53%, when the wave length scale was set a t 405 mp. From Table 111, B, the total uroporphyrin content was 4.2 y. It crystallized in fine needles, melting point 280-285" C. The fainter second, third, and fourth zones were also eluted out, and checked by paper chromatography (1) with markers of knomn quality. The first zone, coproporphyrins consisted of about 70% I isomer and 30% I11 isomer. The second. third, and fourth zones behaved exactly like esters of penta-, hem-, and heptacarboxylic porphyrins, respectively, isolated from pathological urines. The fifth zone uroporphyrins consisted of 80% uroporphyrin I methyl ester and 20% of the I11 isomer. Porphyria Urine. Two consecutive 24-hour urine samples of 1135 and 2100 ml. were collected from a patient xx-ith intermittent acute porphyria. Both samples showed t h e presence of porphobilinogen (8). Both were found to contain 42 y of coproporphyrins per 100 ml. of sample. The amount of uroporphyrins was 24 y per 100 ml. in the first sample and 48 y per 100 ml. in the second sample. Apparently the daily fluctuation of porphyrin level was considerable. The porphyrins n-ere obtained in crystalline forms. Methyl esters of coproporphyrins containing 20% of I and 80% of I11 isomers, melted a t 155-163" C., and those of uroporphyrins containing 50% each of I and I11 isomers melted a t 268-276' C. The chromatogram is shown in Figure 1,B. This case has indicated that a n abnormal porphyrin pattern of a urine sample can be detected before other manifestations of porphyria symptoms, which is useful in clinical diagnosis.
The findings from other porphyria cases, given in Figure 1 as references, were determined in the same way. Recovery Experiment. A mixture of 285 y of uroporphyrin ester, melting point 2817-289" C., and 105 y of coproporphyrin I methyl ester melting point 254" C., was hydrolyzed overnight with 25y0 hydrochloric acid to free porphyrins, and then immediately introduced into 250 ml. of the deporphyrinated filtrate of the normal urine sample. A recovery of 270 y of uroester, melting point 286-290" C., and 101 y of coproporphyrin I methyl ester melting point 253-254"C., was obtained. It amounted to about 95 to 96% over-all recovery. For comparison, porphyrin mixtures of the same composition mere directly chromatographed on calcium carbonate and magnesium oxide columns with benzene-chloroform (10 to 6), and benzene-methanol (100 to 4) as the respective developers (5'). In both instances, a longer period was needed for the development, and the separation of porphyrin esters was incomplete. The loss due t o chromatography alone was over 10%. From the fraction of the coproporphyrin of the magnesium oxide chromatogram two kinds of crystals were observed under a microscope, mainly long needles of melting point 235-240" C., and some hairlike crystals melting point 275282" C. By paper chromatography it was confirmed that the coproporphyrin fraction y a s contaminated m-ith uroporphyrin. With these calibration data and chromatographic patterns, determinations of practically all kinds of porphyrins in urine samples, even without any authentic porphyrin standard, may be
made. The crystalline products obtained can be saved for further identification and experiment,ation. ACKNOWLEDGMENT
The authors are grateful to H. L. Mason of the Mayo Clinic, Rochester, Minn., R. I f . Halpern of Beverly Hills, Calif., and G. R. Kingsley of Veterans Administration Los Angeles, Calif., for samples B, E, and F listed in Figure 1
1.
LITERATURE CITED
(1) Chu, T. C., Chu, E. J.-H., J . Bid. Chem. 227, 506 (1957). ( 2 ) Chu, T. C., Chu, E. J.-H., unpublished
data. (3) Eriksen, L., Scand. J . Clin. & Lab. Invest. 4, 55 (1952). (4) Falk, J. E., "Ciba Foundation Symposium on Porphyrin Biosynthesis and 3letabolism," p. 63, Little, Brown, Boston, 1955. (5) Formijne, P., Poulie, N. J., Ibid., p. 3Afi - --. (6) Markovitz, hf., J . Lab. & Clin. Med. 50, 367 (1957). ( 7 ) Kicholas, R. E. H., Biochem. J. 48, 311 (1951). (8) Schn-artz, S., Vet. Administration Tech. Bull. TB 10-94 (1953). (9) Watson, C. J., Pimenta de Mello, R., Schwartz, S., Ha-vkinson, V. E., Bossenmaier. I.. J . Lab. & Clin. Med. 37, 831 (195i). ' (10) Willstatter, R., Mieg, IT., A n n . Chem. 350, l(1906). (11) Zondag, H. A, Van Kampen, E. J., Clin. Chiin. Acta 1, 133 (1956).
