1222
A N A L Y T I C A L CHEMISTRY
the complexity of the serum system, u-hich contains substances like tryptophan in proteins with which perchloric acid reacts to give a greenish fluorescence, makes i t impossible t o carry out the analysis by the perchloric acid color reagent. When sulfuric acid color reagent is added to a mixture of water and phosphoric acid, deep green solutions result and have different absorption peaks depending on the presence of proteins. The curves obtained for water and gelatin are similar with a peak a t 670 mp, although the intensity of the gelatin solution is a little greater than that of the water solution These are shown in curves A and B of Figure 3. C, D,and E in the same figure were obtained with human albumin alone and human albumin in which bilirubin was added in normal serum (bilirubin content of 0.4 mg. per 100 ml.), and jaundiced serum, respectively. The hyperchromic effect of the proteins in C and D is very marked, for the intensity of color is much greater than the n-ater or gelatin solutions. All of the green chromophores are stable and obey Beer’s law over the range of 0 to 20 mg. per 100 ml. of serum. Additions of bilirubin t o several sera show good recovery when compared to standards prepared by the addition of bilirubin to normal serum which s h o w no bilirubin by diazo reaction (Table 11). Measurements were made a t 600 mp where both sample and standards had similar abmrption peaks.
LITERATURE CITED
Bergh, A. -4.H. van den, and Muller, P., “Handbuch der biologeschen Arbeitsmethoden,” Abt. IV, T1. I, p. 901 Urban & Schww-aenburg,Berlin, 1927. Ehrlich, P., Zentr. Kliniken, 45,721 (1883); 2. anal. Chew?.,22, 301 (1883). Hausser, K. W., et a/., 2. physik. Chem., 29B,363 (1935). Hersfeld, E., Biochem. Z., 251, 394 (1932). Hosley, R. J., A m . J . Clin. Pathol., 19,884 (1949). Jendrassik. L., and Grof, P., Biochem. Z., 297,81 (1938). Lernberg, R., and Legge, J. W., “Hematin Compounds and Bile Pigments,” Sew York, Interscience Publishers, 1939. hIalloy, H. T., and Evelyn, K. A , J . B i d . Chem., 119, 481 (1937). Ibid., 122,597 (1937). Meulengracht, E., Deut. Arch. klin. Med., 132,285 (1920). Newberger, R. A , , J . Lab. Clin. Med., 22, 1192 (1937). Rabinowitch, I. XI.,J . Biol. Chem., 97,163 (1932). Scott, L. D., Brit.J . Ezptl. Pathol., 22,17 (1941). Sheard, C., Baldes, E. J., hlann, F. C., and Bollnian, J. L.. A m . J . Physiol., 76,577 (1926). With, T. K., 2 . physiol. Chem., 278, 120 (1943). Zlatkis, .1.,Zak, B., and Boyle, 8.J., J . Lab. Clin. M e d . , 41. 486 (1953). RECEIVED for review October 27, 1953. Accepted -4pril 3, 1954. Presented before the Division of Riological Chemistry a t t h e 123rd M w t i n g of the A I I E R I C W CHEMICAL SOCIETY, Lou Angeler, Calif.
Determination of Tetraethyllead in Gasoline By Titration with Ethylenediamine Tetraacetate 0.I . MILNER and G. F. SHIPMAN Research a n d D e v e l o p m e n t Department, Socony- Vacuum Laboratories, faulsboro,
S
OME widely used methods for the determination
of tetraethyllead in gasoline are the ASTM method ( I ) , and polarographic ( 2 , 1 4 ) or x-ray (6, 1 1 ) procedures. I n the ASTM procedure, the lead is extracted with concentrated hydrochloric acid and determined gravimetrically as lead chromate. This method is precise and accurate, but is rather long Tlie polarographic procedure, in which lead is first extracted as in the gravimetric method, is equal t o the gravimetric in precision and accuracy, and is far more rapid, but is somewhat specialized for many small control laboratories. The x-ray method eliminates the preliminary extraction and is extremely rapid, but is quite specialized and requires expensive instrumentation; furthermore, the sulfur content of the sample must be knonn, 80 that an appropriate rorrection can be applied. This paper evaluates a simple rapid titration as compared n i t h the lengthy gravimetric method for the determination of the extracted lead Disodium ethylenediaminetetraacetic acid (Versene, E D T d , Sequestrene) is a powerful complexing agent for many metals. I t a a s first used by Schwarxenbach and coworkers and was applied to the titration of magnesium n i t h Eriochrome Black T as an indicator (3, 4,7 , 8, 12, 15). Manns et al. have modified the titration to permit the determination of barium ( I S ) . This proposed procedure parallels earlier work with respect t o the titration. It is based on a technique devised b y Flaschka (9) in which the lead is titrated a t p H 10 in ammoniacal tartrate medium. The indistinct purple-to-blue end point, obtained b y Flaschka, is improved b y adding a known amount of standard magnesium solution. The stability constants of the numerous complexes in the system are such that the color change a t the end point is the same as in the titration of magnesium alone-namely, pink to blue. After the work described in this paper was completed, Grun-
