200 THE CARBOHYDRATES 



4. The middle lamella, which consists of compounds of 

 pectic acid, may be differentiated from the other pectic sub- 

 stances which are mixed with the cellulose of the cell walls by 

 the following method : A thin section is placed in a 20-25 per 

 cent solution of hydrochloric acid in alcohol for twenty-four 

 hours ; the section is then washed with water and treated with 

 methylene-blue or phenosafranin. The middle lamella stains 

 much more deeply than the rest of the wall. 



5. If, after the above treatment with acid alcohol, the 

 section be washed in a 10 per cent solution of ammonia, it is 

 found that the cells separate with ease one from the other. 

 According to Mangin, the combined pectic acid is freed from 

 its bases by the treatment with acid alcohol, and is then 

 dissolved by the ammonia. A recombination of the pectic 

 acid may be brought about by treatment with baryta water, 

 and after this process the cells will not separate one from the 

 other. 



6. Mehta * finds that pectic compounds stain deeply with 

 the following dyes : Alcoholic malachite green, aqueous 

 Congo red, alcoholic eosin, alcoholic safranin. aqueous gossy- 

 pimin, aqueous iodine green, and aqueous ruthenium red. 

 None of the stains, however, is specific for pectic substances ; 

 thus ruthenium red stains oxycelluloses, hemicellulose, gums, 

 galactans, etc. The procedure adopted by Mehta is to dissolve 

 out the various constituents of the cell wall with appropriate 

 reagents and then to compare their staining reactions. The 

 sections are placed in a test tube with the reagent which is 

 then heated for six to eight hours in a boiling water bath, 

 the liquid being decanted off every hour and replaced by fresh 

 reagent. The sections are then washed with hot distilled 

 water, stained for two hours, washed with 90 per cent alcohol 

 to remove excess of stain, dehydrated with absolute alcohol, 

 and mounted in cedar-wood oil. The reagents used were 

 0-5 per cent ammonium oxalate, 0-5 per cent ammonium 

 oxalate in 3 per cent ammonia solution, 4 per cent sodium 

 hydroxide, 3 per cent hydrochloric acid, or 95 per cent 

 alcohol. 



* Mehta : " Biochem. Journ.," 1925, 19, 979. 



