ANALYTICAL EDITION
February, 1944
137
2.5 hours against 11 cc. of distilled water. Free amino acids were determined on aliquot portions of the dialyzate by the ninhydrin reaction ( 1 ) and were compared with amino acid concentrations found by analyses of plasma filtrates obtained by protein precipitation with picric acid (1) (see Table I). With a plasma-water ratio of 2 to 11 a 5 4 0 . aliquQt of the dialyzate contains the amino acids from 0.77cc. of plasma. If, however, a more concentrated dialyzate is required, the ratio of plasma to water could be made 12 to 6, so that a 5-cc. aliquot of the dialyzate would contain amino acids from 3.33 cc. of plasma. Another experiment exemplifies separation of free amino acids from egg albumin.
.$0 . 5 r 0
I
l
l
l
30
60
l
l
I
I
I
I
90 120 150 Time in minute3
Figure 3.
I
l
180
i 210
Time of Rocking
Maximum constant value obtained a k a P boun' dirlysir indicates equilibrium d dlffurlblc amino rcidr
Table
1.
Dialyzate 1
2 3
Av. Picrio said filtrate
Free Amino Acids Alpha-Amino Acid Nitrogen Mg./iOO CC. plaama 4.64 4.71 4.66 4.67 4.67
Table II.
Amino Acids in Egg Albumin Alpha-Amino Acid Nitrogen M g . / 1 0 0 cc. plasma
From analysis of dialyzate of egg albumin plus added amino acids From analysis of dialyzate egg albumin Added amino acid N by difference Amount of added amino acid N
2.88 0.61 2.37 2.44
To a 7% e g albumin solution was added an amino acid mixture obtainej by the hydrolysis of edestin with strong hydrochloric acid. The amount of amino acid alpha-nitro en added, calculated from results of a ninhydrin-carbon dioxije analysis on the hydrolyzate (4),was 2.44 m per 100 cc. After 2.5 hours of dialysis aliquot portions of t%e dialyzate were analyzed (Table 11). LITERATURE CITED
The use of the dialysis cell in quantitative analytical analysis is demonstrated by the following experiments: To illustrate the precise quantitative analytical data that can be obtained by means of the apparatus, three separate dialysis units were set up with 2-cc. portions of human plasma dialyzed
(1) Hamilton, P. B., and Van Slyke, D. D., J . B i d . Chem.. 150, 231 (1943). (2) Kunitz, M., and Simms, H. S., J . Gen. Physiot., 11, 641 (1928). (3) Northrup, J. H., and Kunitz, M., Ibid., 9, 351 (1926). (4) Van Slyke, D. D., Dillon, R. T., MacFadyen, D. A., and Hamilton, P. B., J. Bid. Chem., 141, 627 (1941).
Determination of Small Amounts of Molybdenum in Plants and Soils M. L. NICHOLS AND LEWIS H. ROGERS, Cornell University, Ithaca, N. Y. Spectrographic, colorimetric, and polarographic procedures For the determination of small amounts of molybdenum in plants and soils have been studied. It i s concluded that, for the ordinary laboratory, the colorimetric procedure i s superior if reasonable amounts of sample (1 gram or more of soils, 10 grams or more OF air-dried plant material) are available. However, iF only small amounts of sample are available (100 mg. of soil, 1 gram of air-dried plant material), the spectrographic procedure is recommended. The polarographic procedure has no particular advantages over the other two.
.
