The herbicide fluroxypyr 1-methylheptyl ester was separated from its acid form (fluroxypyr) and two soil metabolites (a pyridinol and a methoxypyridine) by high-performance liquid chromatography (HPLC) with the acid of Drylab G. A difference in retention time of at least 6 min between each compound was achieved, to allow complete separation of radiolabeled components during collection of 1-min fractions. The total elution time was 31 min. Actual HPLC retention times differed from DryLab G predictions by 0.5 min or less.

High-performance liquid chromatographic (HPLC) methods for analyzing new drugs and their synthetic intermediates are needed as the synthesis is optimized and scaled up from making milligram amounts for initial evaluation of biological activity to producing kilogram amounts of the drug for thorough testing purposes. The most efficient solution is a single HPLC method that can be used for each step of the synthesis. A practical approach for the development of a single HPLC method is the use of computer-assisted method development to maximize the resolution within a reasonable analysis time. The computer program DryLab I was used in the development of an HPLC assay for the synthetic intermediates of a leukotriene inhibitor. The use of DryLab I with binary mixtures of organic solvents in the organic portion of reversed-phase HPLC systems is reported. With the retention data from two initial analyses, resolution can be optimized as a function of solvent strength.

Using DryLab G to identify a "hidden" band in a protein separation. Complex chromatograms that result from reversed-pahse gradient elution often exhibit changes in band order when the gradient steepness is changed. This complicates the interpretation of the resulting separation, and prevents the application of computer simulation for method development. A simple procedure based on normalized band areas was used to match bands between runs where the gradient steepness has been changed. In one example involving a Thermus aquaticus ribosomal protein sample, it was possible to find an additional band that was not apparent in tw initial experimental runs with different gradient slopes.

A method for prediction ion-exchange isocratic capacity factors from two initial gradient runs is developed. This does not assume so-called linear solvent strength (LSS) conditions, which cause significant errors in k'(C) vs. C relationships in ion-exchange chromatography. The errors associated with this approach and the LSS model are examined. The present approach allows a more accurate prediction of isocratic capacity factors for ion-exchange chromatography. Experimental application of the method to a variety of compounds, including peptides, polynucleotides and polysaccharides, separated by ion-exchange chromatography is described.

Optimization of high-performance liquid chromatography for application to a cough medication (Tussagesic) and its decomposition and byproducts was performed. Special emphasis was placed on the optimization of all parameters that relate to the chemical selectivity of the separation process itself and on the final proof of the ruggedness of the optimized system. All analytes can be reliably determined and the response surface can be represented graphically. This provides a means for improved transfer of methods between laboratories and for efficient system documentation.

Computer simulation (DryLab software) as an aid for the development of gradient high-performance liquid chromatographic methods is reviewed. Several examples of its application are presented and the accuracy of such predictions is discussed.