Chiral Separation of the Pesticide Diniconazole by Supercritical Fluid Chromatography (SFC)

Matt Przybyciel, Principal Scientist, ES Industries , David A. Kohler, President, ES Industries , Cory E. Muraco, Global Franchise Product Manager, Liquid

Abstract

Diniconazole is a broad-spectrum triazole fungicide used to prevent plant diseases such as powdery mildew, bunt, rust, smut, and septoria leaf spot. Diniconazole is a chiral chemical with one chiral center and is sold commercially as a mixture of R- and S-enantiomers. It has been reported that R-(-)-diniconazole has higher bactericidal activity, while S-(+)-diniconazole shows higher plant growth regulator activity. It is important to know if environmental factors have altered the ratio of R- and S-diniconazole. Chiral Supercritical Fluid Chromatography (SFC) and mass spectrometry detection have been used as a tool to study the different ratios of diniconazole in various plant materials. For that study a ChromegaChiral™ CCA column was used providing baseline resolution of the two diniconazole enantiomers, however, they are closely separated. In this study 14 different chiral stationary phases were evaluated for the chiral SFC separation. From this evaluation we discovered that a better separation can be achieved by using a newly developed ChromegaChiral stationary phase for a high resolution separation of diniconazole enantiomers.

Introduction

A variety of diseases can attack agricultural crops for many reasons including changes in climatic conditions. These include many fungal diseases such as smut and rust, that can significantly affect crop yields and agricultural output unless they are controlled or eliminated. The application of chemical pesticides can effectively eliminate plant pests, however, with increasing doses and accumulated application, they may potentially impact human health.1 Additionally, about 30% of the commonly used chemical pesticides are chiral, however most of these are sold and used as racemic mixtures.2 A racemic mixture, or racemate, is one that has equal amounts of left- and righthanded enantiomers of a chiral molecule. Enantiomers of the same chemical have identical physicochemical properties, but may exhibit differences in biological activity, pharmacokinetics, pharmacodynamics, and toxicity.3 One widely used pesticide used to control plant fungal diseases is diniconazole which is sold and used as a racemic mixture (Figure 1).

Chemical structure of R- and S- Diniconazole, respectively.

Figure 1. Chemical structure of R- and S- Diniconazole, respectively.

Diniconazole is a broad-spectrum triazole fungicide used to prevent diseases in fruits, vegetables, wheat and tea plants. Its mode of action includes the suppression of 14-α-demethylation in the biosynthesis of ergosterol causing a deficiency of ergosterol which prevents the occurrence of rust and smut disease.4 Diniconazole is a chiral chemical with one chiral center and is sold commercially as a mixture of R- and S- enantiomers. It has been reported that R-(-)-diniconazole has higher bactericidal activity, while S-(+)-diniconazole shows higher plant growth regulator activity. In many cases it is important to know if environmental factors have altered the ratio of R- and S-diniconazole residues when applied on agricultural crops.5

Supercritical fluid chromatography (SFC) is a powerful chromatographic technique for the separation, isolation, and analysis of complex mixtures from many different samples. Many chemicals can potentially be used as a supercritical mobile phase for SFC; however, virtually all current practitioners of SFC use carbon dioxide (CO2) which offers several advantages, particularly when compared to liquid chromatography.6 The use of CO2 as the primary component of the mobile phase is one of the key features that benefits the preparative SFC chromatographer, and, since the CO2 used for SFC is recovered from the atmosphere, it is considered a “Green” solvent. Carbon dioxide is miscible with a wide range of organic solvents, is nonflammable, and has low UV absorbance at low wavelengths.7-9 Other advantages of SFC as a technique are the diffusion coefficients of solutes in the SFC mobile phases that have been shown to be 3-10 times higher than in normal liquids potentially allowing for very rapid separations, and …

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