EVERAL recent papers (1, 4, 20) have indicated that molybdenum will need to be considered in future studies on the role of various elements in plant and animal nutrition. In one case (1) it was thought to be essential for plant growth; in another case (4) excessive quantities had a deleterious effect on cattle. The work reported here was undertaken as a result of two observations on the occurrence of this element in plants and soils. In one study (16) it was found that many mineral soils in Florida contained no spectrographically detectable molybdenum. In another study, certain Florida muck soils and some plants grown thereon showed readily detectable quantities of this element (unpublished data). These data made it desirable to study the methods of analysis for very small amounts
S
of molybdenum with respect to their precision, sensitivity, and other factors. The methods for the quantitative determination of molybdenum include gravimetric, volumetric, spectrographic, colorimetric, and polarographic procedures. The three latter methods should be more suitable for the determination of very small amounts. Spectrographically, molybdenum may be determined by a variety of procedures, but there is a growing tendency among workers in this country to use microphotometric methods, with either an internal standard line of the matrix material or an added line standard. The essential feature of this latter method is the introduction into the sample in constant known amounts of an element not originally present. This added element furnishes spectrum lines of constant intensity which, measured with a microphotometer in comparison to the line intensities of the unknown element, give a method of determination. Colorimetrically, molybdenum is most often determined by the yellow-amber color of its thiocyanate, either directly in the original solution or by first extracting with an organic solvent, immiscible with vvater (6, 16). Several reducing agents have been used, but stannous chloride has been employed more often than the others. The reaction is affected by several compljcat-
138
I N D U S T R I A L A N D E N G I N E E R I N G CHEMISTRY
ing, grinding, and mixing thoroughly t o emure homogeneity. Five standards containing from 0.1 to 0.001% of molybdenum and earh containing 0.017, of beryllium were thus prepared. Spectrograms of these standards were prepared using 0.794-
0.10 2 3
z
w
0.08 0.06 0.04
0
m
3
0.02
B t-
Vol. 16, No. 2
0.01
z 0.008 0.006 0.004 I
A-0.01 PER CENT SERYLLIUM
0.002 0.00 I
0
04
08 1.2 16 3170 R A T I O 3131
Figure 1.
20
24
26
Soil Working Curve
ing factors, including the interfeience due to the pink complex formation of the thiocyanate ion (which fades rapidly), the fading of the molybdenum complex itself, and the effect of the concentration of hydrochloric acid and other electrolytes on the equilibrium (6, 6, 17). To overcome these effects, most authors have specified strict adherence to a definite procedure and to certain concentrations of reagents which have been found satisfactory. The polarographic determination of molybdenum has been studied by Uhl (2d), Stackelburg (18), Thanheiser and Willems ( S I ) , Kanevsky and Shvartsburd ( 8 ) ,and Hokhshtein (7). Ko satisfactory curves were obtained by these workers in neutral or alkaline solution but phosphoric, sulfuric, or nitric acid solutions were used successfully. Uhl used a nitric acid-ammonium nitrate solution in which the effect of the nitrate and hydrogen-ion concentration 011 tlia step height WRS overcome by the addhion of lactic acid. SPECTROGRAPHIC METHOD
A quartz Littrow spectrograph and a uonrecording miorophotometer were used. The spectrograph had a linear dispersion of about 5 d. per mm. a t 3200 d. Commercial ammonium molybdate LIS analyzed spectrographically and found to contain traces of calcium, copper, and iron but no zinc, strontium, lithium, cerium, barium, vanadium, yttrium, chromium, manganese, tungsten, aluminium, nickel, zirconium, silver, cadmium, titanium, beryllium, bismuth, antimony, arsenic, tin, or lead. A solution of this ammonium molybdate was prepared and btandardized by precipitation as lead molybdate. The solution after standardization was diluted to 0.1 mg. of molybdenum per ml. and other dilutions were prepared only as needed, since it has been stated that 0.001% molybdenum solutions show only 0.2 to 0.4 of their original strength after long storage in glass, owing to base-exchange and advorption (11). A beryllium solution was prepared by dissolving 5.185 grams of beryllium nitrate trihydrate in water and