Determination of Sulfur in Urine Rapid Photometric Method PAULW. BOUTWELL AKD EDWARD W. TOEPFER, Beloit College, Beloit, W i s . t o t h e determination of s m a l l THE A P P L I C A T I O N of the sulfur photomequantities of sulfur in biological and ( I 1 ) , the use ter to the rapid determination of sulfur i n material has been confirmed in of the sulfur photometer for urine is described. A method involving the earlier work (11). I n the hands the rapid determination of sulfur in foods and biological materiuse of nitric and perchloric acids has been used of an experienced operator, the alsisdescribed. Itwasthought in the oxidation of urine for total sulfur and photometer makes possible the determination of sulfates in from that if this method could be suemav be followed bv either the Dhotometric or 10 to 15 minutes with an error of cessfully to the detergravimetric determination of the sulfur. This about 0.03 mg. in 2 to 3 mg. of mination of the distribution of method of oxidation, together with the use of sulfur, the optimum amount for sulfur in urine, it should offer the sulfur photometer, possesses advantages a d v a n t a g e s O v e r the u s u a l the determination. m e t h o d in speed and ease of in speed and ease of operation, and, in the hands DETERMINATION O F INORGANIC operation. I c i s the purpose of of a n experienced operator, offers a reliable SULFATES this Paper t o demibe such a and rapid routine method of analysis. The preparation of the soluphotometric method t o g e t h e r tion for the photometric deterwith a method for the oxidation mination of inorganic sulfates followed essentially the method of the sulfur for the determination of total sulfur. Jackson (6) and, in earlier papers, Hinds ( 5 ) describe the of Folin (4). Twenty-five cubic centimeters of urine and 10 cc. use of a crude photometer for the determination of sulfates. of 1 to 4 hydrochloric acid were diluted to 250 cc. in a volumetBoth authors refer t o the application of their method to ric flask. From this solution an aliquot was taken, usually the rapid estimation of inorganic sulfates in urine. The 25 cc., containing from 2 to 3 mg. of sulfur. To the aliquot use of the photometer in the determination of sulfur in coal were added 25 cc. of salt solution containing 120 grams of is described by Parr and McClure (9). The sulfur photome- sodium chloride per liter. The resulting solution was made ter has been improved and placed on the market as the up to 100 cc. The colloidal barium sulfate necessary for “Burgess-Parr Sulfur Photometer.1” In the directions (2) the photometric determination was formed by the addition for its use, its application to the determination of sulfur of 0.3 to 0.5 gram of sized barium chloride crystals (20 to in foods is given, following the oxidation of the sample 30 mesh) and then mixed by shaking. After standing for 5 with sodium peroxide in the sulfur bomb. I n attempting minutes with occasional shaking, the readings were taken to adapt this method to the determination of total sulfur on the photometer. The directions (2) for the use of the in urine, a few trials proved that the sodium peroxide bomb instrument were carefully followed. With practice in readfusion was not suited for the oxidation. Benedict’s ( 1 ) ing the instrument and under correctly controlled condimethod using copper nitrate and potassium chlorate could tions, excellent checks can be obtained. The photometric results were checked by Folin’s (4) not be used because of the color of the solution if it was to be followed by the photometric method. Dennis and Reed gravimetric method, and the comparisons are shown in (3’) have avoided this difficulty in their nephelometric method Table 11.. by the substitution of zinc nitrate for copper nitrate in their DETERMINATION OF TOTAL SULFATES oxidation mixture. The authors have used a wet method For the determination of total sulfates, the urine was hydroof oxidation in a small Pyrex Kjeldahl flask involving the use of fuming nitric acid and 70 per cent perchloric acid as lyzed according to Folin’s (4)method. Twenty-five cubic censuggested by LeMatte, Boinot, and Kahane ( 7 ) for the timeters of urine and 20 cc. of 1 to 4 hydrochloric acid were oxidation of sulfur in biological material. This method placed in a Pyrex Erlenmeyer flask covered with a small watch was improved and used successfully with no appreciable glass and boiled gently for 30 minutes. The solution was loss of sulfur by Wolesensky (12) and by Macliay (8) in the cooled, transferred t o a 250-cc. volumetric flask, and the condetermination of sulfur in rubber. Zahnd and Clarke (13) tents