RECEIVED for review February 26, 1958. Accepted June 6, 1958. Work supported by research grant, il-lOOO(C7), from the Kational Institute of Arthritis and Metabolic Diseases, National Institutes of Health, Public Health Service.
Determination of Sulfur Oxides in Stack Gases EDWIN 8. SEIDMAN
Shell Oil Co., Wilmingfon, Calif.
b Sulfur trioxide in stack gases as low as 0.001 % gas volume may be determined in the presence of as much as 0.370 gas volume sulfur dioxide. Ammonia and/or nitrogen oxides do not interfere. Sulfur trioxide is absorbed quantitatively in an 8Oy0isopropyl alcohol solution, which inhibits oxidation of sulfur dioxide. The sulfate i s titrated with 0.01N barium chloride in a solution of 80% isopropyl alcohol, using Thorin indicator. The end point i s sharp and reproducible and the titration is rapid. A modification of this procedure also has been used for the determination of total sulfur oxide. 1680 *
ANALYTICAL CHEMISTRY
I
rapid and accurate methods for the determination of sulfur dioxide and trioxide content of combustion gases has been stimulated by atmospheric pollution studies in several major cities. The standard Anierican Petroleum Institute methods used by the petroleum industry (1, 2 ) are subject to interference from ammonia and nitrogen oxides, both of which are known to be present, for example, in catalytic cracking flue gases. The API methods require absorption of the gas sample in a known amount of standard base. After absorpI~TEREST IK
tion, excess base is titrated and total sulfur oxides are obtained by difference. The sulfur trioxide is then precipitated as benzidine sulfate, redissolved, and titrated. Sulfur dioxide is determined by the difference. Even when no interfering substances are present. several difficulties have been encountered in applying the API methods to refinery stack gases. First, the methods involve small differences between large numbers. I n addition, the colorimetric titration end point is frequently obscured by dark-colored oxidation products from the inhibitors present in the absorber solution. (Ex-
ploratory potentiometric titrations yielded curves which were too flat t o permit establishment of a n inflection point.) Finally, benzidine sulfate is soluble in water t o a degree that severely curtails washing of the precipitate. The method described herein depends upon the titration of sulfate with 0.01N barium chloride in a solution of 807, isopropyl alcohol with Thorin indicator [o-(2- hydroxy- 3,6- disulfo - 1-naphthylaz0)benzenearsonic acid disodium salt] as developed by Fritz and coIi-orkers (6, 7 ) . The indicator gives a sharp color change of yellow to pink a t the end point. According to Fritz, high concentration of a nonaqueous solvent is necessary for the barium titration of sulfate with Thorin. Ethyl alcohol, isopropyl alcohol, or methanol map be used; the concentration of alcohol is bO t o 907, by volume in a p H range of 2.5 to 4.0 ( 7 ) . I n addition, Flint ( 5 ) found that isopropyl alcohol is best for absorbing sulfur trioxide, because absorption is complete. Further, the small amount of sulfur dioxide absorbed n ith sulfur trioxide in isopropyl alcohol does not interfere in the titration. Thus, the absorber solution may be titrated directly. A modification of the method can be used for determining total sulfur oxides. The gas stream is scrubbed through absorbers containing an aqueous solution of sodium hydroxide and hydrogen peroxide. After destruction of excess peroxide, sodium ion, which interferes in the titration, is removed with a cation exchange resin in the hydrogen form. The resulting solution is diluted n ith isopropyl alcohol and titrated with barium ion. REAGENTS AND SOLUTIONS
Hydrogen peroxide, 3% solution. Sodium hydroxide, about 0.2N. Isopropyl alcohol, reagent grade, and a solution of 80% by volume isopropyl alcoho1--20% demineralized water. Barium chloride, 0.0100N. Cation exchange resin. Dowex 50, cross-linking 16, 50 to 100 mesh (or ion exchange resin, H C R , hydrogen form, Hach Chemical Co., Ames, I o ~ a ) . Thorin indicator, Distillation Products Industries, Rochester 3, S. Y.; 0.2% solution in mater. PROCEDURES
Sulfur Trioxide. Three absorbers are connected in series with Tygon (or similar) tubing as shown in Figure 1. Each of t h ~absorbers is filled n i t h 100 ml. of 8001,isopropyl alcohol and immersed in a n ice-water slurry. A 1-cubic foot gas sample is drawn in 20 minutes according to dnierican Petroleum Institute Method 751. After the sampling, the absorbers are rinsed with 80% alcohol into a volumetric flask and made up to 500 ml. with the 807, alcohol. T o a 25-m1. aliquot is
BLEED LINE
OUTLET
SAMPLE LINE
"IU'
POSITIVE PRESSURE
-
I'
Figure 1.