diluting to 250 ml. This solutioii contained approximately 1 mp. of beryllium per ml. It was not standardized, but measured amounts of the same solution were used throughout this study. Dilutions of 1 to 10 and 1to 100 were made to facilitate the addition of timall quantities. Two types of base material for soil analysis were used: silica (powdered sand) and a thoroughly ground and homogenized “synthetic soil”, each gram of which contained 0.7 gram of the above silica and 0.1 gram each of ferric oxide, aluminium oxide, and calcium carbonate. Both were analyzed spectrographically for molybdenum and beryllium and none was detected. Spectrographic standards were prepared from both base materials by adding the molybdenum and beryllium solutions, dry-
(’m..(5/~e-inch)graphite electrodes which had been cut and bored, purified by the procedure of Standen and Kovach (19), and prehurned for 2 minutes. Five to 7 mg. of the standard were put into the ?up of the lower electrode with a small, half-cylinder-shaped, platinum spatula. The standard was not weighed, since, with a little practice, this amount can be estimated with some facility and an exact amount is not necessary, since the line standard had been added in known concentration. This approximate weight served as a control oil the time of burning, etc. Using a 15-micron slit and Type 111-0 Eastman spectroscopic plates the arc (direct current, 250 volts, 12 amperes) was struck (the shutter heing opened just before striking the arc) and maintained until the sample was completely volatilized. The arc was focused OD the slit with a quartz condensing lens and arc wandering was corrected manually. After processing, the galvanometer de!ection ratios of the molybdenum 3170 A. and beryllium 3131 A. lines were determined with the microphotometer. The average values thus obtained were used to plot the working curves in Figure 1. Working curves for plant materials (Figure 2) were prepared in a similar manner, except that since the matrix material of plant ash is different from soils, a base material was made up so that each gram contained 0.806 gram of calcium carbonate, 0.030 gram of magnesium oxide, 0.117 gram of potassium sulfate, 0.018 gram of sodiuni chloride, and 0.029 gram silicic acid. All the materials were found spectrographically to be free from beryllium and molybdenum. Ten samples of plants and soils \\ere analyLed in the following manner. The material was dried a t 110’ C., then ashed at 450’ C. (at 550’ C. there is danger of loss of molgbdenum by volatiliAation), carefully pulverized, and mixed in an agate mortar. A I-gram portion of the ash was weighed out (100 mg. would be sufficient), sufficient beryllium solution was added to give O . O l ~ o of beryllium, and the material was dried and again homogenized. Quintuplicate spectrogramr were prepared and measured with the microphotometer, and the proportion of molybdenum determined from the appropriate curve. The result3 of the analyses are given in column 3 of Table I.
Table 1.
Analysis of Soils and Plants , -
Nature
Spectrographic
31f
Sample
hat1
%
76
A
Tiace (below calibration) None
0 00ofi.-,
0.0004
oIOOOa
Tracr
0.0015
0.0011
0,000Y
% Woody peal 10-9 inchwi
92.84
I I -. Molybdenunl’ ColoriPolarographic metric
52.95” Brighton peat (Winches) 8.81 Brighton peat (6-18 inoliesi Everglades peat 11.41 (0-8 inrhes) 48.63 Okeechobee murk (8-20 inches) 10.25 Dallis grass 5.07 Para $raw 13.04 Napier grasa 4.47 Sugar can? Ivavch 3.34 Raw grass a On ash basis. 6 Probahly due t o previous burning
0.0026
0.0021
0,0017
0.0022
0.0019
0,OOlB
0.0085 0.037
0.0031
0.0026 0.031 0.0035
0,005i
0,0028 0.00311
0.032 0.0042 0,0023 0.0025
0.0020 0 0019
Neglec!ting the time for ashing, preparation of the sample requires ;Ibout 30 rhinutes, and the additional time required for each determination on a routine baais is estimated a t about 10 minutes. There is also the possihility of simultaneous determination of several elements, which would decrease the time per det.ermiiiation. The factors affecting the precision of this procedure are ( I ) tionuniformity of the sample, (2) contamination, (3) variat,ion of exposure conditions such as wandering of the arc and change in line voltage, and (4)the photometric error. Twenty replicate analyses of the sample of sugar cane leaves showed 5 probable error of a single determination of approximately 10% of the mean. The use of solutions of both the standards and unknowns might possibly increase the precision, but would increase the time required for analysis and the danger of contaminat iim.