N. 1.
nald ( 1 0 ) published a method in which the same titrant is used; however, the two methods are quite different in other respects. A single determination (including the extraction) can be completed in 1 hour by the method described below. By making several determinations simultaneously the working time per determination can be reduced to considerably lese than l hour. RECOMMENDED METHOD
Preparation of Indicator and Standard Solutions. Eriochrome Black T Indicator. Prepare by grinding 0.2 gram of the dye (Eastman Kodak Co., Rochester, X. Y., P6361) with 100 grams of ammonium chloride to a fineness of 40 to 50 mesh. Store in a tightly stoppered bottle. Standard Lead Solution. Prepare a 0.05.t’ solution from reagent-grade lead nitrate crystals which were crushed and dried a t 105” C. before weighing. Standard Magnesium Chloride Solution. Prepare an approximatelv 0.05N solution from the hexahvdrated salt. To a meas;red portion of the solution add 0.3- gram of ammonium chloride, 3.0 ml. of concentrated ammonium hydroxide, and 75 mg. of prepared indicator. Titrate with 0.055 disodium ethylenediamine tetraacetate solution, and express the strength in terms of volume of tetraacetate equivalent t o 1 ml. of magnesium solution. Standard Disodium Ethylenediamine Tetraacetate Solution. Prepare a 0.055 solution from the dihydrated salt and standardize against the standard lead solution as follows: T o a measured volume of standard lead solution, add 2 grams of tartaric acid, 0.3 gram of ammonium chloride, 7 ml. of concentrated ammonium hydroxide, and 75 mg. of prepared indicator. Pipet exactly 1.00 ml. of standard magnesium solution and titrate with the tetraacetate solution. Calculate the normality after subtracting the volume of tetraacetate solution equivalent t o the magnesium added. (The reagent, disodium Versenate analytical reagent, is available from Bersworth Chemical Co., Framingham, Mass.) Procedure. Proceed as in the ASTM method ( I ) , obtaining the combined acid extract and washings in a 300-ml. Erlenmeyer.
1223,
V O L U M E 26, NO. 7, J U L Y 1 9 5 4 flask. [The extraction method was originally described by Calingaert and Gambrill(6)l. S d d 2 grams of tartaric acid and neutralize with concentrated ammonium hydroxide. As the end point is approached, test with litmus paper after each 1- or 2-ml. addition; approximately 25 ml. will be required. Add 15 ml. more of ammonium hydroxide, 0.3 gram of prepared indicator, and exactly 1.00 ml. of standard magnesium solution. Titrate with the standard ethylenediamine tetraacet at e .solution.
T a b l e I.
D e t e r m i n a t i o n of T e t r a e t h y l l e a d in Gasoline (Results in ml. of TEL/gallon) .4STM D 5 2 6 4 8 T Versene Method Difference (Analyst B) (.lnalyst A) betaeen Averages .4t Found Range Av. Range
r2 2 0 0 3 3 2 2 2 4 4 2 2 2 2 4
4
2 2 2
93
95 488 503 80 82 92 92 93 51
56
93 94 99 99 47 53 93 93 93
2 95
0 00
2 950
0 015
0 500
0 02
3 810
0 01
2 923
0 05
4 535
0 01
2 935
0 00
2 990
0 06
4 500
0 00
2 930
0 02
2 940
2 92 7 92 0 488 0 444 3.82 3 81 2.93 2.98
4 59 2 52 2 52 4 69 4 71 2 93 2 92
Q 01
4 585
0 00
2 520
0 02
4 700
0 01
2 925
2 920
t0.030
0 044
0 466
+O
0 01
3 815
-0.005
0 02
2 940
-0.017
4.48 4.48 2.91
0.00
4.480
tO.055
...
..
...
...
2.99 3.01 4.50
0.02
3.000
-0.010
..
...
...
2.92 3.02 2.96 3.03 4 00 3.90 4.51 4.57 2.53 2.54 4.69 4.72 2.93 2.97
0.01
2,970
-0.040
0.07
2.995
-0.055
0.10
3.950
0.06
4.540
f0.045
0.01
2.535
-0
0.03
4.705
-0.005
0.04
2.950
-0.025
...
