Determination of Anionic Detergents in Surface Waters and Sewage with Methyl Green W. ALLAN MOORE and RAYMOND A. KOLBESON Robert A. raft Sanitary Engineering Center, Public Health Service, U. S. Department o f Health, Education, and W e l f a r e , Cincinnati, O h i o
up every 4 weeks. Prepare a n intermediate solution, which should be made up a t least every 2 weeks by diluting 100 ml. of the stock solution to 1 liter. Dilute 50 ml. of this intermediate solution t o 1 liter; 1 ml. contains 5 y of sodium lauryl sulfate From this standard solution set up a series of concentrations from 5 to 100 -;, dilute to 20 ml., and form the complex according t o the procedure given below. Read the absorbance of these standards a t 615 mp against a blank containing all reagents except sodium laur t.1 sulfate.
The methyl green procedure was designed to eliminate or low-erinterferences encountered in the determination of anionic detergents by the methylene blue method. Interferences from thiocyanate, nitrate, and nitrite ions were reduced greatly. The methyl green procedure is not affected by high concentration of sulfate, chloride, sulfite, and phosphate. Recoveries of alkyl sulfate added to river and lake waters as well as to domestic sewage w-ere more consistent and more accurate when the methyl green procedure was used.
Analysis of Samples. .4 sewage sample generally requires a volumetric dilution prior t o the determination, owing to its high detergent content. To the sample (20 mi. or aliquot diluted to 20 ml.) in a 125ml. se aratory funnel add the required buffer (for most samples 10 mf is usually sufficient) and 2 ml. of 0.5% methyl green solution. Swirl the funnel t o ensure complete contact of dye with sample. -4dd 40 ml. of benzene (accurately measured) and shake for 1 minute, or approximately 250 times. Allow to settle, then sn irl t o dissolve the complex completely in the benzene. Remove the water layer with care, so as not to lose any of the benzene la) er. Add 15 ml. of distilled water buffered at pH 2.5 to the benzene layer and shake 75 t o 100 times. Allow to settle, then swirl the funnel. After allowing water t o settle out of the benzene layer (20 t o 30 minutes), remove a sample with a clean, dry medicine dropper, and determine the absorbance a t 615 mM. Insertion of benzene into the cells by dropper usually introduces some air into the solvent, which escapes after about 1 minute. Read the concentration of detergent in terms of sodium lauryl sulfate from the standard curve. Wash all glassware in acid to remove any complex adhermg to it. Both the complex and dye are very soluble in chloroform and are removed from spectrophotometer cells by a chloroform washing after each reading. This is necessary because the complex and dye adhere readily to glass.
T
H E literature shows that anionic detergents represent 90% of all detergents used. Several methods (1,2,5,9,12,13,15, 1 6 ) have been employed in the quantitative estimation of anionic detergents in sen age, but the Jones method ( 1 1 )with its modifications ( 3 , 4,6-8, 14) appears t o be the predominantly popular procedure. Hon-ever, interference of thiocyanates, nitrates, nitrites, protein, and compounds found in urine causes inaccuracy. Thiq paper presents a method which is more rapid and less affected by these interfering substances. It is designed only for anionic detergents and uses the principle of complexine the detergent with methyl green dye, extracting the complex with benzene, and measuring the absorbance of the benzene solution for comparison against a standard curve previously prepared. Sulfonated or sulfated surface active agents react a i t h methyl green t o form the same purple complex. hIethyl green was chosen from a group of 13 dyes, which were all tested for complex formation with detergents. Meldola blue, Capri blue, Alizarin green, Alizarin blue, patent blue, crystal violet, Victoria blue, and Azo carmine G proved unsatisfactory because they did not form the complex, were soluble in the extracting solvents, or were subject to interference from thiocyanate ions. The methylene blue method ( 1 1 ) as used in this study is essentially the modified Jones method of Faust ( 7 ) .