A N A L Y T I C A L EDITION
February, 1944
The factors affecting the accuracy itre (1) incomplete burning and (2) the influence of varying major constituents on the volatility and excitability of the molybdenum and beryllium atoms. I n this work the sample was always completely burned and standards approximating the composition of the samples were used. The lower limit of detectability of molybdenum depends to Bome extent on the material being analyzed. The absolute aensitivity of the 3170 A. line appears to be about 0.05 microgram under the conditions used here; hence, with a 10-mg. sample approximately 0.0005 % of molybdenum could be detected. Chemical concentration could increase the sensitivity, but would partially vitiate the advantages of a spectrographic method. COLORIMETRIC M E T H O D
C:ertaiu features of Sandell's procedure (1 7) have been conibined with some of those of Hoffman and Lundell (6) in this work. A solution of sexivalent molybdenum in a volume of 50 ml. was prepared, to which were added 7 ml. of conccntrated hydrochloric acid. To this solution, contained in a separatory funnel, 3 ml. of 10% potassium thiocyanate solution and 3 ml. of stannous chloride solution (10 grams of stannous chloride dihydrate in 100 ml. of 1 to 9 hydrochloric acid, prepared daily) were added in the order given, and mixed well after each addition. After 1 or 2 minutes the solution was extracted first with 10 ml. of ether (pretreated with one tenth its volume of e ual amounts of the potassium thiocyanate and stannous chlorile solutions), then with 5-ml. portions of ether until no additional color was observed in the ether extract. The combined extracts were run into a 25-ml. volumetric flask (for very small quantities a 10-ml. volumetric flask was used) and diluted to volume with the treated ether. The solution was compared in a Duboscq colorimeter with a standard which was prepared simultaneously and in the same manner from the standard ammonium molybdate solution. Experiments conducted with various interfering ions which would normally be present in solutions of plant or soil samples confirmed the findings of Hoffman and Lundell that the presence of iron affects the molybdenum thiocyanate complex color. This is an enhancing effect which reaches a maximum when about 2 mg. of iron are present, and additional iron has no further effect. Hoffman and Lundell recommend the addition of 10 mg. of ferric iron to every determination before reduction and extraction and this procedure was adopted in all cases where the iron was not already present. Other ions, including Ca++ (200 mg.), K+ (260 mg.), Na+ (200 mg.), Mg++ (20 mg.), Mn++ (2 mg.), Pb++ (0.2 mg.), Zn++ (0.2 mg.), Cu++ (0.2 mg.), Zr++++ (1 mg.), Ti+++ (1 mg.), Sod-- (80 mg.), Cr#07-- (0.7 mg.), PO,--.(100 mg.), B407-- (2 mg.), and SiOa-- (80 mg.) were added in the above amounts to both 500 and 50 micrograms of molyb-
139
denurn anti were fouiid to give no iiiterference in these proportions which are ordinarily encountered (83) in plant and soit samples. Other experiments confirmed previous statements on the instability of the molybdenum thiocyanate color. Standards and unknowns were therefore prepared simultaneously and compared immediately. A calibration curve was prepared (g4) by plotting the average reading-i.e., the arithmetic mean of 10 or more readings, with one cup set a t some predetermined heightagainst the ratio of the solution concentrations in the two cups. Although a straight-line relationship exists, the indicated lack of conformity to Beer's law shows that the standards and unknowns should have a difference in concentration of not more than 25%. The following procedure was finally adopted for ttrialyfiis of the samples: The material was first ignited a t 450" C. and then carefully mixed. One gram of this ash was weighed out, and 10 ml. of (1 to 4) hydrochloric acid were added and heated. The solution was filtered through an ashless filter paper, washed several times with water, and the filter paper and residue were transferred to a platinum dish and ignited a t 450" C. A few milliliters of water, several drops of sulfuric acid, and about 10 ml. of hydrofluoric :wid were added, evaporated on a steam bath, and ignited. . Water and hydrofluoric acid were added again and the evaporation was repeated. With plant materials there was often a residue of organic matter which was removed by again igniting the dish a t 450" C. A few milliliters