Bioassay development and

In order to accelerate the process of target identification and target selection prior to bioassay development and high-throughput screening, chip hybridiza-... 

Improved Methods for Collection, Bioassay, Isolation, and Characterization of Compounds. Techniques used to characterize natural products are evolving rapidly as more sophisticated instrumentation is developed. Plant physiologists and chemists should work closely together on this aspect, since rapid and reproducable bioassays are essential at each step. There is no standard technique that will work effectively for every compound. Briefly, isolation of a compound involves extraction or collection in a appropriate solvent or adsorbant. Commonly used extraction solvents for plants are water or aqueous methanol in which either dried or live plant parts are soaked. After extracting the material for varying lengths of time, the exuded material is filtered or centrifuged before bioassay. Soil extraction is more difficult, since certain solvents (e.g. bases) may produce artifacts. 

Several chemical-assay methods (15,23,50) for tetraethyl pyrophosphate were recently developed and applied by seven collaborating laboratories to samples of representative commercial products and to a sample of purified tetraethyl pyrophosphate which served as a common standard. Concordant results, which correlated well with bioassay results,... 

During the 1970 s and early 1980 s a large number of test methods were developed to measure the toxic potency of the smoke produced from burning materials. The ones most widely used are in refs. 29-32. These tests differ in several respects the conditions under which the material is burnt, the characteristics of the air flow (i.e. static or dynamic), the type of method used to evaluate smoke toxicity (i.e. analytical or bioassay), the animal model used for bioassay tests, and the end point determined. As a consequence of all these differences the tests result in a tremendous variation of ranking for the smoke of various materials. A case in point was made in a study of the toxic potency of 14 materials by two methods [33]. It showed (Table I) that the material ranked most toxic by one of the protocols used was ranked least toxic by the other protocol Although neither of these protocols is in common use in the late 1980 s, it illustrates some of the shortcomings associated with small scale toxic potency of smoke tests. 

Bioassays have been likened to analytical machines insofar as pharmacologists use them to assign biological properties to compounds in the same way a chemist measures the physical-chemical properties of molecules. If the fundamental role of the medicinal chemist is to optimize the pharmaceutical properties of so-called lead compounds by structural modification, then the role of the pharmacologist in the drug discovery process is to select, develop, and apply bioassays to provide relevant robust data that inform the medicinal chemist of the impact of the modifications he makes. 

A reduction in cutinase production should result in the PNB-1 mutant being less virulent. The pathogenesis of the two strains were evaluated in a pea stem bioassay developed by Kolattukudy and coworkers in which infection by Fusarium solani results in wound formation within three days on the epicotyl of pea sedlings (15). The virulence of T-8 had previously been shown to be reduced in this assay by the addition of inhibitors of cutinase or by rabbit anticutinase antibodies (15-18), indicating that cutinase played an important role in pathogenesis. When the cutinase-defective mutant was evaluated in the bioassay, the mutant exhibited a 55% reduction (p < 0.05) in virulence compared with the T-8 parental strain and the addition of purified cutinase at 1 mg/ml to the mutant enhanced wound formation to 80% of that of the parent (p > 0.5). These data further support the notion that the mutant was defective in cutinase. 

Nishioka, M. G., C. C. Chuang, and B. A. Petersen, Development and Quantitative Evaluation of a Compound Class Fractionation Scheme for Bioassay-Directed Characterization of Ambient Air Particulate Matter, Environ. Ini., 11, 137-146 (1985). 

Although some investigators have tested adsorption and desorption separately, most have reported the efficiency of the total process. Separate efficiencies are useful for methods development and are of academic interest, but the total efficiency is primary for analytical and bioassay purposes. 

Standardized ecotoxicity tests (bioassays) have been developed and optimized over the last few years and encompass the effects on bacteria, daphnia and fish (DIN 38 412, parts 30, 31 and 34). These tests are designed to assess the toxicity on aquatic organisms. They are quick to perform, easy to handle and comparatively inexpensive, with the goal of allowing the toxicity of a complex water matrix to be estimated. However, they use pre-concentration steps so that it is possible that not all byproducts are recovered (which itself is hard to prove). 

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