ASCORBIC ACID IN TOMATOES 17 



Preparation of sample from tomato juice. 



1. Place 25 cc. of tomato juice in each centrifuge tube. Add 10 cc. of acid 

 mixture to each sample. 



2. Centrifuge at 1800 revolutions per minute for 15 minutes. 



3. Decant serum into a 200 cc. volumetric flask. 



4. Cover residue in centrifuge tubes with acid mixture and extract a second 

 time by centrifuging for same period of time and speed. 



5. Decant serum into previous extraction. Make up to 200 cc. volume with 

 acid mixture. 



6. Titrate 25 cc. aliquot portions of this 200 cc. volume with the 2, 6-di- 

 chlorophenolindophenol dye. The pink end point should persist for one 

 minute. 



7. Make three separate titrations. 



Example oj Calculation 



Amount of tomato juice — 25 cc. 



Amount of tomato juice in 25 cc. aliquot — 3.13 cc. 



0.68 cc. indicator = 3.13 cc. tomato juice. 



1 cc. indicator = 4.60 cc. tomato juice. 



4.60 cc. tomato juice =0.445 mg. ascorbic acid (from standardization of 



the indicator). 

 1 cc. tomato juice =0.096 mg. ascorbic acid. 



COMPARISON OF DYE, MODIFIED IODINE, AND BIOASSAY 

 METHOD FOR DETERMINING ASCORBIC ACID 



Tests were made to show how close a correlation existed between the chemical 

 titration methods and the more time-consuming animal-feeding method. 



The contents of two cases of 14-ounce cans of a commercial brand of tomato 

 juice were thoroughly mixed in one batch, heated to 170° F., sealed in No. 1 

 tin containers and sterilized in boiling water for 15 minutes. As it took no more 

 than ten minutes to seal the entire lot of cans, they were considered uniform in 

 every particular. Slight differences in headspace and temperature may have 

 occurred. Three guinea pigs were placed on each feeding level of 4.50, 4.75, 

 5.00, 5.25, 5.50, 5.75, 6.00, 6.50, 7.00, 7.50, and 8.00 cc. daily. With the three 

 negative controls and the three positive controls, this made a total of 39 animals 

 on the experiment. Daily titrations by both the iodine and the dye methods 

 were made on the identical cans from which the animals were fed. Tomato juice 

 supplemented the basal diet of Sherman, La Mer, and Campbell (1922) over the 

 90-day feeding period. The results of the chemical titrations and the animal 

 feedings are reported in Tables 2 and 3. The pertinent animal data are reported 

 in Figure 1. 



The results show that the daily protective level for guinea pigs on this recanned 

 tomato juice was 6.7 cc. Unpublished data in this laboratory show that 0.6 

 mg. of pure ascorbic acid is the daily protective level of a young, growing 300 to 

 400 gram guinea pig. Calculating from this, it is seen that this tomato juice 

 contained 0.09 milligrams of ascorbic acid per cc. or 51 units per ounce. By the 

 2, 6-dichlorophenolindophenol chemical method, this sample of tomato juice 

 was shown to contain 51 units per ounce. The modified iodine method gave 

 74 units per ounce. 



This experiment shows that the chemical method, as described in detail, is 

 reliable for determining ascorbic acid. The 2, 6-dichlorophenolindophenol dye 