of water and 1 ml. of hydrochloric acid were added, warmed, and the solution was made up to 50 ml. With soils there was sometimes a residue which was filtered off. The hydrochloric acid concentration was adjusted, 10 mg. ,of iron mere added if not already present, 3 ml. of potassium thiocyanate solution and 3 ml. of stannous chloride solution were added, the solution was extracted with several .portions of ether, diluted to volume, and the color compared with a standard of approximately the same concentration prepared simultaneously. Using this procedure, duplicate analyses were made on the ten fiamples previously analyzed spectrographically. The results are given in column 4 of Table I. The procedure used volatilizes the silicoii G S silicon tetrafluoride because it was found that molybdenum is often retained in t,he silica residue if a sodium carbonate fusion is used. The fusion also has the disadvantage of introducing some platinum into the solution, giving an interfering color which is especially serious when only a few micrograms of molybdenum are present. Xu molybdenum could be detected spectrographically in the residue from the hydrofluoric-sulfuric acid treatment nor was there any appreciable loss of molybdenum by bhia treatment, but the met,hod is sensitive only to about 0.05 microgram. To determine the accuracy and precision of the method a synthetic ash solut,ion was prepared containing 100 mg. of Kf and Gaff, 50 mg. of Mg++, 10 mg. of Na+ and Fe+++, 2 mg. of Al+++ and Mn++, and 1 mg. of Co++, Zn++, Cu++, and Ni+t, all as the chlorides, and 20 mg. of phosphorus as orthophosphoric acid. Known amounts of molybdenum were added and the analysis was carried out as previously described. Under these ideal conditions an accuracy of 5y0 and a probable error of 2.9% were secured. It is estimated that with plant or mil samples an accuracy of 15% could reasonably be expected. The lower limit of detectability with this procedure is approximately 1 microgram, but the precision is rather poor with this amount, in 10 ml. It is estimated that eight to ten det'erminatfions could be made in one day, although the speed and sensitivity might be increased by using photoelectric or microcolorimeteru. The advantages of this procedure are that no special apparatus is required arid very small proportions of molybdenum may he determined. On the other hand, the residual solution is probably not suitable for other determinations and each deterrniiiatioii requires relatively large amounts of sample (1 grani) which might not be available in all cases.
0
0.3
0.6
0.9
1.2
i.5
-
3170 RATIO 3131 Figure 2.
Plant Working Curve
1.8
POLAROGRAPHIC METHOD
The polarograph used was a photographic recording Heyrovski. instrument manufactured in Prague. The dropping electrode was made by sealing a 2.5-em. length of Pyrex "marine
140
I N D U S T R I A L A N D E N G I N E E R I N G CHEMISTRY
barometer tubing” with an internal diameter of about 0.05 mm. into the bottom of a 50-ml. distilling flask with a 2.5-cm. length of Pyrex tubing. The distilling flask was connected to a manometer and leveling bulb containing mercury to maintain a suitable pressure and dropping rate ( l a ) . Previous investigators, using fairly large amounts of molybdenum, have obtained satisfactory curves only in 18 N sulfuric acid (!8), 33% phosphoric acid (8), and ammonium nitrate-nitric acid (82) solutions. Preliminary experiments were made with known amounts of molybdenum from the standardized ammonium molybdate solution and various regulating solutions by removing oxygen with nitrogen and analyzing polarographically with suitable galvanometer sensitivity. Regulating solutions of 0.4 to 70y0phosphoric acid, 18 N sulfuric acid, sodium sulfate-sulfuric acid pH 1.5, sodium acetate-acetic acid pH 3.6, ammonium chloride-hydrochloric acid pH 1.1, 0.05 M potassium chloride, and ammonium nitratenitric acid pH 0.5 to 4.7 were used. Satisfactory curves were obtained with 20 to 35% phosphoric acid and various ammonium nitrate-nitric acid mixtures. In the latter case the step heights were markedly affected by changes in either hydrogen- or nitrate-ion concentration. The solution giving the most easily measured curves with sharper and better defined upper and lower “breaks” was one of pH approximately 1 and a nitrate concentration of 1.0 N . Uhl (bd) recommended the use of lactic acid with this solvtion, but its addition gave unsatisfactory curves with maxima.
