Ajit S. Shah

EPSP Synthase: The Design and Synthesis of Bisubstrate Inhibitors Incorporating Novel 3-Phosphate Mimics

Abstract Novel aromatic bisubstrate inhibitors of the enzyme EPSP (5-enolpyruvoylshikimate-3-phosphate) synthase (EC 2.5.1.19) have been designed and synthesized as structural analogs of the single, catalytic intermediate 1 utilized by the enzyme. These aromatic inhibitors incorporate novel α-hydroxyphosphonates, malonate ethers and α-hydroxymalonates as replacements for the hydrolytically labile 3-phosphate group. These 3-phosphate mimics were much preferred to the corresponding methylene and vinylic phosphonates, malonates and phosphonomethyl ethers.

The thermal stability of ethyl diazoacetate

Ethyl diazoacetate (EDA) is an important organic chemistry synthon, but its use as an industrial intermediate is limited due in part to the safety concerns associated with its instability and high reactivity. In a continuing effort to understand the risk of employing EDA as an industrial reagent, the thermal stability of EDA was studied. Reported are the results of accelerating rate and differential scanning calorimetry (DSC) experiments.

Detonation properties of ethyl diazoacetate

Ethyl diazoacetate is an important organic chemistry synthon, but its use as an industrial intermediate is limited due in part to the safety concerns associated with its instability and high reactivity. In a continuing effort to understand the risk of employing ethyl diazoacetate as an industrial reagent, detonation tests were completed. The results of these studies indicate that ethyl diazoacetate does not show detonation properties.

Understanding the large-scale chemistry of ethyl diazoacetate via reaction calorimetry

Ethyl diazoacetate is an important organic chemistry synthon, but its use as an industrial intermediate is limited due in part to the safety concerns associated with its instability and high reactivity. Reported are the reaction calorimetry results obtained from a new ethyl diazoacetate preparative method, the results from its use in a novel, exothermic reaction and the application of this data toward process development, a successful pilot plant campaign and full-scale reactor design. The results indicate that, given the proper engineering design, in situ preparation and use of ethyl diazoacetate solutions can be used at production scale without significant risk.

New aromatic inhibitors of EPSP synthase incorporating hydroxymalonates as novel 3-phosphate replacements

A new, aromatic analogue of the EPSP synthase enzyme reaction intermediate 1 has been identified, which contains a 3-hydroxymalonate moiety in place of the usual 3-phosphate group. This simplified inhibitor was readily prepared in five steps from ethyl 3,4-dihydroxybenzoate. The resulting tetrahedral intermediate mimic 9 is an effective, competitive inhibitor versus S3P with an apparent Ki of 0.57 +/- 0.06 microM. This result demonstrates that 3-hydroxymalonates exhibit potencies comparable to aromatic inhibitors containing the previously identified 3-malonate ether replacements and can thus function as suitable 3-phosphate mimics in this system. These new compounds provide another example in which a simple benzene ring can be used effectively in place of the more complex shikimate ring in the design of EPSP synthase inhibitors. Furthermore, the greater potency of 9 versus the glycolate derivative 10 and the 5-deoxy-analog 11, again confirms the requirement for multiple anionic charges at the dihydroxybenzoate 5-position in order to attain effective inhibition of this enzyme.

Application of reaction calorimetry toward understanding the large scale chemistry of ethyl diazoacetate

Ethyl diazoacetate is an important organic chemistry synthon, but its use as an industrial intermediate is limited due in part to the safety concerns associated with its instability and high reactivity. Reported are the reaction calorimetry results obtained from a new ethyl diazoacetate preparative method, the results from its use in a novel, exothermic reaction and the application of this data toward process development, a successful pilot plant campaign and full-scale reactor design. The results indicate that, given the proper engineering design, in situ preparation and use of ethyl diazoacetate solutions can be used at production scale without significant risk.