The tubes containing the extracts 

 and standards were sterilized at 15 

 poxinds pressure for 15 minutes. The 

 tubes were cooled and inoculated 

 aseptically with the organism used for 

 the vitamxn being assayed. After 

 inoculation, the tubes were incubated 

 for 65-70 hours at 37° C. In order to 

 stop bacterial growth at the end of this 

 period, the tubes were heated in an auto- 

 clave until 15 pounds pressure was obtain- 

 ed in the chamber. The acid produced in 

 each tube by the growth of tne bacteria 

 irfas titrated electrometrically with O.IN 

 sodixom hydroxide (figure II-3). 



Results of three to six assays were 

 averaged to obtain the data reported in 

 tables II-l, 2, 3» and 4. 



The analyses for moisture, ash, protein, 

 and fat were made by standard procedures des- 

 cribed by the Association of Official Agricul- 

 tural Chemists (1950). The thiamine content 

 was determined by the method described by 



Figure II-2. Adding water 

 to assay tubes using 

 automatic dispenser. 



the Association of Vitamin 

 Chemists (1947). 



Some of the specicil fish 

 meals used in the feeding tests 

 were prepared in the Seattle 

 Technological Laboratory. 



The methods used for pre- 

 paring the various fish meals 

 were as follows: 



1. Steam-heated, vacutmi- 

 dried meals : The raw material 

 was fed into a Stokes rotary 

 steam-jacketed drier. Steam 

 pressure was applied in the 

 jacket and air was exhausted 

 from the chamber until 



Figure II-3. Titration equipment 

 used to determine the acid pro- 

 duced during the growth period. 



16 



