Fluorometric Determination of Selenium in Effluent Streams with 2,3-Diaminonaphthalene James A. Raihle Analytical Research Department, Abbott Laboratories, North Chicago, IL 60064
Selenious acid reacts with 2,3-diaminonaphthalene in acid solution to form the strongly fluorescent naphtho-[2,3-6]-2selena-l,3-diazole. Elemental selenium is converted to selenious acid by the action of a bromine-bromide redox buffer prior to the addition of the 2,3-diaminonaphthalene. Analyses of elemental selenium show a conversion equivalent to 2500 ppb selenium in effluent streams. Selenates are not reduced to selenites under the conditions of this analysis, hence the method is specific only for elemental selenium and selenium in the four-valence state. Common substances, particularly nitrite, have been found to be noninterfering at a level of 0.05mM. Plots of fluorescent intensity vs. concentration in the region of 0.005 to 0.2 pg selenium(1V) are linear and practically free from reagent interference at an excitation frequency of 366 nm and a fluorescence emission maximum of 522 nm in both standard solutions and spiked effluent samples.
M
ore stringent controls on trace levels of toxic elements released into freshwater supplies are rendering classical methods obsolete. The current level of selenium tolerated in Illinois is 10 parts per billion (ppb). Most classical methods are ineffective at this level owing to limitations on the volume of sample, time of analysis, or removal of interfering substances. Cheng’s (1956) colorimetric determination of microgram amounts of selenium with 3,3’-diaminobenzidine can be adapted to fluorescent measurement but has neither the fluorescent intensity nor the extractability into organic solvents from an acidic media as the reaction of selenium with 2,3-diaminonaphthalene. X-ray fluorescence methods involving addition of metals to pyrrolidine dithiocarbamate (Marcie, 1967) or coprecipitation (Luke, 1968) were abandoned owing to the large amounts of interfering residues obtained from the effluent streams. Modern analytical techniques also have their inherent drawbacks. The minimum level detectable by atomic absorption has been reported as 50 ppb (Atomic Absorption Newsletter, 1968). According to Gulf General Atomic Inc., neutron activation analysis at the 10-ppb level requires an extended irradiation time (several days) (Kovar, 1970). A delay of at least four weeks is encountered in receiving data, since the samples must be analyzed at an outside laboratory. The formation of the piazselenol, naphtho-[2,3-d]-2-~elena1,3-diazole, by the reaction of 2,3-diaminonaphthalene with selenium(1V) is applicable to unfiltered effluent samples down to the 0.5-ppb level (Parker and Harvey, 1962). Shimoishi and Toei (1970) reported that elemental selenium in concentrated sulfuric acid could be quantitatively oxidized to selenium(1V) with a bromine-bromide redox buffer. This technique was successfully incorporated into the procedure of Parker and Harvey to oxidize and detect selenium metal. Experimental Reagents. All chemicals were reagent grade and except for the 2,3-diaminonaphthalenewere used as received. STOCKBROMINE-BROMIDE REDOXBUFFER SOLUTION. (0.1M bromine-0.2M bromide) Weigh 45.7 grams of saturated
bromine water and 2.38 grams of potassium bromide into a 100-ml volumetric flask, dilute to volume with distilled water. Working buffer solution prepared immediately before use by a 1 :100 dilution with distilled water. 2,3-DIAMINONAPHTHALENE. This reagent is prepared daily using the procedure of Parker and Harvey. Fifty milligrams of 2,3-diaminonaphthalene (Aldrich Chemical No. 13,653-0) is placed in a 100-ml volumetric flask and dissolved in 50 ml of 0.2N HCI. The solution is placed in a 5OoC bath for 20 min, transferred to a 125-ml separatory funnel, allowed to cool, and then is extracted twice with 10-ml portions of cyclohexane to remove fluorescent impurities. Apparatus. An Aminco-Bowman Spectrophotofluorometer equipped with 1-mm slits on exit and entrance ports is used with the slit at the photocell adjusted to 2 mm. One centimeter square silica cells are used. Procedure
Pipet 10.0 ml of effluent to a 125-ml erlenmeyer flask containing 10 ml of 0.2N hydrochloric acid and 3.0 ml of the millimolar bromine-bromide redox buffer at 50OC. Cover the flask with a 50-ml beaker and maintain at 5OoC for 30 min. Add 5.0 ml of the purified 2,3-diaminonaphthalene reagent, mix, and maintain at 5OoC for an additional 30 min. Transfer to a 125-ml separatory funnel and allow to cool to room temperature. Extract twice with 5-ml portions of spectrographic grade cyclohexane (Eastman S702) collecting the cyclohexane in a second separatory funnel. Each extraction should be shaken gently for 60 sec and allowed to separate. Extract the cyclohexane twice with 25-ml portions of 0.2NHC1 (room temperature), and collect the cyclohexane in a 25-ml volumetric flask. Dilute to volume with cyclohexane. Concomitantly prepare a reagent blank and a standard of sodium selenite equivalent to 10 ppb selenium. Determine the fluorescent intensity of each solution at 522 nm when excited at 366 nm on the Aminco-Bowman spectrophotofluorometer. Adjust the sensitivity of the instrument so the fluorescent intensity of the standard is approximately 50 %. The fluorescent intensity of the standard solution should be approximately 25 times that of the blank. If significantly less, the assay should be repeated. Results and Discussion
Plots of fluorescent intensity vs. concentration between 0.005 and 0.20 pg selenite obey Beer’s Law and are practically free from reagent interference in both standard solutions and spiked effluent samples. The re1 std dev in this range is 10% and is adequate for routine analysis. Effluent samples were spiked with 0.020, 0.050, and 0.100 pg selenite. The recoveries, 0.022, 0.053, and 0.095, were all within the relative standard deviation. Except for purification of the 2,3-diaminonaphthalene reagent, no unusual precautions were necessary. Parker and Harvey (1962) suggested that the analysis be performed in the absence of light. Laboratory experiments have shown that there is essentially no difference when the reaction and extraction are carried out in the normal laboratory environment and the fluorescence determined within 4 hr. The extracted cyclohexane containing the piazselenol can be stored in the dark and read within 24 hr without affecting the end result. Volume 6, Number 7, July 1972 621
The normal composition of the efluent samples at Abbott Laboratories is within the maximum tolerated impurity levels as listed in the Illlnois State Standards (1968). These tolerances are partially summarized in Table I and do not interfere in the assay. Wiersma (1970) employed 2,3-diaminonaphthalene as a fluorometric reagent for nitrite ion and in addition performed
Table I. Illinois Effluent Standards Constituenta Standard 2 . 5 ppm Ammonia nitrogen 1 . 0 ppm Arsenic 40 mg/l. BOD 0.05 ppm Chromium (VI) 1 . 0 ppm Chromium (111) 0.025 ppm Cyanide 0 . 5 ppb Mercury Nitrate 45 PPm 6-1 0 PH 0 . 2 ppm Phenol Selenium 10 PPb 45 mg/l. Total suspended solids 750 mg/l. Total dissolved solids Illinois Sanitary Water Board.