4 02 4 58
0 00
034
... 015
-
Calculation. Calculate thP milliliters of tetraethyllead per gallon a t 60’ F. by means of the equation: Tetraethyllead, ml. per gallon
=
( A - F ) N (0.1036) V1.057 3785.3
- 371 (a - F ) S V
where
-4
volume of disodium ethylenediamine tetraacetate solution used in the titration. .Y = normality of disodium ethylenediamine tetraacetate solution. F = volume of disodium ethylenediamine tetraacetate solution equivalent to 1.00 ml. of magnesium chloride solution. T 7 = volume of sample at 60’ F., in ml. =
The factor 1.057, representing the grams of lead per ml. of tetraethyllead, v a s calculated using 1.65 as the density of tetraethyllead. The factor 3785.3 represents the number of milliliters in a U. S. gallon.
If the sample was measured a t a temperature other than 60’ F., use the appropriate corrected volume, V , as given in the ASTM method ( I ) , section 6b. INTERFERENCES
Ordinarily, lead is the only metal that will be encountered in the acid extract. If the sample contains other metals, they may
be titrated in the acid extract with the lead. If more than a few parts per million of other metals are present, add 0.1 gram of potassium cyanide after the excess ammonium hydroxide is added, and proceed as usual. The cyanide eliminates the interference of nickel, zinc, copper, etc., but not magnesium and calcium. On a few occasions, end-point difficulties were encountered i n gasolines containing a blue dye. The interference was removed by adding a few drops of bromine water t o the acid eitract and boiling t o expel the escess bromine prior t o the addition of tartaric acid and ammonia. EVALUATION O F METHOD
The results obtained on fifteen commercial gasolines by the
ASTN method ( 1 ) and this method are given in Table I. To determine if the two methods were comparable-that is, if the average difference between the methods was greater than t h e esperimental error-two separate statistical t tests were performed. In the first test, the data were treated simply as 30 trials for the volumetric method and 28 trials for the gravimetric method. In the second test, averages ivere compared for those samples on which an equal number of trials had been made by the two methods. Both methods of testing gave t values of almost zero, indicating that any difference betn-een the txvo analytical methods can be ascribed only to random variations. The standard deviations calculated from the duplicate measurements for the volumetric and ASTM methods are 0.019 and 0.036 for a range of 0.5 t o 5.0 ml. of tetraethyllead per gallon. The expected deviation of duplicates from their mean, based on tn-ice the standard deviation (0.05 probability), is 0.038 for the proposed method and 0.072 for the ASTM gravimetric method. A comparison of the variances by means of an F test gave a value of 4.8, exceeding the critical value F (0.99) ( I S , 16) = 4.1; this indicates that the observed difference in precision is real if equal operator precision is assumed. ACKNOWLEDGMENT
The authors Fish to express their appreciation to H. H. Dellinger and F. E. McLane for making available the results obtained in Paulsboro Works Laboratory, and to J. M. Wolfram and A. S. Hopkins who performed the analyses reported in Table I. LITERATURE CITED
-&m. Soc. Testing Xaterials, “-4STlI Standards on Petroleum Products and Lubricants,” Method D 526-48T, 1952. Ibid., D 1269-53T, 1953. Banewicz, J. J., and Kenner, C. T.,- ~ N A L . CHEM.,24, 1186
(1952). Banks, J., Analyst, 77,484 (1952). Calingaert, George, and Gambrill, C. AI., ISD. EKG.CHmf., ANAL.ED.,11, 324 (1939). Calingaert, George, Lamb, F. W., Miller, H. L., and Soakes, G. E., 45.41,.CHEM.,22, 1238 (1950). Cheng, K. L., Kurtz, Touby, and Bray, K. H., Ibid., 24, 1640 (1952). Diehl, Harvey, Goetz, C. A., and Hach, C. C., J . A m . V a t c r Works Assoc., 42, 40 (1950). Flaschka, H., Mikrochemie P E T . ;Ilibrochim. Acta, 39, 315 (1952). Grunwald, A., ErdBZ u. Kohle, 6 , 550 (1953). Hughes, H. K., and Hochgesang, F. P., AKAL. CHEK, 22, 1248 (1950). Jenness, Robert, Ihid., 25, 966 (1953). Manns, T. J., Reschovsky, 11.U., and Certa, A. J., I b i d . , 2 4 , 908 (1952). Socony-Vacuum Laboratories, Analytical Method S V l I 85-50, July 20, 1950. Willson, A. E., ANAL.CHEW,22, 1571 (1950). RECICIVED for review February 3, 1964. Accepted April 8, 1954.
