March 15.1933
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limiting factor is the upper limit of the standard rather than average being 0.834. These results indicate the variation the amount of potassium iodide present. that may be expected from the analysis of single kernels owing ANALYSISOF C. P. THALLIUM SALTS. The method devel- to unequal distribution of the thallium. A 3-gram sample of oped is essentially for samples of low thallium content, but is the same material (approximately seventy-five kernels) was satisfactory and extremely rapid for the assay of high per- pulverized and thoroughly mixed. Two analyses from this centage salts where an accuracy greater than *2 per cent is sample gave values of 0.79 and 0.84 per cent, the average not required. Analysis of the e. P. acetate, chloride, nitrate, being 0.815. Three blank tests run on strychnine-coated and carbonate gave values varying from the theoretical thal- barley did not liberate a measurable quantity of iodine in any lium percentage by 0.13 to 1.76 per cent. At the other ex- case. treme, analyses of small samples containing 0.04 to 0.1 mg. A dog was given thallium sulfate by capsule, equivalent to gave values in error by 3 to 5 per cent. 25 mg. of thallium per kg. of body weight. The animal was INTERFERENCE BY OTHER METALS. Analyses were conplaced in a metabolism cage, and the total urine for each day ducted on aliquots containing 0.5 mg. of thallium to which collected. Triple analyses on the samples for the first two were added 5 mg. of another metal in the form of a soluble days show the checks obtained: first day, 24.4 * 0.4 mg.; salt. These tests were made on copper, lead, iron, arsenic, second day, 13.9 * 0.3 mg. Blank tests made on normal mercury, tungsten, and molybdenum, the complete analysis urine gave zero values. being used exclusive of the preliminary oxidation with potasThe following data are illustrative of results obtained' on sium chlorate. The values obtained were in all cases within tissues from thallium-poisoned game birds on which a com0.01 to 0.02 mg. of the true value, indicating no appreciable plete report will appear elsewhere. The values shown are interference by the metals tested. Attempts to assay thal- on heart muscle from thallium-poisoned geese : lium chromate by the colorimetric method were not successTIME TIME THALLIUMUNTIL THALLIUMTHALLIUMUNTIL THALLIUM ful, however. No method was found to eliminate satisfacDOSAQE DEATH FOUND DOsAQE DEATH FOUND torily the chromate interference. Mo./ko. Daw Mob. Mg./kg. Days Mg./kg. PREPARED SAMPLES AND BLANKS. A number of blank tests 1 33.2 32 40 8 20.0 28 2 25.3 20 13 10.1 were made on 20-gram samples of beef heart and liver, the iodine values obtained ranging from 0.01 to 0.02 mg. of thalLITERATURE CITED lium. Prepared samples containing 0.5 mg. of thallium per 20 grams of meat were carried through the complete analysis (1) Baldeschwieler, E . L., IND.ENG.CHEM.,Anal. Ed., 4,101 (1932). (2) Fridli, R., Deut. 2. ges. gericht. Med., 15, 478 (1930). with errors of 1 to 5 per cent. G. H., Levine, Max, and Buchanan, J. H., IND.ENG. ILLUSTRATIVE RESULTSON GRAIN,URINE,AND TISSUES. (3) Nelson, CHEM.,Anal. Ed., 4,56 (1932). Thallium-coated wheat, prepared on the ratio of one pound (4) Noyes, A. A., Bray, W. C., and Spear, E . B., J. Am. Chem. SOC., 30, 515 (Procedure 65a), 517 (Procedure 65d) (1908). of thallium sulfate to 100 pounds of wheat, and thus containing 0.81 per cent of metallic thallium, was used for analysis. RECEIVED September 1, 1932. These experiments were carried out with Five analyses on individual kernels made by direct nitric the codperation of the Hooper Foundation for Medical Research, San Franacid oxidation gave values from 0.605 to 1.11 per cent, the cisco, Calif.
Microdetermination of Calcium in Sea Water PAUL L. KIRKAND ERIKG. MOBERG Division of Biochemistry, University of California Medical School, Berkeley, Calif., and Scripps Institution of Oceanography, La Jolla, Calif.
