiral pesticides?isofenphos-methyl,isocarbophos,flufiprole,fipronil and ethiprole?were baseline separated on experimental contidions while isofenphos only obtained partial separation.Chiralpak AD-3 column had better chiral separation ability than others for chiral pesticides containing S/P.When using the different modifier of the same concentration,capacity factor of pesticides except flufiprole decreased in order of 2-propanol,ethanol,methanol,while capacity factor of flufiprole increased in order of 2-propanol,ethanol,methanol.When using the same modifier,the capacity factor and resolution decreased gradually on the whole with increasing percentage of a given modifier.Thermodynamic study of enantiomeric separation showed that the separational mechanisms of chiral pesticides have not changed under the temperature range assayed.The enantiomeric separations of five chiral pesticides were“enthalpy driven”process and the separation factor decreased as the temperature increased.As the back pressure increased,the capacity factor and resolution of chiral pesticides decreased slightly,namely the elution rate accelerated.But the separation factor basical kept within a certain range.The half-lives time(t_1/2)of flufiprole in RLMs and HLMs was 7.22and 21.00 min for R-?+?-flufiprole,while 11.75 and 17.75 min were obtained for?S?-?-?-flufiprole by regression analysis,respectively.The changes of the eliminated rate of flufiprole with substrate concentration had a similar form with Michaelis-Menten equation.The V_max of R-?+?-flufiprole?3906.3±80.57?M/min/mg?was about 3-fold of S-?-?-flufiprole?1430.6±52.93?M/min/mg?in rat liver microsomes?RLMs?,while the V_max of S-?-?-flufiprole was about 2-fold of R-?+?-flufiprole in human liver microsomes?HLMs?.For the CL_int,the CL_intnt of rac-flufiprole also showed opposite enantioselectivy in RLMs and HLMs.Besides,the CL_int of R-?+?-flufiprole and S-?-?-flufiprole in RLMs was higher than that in HLMs.