474 Prof. A. B. Macallum. Detection and Localisation 



tions to remain a long time in this fluid for this purpose. I have, as 

 already stated, found that the long continued action of the reagent 

 has this result, and that the conversion is more marked if the reagent 

 is allowed to act at a slightly increased temperature. One cannot be 

 absolutely certain that the anhydrous forms of phosphoric acid when 

 liberated, and before being converted into orthophosphoric acid, da 

 not diffuse through the tissue elements, but in a number of experi- 

 ments made to decide this point, I ascertained that if such diffusion 

 did occur, it was in such minute amounts as to be unobservable. A 

 risk of diffusion is incurred when a tissue, very rich in orthophos- 

 phates, is acted on by the reagent. A part of the phosphoric acid 

 in this case, except in very thin sections, diffuses and forms a slight 

 deposit of phospho-molybdate crystals on the preparation. Prepara- 

 tions of renal tubules and the cat's placenta illustrate this well. 



Owing to the abundance and general distribution of lecithin in 

 animal and vegetable tissues, it is necessary to extract this com- 

 pound from them in order to ascertain the distribution of the other 

 phosphorus compounds. Bitto* has shown that the extraction can 

 be regarded as complete only when the material, first treated with 

 ether, has been acted on by boiling ethyl alcohol thirty times, each 

 period of extraction lasting about ten minutes. Adopting this pro- 

 cess, I subjected samples from all the material used to extraction in 

 a Soxhlet apparatus for five hours, the condensed but still hot alcohol 

 being siphoned off every 6 10 minutes. This treatment is specially 

 necessary in the case of nerve tissues in which it makes a marked 

 difference in the phospho-molybdate reaction. 



A much more difficult problem is that of the removal of the in- 

 organic phosphates from tissues. Jollyf used acetic acid of 20 per 

 cent, strength for this purpose, claiming that this fluid removes all 

 the phosphates except that of iron. It does indeed remove a large 

 part of them, but not those which may be in the nuclear elements. 

 In order, therefore, not to confuse the inorganic phosphorus with 

 that of organic combinations, I have always endeavoured to de- 

 termine in any given material what extent of molybdo-phosphate 

 reaction may be obtained in the first ten minutes after the nitric- 

 molybdate reagent is added. This reaction indicates whether the 

 tissues are rich or poor in inorganic phosphates, and it may be com- 

 pared with what may be obtained after a longer stay in the reagent, 

 any enhancement in the reaction thus demonstrating the phosphorus 

 of organic compounds. 



* " Ueber die Bestimmung des Lecithingehaltes der Pflanzenbestandtheile,' 

 ' Zeit. fur Physiol. Chemie,' vol. 19, p. 488, 1894. 

 f ' Comptes Keudus,' vol. 125, p. 538, 1897. 