F
OR many investigations in oceanography and marine
biology information concerning the calcium content of sea water is required. Previous workers have determined calcium in sea water by precipitating with oxalate, igniting, and weighing as the oxide. A summary of the results obtained is given by Thompson and Wright (C), who also determined the concentration of calcium in water from the Puget Sound and the Gulf of Alaska. They found that in these regions calcium bears a constant ratio to the chloride, and computations made from the data of other investigators indicated that this is the case also in other parts of the sea. It is especially noteworthy that 77 samples collected by the Challenger Expedition from various parts of the world and analyzed by Dittmar (1) gave an average calcium-chloride ratio but slightly higher than that obtained by Thompson and Wright. It thus appears that for most purposes the calcium content of normal sea water can be calculated with sufficient accuracy from the chlorinity or from some other easily determined component or property of the water directly related to the chlorinity. However, there is often need for determining calcium in connection with experimental studies
in which sea water is used and in investigating water from localities where calcium salts may be dissolved or precipitated. For many such investigations, which usually call for numerous analyses, the standard gravimetric method is too time-consuming and requires larger samples than are often available. For that reason the applicability to seawater analysis of the microcalcium method described by Kirk and Schmidt (8) was studied and it was found to give satisfactory results after certain modifications were made. Because of these modifications and because of the necessity for close attention to certain details, a rather detailed description of the technic finally adopted is given in this paper. Nearly all the chemical constituents of sea water vary with depth and this is true also of a number of factors which affect the solubility of calcium salts. By several workers calcium has been determined in water from various depths of the sea, but there is no record of a detailed study of its vertical distribution in any locality. After perfecting the analytical method, such a study was consequently made and the results, which also indicate the order of accuracy of the method, are included in this paper.
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96
METHOD APPARATUS AND REAGENTS. The apparatus needed for the determination of calcium in sea water is the same as that used by Kirk and Schmidt ( 2 ) . It consists primarily of a set of microfilters (obtainable from the Central Scientific Co., Chicago, Ill.) which have been described in detail in the above-mentioned article. These are inserted in rubber stoppers which have been cut to half their usual thickness and placed in a battery of suction flasks attached to a vacuum manifold. I n addition, beakers of 150 ml. capacity, with watch glasses and small stirring rods to fit, are required.
Vacuum
To Ki?lnOy B o t t l e
FIGURE1 Two special wash bottles are desirable, one for ammonia and one for 2 N sulfuric acid. The ammonia bottle should be fitted with a plain rubber bulb having no intake valve, thus making it easy to stop the flow of ammonia by simply releasing the pressure on the bulb. The acid wash bottle is conveniently made from an Erlenmeyer flask of about 200 ml. capacity and should have an outlet tube with a fairly fine tip and an air intake extending to the bottom of the bottle. This bottle is used by inverting it, thus allowing the acid to flow out in a fine stream, and it should be wrapped with asbestos or some other material which is a relatively poor conductor of heat, since the acid is heated before use. For the titrations, a 10-ml. buret calibrated in 0.02 ml. is desirable. I n order to avoid contact of permanganate with rubber, and to simplify the filling of the buret, a convenient attachment is shown in Figure 1. I t s use is practically selfexplanatory. If a vacuum line is available, the tube T is attached to it, the vacuum cock opened slightly, and the outlet 0 is closed with the fingers to fill the buret. Simply removing the finger stops the flow. The reagents needed are as follows: 1. Potassium permanganate solution, approximately 0.06 N , kept in a dark bottle. This solution is made b dissolving about 2.0 grams of potassium permanganate in each$ter of solution and allowing to stand a t least 24 hours in a clean glassstoppered bottle t o permit the manganic oxide to settle out. The solution is then filtered through sintered glass into the stock bottle, or if such filters are not available, it may be carefully si honed into the bottle, being careful not to disturb the layer ofmanganic oxide, and discarding the first solution coming through the siphon and the last inch in the bottom of the original bottle. The permanganate must never come in contact with rubber or other organic matter. It is standardized against pure sodium oxalate or oxalic acid. The sodium oxalate obtainable from the Bureau of Standards is to be preferred. Permanganate approximately 0.06 N is chosen because about 9 ml. of this solution will titrate the calcium from 25 ml. of sea water. 2. Ammonium oxalate solution, 4 er cent. 3. Hydrochloric acid solution ma& by diluting 1 volume of concentrated acid with 3 volumes of water. 4. Ammonium hydroxide solution, approximately 2 N .
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5. Sulfuric acid solution, approximately 2 N . 6. Ammonium hydroxide solution, about 1 to 6, saturated with calcium oxalate. This is filtered before use. 7. Bromocresol purple solution, 0.5 per cent. 8. Asbestos] preferabIy short fiber, Italian, washed and ignited (obtainahle from the J. T. Baker Chemical Co., Phillipsburg, N. J.). Before use this is rubbed in a mortar to shorten the fibers further, after which it is heated with dilute sulfuric acid. Potassium permanganate is added to a faint permanent pink, followed by more heating. When it no longer discolors permanganate it is thoroughly washed with water and final1 suspended in a Aask of water. Such asbestos gives no blanz titration with permanganate.