DI SCUSSIOR'
Standard. The anionic detergents found in sewage vary in molecular weight, from approximately 250 to 350. Several available compounds were considered for a standard, but sodium lauryl sulfate with a molecular weight of 288 was chosen because it is procurable in pure form and is readily stable in 100 p.p.m. solution for 3 to 4 reeks. Solutions containing 100 p,p.m. of sodium lauryl sulfate and a specific preservative were tested for a period of 6 weeks. The preservatives used were 500, 1000, and 2000 p.p.m. of sodium borate, 1 p . p m of copper, G p.p.m of sodium azide, 100 p.p.m. of mercuric chloride, and 4000 p.p.m. of salicylic acid. The mercuric chloride and sslicylic acid showed signs of interference a t the concentration used and Kere discont'inued. A sample not containing preservative v a s used as a control. After all samples had been tested at intervals of 3 days for a month, very little deterioration was observed. Between the fourt'h and sixth week, the sample n-ithout preservative sho\ved a 20% breakdown, whereas the ssmples containing 500 p.p.m. of sodium borate and other preservatives showed a 10% loss of detergent. Questions have arisen as to whether or not the vast number of detergents on the market all form color complexes having maximum absorptions a t the same wave length. Gowdy ( 8 ) states that alkyl sulfates and alkyl aryl sulfonates represent the largest proportion of the synthetic detergents found in s e ~ a g e . On the basis of this statement, absorption curves n.ere determined for an alkyl arj,l sulfonate and sodium laurj-l sulfate. .Zs shown in
APP4R4TUS AND REAGENTS
Apparatus. Any spectrophotometer capable of accommodating 1.0-cm. cells and possessing a wave-length selection a t 615 mg may be used. AiBeckman Model B was used in making all measurements in this study. Pear-shaped separatory funnels (125-ml. ), a 40.0-ml. volumetric pipet, and medicine droppers were also used. Reagents. Methyl Green. Dissolve 0.5 gram of certified methyl green (obtainable from Hartman-Leddon, Go., Philadelphia, Pa., Catalog No. 270) in distilled water and dilute t o 100 ml. Benzene, reagent grade. Chloroform. Glycine-Hydrochloric Acid Buffer (pH 2.5). Dissolve 7.5 grams of glycine and 5.8 grams of sodium chloride in distilled water and dilute to 1 liter. Dilute this solution with approximately 0 . 1 S hydrochloric acid t o produce a p H of 2.5. (The stability of this buffer is increased greatly by adding a few drops of benzene and keeping it refrigerated.) Sodium lauryl sulfate, reagent grade (obtainable from Matheson, Coleman and Bell, Xorwood, Ohio, Catalog No. 7003). PROCEDURE
Standard Curve. Prepare a stock solution of sodium lauryl sulfate containing 1000 p.p.m. by dissolving 1.0 gram in distilled water and diluting t o 1 liter. This stock solution should be made 161
ANALYTICAL CHEMISTRY
162
Fignre 1, maximum absorption occurs a t the same wave length (615 mp) for each compound, COSCESTRATIOS RAXGE. Figure 2 represents a standard curve using concentrations from 0 to 125 y of sodium lauryl sulfate in 20 ml. of distilled water. The curve conforms to Beer’s law to approximately 60 y. Should concentrations lower than 5 y be encountered, the sample size can be increased or the readings taken using a 5.0-em. cell Tvith reference to a standard curve made up for the larger cell. DYE COSCESTRATIOS.The 2 nil. of 0.5% methyl green used for complexing the detergent is necessary in such high concentration because of a chemical phenomenon which has not as yet been investigated. The reaction is not stoichiometric, in the sense that 1 mole of added dye reacts with 1 mole of detergent. Sufficient dye must be added to the sample to develop an equilibrium at which the absorbance readings are constant. This was determined for a maximum permissible detergent concentration of 75 y, to which the method is applicable. ( A sample containing more than this amount of detergent should be diluted.) To solutions containing 75 y of sodium lauryl sulfate, 2 ml.
Table 1. Dye Concentration Kecessarg for Stabilization of Color Concentration of Methyl Green,
%
0.1 0.2 0.3 0.4 0.5
Absorbance 0.140 0.225 0,260 0 270 0.271
of methyl green were added, in concentrations varying from 0.1 to 0.5%. As shown in Table I, the necessary dye concentration is 2 ml. of 0.4% methyl green. I n this study, however, the 0.5% concentration was used to ensure an adequate excess.
