222 BELSER [CHAP. 9 



system will be used to illustrate the basic concepts of the technique. One 

 organism which has been employed widely in sea-water bioassay is E. gracilis. 

 This alga will not grow in a medium devoid of vitamin B12 (Provasoli and 

 Pinter, 1953). If vitamin B12 is added to the basal medium, the alga grows, 

 and the amount of growth is directly proportional to the quantity of vitamin 

 B12 added to the medium within certain concentration limits. 



Thus, if one wishes to test an unknown sample of sea-water for the presence 

 of vitamin B12, it is a simple matter to add an aliquot of the sample to the 

 basal medium, inoculate with Euglena and wait until terminal growth is reached. 

 If growth occurs, B12 is present, and the amount of growth (i.e. increase in 

 biomass) will be proportional to the amount of B12 present. 



With slight modifications in the method and using a variety of organisms 

 as the reagents, it is presently possible to assay for a large number of organic 

 compounds. In the following sections, these organic compounds will be con- 

 sidered in their natural groupings ; the organisms involved, the sensitivity with 

 which they can be detected, and any special conditions or techniques required 

 to detect them, will be discussed. 



2. Vitamins 



The group of organic compounds which have been searched for most diligently 

 by biologists, using the bioassay technique in sea-water samples, is the vitamins. 

 Certain of the vitamins have been implicated in the growth of marine algae 

 (Lewin, 1959) and, for this reason, have been screened for much more stringently 

 than most other compounds. 



One vitamin which has occupied the attention of a great many investigators 

 is vitamin Bi 2 . This compound is highly potent biologically in millimicrogram 

 quantities, and the assays for it are extremely sensitive (Hutner et al., 1958). 

 Sea-water is being routinely assayed for vitamin B12 in Great Britain (Daisley 

 and Fisher, 1958; Droop, 1955), the United States (Provosali and Pinter, 

 1953), Japan (Kashiwada, Kakimoto and Kanazawa, 1959) and Norway (Larsen 

 and Haug, 1959). The assay for vitamin B12 is a rather complicated one, 

 inasmuch as vitamin B12 activity resides in quite a number of similar chemical 

 compounds. These analogues have been characterized chemically and bio- 

 logically in some cases, and only biologically in others. In Table I the vitamin 

 activity of the various compounds is characterized biologically. 



The type of assay used will be largely determined by whether one is interested 

 in vitamin B12 itself (cyanocobalamin) or in vitamin B12 activity, since many 

 of the analogues may be used by organisms in place of the vitamin itself. It has 

 been concluded by a number of people (Daisley, 1959 ; Droop, 1955) in the field 

 that a good bioassay for this vitamin should include a number of organisms, so 

 that full information regarding the vitamin B12 activity of a sample may be 

 obtained. Table II lists some of the organisms which are currently used, and 