PROCEDURE. A sample of sea water is measured with a 25-ml. pipet into a 150-ml. beaker. About 10 ml. of hydrochloric acid solution are added, followed by 20 ml. of ammonium oxalate solution. One drop of bromocresol purple is added and the solution heated on an electric hot plate. When the solution begins to steam, 2 N ammonium hydroxide is added slowly with stirring to the turning point of the indicator. The last portions should be added drop by drop. The hot solution is removed from the hot plate and covered with a watch glass. The solution, which should be faintly purple when cool, is allowed to stand about 4 hours and is then filtered through the microfilters mentioned above. A filter is prepared by first placing the platinum disk on the shoulder of the glass portion, making sure it is squarely seated. Asbestos, treated as described above, is next run in, preferably from a pipet with an enlarged orifice. Enough is used to form a layer about 1 mm. thick. This is washed under vacuum two or three times. In case the filter so formed filters unusually slowly, it is advisable to add a few milliliters of alcohol, followed by a little ether. Water is again used following the ether. This treatment will increase the speed of filtration in nearly every case. The outside of the lip of the beaker containing the solution is greased with a little vaseline to prevent loss of precipitate over the edge, after which the solution is filtered under a mild vacuum. The inside of the beaker .is rinsed with one portion of dilute ammonia saturated with calcium oxalate. (This solution should contain no excess oxalate and is conveniently used in the ammonia wash bottle described above.) This wash solution is passed through the filter. The filter pad and disk are now pushed out of the filter into the beaker by means of the stirring rod which should be small enough in diameter to be inserted easily through the bottom of the filter. The calcium oxalate which adheres to the sides of the filter is washed out first with a little water, then with hot 2 N sulfuric acid from the acid wash bottle, and finally with a little more water. The sides of the beaker are washed down with the hot acid and the solution is ready for its second precipitation. This is carried out exactly as before by adding 20 ml. of ammonium oxalate, one drop of bromocresol purple and then ammonia to the hot solution to the turning point of the indicator. The hydrochloric acid is omitted, since the solution is already strongly acid. The solution is allowed to stand 4 hours and is filtered again exactly as before. This brings the total asbestos in the filter to twice the usual amount, but does not change the speed of filtration appreciably, The washing of the second precipitate must be carried out carefully. The authors habitually use three rinsings of the beaker and two further washings of the filter with the dilute ammonia saturated with calcium oxalate. The precipitate, filter pad, and disk are transferred to the precipitation beaker as before with hot sulfuric acid and are then ready to be titrated with the permanganate solution. For the titration, the precautions are the usual ones for permanganate titrations. The solution should be a t about
INDUSTRIAL AND ENGINEERING CHEMISTRY
March 15, 1933
70' C., especially at the end point, and the permanganate is not added faster than it can be decolorized. The first permanent pink is the end point.
DISCUSSION OF METHOD Sea water contains approximately three times as much magnesium as calcium. With so large an excess of magnesium it has so far proved impossible to adjust the conditions so as completely to separate the two elements in one precipitation. The amount of magnesium oxalate occluded on the calcium oxalate after the first precipitation represents about 6 to 8 per cent of the total oxalate precipitated. Thompson and Wright (4) used triplicate precipitations and present data to show that this is essential. They do not state their method of precipitation, but presumably they followed the usual procedure of making the solution definitely alkaline with ammonia. If so, the necessity for three precipitations can readily be explained, since in alkaline solution much larger quantities of magnesium oxalate are occluded than in neutral or slightly acid solutions. The authors have repeatedly used two and three precipitations on identical samples of sea water and find identical values for the titration within the limit of accuracy of the method. The presence of ammonium chloride has long been thought to aid in preventing the occlusion of magnesium oxalate. The authors have not specifically checked this point, though in a recent publication Popoff, Waldbauer, and McCann (3) state that ammonium chloride has no influence, and also point out that long digestion in alkaline solution increases the magnesium content of the precipitate. The present authors have not used an alkaline solution nor long digestion, inasmuch as a t the pH chosen the size of crystals is very satisfactory and long digestion is entirely unnecessary. Like the above-mentioned investigators, they have found that the amount of ammonium oxalate added is the most important factor. They have used 5, 10, and 20 ml. of ammonium oxalate, and get consistent results and complete recovery only when 20 ml. are used.