made up to volume. Asuitable aliquot was taken, usually report the loss of sulfur in the use of nitric and perchloric 25 cc., containing about 2 to 3 mg. of sulfur. To this were acids. Their method of carrying out the oxidation differs added 25 cc. of a salt solution containing 3 grams of sodium from the one which the authors (11) have used, wherein chloride, the volume made up to 100 cc., and the photometer no appreciable loss of sulfur in the oxidation of various types readings made as before. The results were checked by the of biological material has been experienced. Its applica- usual gravimetric method (4). The comparisons are shown tion to the oxidation of sulfur in urine has proved successful, in Table 11. In the course of the hydrolysis of the ethereal sulfates After oxidation, the resulting solution is easily prepared for either the gravimetric or photometric determination of with hydrochloric acid, there is a gradual darkening of the sulfates. It should be noted that any error in the precipi- urine. It was first thought that this darkening would make tation of barium sulfate in the presence of nitric and per- the solution unfit for photometric determinations. The chloric acids is avoided by the destruction of the excess of color fades out considerably on dilution and does not affect the accuracy of the readings. This was shown by adding these acids a t the end of the oxidation. The reliability of the photometric method when applied known amounts of a standard solution of potassium sulfate to a solution of hydrolyzed urine prepared as described * Since the oompletion of this paper, the form of the photometer haB been above in which the total sulfates had been determined, and simplified t o increase its speed and accuracy and renamed the ”Burgessnoting the recovery of sulfur obtained. The results are Parr Turbidimeter.” The new form is now being used in this laboratory given in Table I. in preference to the old In the continuation ot our work.
I
N A recent paper by Toepfer
117
A N A I,Y T I C A L E D I T I O N
118
Vol. 4, No. 1
UNTABLEI. PHOTOMETRIC RESULTSFOR TOTAL SULFATES AFFECTED BY DARKENING OF HYDROLYZED URINE
quent photometer determination. The solution was made up to 100 cc. and suitable aliquots were taken for the phoSULFUR tometer readings. The oxidations can be completed and the IN SOLN. SULFUR SULFUR SULFUR SULFUR TAKEN ADDED PRESENT FOUND ERROR RECOVERED solutions prepared for the readings in from 1 to 1.5 hours. Mg. Mg. ’ Mg. Mg. Mg. % A blank for the perchloric acid method of oxidation was 100.44 1.82 +o.oos 1,812 1,000 0.812 determined according to the “true blank” method of Stock2.03 $0.018 100,89 2.012 1.200 0.812 2.35 $0.138 106.22 2.212 1.400 0.812 holm and Koch ( I O ) . The blank was equivalent to 0.002 2.41 -0.002 99.91 2.412 1.600 0,812 gram of barium sulfate or 0.28 mg. of sulfur, for which the 2.66 +0.048 101.83 2.612 1.800 0.812 2.83 $0.018 2.812 100.64 2.000 0.812 corresponding results in Table I1 have been corrected. The 3.04 4-0.028 3.012 100,92 2.200 0.812 results for total sulfur were checked in two ways. First, TABLE11. COMPARISONOF PHOTOXETRIC AND GRAVIMETRICthey were compared with gravimetric determinations made RESULTSFOR INORGANIC SULFATES,TOTALSULFATES,AND on the solution following the perchloric acid oxidation of TOTAL SULFUR the urine. Second, both the photometric and gravimetric VOL. WT. OF Av. S results following the perchloric acid oxidation were comIN ALI- WT, OF WT. OF S PER PER SAM- DETERMIpared with the results obtained by Benedict’s ( I ) gravimetric LITER LITER METHODVOL. QUOT BaSOi S P L ~ NATION Cc. Cc. Gram Mg. Gram Gram method. Blank determinations were made for Benedict’s 1 Inorg. so4 Photo. 25 2.5 1.78 0.712 method by the true blank method. The blank averaged Photo. 25 2 . 5 1.78 0.712 0.712 Grav. 25 0.1303 17.90 0.716 0,0044 gram of barium sulfate, for which the corresponding Grav. 25 0.1288 17.70 0.708 0.712 figures in Table I1 have been corrected. Total so4 Photo. 26 2.5 1.91 0.764 The comparative results shown in Table I1 were all made Photo. 25 2 . 5 1.91 0.764 0.764 on 24-hour samples of urine. The volumes were made up Grav. 25 0.1372 18.84 0.754 Grav. 25 0.1365 18.72 0.749 0.752 to 1000 cc. in each case. The results are calculated t o Total S Photo. 5 2.5 2.06 0.824 grams of sulfur per liter and represent the output of inorHC104 5 2.5 2.06 0.824 ganic sulfates, total sulfates, and total sulfur for the ‘245 2.5 2.06 0.824 0.824 Grav. 10 0 0592 8.13 0.813 hour periods. HClOn 10 0.0588 8.08 0.808 0,811 Grav. Benedict
2
8
10 10
0.0606 0.0616
8.32 0 832 8.46 0.846
0 839
Inorg. SO4 Photo. Photo. Grav. Grav.
