5 ml. is transferred to a test tube. T o the chloroform extract is added 0.2 ml. of laboratory reagent grade ethanolamine. The tube is shaken, and the intensity of the yellow color which develops is read within 30 minutes in a spectrophotometer at 385 mp. The blank is prepared from a sample of the water obtained before the addition of the compound. The concentration is obtained from a standard curve which is prepared from readings corresponding to 0 to 1.5 p.p.m. of 5,2’-dichloro-4’nitrosalicylicanilide in water. Concentrations u p to 20 p.p.m. of the compound in water are determined by a modification of .the above procedure. A 10-ml. sample of the filtered water is placed in a 6 X 5/8 inch glass-stoppered test tube, 0.5 ml. of 2N sulfuric acid, and 5 ml. of analytical reagent grade chloroform are added. The tube is stoppered and treated as described above. A 3-ml. aliquot of the chloroform extract is mixed with 0.1 ml. of laboratory reagent grade ethanolamine, and the determination is carried out as before. As 5,2’-dichloro-4’-nitrosalicylicanilide is not very soluble in water, and because the compound is supplied as the ethanolamine salt in the Bayer 73 formulation, it was necessary to dissolve the pure compound in excess ethanolamine to prepare the standard solution. The standard solution is prepared by dissolving 5 mg. of pure 5,2’ - dichloro - 4’ nitrosalicylicanilide in 0.2 ml. of ethanolamine with gentle warming on a water bath, then adjusting to a volume of 100 ml. with distilled water. A series of standards is prepared from dilutions of this solution. The standards are read in a spectrophotometer against a blank prepared from distilled water. The samples should be shaken uniformly during the extraction process. Shaking for 30 seconds was sufficient t o extract the compound, nor did
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further shaking increase the amount recovered. The color obtained by adding ethanolamine to the chloroform extract must be examined in the spectrophotometer within 30 minutes as it slowly diminishes in intensity after this time. Table I shows a series of recoveries of 5,2’-dichloro-4’-nitrosalicylicanilide obtained from samples of creek water to which the compound had been added in the laboratory. Method B. A 100-ml. sample of the filtered water t o be examined is placed in a 250-ml. reagent bottle, 5 ml. of saturated sodium bicarbonate solution, 5 ml. of aqueous solution of 0.02y0 safranin 0 (certified dye C.1.841 from Matheson, Coleman and Bell, Ohio), and 10 ml. of analytical reagent grade chloroform are added. The bottle is closed with a glass stopper, shaken vigorously for 30 seconds, and let stand for 5 minutes to separate the layers. The chloroform extract is removed by a capillary pipet and 4 ml. is transferred to a spectrophotometer cell. The color is examined in a spectrophotometer a t 515 mp. The blank is prepared from a sample of the water obtained before the addition of the compound. The concentration is obtained from a standard curve constructed to cover the range of concentrations of 5,2’-dichloro-4’-nitrosalicylicanilide 0 to 1.0 p.p.m. in water. Concentrations 0 to 10 p.p.m. of the compound in water are determined b y a modification of the above procedure. A 5-ml. sample of the water is placed in a 6 X 5 / 8 inch glass-stoppered test tube; 0.5 ml. of saturated sodium bicarbonate solution, 0.5 ml. of aqueous solution of 0 02% safranin 0, and 10 ml. of analytical reagent grade chloroform are added. The tube is stoppered and treated as described above, Pure 5,2‘-dichloro-4’-nitrosalicylicanilide is used in the preparation of the standard curve as described under Method A.
The samples should be shaken uniformly during the extraction process. .4dditional shaking after 30 seconds did not increase the amount of molluscicide recovered. The intensity of the color was esamined in a spectrophotometer a t 15-minute intervals for 2 hours without any variation being found, provided that compensation was made for loss of the solvent by evaporation. Table I lists a series of recoveries of 5,2’-dichloro-4’-nitrosalicylicanilideobtained from samples of creek water to which the compound had been added in the laboratory. ACKNOWLEDGMENT
The author thanks Colin C. Reece for valuable technical assistance. The sample of Bayer 73 was obtained through the courtesy of Farbenfabriken Bayer Aktiengesellschaft, Leverkusen, Germany. LITERATURE CITED
(1) Farbenfabriken Bayer Aktiengesell-
schaft Leverkusen-Bayerwerk Biological Institute. “Leaflet on the Molluscicide Bayer f 3 ’ 7(1959), Mimeographed. (2) Foster, R., Teesdale, C., Poulton, G.
F.,Bull. World Health Organ. 22, 543
(1960). ( 3 ) Gonnert, R., Pflanzenschutz Nachrichten “Buyer” 15 (I), 4 (1962). ( 4 ) Haskins, W. T., ANAL. CHEM. 23, 1672 (1951).
KEITHJ. FARRINGTON Division of Animal Health Commonwealth Scientific and Industrial Research Organization McMaster Laboratory, Glebe, N.S.W. Australia
Interference in Determination of Lead by Dithizone Extraction SIR: Recently serious interference was experienced b y this laboratory with the determination of lead in blood employing the method of Cholak, Hubbard, and Burkey (I, 2). As a result, a quantitative extraction of lead could not be accomplished. T o determine the cause of this interference, the reagents were individually replaced until it r a s determined that the difficulty arose from use of a commercially prepared O.O2Y0 solution of phenol red (Fisher 5-985 M), a supply of which had recently been received b y this laboratory for use in the lead procedure. Additional batches of this phenol red were tested and produced the same interference. Tests utilizing some of the other
commercially prepared phenol red solutions resulted in no loss of lead. Correspondence with the manufacturer (3) (Fisher Scientific Co.) revealed that a chelating agent is included in the preparation of this particular indicator solution, and therefore it is not suitable for analysis of lead by dithisone extraction. Two drops of this phenol red will remove as much as 4oy0 of the lead in the sample when the citrate-cyanide-hydroxylamine solution containing the indicator is acidified and then returned to a p H of 8.5 prior to extraction. Because this procedure is used widely for the analysis of lead in air and biological materials, we call this experience
to the attention of analytical chemists who may purchase phenol red in solution form. LITERATURE CITED
(1) American Public Health Assoc., Inc., “Methods for Determining Lead in Air and in Biological Materials,” p. 36, 1955. (2) Cholak, J., Hubbard, D. M., Burkey, R. E., ANAL.CHEM.20,671 (1948). (3) Murray, P., Fisher Scientific Go., Fair Lawn, N. J., personal comrnunication, May 1962.
KENNETH M. HALLaM Division of Chemistry Bureau of Laboratories State of Maryland Department of Healtk Baltimore, Md. VOL. 34, NO. 10, SEPTEMBER 1962
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