Some Nutritional Relationships Amo7ig Microbes 139 



Production of B vitamins by selected marine bacteria was 

 studied at intervals over a period of twenty-four hours for the 

 purpose of determining whether these substances are excreted 

 during the period of growth. Isolates P9 and P85 from Puerto 

 Rico, and B739 from Long Island Sound, were inoculated into 

 a series of flasks, each containing 10 ml of medium B (see Table 

 1 ) . Growth was allowed to take place at 30 C on a rotary shaker. 

 At different time intervals, the optical density of the growing 

 cultures was determined at wavelength 600m/i, and samples of 

 the culture broths were taken for vitamin assays. An aliquot was 

 centrifuged to remove the bacteria, and the supernatant was 

 collected and heated to 100 C for three minutes. Another portion 

 of tlie whole culture was also heated in the same way, to kill the 

 cells and liberate bound vitamins. Assays on the two portions 

 gave indications of the total and excreted B vitamins present at 

 different times during the growth period. The experiments were 

 repeated on different days, with similar results. Typical data 

 obtained for cultures P85 and B739 are shown in Figures 1 and 

 2. Antibiotic production and synthesis of five B vitamins by P9 

 are presented in Figure 3. 



It became apparent from the experimental data that appre- 

 ciable amounts of B vitamins are synthesized by some cultures 

 of marine bacteria, concurrent with the rise in the growth curve. 

 The vitamins biotin, riboflavin, nicotinic acid, and pantothenate 

 are produced and excreted into the medium during the period of 

 rapid gi'owth. Other vitamins, such as Bi^ and sometimes thia- 

 mine, are produced and retained witliin the cells, but can be re- 

 leased by denaturation of the cell constituents through heating 

 at boiling temperature for a brief period (3-5 min.). The beers 

 of culture P9 were inhibitory to the vitamin B12 assay organism, 

 making this test impossible. The chief difference in the vitamin 

 data obtained with culture P9 and the others, is tliat thiamine 

 was excreted into the medium along with the other \itamins. 



Production of B vitamins during growth of pure cultures of 

 green algae, blue-green algae, and diatoms, isolated from marine 

 muds, has also been studied. Simple demonstrations of vitamin 

 excretion may be obtained by placing portions of an algal culture, 

 growing in vitamin-free ASP-2 agar medium (9), on the surface 