25 25 25 25
3.5 3.5
2.17 2.17 0.1152 15.82 0.1152 15.82
0.620 0.620 0.620 0.633 0.633 0.633
Total SO, Photo. Photo. Grav. Grav.
25 25 25 25
3.5 3.5
2.41 2.39 0.1242 17.06 0.1232 16.92
0.688 0.683 0.685 0.682 0.677 0.680
Total S
Grav. HClO4 Grav. Benedict
5 5 5 5 5 10 10
2.5 2.5 2.5
1.83 1.84 1.82 3.78 3.68 7.86 7.66
0.732 0.736 0.728 0.732 0.755 0.736 0.746 0.786 0.766 0.776
Photo. Photo. Grav. Grav.
25 25 25 25
3.5 3.5
2.08 2.06 0 1076 14.78 0.1076 14.78
0.594 0.588 0.591 0.591 0.591 0.591
Photo. Photo. Grav. Grav.
25 25 25 25
3 5 3.5
2.18 2.18 0.1165 16.00 0,1160 15.93
0.622 0.622 0.622 0.640 0.637 0.638
Photo. HC104 Grav. HC104 Grav. Benedict
5 5 5 5 10 10
3.75 3.75
2.47 2.47 3.41 3.32 6.73 6.70
0.659 0.659 0.659 0.683 0.664 0.673 0.673 0.670 0.671
Inorg.
so4
Totals04
Total
S
Photo. HClOk
’
0.0275 0.0268 0.0572 0.0658
0.0249 0.0242 0.0490 0.0488
DETERMINATION OF TOTAL SULFUR
,
For the determination of total sulfur, 5 cc. of urine were measured into a 300-cc. Pyrex Kjeldahl flask by means of an Ostwald pipet, and 2 cc. of fuming nitric acid (sp. gr. 1.50) and 1 cc. of 70 per cent perchloric acid were added. The solution was heated carefully over a low flame until the first reaction had practically ceased. It was then boiled gently until white fumes appeared. At this point the solution should be colorless or nearly so. If not, more perchloric acid was added and the boiling continued. The solution was then cooled and 2 grams of sodium chloride were added. This released chlorine and nitrogen dioxide from the destruction of the excess of nitric and perchloric acids. The flask was heated until the evolution of the fumes had ceased and the solution was colorless. It was then diluted with 20 to 30 cc. of water and made just basic with 4 N sodium hydroxide, then neutralized and made acid with 1 cc. in excess of concentrated hydrochloric acid. This treatment insures the correct salt content and acidity for the subse-
RETENTION OF SULFURIN PERCHLORIC ACID METHODOB OXIDATION In order to see if there was any loss of sulfur during the perchloric acid method of oxidation, aliquots of a urine of known total sulfur content were oxidized, and the per cent. of sulfur recovered after correcting for the blank was determined. The results which are given in Table I11 show that there is no apparent loss of sulfur during the oxidation in the Kjeldahl flasks.