It was hoped that it would be possible to dissolve the plant or soil material in a suitable acid, adjust the pH and nitrate-ion concentration, and analyze the 8olution directly with the polarograph. A study of various other ions, in proportions such as might be expected in solutions of plants and soils, showed that potassium, calcium, sodium, and magnesium nitrates had no effect. Khile small amounts of potassium chloride (1 and 5 ml. of 1 M potassium chloride in 50 ml. of solution containing 5 microgram of molybdenum per ml.) did not have a great effect upon the character of the polarogram, larger amounts (10 and 25 ml.) affected the limiting current angle markedly. This may be due to the chloride ion, since equivalent amounts of potassium sulfate had no such effect. The presence of phosphoric acid reduced the step height and small amounts of iron gave a wave just preceding and close enough to that of the molybdenum to cause interference. The interference of one element with another may sometimes be eliminated by complex formation (IO) and Uhl (Zb) stated that he could overcome the interference of iron by the addition of oxalate but not with citrate or fluoride. All were found unsatisfactory, as the first two either reduced the step height or introduced maxima and the fluoride eliminated the step altogether. Attempts to remove the iron by precipitation with sodium hydroxide ( l a ) or ammonium hydroxide (14) gave unsatisfactory results. The molybdenum was therefore separated by precipitation with alpha-benzoinoxime (9), dissolved, and the solution analyzed polarographically. The best conditions for precipitation of molybdenum with alpha-benzoinoxime are in solution3 containing 57, nitric, hydrochloric, or sulfuric acid with an excess of reagent and below 10” C. Vanadium, chromium, tungsten, palladium, columbium, gold, or tantalum interferes but the last five seldom are encountered in plants and soils and the first two cause no trouble if they are reduced with a little sulfurous acid before the benzoinoxime is added. Since no data were available on the precipitation of less than 50 micrograms of molybdenum in 200 ml. of solution, varying amounts of molybdenum were put in 50 mi. of solution containing 5% hydrochloric acid and analyzed. The solution was cooled below 10’ C., and 2 ml. of a 27?, alcoholic benzoinoxime solution and a few drops of bromine water were added. After standing 10 to 15 minutes with occasional stirring, the precipitate was filtered off, ignited a t 450” C., the residue dissolved in 3 drops of N sodium hydroxide, 1 ml. 2 N nitric ac,id, and 2 ml. of 4 N ammonium nitrate added, the solution diluted to 10 ml., oxygen removed, and analyzed polarographically. The wave heights were evaluated by measuring, at the half-wave potential, the vertical distance between straight-line extensions of the principal slope lines through the centers of oscillation amplitudes before and after the current step. The resultc: varied from 0.9%
Vol. 16, No. 2
