Racemic difluoromethylornithine (D/L-DFMO) is an inhibitor of ODC (ornithine decarboxylase), the first enzyme in eukaryotic polyamine biosynthesis. D/L-DFMO is an effective anti-parasitic agent and inhibitor of mammalian cell growth and development. Purified human ODC-catalysed ornithine decarboxylation is highly stereospecific. However, both DFMO enantiomers suppressed ODC activity in a time- and concentration-dependent manner. ODC activity failed to recover after treatment with either L- or D-DFMO and dialysis to remove free inhibitor. The inhibitor dissociation constant (K(D)) values for the formation of enzyme-inhibitor complexes were 28.3+/-3.4, 1.3+/-0.3 and 2.2+/-0.4 microM respectively for D-, L- and D/L-DFMO. The differences in these K(D) values were statistically significant ( P <0.05). The inhibitor inactivation constants (K(inact)) for the irreversible step were 0.25+/-0.03, 0.15+/-0.03 and 0.15+/-0.03 min(-1) respectively for D-, L- and D/L-DFMO. These latter values were not statistically significantly different ( P >0.1). D-DFMO was a more potent inhibitor (IC50 approximately 7.5 microM) when compared with D-ornithine (IC50 approximately 1.5 mM) of ODC-catalysed L-ornithine decarboxylation. Treatment of human colon tumour-derived HCT116 cells with either L- or D-DFMO decreased the cellular polyamine contents in a concentration-dependent manner. These results show that both enantiomers of DFMO irreversibly inactivate ODC and suggest that this inactivation occurs by a common mechanism. Both enantiomers form enzyme-inhibitor complexes with ODC, but the probability of formation of these complexes is 20 times greater for L-DFMO when compared with D-DFMO. The rate of the irreversible reaction in ODC inactivation is similar for the L- and D-enantiomer. This unexpected similarity between DFMO enantiomers, in contrast with the high degree of stereospecificity of the substrate ornithine, appears to be due to the alpha-substituent of the inhibitor. The D-enantiomer may have advantages, such as decreased normal tissue toxicity, over L- or D/L-DFMO in some clinical applications.

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Syntheses of the enantiomers of malathion, malaoxon, and isomalathion are reported herein. Malathion enantiomers were prepared from (R)- or (S)-malic acid in three steps. Enantiomers of malathion were converted to the corresponding enantiomers of malaoxon in 52% yield by oxidation with monoperoxyphthalic acid, magnesium salt. The four isomalathion stereoisomers were prepared via two independent pathways using strychnine to resolve the asymmetric phosphorus moiety. The absolute configurations of the four stereoisomers of isomalathion were determined by X-ray crystallographic analysis of an alkaloid salt precursor. A high-performance liquid chromatography technique was developed to resolve the four stereoisomers of isomalathion, and to determine their stereoisomeric ratios.

An industrial process for the purification of metric tons of enantiomerically pure difluoromethylornithine (DFMO HCl) is described. The amino acid DFMO HCl is cyclized to form the lactam, which is acylated with pivaloyl chloride to form rac-Npivaloyl-DFMO lactam (4). The lactam 4 provides enhanced separation compared to a direct resolution of racemic DFMO HCl (1). A hybrid chiral resolution process is proposed to separate the enantiomers of 4. This process involves a multicolumn continuous enantioselective chromatographic process (VARICOL) coupled with enantioselective crystallization of (D)-N-pivaloyl-DFMO lactam 5. The interest of this hybrid process is based on a favorable eutectic point providing a higher productivity of the VARICOL process and lower purification costs than the chromatographic process alone. A final chemical modification (hydrolysis) is used to form the single enantiomers of both (D)-DFMO ( 6) and (L)-DFMO in high chemical purity and enantiomeric excess. A global optimization approach is applied to design an economical industrial process, which is based on a parametric study of the VARICOL process and enantioselective crystallization to obtain maximum recovery and purity while significantly lowering the cost of manufacturing the single enantiomers. A detailed description of the global process optimization is presented.

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