TABLE111. PHOTOMETRIC DETERMINATIOXS SHOWINGRETENTION OF SULFUR DURING PERCHLORIC ACID OXIDATION TOTAL 9 PRESENT TOTAL9 FOUND Mg. Mg. 2.16 2 15 2 15 2.17 2 15 2 15 2 15 2.15 2 2 2 2
84 84 84 84
2 65 2.65 2.65 2.65
ERROR
S RECOVERED
Mg
%
-40.01 $0.02 0.00 0.00
100 4 100 9 100.0 100.0
87 85 84 84
+ O 03
101 100 100 100
2.66 2.62 2.62 2.67
+o 01
2 2 2 2
+o 01
0 00 $0 01 -0 03 -0 03 +o 02
0
3 0 3
100 2 98.9 98.9 100.7
APPLICATION OF METHODTO ROUTINE ANALYSIS The photometric method was tried out on a series of 24hour samples of urine with the results shown in Table IV, expressed in grams of sulfur per 24-hour period. The usual diet, which was rather low in protein, was observed throughout except that for April 16, the diet was high in sulfur and for April 27 it was somewhat higher in protein. The data have small interest aside from showing the consistency of the results which may be expected by this method. TABLEIv.
NITROGEN CONTENT AND SULFUR DISTRIBUTION
TOTALINORGANIC TOTAL TOTAL ETHEREAL NEUTRAL SAMPLE N so4 so4 S so4 s Grams Gram Gram Gram Gram Gram 0.082 April 15 ii:i4 0.127 April 16 0.088 April 17 10.77 0.087 11.67 April 18 9.77 April 24 0.062 April 27 13.10 0.098 DATBOF
January 15, 3932
INDUSTRIAL AND ENGINEERING CHEMISTRY LITERATURE CITED
(1) Benedict, 8. R., J. Biol. Chem., 6 , 363 (1909). ( 2 ) Burgess-Parr Co., Moline, Ill., Booklet 108. (3) Dennis, W., and Reed, L., J. Biol. Chem., 71, 205 (1926). (4) Folin, O., Ibid., 1, 131 (1905). (5) Hinds, J. I. D., J . Am. Chem. SOC.,18, 661 (1896); 22, 269 (1900). (6) Jackson, D. D., Ibid., 23, 799 (1901). (7) LeMatte, L., Boinot, G., and Kahane, E., J pharm. ehim., 5 , 325 (1927); Compt. rend. SOC biol., 96, 1211 (1927)
119
(8) Mackay, J. G., J . Soc. Chem. Ind., 49, 233 (1930). (9) Parr, S. W., and McClure, C. H., J. Am. Chem. floc., 26, 1139 (1904). (10) Stockholm, M., and Koch, F. C., Ibid., 45, 1953 (1923). (11) Toepfer, E. W., and Boutwell, P. W., IND. ENQ.CREM.,Anal. Ed., 2, 118 (1930). (12) Wnlesensky, E., IND. ENQ.CHEM.,20, 1234 (1928). (13) Zahnd, H., and Clarke, H. T., J. Am. Chem. Soc., 52 3275 (1930). RECEIYED August 28, 1931.
Emulsive Capacity of Sulfonated Oils Miscibility of Sulfonated Oils and Neutral Oils RALPHHART,The Hart Products Corp., 1440 Broadway, New York, N . Y.