SODIUM LAURYL
,300
SULFATE
-
0 w
z
U
m
K Y) 0 m 4
D O D E C Y L B E NZENE
I00
5 84
I 590
I 600 WAVELENGTH
-
SULFONATE
I
I
I
I
610
620
630
635
- MILLIMICRONS
Figure 1. Absorption curves
EXTR.4CTING SOLVENTS. Several solvents were tried for the extraction of the complex, but the solubility of the dye in most of them precluded their use. Only xylene and benzene extracted the complex without dissolving the dye. Benzene proved to be Table 11. Effect of pA on Color Formation the better extractant, because the complex was more soluble in Absorbance PH it and it did not tend to form emulsions with strong sewage sam3.1 0.151 3.0 ples. 0.157 0.171 2.9 EFFECT OF pH. The intensity of the color formation is depend0.157 2.8 2.7 0.157 ent upon the hydrogen ion concentration of the sample, as 2.6 0.167 2.5 0.168 shown in Table 11. The results given in this table show that 0.166 2.4 even though a p H of 2.9 produces maximum color formation, 0.159 2.2 2.1 0.143 less rigid control ie necessary at p H 2.5. To maintain the latter pH, a glycine-hydrochloric acid buffer was used. TIMEFOR COLORDEVELOPXEST. The time required for comTable 111. Effect of Interfering Substances on Methyl plexing the detergent with the methyl green dye is very short. Green and Methylene Blue Methods Several samples containing 50 y of sodium lauryl sulfate, buffer Recovery solution, and dye were allowed to stand for different time interN e t h v l Green Methvlene Blue .\l;tllod hfethod Concentration, interfering vals prior to being extracted. The fact that there was no difSubstance P.P.11. P.p.in. 72 P.p.rn. % ference in the absorbances obtained on immediate extraction, and on extraction after 15 minutes, shows that the reaction between the dye and detergent is rapid. COLORSTABILITY. The absorbance of the benzene-soluble comdex formed with methyl green and a dilute solution of detergent was read a t various Table 11’. Recovery of Detergent from Sewage time intervals to determine if Methyl Green Method Methylene Blue Method color change was occurring Detergent Detergent Detergent Detergent Detergent Detergent with respect to time. The Sainple In added, recovered, 9X In added, recovered. % KO. Sewage p.p.m. p.p.m. recovered Sewage p.p.rn. p.p.in. recovered initial reading was taken after a 25-minute lapse in time to 0.5 0.5 100 0.5 0.53 105 1 18.0 24.7 allow the water to settle out of 2.0 2 01 101 2.0 2.12 106 the benzene layer after it had 0.5 0.5 100 0.5 0.18 36 9 14.1 21.1 been washed. Even after 90 2.0 2 0 100 2.0 1.16 58 minutes, there was no change 3 19.4 1.0 1.05 105 21.5 1.0 1.12 112 in the absorbance of the color .. 2.5 2 4 9fi 2 . 5 Less than blank 1 4.2 8.0 complex. 50 48 96 50 5-1 104 Faust (7) has recommended 0.3 0 27 90 0.3 0.43 143 that the complex formed using 5 3.3 4.0 1.5 1.5 100 1.5 1.67 111 the methylene blue method be read immediately after wash-
V O L U M E 28, NO, 2, F E B R U A R Y 1 9 5 6
163 400
ing, because of its fading properties. ComI I I I I I I I I 1 I plexes formed by both methods using sewage were allowed to stand for a 24-hour period and compared. The fading of the methyl green com300 plex nas negligible as compared to that of the .. niethl lene hlue complex. EXTRACTION OF COMPLEX.Benzene was used w u 4 z for all extractions of the methyl green-detergent complex. The use of this lighter than water 0 solvent makes it difficult to use more than one m 4 zco extraction because of the error involved in transfer. T h e effectiveness of a single extraction is ehonn in Figure 3; 95% of the complex is renioved in a single extraction with benzene. I n thip same figure, a curve is also given for the methj lene blue method; using chloroform as the I I I I I I I solvent, a single extraction removes about 84% $0 60 70 80 90 IO0 110 M I C R O G R A M S OF SODIUM L A U R Y L S U F A T E of the color complex. Five extractions woqld be necessary to remove 95% of the complex, Figure 2. Standard curve for sodium lauryl sulfate which is accomplished with one extraction in the methyl green procedure. EFFECTOF WASHIXGBESZEXEA N D CHLOROFORM. The solvents containing the complex are washed with water to remnvo I I I I I interferences which form water-soluble dye complexes. The nater wash is also used in the methylene hlue method to remove 0 2 excess dye. Samples containing 50 y of sodium lauryl sulfate were run by both methods, The results obtained show that the chloroform extract of the dye complex in the methylene blue method is affected more hy the water washings than is the benzene extract z in the methyl green method, probably because chloroform i+ W W a E approximately 12 times as soluble in water as is benzene. I ~ ~ T REERM O V ~monf L BENZENE.Washing the benzene > J with water causes a small amount of water to he temporarily II W suspended in it. Several types of drying materials nere used to z absorb the suspended water, but all failed because of the affinity W 00 I of the complex for filter paper, cotton, glass wool, and chemical m U drying agents. The water was efficiently removed by alloffing it to settle out for 20 to 30 minutes. 0 m Precision and Accuracy. Ten replicates of sodium lauryl 4 sulfate \
