PRELIMINARY SCREENING METHODS 



All compounds were tested at an initial concentration of 5 p. p. m. Tests were conducted for a 

 24-hour period at a water temperature of 55 deg. F. 



Observations on the toxic effect of compounds were made on larvae of the sea lamprey 

 (Petromyzon marinus) and also on rainbow trout (Salmo gairdnerii) and bluegill sunfish ( Lepomis macrochi - 

 rus) . Larval lampreys varied from 3 to 5 inches in total length. Test fishes were of fingerling size, 4 inches 

 or slightly less in length; every effort was made to keep size variation at a minimum. Larval lampreys were 

 collected by means of an electric shocker in the Ocqueoc River, Presque Isle County, Michigan, and were 

 held in running water in aquaria and small "races" under conditions which simulated their natural stream 

 habitat. Test fishes were obtained from the stocks of local State and Federal fish hatcheries and were held 

 in large raceways. These specimens were maintained in the best possible physical condition until used in 

 the laboratory. 



Generally, two specimens of each of the three species were used in each test. Due to periodic 

 difficulties in obtaining supplies of bluegills and rainbow trout, some compounds were tested using lamprey 

 larvae and only one of these fishes; others were tested using only the larvae. The aggregate test animals 

 available, usually six in number, were placed together in a 10-liter glass battery jar containing 5 liters of 

 water. These jars were provided with aeration through standard stone air-breakers (at near oxygen saturation, 

 as determined by repeated tests) and were maintained at a constant temperature by immersion in specially- 

 constructed constant temperature troughs. These troughs were modified from a design described by 

 Lagler (1953). Water temperature was maintained within the limits of +1. deg. F. Four such constant 

 temperature units were utilized, each having a capacity of thirteen 10-liter battery jars (Figure 1). Twelve 

 of these test jars (each containing a substance being assayed) were included with one control jar in each 

 trough. Fish and larvae in the control jar were exposed only to the water and physical conditions of the 

 typical test container. 



Water used in all tests was drawn from a supply pumped directly from Hammond Bay of Lake 

 Huron. The suction line intake of this pumping system was located 250 feet offshore at a depth of about 

 9 feet. Water from this source was consistently clear and of a relatively uniform quality. During a typical 

 year while tests were being conducted (November 1953 to December 1954), pH varied from 7.5 to 8. 2, 

 dissolved oxygen from 8. 6 to 13. 7 p. p. m. , and free CO 2 from 5. to 9. p. p. m. Further data on the 

 physical and chemical characteristics of northern Lake Huron water has been presented in a recent report 

 by Ayers, Anderson, Chandler, and Lauff (1956). 



Chemicals were weighed in calibrated weighing bottles to the nearest milligram on a Volland 

 Speedigram balance. Solubility of each compound was then determined in water, acetone, and ethyl 

 alcohol (absolute) . Five cubic centimeters of the indicated solvent was added to each sample. Each 

 concentrated mixture was next added to a predetermined volume of water (as required by actual weight of 

 sample and desired concentration) and agitated with a Power-Stir to produce a more dilute solution. 

 Emulsions or suspensions of insoluble compounds were made with the aid of a Waring blender. These 

 prepared solutions, emulsions, or suspensions were added to the test containers in which the experimental 

 animals had already been placed. The resultant volume in each test jar varied from 5800 to 6200 cc. 



Knowledge of the degree of purity of many chemical samples was not available to us. All 

 samples were therefore treated as "pure" preparations and the solutions at routine test concentrations were 

 made accordingly. The specific content of some formulated materials was known while for others only 