I/
Sampling train
added 25 ml. of 80% alcohol and 3 or 4 drops of Thorin indicator. The solution is titrated with 0.01N barium chloride to the pink end point. The determination should be completed within 1 hour after sampling is complete to eliminate errors due to oxidation of sulfite to sulfate. Total Sulfur Oxides. Two absorbers are connected in series with Tygon tubing. To each absorber 25 mi. of 3% hydrogen peroxide and 25 ml. of 0.2N sodium hydroxide are added. T h e gas is sampled as previously described. T h e solution from the absorbers is rinsed with water into a beaker and boiled for 30 minutes to remove peroxide. The solution is then acidified with hydrochloric acid, boiled for 5 minutes t o remoi-e carbonate, cooled, and made u p to 500 ml. About 7 5 ml. of this solution is passed through a dispenser containing cation exchange resin and discarded. The dispenser is 15 X 3 em. with a 10-em. column of resin above a glass woo1 plug. An additional portion of the solution is then passed through the resin, and a 10-ml. aliquot is added to 40 ml. of reagent grade isopropyl alcohol containing 3 or 4 drops of Thorin. The resulting solution is titrated with 0.01N barium chloride to the pink end point. RESULTS AND DISCUSSION
Although sampling procedure is of fundamental importance in stack gas analysis, the current study n-as restricted to developing improved methods for absorber solution analysis, as sampling has been studied by others (3-5). Total sulfur oxides were investigated in the range of 0.02 to 0.307, gas volume and sulfur trioxide in the range of 0.001 to 0,0207, gas volume. Two independent methods have substantiated isopropyl alcohol absorption of sulfur trioxide (8, 9 ) . Determination of Sulfur Trioxide. Fritz ( 7 ) has reported t h a t sulfite interferes seriously in t h e titration of sulfate. It has been t h e authoi's eyperience, however, t h a t t h e concentration of sulfite in a n isopropyl alcohol absorber used for scrubbing refinery stack gases does not interfere. T o verify this observation, known synthetic absorber solutions n-ere made u p containing sulfur di-
oxide a n d sulfur trioxide in SOTo alcohol. Aliquots were added to 80% alcohol and titrated with 0.01N barium chloride. There '\vas no significant difference between theory and the actual amount of barium chloride required a t the 95% confidence level (Table I).
Table I.
Effect of Presence of Sulfite" M1.0.005MBarium Chloride
Sample
Theory
Actual
1
1.43
B
0.86
1.32 1.35 1.42 1.40 0.85 0.88 0.89 0.85
a
SO2 t o SO, mole ratio about 5 to 1.
As a precaution against oxidation of sulfur dioxide to sulfur trioxide, the absorbers should be immersed in an ice-n ater slurry during sampling. I n order to check further if any significant oxidation of sulfur dioxide occurs, several absorber solutions were allowed to stand a t room temperature and sulfate -cas determined after various intervals. After 2.5 to 3 hours there was a slight increase in the niilliequivalents of sulfate found; plots of the data showed that the increase in sulfate was linear with time. The sloiv rate of oxidation of sulfur dioxide is further indication of the fact that its presence does not interfere or give high results. Determination of Total Sulfur Oxides. Both gravimetric determination of total sulfur oxide as barium sulfate and amperometric titration n i t h lead ion have substantiated the Thorin titration. A statistical analysis of 33 samples run both by Thorin titration and gravimetrically gave no significant difference between the two methods at the 95% confidence level. The standard deviation of the method, including sampling technique, is 1.5% of the mean value, 0.257, volume total sulfur oxides, equivalent to 0.0019 meq. of sulfur tiioxide. The standard VOL. 30, NO. 10, OCTOBER 1958
1681
Table II. Effect of Removal of Sodium Ion by Ion Exchange Resin
Sample A
Meq. Sulfate After resin Before resin 0.1105 0.1111 0.1105 0,1038
B
0.1038 0.1038 Av. 0.1072
0.1074 0,1068 0.1068
n . inno
0 . io06
0.1006 0.1037
deviation of the Thorin titration alone, as determined by triplicate titrations on 21 samples, is 0.4% of the mean, or 0.0006 meq. of sulfur trioxide. The use of acid or neutral peroxide for the total sulfur oxide absorber
solution would simplify the procedure by eliminating the ion exchange step, However, it is preferred that the peroxide be destroyed in order to prevent deleterious effects upon the organic ion exchange resin. The presence of caustic is necessary to prevent stripping of sulfur dioxide by high concentrations of carbon dioxide. The presence of sodium in the solution caused significant interference, thus substantiating the necessity for its removal by the resin (Table 11). ACKNOWLEDGMENT
The author is pleased to acknowledge the assistance of C. C. Graf in the initial phases of the work. Appreciation is expressed to the Shell Oil Co. for permission to publish this paper.