EXPERIMENTAL I n order to determine the absolute accuracy of which this method is capable, the authors prepared from reagent chemicals a solution to contain calcium, magnesium, and sodium, in practically the same concentrations as in sea water. (Some difficulty was experienced in obtaining calcium-free magnesium sulfate.) Of this solution five identical samples were analyzed. The results, which are shown in Table I , indi7 cent. cate a maximum error of ~ 0 . per TABLE I. ANALYSISOF A SOLUTION OF KNOWN CALCIUM CONTENT CALCIUM
PR~SENT Mu. 5.64 5.64 5.64 5.64 5.64 Average Maximum
CALCIUM FOUND
Mg. 5.66 5.60 5.68 5.66 5.63 5.646
DIBPERENCB Mg. % $0.02 $0.35 -0.04 -0.71 +0.04 $0.71 +0.02 $0.35 -0.01 -0.18 50.026 k0.46 *0.04 &0.71
The method was further checked by analyzing a set of sea-water samples to half of which had been added known and varying quantities of calcium. Table I1 shows the results. It may be noted that six analyses of the original water gave an average calcium content of 10.40 mg. per sample with a maximum range of 0.07 mg. The samples to which calcium had been added gave practically the same average after correcting for the added calcium.
97
TABLE11. ANALYSISOF SEA WATERCONTAININQ VARYING QUANTITIES OF CALCIUM (All calcium values for 25-ml.samples) TOTAL CALCIUM ORIGINAL CALCIUM CALCIUM FOUNDMINUS CALCIUM ADDED FOUND CALCIUM ADDED Mg.
Mg.
10.06 10.04 10.02 10.03 10.02 10.09
1.13 2.26 2.26 2.26 2.26 5.64 5.64
MO.
Mg.
11.16 12.32 12.30 12.24 12.31 15.61 15.60
10.03 10.06 10.04 9.98 10.05 9.97" 9.96O Average 10.043 10.032 a Values omit,ted in computing average because upper limit of recoverable quantity of calcium is apparently reached at about this point.
To ascertain the variation in the calcium content with depth along the coast of southern California and to obtain data for comparing with results obtained by other methods and for other localities, calcium was determined on seventeen water samples collected on July 14, 1931, from different depths a t a point ten miles seaward from the Scripps Institution pier. The samples were also analyzed for chloride and the results are shown in Table 111. This table gives further information regarding the relative accuracy of the calcium method, the maximum deviation from the average of two determinations being 2.4 mg. per kg., corresponding to about *0.5 per cent. For all but five of the seventeen samples the deviation is less than 1.0 mg. per kg. TABLE111. CALCIUM IN SEAWATERFROM VARIOUS DEPTHS -CALCIUM-DEPTH C1 Meters C . / k g . 0 18.78 5 18.78 10 18.77 15 18.72 20 18.70 25 18.67 30 18.62 35 18.60 40 18.61 50 18.61 75 18.70 100 18.78 150 18.86 200 18.96 250 19.04 300 19.01 400 19.00 Av. 18.77 Max. 19.04 Mjn. 18.60 Diff. 0.44
Deviation from av
A B Av. G./kg. C./ko. G./ks. G./kg. 0.4055 0.4048 0.4052 0.0004. 0.4040 0.4063 0,40520.0012 0.4063 0.4075 0.4069 0.0006 0.4056 0.4064 0.4060 0.0004 0.4058 0.4067 0,40620.0006 0.3974 0.4017 0.3996 0.0022 0.4014 0.4017 0.4016 0.0002 0.3986 0.3982 0.3984 0.0002 0.4010 0.4010 0.4010 0. 0.4014 0.4017 0.4016 0.0002 0.4048 0.4056 0.4052 0.0004 0.4001 0.4028 0,40150.0014 0.4071 0.4024 0.4048 0.0024 0.4051 0.4062 0.4057 0.0006 0.4116 0.4089 0.4103 0.0014 0.4062 0.4081 0.4072 0.0010 0.4089 0.4105 0.4097 0.0008 . . . . 0.4045 0.0008 . . . . . . . . 0.4103 0.0024 ,.., , . . . 0.3984 0. ... . . 0,01190.0024
.... . ..
--A
RATIOCa:ClB Av.