s
U L F O N A T E D oils find de-emulsifying effect upon the T H E EMULSIVE C A P A C I T Y of a sulfonvaried and extensive apmixture. ated oil is determined by mixing it with various plication in the industries, amounts of olive oil, clearing with oleic acid, and PROCEDURE being used as mordants, dye asJinally testing the stability of the emulsion. To d e t e r m i n e the emulsive s i s t a n t s , detergents, softeners, The emulsive capacity is useful in formulating capacity, the writer proposes to wetting-out agents, etc. They find the maximum amount of are particularly u s e f u l t o t h e commercial products, and serves also as a criterion manufacturer of specialty oils olive oil which, when m i x e d of quality and as a means of identijcation. for making soluble or emulsifyw i t h 100 g r a m s of t h e s u l Judging by the amount of oleic acid required ing immiscible liquids, such as fonated oil under examination, to produce uniform mixtures, sulfonated oils are fatty oils, m i n e r a l oils, and gives an emulsion of a definite more miscible with mineral than with fatty oils, s t a b i l i t y . A m i x t u r e of the solvents. Many bleach or kier oils on the market are mixtures two oils is usually cloudy but and both fatty and mineral oils are less miscible becomes clear u p o n adding of s u l f o n a t e d oil and pine oil with sulfonated castor than with sulfonated olive oleic acid. The test c o n s i s t s or another solvent; wool, silk, oil. Sulfonated oils are better emulsifirs for of two p a r t s : p r e l i m i n a r y and rayon oils are often mixmineral than for fatty oils; in this respect, acid emulsions, in which the sult u r e s of s u l f o n a t e d o i l a n d sulfonated castor oil is inferior to acid sulfonated mineral, neat’s-foot, olive oil, or f o n a t e d oil varies by 10 per a similar lubricant. Sulfonated cent intervals; and final emulolive oil. Complete neutralization of a sulfonated oil also forms the base in the sions, in which the sulfonated oil decreases its miscibility with neutral oils. so-called soluble cutting oils for oil varies by 1 or 2 per cent of Completely neutralized sulfonated oil, with the metals, and finally s u l f o n a t e d the mixtures. proper addition of alcohol, is a better emuls8er cod or other fish oil is used conPRELIMINARY E M U LSI o N S. for mineral oils than the acid oil but is a poorer siderably in treating leather. Ten grams of a mixture of the The emulsifying property of sulfonated oil and olive oil (9 emulsiJier for fatty oils. s u l f o n a t e d oils depends to a grams and 1 gram, respectively; great extent upon the nature of the raw oil, method of sulfona- 8 grams and 2 grams, respectively, etc.) are thoroughly mixed tion, neutralization of the finished product, moisture content, in a 100-cc. glass beaker, using preferably a thermometer as a etc. Geronazzo (2), in discussing leather oils, states that the stirring rod, and titrated a t 40” C. with oleic acid until the quality of a sulfonated oil is related directly to the duration of mixture just turns clear. Next 0.1 cc. of excess oleic acid is its own emulsion and to its “emulsive capacity”-i. e, the prop- added, and a 5 per cent emulsion made, of which 100 cc. are erty of retaining a definite quantity of another fatty sub- allowed to stand for exactly 2 hours in a 4-ounce oil bottle. The stance in a homogeneousemulsion. Rumcke ( I ) concludesthat titration with oleic acid is very sharp and sensitive to less than sulfonated oils with the greatest carrying capacity for mineral 0.25 per cent of the mixture. A large excess of oleic acid, for oils give the best results in practice. I n spite of its evident im- reasons already mentioned, must be avoided. After the 2 portance there seems to be no detailed procedure in the litera- hours, the emulsions are closely examined and note made of turefor determining the emulsifying capacity of sulfonated oils; those showing free oil on the surface. With the preliminary hence the simple method outlined in this paper may be timely. emulsionsthere is no difficulty in separating the good emulsions This test is of importance not only in formulating commercial from the poor ones, as the change is very marked. products, but is useful also as a criterion of the quality of the FINAL EMULSIONS.Another series of mixtures is now made oil and as a means of identification. beginning with the first satisfactory emulsion containing the A sulfonated-oil mixture to be marketable must be uniform least amount of sulfonated oil, but instead of 10 grams, a and clear and remain so indefinitely. It has also been found quantity ten times as much is used, or 100 grams. Six that a clear oil gives a better emulsion than a cloudy product, mixtures are made, each consecutive one containing 2 grams which may separate on standing. Homogeneity is usually less of the sulfonated oil. In each case after clearing, 0.5 gram attained by adding oleic acid (red oil) to the mixture (3). An excess of oleic acid is added. Emulsions are made as above, excess of oleic acid, however, must be avoided as it may exert a and the first emulsion next to the last one showing free oil is
