VITAMIN C 219 



for decolorizing iodine was not observed after decitrated lemon juice 

 had been heated anaerobically at 58° or 85° C. 



These observations suggested the possibility that the decrease in 

 stability of vitamin C produced by autoclaving is due to the formation 

 of a substance which hastens the destruction of the vitamin rather 

 than to the destruction of a thermolabile stabilizing factor. This sug- 

 gestion received confirmation in the decreased antiscorbutic properties 

 and capacity to reduce phenolindophenol of autoclaved decitrated lemon 

 juice when mixed with unheated juice and stored for 7 days. The 

 addition of quinol to decitrated lemon juice had the same effect. 



The use of butyl alcohol to remove the yellow coloring matter and 

 wax from lemon juice, in the process of preparing vitamin C con- 

 centrate has been introduced by Grettie and King (1929). Their 

 method, based in other respects upon the procedure of Zilva with 

 modifications, is essentially as follows : 



The juice is decitrated with basic lead carbonate instead of with chalk as in 

 the Zilva method. After stirring slowly until the evolution of carbon dioxide has 

 ceased, 2 or 3 hours, and cooling, the crystalline precipitate is filtered oflf and the 

 filtrate, without concentration or alcohol treatment as in the Zilva method, is 

 fermented with yeast. This required, in their experiments, about 24 hours, but 

 could be hastened by adding a few drops of 20 per cent phosphoric acid and 

 centrifuging before the yeast was added. After the fermentation is complete, the 

 yeast is centrifuged out and 17 cubic centimeters of a saturated solution of neutral 

 lead acetate added to each 100 cubic centimeters of the decitrated juice. The white 

 inactive precipitate is removed by centrifuging, the filtrate brought to a pH of 

 7.2 to 7.4 with dilute ammonia, and the active precipitate removed by centrifug- 

 ing. This is dissolved in dilute acetic acid made up to half the original volume 

 with water and the precipitation with ammonia repeated. The second precipitate 

 is treated with dilute hydrochloric acid in suflficient excess to convert the yellow 

 lead precipitate to white lead chloride and is then extracted three times with 10 

 cubic centimeters portions of butyl alcohol which precipitates the waxy matter. 

 Sufficient ethyl alcohol is then added to bring the alcohol concentration to 90 per 

 cent, the precipitated lead chloride is removed, and the filtrate is evaporated to 

 dryness and made up to one-fifth the original volume of the lemon juice with 

 water containing traces of acetic acid. 



The extract thus prepared was found to retain practically all of 

 the antiscorbutic value of the original lemon juice. In an attempt to 

 purify it still further acetone was added to a concentrated water solu- 

 tion of the extract. By precipitating the ammonium chloride present 

 this reduced the total solids to from 0.5 to 1 milligram per cubic centi- 

 meter of lemon juice equivalent. The filtrate from this precipitation 

 was carefully dried in vacuo and the dried material extracted with 50 

 cubic centimeters of absolute acetone for 24 hours. Feeding tests showed 

 that the active material is definitely soluble in absolute acetone. Analyses 

 of the extract gave a content of 0.38 to 0.6 milligram of the total 

 solids per cubic centimeter of lemon juice. Total nitrogen constituted 



