Extraction and Measurement Techniques 17 



the frozen plant material is dried in vacuo without ever experiencing 

 temperatures above freezing. The dry material can then be powdered 

 and extracted with wet ether. 



25" C. 



TEMPERATURE DURING EXTRACTION 



Fig. 4. The effects of temperature upon auxin yield during extraction of tobacco 

 ovaries witfi wet ether (Wildman and Mnir, 1949). 



Wildman and Muir have shown that half-hour extractions carried 

 on successively for two hours can effectively extract free indoleacetic 

 acid from lyophilized tobacco ovary tissue. The technique may be 

 summarized as follows: 



1. Plunge plant material into liquid air or dry ice and acetone 

 for rapid freezing. 



2. Dry by lyophilization. 



3. Grind to 40 mesh in Wiley mill. 



4. Store in vacuo over P2O5, in darkness, until use. 



5. Extract with peroxide-free ether (95%) at 0° C for four half- 

 hour intervals. 



6. Combine ether extracts and reduce in volume to a few ml. 



7. Transfer quantitatively to agar for Avena assay. 



Some plant materials require neither boiling nor lyophilization 

 for solvent extraction of free auxin. Van Overbeek et al (1945) have 

 established a technique for obtaining free auxin using short term 

 extraction. Free auxin will be obtainable in higher concentrations in 

 the first few ether extractions, whereas enzymatically released auxin 

 will continue to appear over a period of time. These workers have 



