Discovery of novel piperonyl derivatives as diapophytoene desaturase inhibitors for the treatment of methicillin-, vancomycin- and linezolid-resistant Staphylococcus aureus infections.

Inhibition of S. aureus diapophytoene desaturase (CrtN) could serve as an alternative approach for addressing the tricky antibiotic resistance by blocking the biosynthesis of carotenoid pigment which shields the bacterium from host oxidant killing. In this study, we designed and synthesized 44 derivatives with piperonyl scaffold targeting CrtN and the structure-activity relationships (SARs) were examined extensively to bring out the discovery of 21b with potent efficacy and better hERG safety profile compared to the first class CrtN inhibitor benzocycloalkane derivative 2. Except the excellent pigment inhibitory activity against wild-type S. aureus, 21b also showed excellent pigment inhibition against four pigmented MRSA strains. In addition, H2O2 killing and human whole blood killing assays proved 21b could sensitize S. aureus to be killed under oxidative stress conditions. Notably, the murine study in vivo validated the efficacy of 21b against pigmented S. aureus Newman, vancomycin-intermediate S. aureus Mu50 and linezolid-resistant S. aureus NRS271.

The interactions of piperonyl butoxide and analogues with the metabolic enzymes FE4 and CYP6CY3 of the green peach aphid Myzus persicae (Hemiptera: Aphididae).

BACKGROUND: Piperonyl butoxide (PBO) is a well-known insecticide synergist capable of interacting with phase 1 metabolic enzymes, specifically esterases and cytochrome P450s. In this study, structure-activity relationship analyses were used to characterise the interaction of around 30 analogues of PBO with the esterase FE4 and the P450 CYP6CY3 from insecticide-resistant Myzus persicae (Sulzer), in order to predict the synthesis of more potent inhibitors. RESULTS: Enzyme inhibition studies were performed against esterase and oxidase activities and, together with in silico modelling, key activity determinants of the analogues were identified and optimised. Novel analogues were then designed and synthesised, some of which showed greater inhibition against both enzymatic systems: specifically, dihydrobenzofuran moieties containing an alkynyl side chain and a butyl side chain against FE4, and benzodioxole derivatives with a propyl/butyl side chain and an alkynyl ether moiety for CYP6CY3. CONCLUSIONS: In vitro assays identified potential candidate synergists with high inhibitory potency. The in vivo confirmation of such results will allow consideration for a possible use in agriculture. © 2016 Society of Chemical Industry.

The use of substituted alkynyl phenoxy derivatives of piperonyl butoxide to control insecticide-resistant pests.

BACKGROUND: Derivatives of piperonyl butoxide with alkynyl side chains were tested in vitro and in vivo against pyrethroid-resistant Meligethes aeneus and imidacloprid-resistant Myzus persicae. RESULTS: Synergists with the alkynyl side chain were more effective inhibitors of P450 activity in vitro than piperonyl butoxide, and demonstrated high levels of synergism in vivo, with up to 290-fold synergism of imidacloprid against imidacloprid-resistant M. persicae. CONCLUSIONS: These 'second-generation' synergists could overcome metabolic resistance in many pest species and possibly enable reduced rates of insecticide application in some cases. © 2016 Society of Chemical Industry.

The interactions between piperonyl butoxide and E4, a resistance-associated esterase from the peach-potato aphid, Myzus persicae Sulzer (Hemiptera: Aphididae).

BACKGROUND: It has been reported previously that piperonyl butoxide (PBO) can inhibit both P450 and esterase activity. Although the method by which PBO combines with cytochrome P450 has been identified, the way in which it acts as an esterase inhibitor has not been established. This paper characterises the interactions between PBO and the resistance-associated esterase in Myzus persicae, E4. RESULTS: After incubation with PBO/analogues, hydrolysis of 1-naphthyl acetate by E4 is increased, but sequestration of azamethiphos is reduced. Rudimentary in silico modelling suggests PBO docks at the lip of the aromatic gorge. CONCLUSIONS: PBO binds with E4 to accelerate small substrates to the active-site triad, while acting as a blockade to larger, insecticidal molecules. Structure-activity studies with analogues of PBO also reveal the essential chemical moieties present in the molecule.

An analogue of piperonyl butoxide facilitates the characterisation of metabolic resistance.