loss with 100 micrograms, to 10% loss with 5 micrograms, and 30% loss with 1 microgram. Known amounts of molybdenum were added to the same synthetic ash solution as used with the colorimetric method, and determinined as outlined above. For comparison, solutions containing the same quantities of molybdenum, ammonium nitrate and nitric acid and 2 micrograms of iron (since the small quantities of iron carried down with the alpha-benzoinoxime caused a slight shift in the half-wave potential), were prepared and analyzed polarographically. A calibration curve for a drop rate of 1.8 seconds and calculated to 1/z0 galvanometer sensitivity shows a straight-line relationship between concentration and step height up to 5 micrograms of molybdenum per ml. The soil and plant samples were prepared for precipitation as described for the colorimetric procedure. The hydrochloric acid concentration was adjusted to 5% and the precipitation made as above, adding a little sulfurous acid in the case of soils. With the soil samples containing a large proportion of iron, the alkaline solution of the residue after ignition was filtered through a small, sintered-glass funnel to remove the iron carried down with the precipitate. The results of the analysis of the plant and soil samples by this method are given in column 5 of Table I. The accuracy of this procedure varies with the amount of molybdenum present and it should not be used for materials containing less than 0.0005% molybdenum unless the samples are larger than 1 gram. It is estimated that 6 t o 8 determinations could be made per day. Because of the separation procedure, molybdenum is the only constituent which can be determined. Some copper is precipitated by the benzoinoxime, and gives a small step just preceding the molybdenum. The corrected half-wave potential was determined t o be -0.42 volt, using a glass-jointed salt bridge as suggested by Kolthoff and Lingane (IO). The marked tendency of molybdenum to form heteropoly acids ( 8 ) , the enhanced reducibility of these molybdenum heteropoly acids (a), and the fact that no step is obtained in hydrochloric acid solution but is in nitric, sulfuric, or phosphoric solutions make it seem likely that the step obtained in this study may be due to the reduction of a nitrate-molybdate heteropoly acid. LITERATURE CITED
(1) Arnon and Stout, Plant $PhysioZ.,14, 599 (1939). (2) Emeleus and Anderson, Modern Aspects of Inorganic Cheinistry”, New York, D. Van Nostrand Co., 1938. (3) Feigl, “Specific and Special Reactions”, New York, Nordeman Publishing Co., 1940. (4) Ferguson, Lewis, and Watson, Nature, 141, 553 (1938). (5) Hiskey and Meloche, J . A m . Chem. SOC.,62, 1565, 1819 (1940) ; 63, 964 (1941). (6) Hoffman and Lundell, Bur. Standards J . Research, 23, 497 (1939). (7) Hokhshtein, J. Gcn. Chem. (U.S.S.R.) 10, 1725 (1940). (8) Kanevsky and Shvartsburd, Zasodskaua Lab., 9, 283 (1940). (9) Knowles, BUT.Sta?dards J. Research, 9, 1 (1932). (10) Kolthoff and Lingane, “Polarography”, New York, Interscience Publishers, 1941. (11) Leutwein, 2. Mineral. Geol., 1940A,129. (12) Lingane and Kolthoff, J . Am. Chem. Soc., 61, 825 (1939). (13) Lundell and Hoffman, “Outlines of Methods of Chemical Analysis”, Kew York, John Wiley & Sons, 1938. (14) Malowan, 2. anal. Chem., 79, 201 (1929). (15) Marmoy, J . SOC.Chem. Ind., 58, 275 (1939). (16) Rogers, Gall, Gaddum, and Barnette, Univ. Fla. 4 g r . Expt Sta., Bull. 341 (1939). (17) Sandell, IND.ENQ.CHEM.,-1N.4L. E D , 8, 336 (1936). (18) Stackelberg, Klinger, Koch, and Krath, Tech. Mitt. K r u p p Forschungsber., 2, 59 (1939). (19) Standen and Kovach, Proc. A m . SOC.Testing Materials, 35, Part 11, 79 (1935). (20) Steinberg, J. Agr. Research, 55, 891 (1937). (21) Thanheiser and Willems, Arch. Eisenhiittenw., 13, 73 (1939). (22j Uhl, 2. anal. Chena., 110,102 (1937). (23) U. S. Dept. Agr.. Yearbookof Agriculture, ”Soils and Minerals”, p. 778, 1938. (24) Yoe, “Photometric Chemical Analysis”, Vol. I, p. 70, New York, John Wiley & Sons, 1928.