LITERATURE CITED
(1) American Petroleum Institute, “Man:
ual on Disposal of Refinery Wastes Method 774-54, Vol. 5, 1st ed., 1954. (2) Zbid., Method 775-54, Vol. 5, 1st ed.,
1954. (3) Corbet>t,P. F., J. Znst. Fuel 24, 24751 (1951). (4) Corbett, P. F., J . SOC.Chem. Znd. 67, 227 (1948). (5) Flint, D:, Zbid., 67, 2 (1948). (6) Fritz, J. S., Freeland, M. Q., ANAL. CHEM.26, 1593 (1954). (7) Fritz, J. S., Yamamura, S. S., Zbid., 27, 1461 (1955). (8) Los Angeles ilir Pollution Control District Method, 1956. (9) McCombie, H. R., private communi-
cation, Shell Chemical Corp., P.O.
Box 431, Pittsburg, Calif.
RECEIVED for review September 13, 1957. Accepted hlay 9,1958.
indirect Flame Spectrophotometric Determination of Sulfate Sulfur W.
M. SHAW
University o f Tennessee Agricultural Experiment Sfation, Knoxville, Tenn.
,A study was made of the flame spectral emission properties of barium with the objective of applying the barium emission to indirect flame spectrophotometric determination of sulfate in waters and oxidized biological materials. The barium emission intensities were measured in oxyhydrogen flame with the Beckman DU spectrophotometer and photomultiplier attachment using the 51 5-mp band head for peak emission, and the 522-mp for cation background correction. Iron, aluminum, and phosphate were eliminated by precipitation in ammonium acetate solution buffered a t p H 4. Calcium, strontium, and manganese gave spectral interferences. Sulfate determinations in waters containing a wide range of cation and phosphate concentrations carried out b y the gravimetric and flame spectrophotometric methods showed satisfactory agreement.
T
determination of sulfate is of widespread interest and application: agriculturally, in irrigation waters, soils, crops, and feeds; industrially, in boiler feed waters; biochemically, in oxidation products of certain amino acids; and medicinally, in blood serum for the detection of renal deficiency. The procedures for the determination of sulfate may be grouped into three classes: gravimetric (17 ) , titrimetric HE
1682
ANALYTICAL CHEMISTRY
(H),and turbidimetric (8). Each has its o m quantitative adaptation, and serves particular requirements of speed and convenience. The accuracies attributed to these methods by the American Public Health Association ( 2 ) are: gravimetric, t o 1 2 % ; turbidimetric and titrimetric, to *lo%. NeJv colorimetric (5) and titrimetric (16, 18) methods hare recently appeared in print. These methods, although of high precision and accuracy, h a r e no direct bearing on this study. The objective vias to develop a flame photometric procedure for the determination of sulfate sulfur. This procedure should be faster than the gravimetric procedure, and more accurate than the simpler titrimetric and turbidimetric procedures, and, a t the same time, eliminate sulfate and phosphate as interferences with the accurate flame photometric determinations of calcium (4). which until recently ( 1 ) hal-e received scant recognition. The procedure should have a wide range of application, including the determination of sulfate in rain n-ater, soil extracts, lysimeter leachings, irrigation waters, and sea water, and total sulfur in ashed plant and biological materials. It was the aim to carry out the sulfate determination on sample preparations used for the determination of other elements. The method is based on the reaction of sulfate ion with barium chloride, re-
sulting in the precipitation of barium sulfate. The barium chloride must be added in standard quantities, because the sulfate determination is based upon the difference between the barium added and the residual barium after precipitation. The rcsidual barium is determined by its flame emission intensity as compared with barium standards. ANALYTICAL PROBLEMS
T h e analytical problems are essentially those of flame spectrophotometric determination of barium in complex miutures. These m a p be resolved into t h e follon-ing parts: 1. Spectrophotometric adjustments for barium emission, including selection of wave length, fuel combination, and suitable barium concentration range. 2. Extent of barium emission interferences from extraneous cations and anions. 3. Methods of correcting for interfering ions or removal, where corrections are not feasible. 4. Practical procedure for the flame photometric determination. 5. Testing for analytical accuracy on substances selected for wide range of sulfate content and interfering elements, using gravimetric determinations as reference. CHEMICALS AND EXPERIMENTAL PROCEDURE
Stock solutions were prepared from analytical grade chemicals of the cations