BACKGROUND: Previous work has demonstrated that piperonyl butoxide (PBO) not only inhibits microsomal oxidases but also resistance-associated esterases. The ability to inhibit both major metabolic resistance enzymes makes it an ideal synergist to enhance xenobiotics but negates the ability to differentiate which enzyme group is responsible for conferring resistance. RESULTS: This study examines an analogue that retains the ability to inhibit esterases but is restricted in its ability to act on microsomal oxidases, thus allowing an informed decision on resistance enzymes to be made when used in conjunction with the parent molecule. CONCLUSION: Using examples of resistant insects with well-characterised resistance mechanisms, a combination of PBO and analogue allows identification of the metabolic mechanism responsible for conferring resistance. The relative potency of PBO as both an esterase inhibitor and an oxidase inhibitor is also discussed.

Benzodioxole derivatives as negative effectors of plant proteases.

Previous work demonstrated that a commercial formulation of piperonyl butoxide (PBO) did inhibit the activity of some plant proteolytic enzymes. In this paper, the effect of pure PBO and nine pure PBO homologues (PBOH) appropriately synthesized toward bromelain and papain was studied in hydrocarbon solution using the bis(2-ethylhexyl)sodium sulfosuccinate (AOT) reverse micellar system. This study establishes that the majority of these compounds show, in vitro, interesting protease inhibition activities. The benzodioxole and dihydrobenzofuran structures, in particular, 5-[2-(2-butoxyethoxy)ethoxymethyl]-benzo[1,3]dioxole (EN 1-40) and 6-[2-(2-butoxyethoxy)ethoxymethyl]-5-propyl-2,3-dihydrobenzofuran (EN 16-5), respectively, appear to be responsible for protease inhibition. Measures of octanol/water partition coefficients on PBO and PBOH have demonstrated that water solubility plays a fundamental role in the expression of protease inhibition activity.

Comparative evaluation of new synergists containing a butynyl-type synergophore group and piperonyl butoxide derivatives.

Cross-substituted derivatives of piperonyl butoxide (PBO) and MB-599 (proposed common name: verbutin) were synthesized and investigated as carbofuran and permethrin synergists against housefly, Musca domestica L. The majority of PBO and MB-599 derivatives were significantly more potent synergists for carbofuran than for permethrin. PBO, the most important representative of this series was not the most potent synergist for carbofuran or for permethrin. Cleavage of the methylenedioxy ring of methylenedioxyphenyl (MDP) polyether compounds resulted in complete loss of synergistic activity with both insecticides, but it could be restored or even improved by incorporating an alkynyl ether moiety into the molecule. The improved synergistic activity was found to be closely associated with the 2-butynyloxymethyl side-chain, suggesting that this can be regarded as a characteristic synergophore group. MB-599, one of the most promising compounds bearing this group showed considerably higher activity with carbofuran (synergist ratio, SR = 37.8) than with PBO (SR = 6.4). There was no significant difference between synergistic activities of MB-599 (SR = 4.6) and PBO (SR = 4) for permethrin.

Residue levels of pyrethrins and piperonyl butoxide in soil and runoff water.

Simultaneous analysis of pyrethrins (Py-I and Py-II) and piperonyl butoxide (PBO) in soil and runoff water samples following field application of a new pyrethrum formulation containing pyrethrins (Py's) and PBO is described. Residues of total Py's and PBO were extracted from soil samples using hexane-acetone (9:1). A solid phase extraction (SPE) column containing C18-octadecyl bonded silica was used to separate Py's and PBO residues from runoff water. Residues in soil and water were quantitated by high performance liquid chromatography (HPLC) equipped with C18-column and a UV detector. Concentration of Py-II in soil was 100 times higher than that of Py-I 1 h following treatment and 9.6 times higher than Py-I in runoff surface water 11 days following treatment. Results indicated that Py's are non-persistent in soil (even though lipophillic) and water when applied at the recommended rate of 6 lbs (5.31 g A.I.) per acre. There was a consistent decrease in total Py's residues as time after spraying increased. Py's residues in soil decreased from 0.91 to 0.11 ppm 4 days following treatment and one month after treatment only 0.002 ppm were detected. The highest concentration of Py's in runoff water was 36.09 ng/liter following the first rainfall (11 days following treatment). PBO initial residues detected in soil samples were low (0.84 microgram/g soil) while no residues of PBO were detected in runoff water.