a
0
z
z
Acknowledgment
Table 11. Effect of Foreign Ions on the Fluorometric Determination of Selenite Ion added” bg Selenite found* None Nitrite Al(II1) Ca(I1) Cd(I1) Cr(II1) Cu(I1) Fe(II1) Ni(I1) Pb(I1) Sn(I1) Zn(I1)
a detailed study into the effects of foreign ion interferences. He reported that Se(IV), Cu(II), Al(III), Bi(TII), Ni(II), Cr(111) and Sn(I1) at the 0.05mM level induced errors of greater than 25 a t the 0.5 pg level of nitrite ion. The effect of foreign ions, in particular, nitrite, on the selenium assay was found to be negligible. Table I1 indicates only aluminum(II1) with an error of 15 was greater than the relative standard deviation. The addition of the bromine-bromide oxidation step oxidizes selenium metal to selenious acid. The maximum level of conversion from selenium metal to selenious acid has been established at 25 pg (2500 ppb). The buffer does not interfere in the fluorescent assay as the fluorescent excitation and emission curves for equal amounts of the piazselenol of sodium selenite determined both with and without the addition of the redox buffer in the initial reaction step are quantitatively identical. The redox buffer does not further oxidize the selenious acid to selenic acid nor does it reduce selenates to selenites. An equivalent of 10 ppb selenate was processed according to the procedure without any resultant fluorescent maximum at 522 nm. The procedure is sensitive to selenium(0) and selenium(1V). If it is desirable to determine all forms of selenium, the digestion method of Watkinson (1966) for the determination of selenium in biological fluids should be applied to the effluent samples.
0.048 0.051 0.055 0.048 0.048 0.048 0.051 0.052 0.052 0.053 0.049 0.046
0.05mM foreign ion added to each sample.
* Theoretical 0.048 pg selenite.
The author is indebted to L. T. Sennello, H. Stelmach, and D. C. Wimer of Abbott Laboratories for their consultation on the development of this technique. Literature Cited Atomic Absorption Newsletter, 7 , 5 (1968). Cheng, K. L., Anal. Chem., 28,1738 (1956). Illinois Sanitary Water Board, Illinois Department of Public Health, Sewage and Industrial Waste Treatment, Requirements and Effluent Criteria, Technical Release T R 20-22, 2nd ed. (revised April 1,1968). Kovar, Lawrence E., Gulf General Atomic, San Diego, Calif., personal communication, 1970. Luke, C. L., Anal, Chim. Acta., 41,237 (1968). Marcie, F. J., ENVIRON. SCI.TECHNOL., 1,164 (1967). Parker, C. A,, Harvey, L. G., Analyst, 87,558 (1962). Shimoishi, Y . ,TGei, K., Talanta, 17,165 (1970). Watkinson, J. H., Anal. Chem., 38,92 (1966). Wiersma, J. H., Anal. Lett., 3,123 (1970). Received for review November 16, 1970. Accepted February 7, 1972.
Ammonia-Nitrogen Removal by Breakpoint Chlorination Thomas A. Pressley,’ Dolloff F. Bishop, and Stephanie G . Roan US. Department of the Interior, Federal Water Quality Administration, Advanced Waste Treatment Research Laboratory, Robert A. Taft Water Research Center, Cincinnati, OH 45268
T
he chief nitrogenous pollutants in municipal waste waters have been categorized (Sawyer and McCarty, 1967; “Standard Methods for the Examination of Water and Wastewater,” 1965) into three main groups: ammonia nitrogen, organic nitrogen, and nitrite and nitrate nitrogen.
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To whom correspondence should be addressed.
622 Environmental Science & Technology
Ammonia nitrogen in waste water is formed by the enzymatic breakdown of urea, proteins, and other nitrogen-containing materials. Most of the organic nitrogen in waste waters is in the form of amino acids, polypeptides, and proteins. Little nitrite and nitrate nitrogen is present in waste waters unless biological oxidation of ammonia to nitrite or nitrate occurs or unless the nitrite and nitrate nitrogen is introduced in industrial or agricultural discharges.
