Vitamin pool 2(v-2) 



pyridoxal phosphate 



pyridoxamine dihydrochloride 



cyanocobalamin 



vitamin A 



acetylcholine chloride 



ascorbic acid 



carotene 



nicotinic acid 



Vitamin pool 3 (V-3) 



calciferol 



tocopherol 

 rutin 

 menadione 



ug/lOO ml of sample 



5.0 



5.0 



0.015 

 10.0 

 10.0 

 10.0 

 10.0 



50.0 



ug/100 ml. of sample 



10.0 

 10.0 

 10.0 

 10.0 



The PI metal stock solution (Provasoli et al, 

 1957) furnished micronutrients and EDTA in 

 the following final concentrations: 



Constituent mg/lOO ml of sample 



These vitamin pools vere prepared in 50°/ o 

 ethyl alcohol and added in 0.1-ml. amounts 

 per 100 ml of sample sea water. Pools of 

 purine and pyrimidines (pp) (adenine, 

 adenosine, adenylic acid, guanine, guanosine, 

 uracil, cytidylic acid thymine, xanthine, and 

 hypoxanthine ) , essential amino acids (EAA) 

 (valine, isoleucine, leucine, threonine, phen- 

 ylalanine, trytophane, lysine, arginine, his- 

 tidine, and methionine), and nonessential amino 

 acids (NKAA) (glutamic acid, aspartic acid, 

 serine, proline, cystine, glycine, alanine, and 

 tyrosine) were made up in double distilled water 

 in one-mg. amounts per 100 ml. of sample (final 

 concentration) . The purine and pyrimidine pool 

 as well as the nonessential amino acid pool was 

 sterilized in the autoclave, whereas the 

 essential amino acid pool was passed through an 

 ultrafine Morton s inter ed-glass filter. 



Organic complexes were added to some samples 

 to give the following final concentrations: 

 soil extract (from Scripps garden soil) 

 (Sweeney, 1951) 1 ml/100 ml of sample; yeast 

 extract (Difco) 0.001 g/100 ml. 



Inorganic substances were added to give the 

 following final concentrations: 

 KHO3...IO ugm at N0o/L of sample j KgHP01f...l ugm 

 at POl^/L of sample} PI Metals... 3 ml/100 ml of 

 sample . 



Na 2 EDTA 



Fe 



B 



Mn 



Zn 



Cu 



Co 



3-0 



0.03 



0.6 



0.12 



0.015 



0.00012 



0.0003 



The 250-ml. glass-stoppered reagent bottles 

 were incubated in a water bath which was 

 illuminated (fluorescent lighting from a 

 battery of long bulbs at an illuminance of 

 1250 1 150 foot-candles) through a glass 

 bottom. The temperature of the water bath 

 was maintained as close to the temperature of 

 the surface sea water as possible. Black 

 bottles (prepared by careful covering with 

 black masking tape) were incubated with the 

 light bottles as controls to distinguish 

 dark uptake of C 1 ^ from photosynthetic fixa- 

 tion. After the bottles were incubated in 

 the presence of added nutrients for approxi- 

 mately four hours, the NaHC-'- 0o was added 

 to the water samples from a sterile ampule 

 and the samples were incubated for two to 

 four additional hours. 



At the end of the incubation period, equal 

 volumes of water (2.0 ml) were removed from 

 each sample bottle after shaking and plated 

 in duplicate, as described above, to determine 

 the numbers of heterotrophic marine bacteria. 

 In some experiments the zero-hour count was 

 subtracted from the final bacterial popula- 

 tion to give the bacterial increase. 



The water remaining in each bottle was filter- 

 ed through an BA Millipore filter (0.1*5 t 

 0.02 microns, Millipore Filter Corporation, 

 Watertown, Mass . ) which retained all of the 

 larger particulate matter; the filter wa6 

 then washed with more than 100 ml of sea water 

 and dried in a desiccator for at least 21*- 

 hours over silica gel. The radioactivity on 

 the filter pad was measured in a proportional 

 flow counter (Nuclear Measurement Corporation, 

 PC-1). 



