Extraction, purification, and estimation of ATP from leaves, floral buds, and immature fruits of cotton
✍ Scribed by Gene Guinn; Marie P. Eidenbock
- Publisher
- Elsevier Science
- Year
- 1972
- Tongue
- English
- Weight
- 509 KB
- Volume
- 50
- Category
- Article
- ISSN
- 0003-2697
No coin nor oath required. For personal study only.
✦ Synopsis
Initial attempts at enzymic quantitation of ATP extracted from young cotton (Gossypium hirsutum L.) bolls were unsuccessful; extracts gave no measurable luminescence in the luciferin-luciferase assay. Furthermore, even the light produced by injecting reagent ATP was immediately quenched by subsequent injection of boll extract.
Other workers have reported partial inhibition of enzymic activity in ATP assays (l-7)) primarily by various salts. Stenlid (5) reported that isoflavones strongly inhibited the luciferin-luciferase reaction. Many plants contain phenolic compounds that are potent inhibitors of enzymatic activity when released from sequestered sites (8,9). The nature and amount of phenols in plants may vary with different tissues (9), environment (10) ,' and age (9,ll). These possible differences in phenols should be considered when using enzymic assays because indicated differences in results could be partly or wholly due to differences in inhibition of the enzymes used.
Loomis and Battaile (8) reported that Polyclar AT2 (an insoluble polyvinylpyrrolidone) effectively sorbs phenolics and we found that it could be used to sorb the inhibitor(s) from cotton boll extracts.
In this paper we compare a number of solvent systems for extraction of ATP and outline a procedure for extraction and purification of ATP from cotton leaves, squares, and bolls so that it can be quantitated by the luciferin-luciferase assay system. The results may also be applicable to other plant tissues.
'Journal Paper No. 1895 of the Arizona Agricultural Experiment Station. 'Mention of a trade name or proprietary product does not constitute a guarantee or warranty of the product by the U. S. Department of Agriculture, and does not imply its approval to the exclusion of other products that may also be suitable.
89