0.02159 0.02156 0.02157 0.02151 0.02163 0.02157 0.02165 0.02171 0.02168 0.02167 0.02171 0.02169 0.02169 0.02175 0.02172 0.02129 0.02152 0.02141 0.02156 0.02157 0.02157 0.02143 0.02141 0.02142 0.02155 0.02155 0.02155 0.02157 0.02169 0.02158 0.02167 0.02165 0.02169 0.02130 0,02145 0.02138 0,02159 0.02134 0.02147 0.021370.02142 0.02148 0.02140 0.02162 0.02155 0.02137 0.02147 0.02142 0.02152 0.02161 0.02167 . . . . . . . . . . 0,02154 . . . . . . . . . . 0.02172 ..... 0.02138 . . . 0 00034
. ..
..... .. . .
The average calcium content of the water was found to be 0.405 gram per kg. A slight variation with depth is indicated, corresponding in general with the variation in chlorinity which is clearly shown in the table. The average calcium-chloride ratio is 0.02154, which is in remarkably close agreement with the ratio, 0.02150, found by the careful work of Thompson and Wright (4). It is to be noted that these authors analyzed water mainly from the Puget Sound and the Gulf of Georgia, where the chlorinity is decidedly lower than along the California coast. Their ratios, which represent composite samples, are somewhat more constant than those in the present investigation, ranging from 0.02139 to 0.02160, as compared with a range from 0.02138 to 0.02172. From their results Thompson and Wright conclude that in sea water calcium occurs in constant ratio to the chlorinity, within the limits of accuracy of the analytical methods. I n general, the same is true of the region investigated here, but in the case of some of the samples the variation in the ratios is not wholly attributable to known experimental errors, and the results a t least suggest that in certain parts
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98
of the sea biological activity or other factors may affect the calcium content of the water to a measurable extent. Further investigation of this subject is desirable and may throw some light on certain geological and biological phenomena taking place in the sea.
SUMMARY A micromethod for determining calcium has been adapted to the analysis of sea water and the technic is described in detail. The method is comparatively rapid and is capable of an accuracy of about 0.5 per cent or better. The results of calcium determinations on sea water from
Vol. 5 , No. 2
various depths are reported. These results confirm the conclusion drawn by previous workers that in sea water calcium is one of the substances present in constant proportion of the total salt.
LITERATURE CITED Dittmar, W., “Voyage of H. M. 8. Challenger,” Phys. Chem., 1, part I, p. 1, Edinburgh, 1884. (2) Kirk, P. L., and Schmidt,C. L. A., J. Bid. Chem., 83,311 (1929). (3) Poooff. S.,Waldbauer, L.. and McCann, D. C., IXD. ENG.CHEM., Anal. Ed., 4, 43 (1932). (4)Thompson, T. G., and Wright, C. C., J. Am. Chem. Soc., 52, 915 (1)
(1930). RIUCEIVPD October 17, 1932.
Relation between Volume of Respiration Chamber and Concentration of Carbon Dioxide in End Sample and in Composite Sample of Air M, KLEIBER,Division of Animal Husbandry, College of Agriculture, University of California, Davis, Calif.
F
OR the measurement of the production of carbon di- carbon dioxide in a sample taken at a certain moment) the oxide and the consumption of oxygen, the subject author developed the equation in an earlier study (1). The of the experiment is either connected to or enclosed present paper shows that in many cases the composition of the in a respiration apparatus. The first method implies the composite sample is more important than that of the momenuse of a mouthpiece with nose clamp, as generally used on tary samples, and carries out the calculation for the influence humans for purposes of clinical research; or connection of the volume of the chamber, the rate of ventilation, and by means of a tracheotomy tube in the case of animals. For the rate of production upon the composition of the composite those experiments with animals in which tracheotomy is to be sample. An equation is given from which investigators in avoided, the second method must be used. The animal is this field can predict the influence of an error in gas analysis enclosed in a respiration chamber which is generally connected upon the error in the result for any apparatus. This is a guide for the selection of the apparatus for a given purpose. to a ventilating system. For the construction of a respiration apparatus for measuring the metabolism of animals, it is of interest to know the RELATIVESIGNIFICANCE OF MOMENTARY AND relation between the volume of the chamber, the rate of COMPOSITE SAMPLES ventilation, the rate of respiratory exchange, and the In an apparatus of the Tigerstedt type, a momentary concentration of the carbon dioxide and oxygen in the - sample is- t a k e n from the chamber. The d e g r e e of air in the chamber at the accuracy of the result of a beginning and at the end of t r i a l m a y t h e n be prea period, and a composite dicted, provided this degree sample which c o n s i s t s of is limited by the accuracy small a m o u n t s of the air of the gas analysis. taken regularly during the With a given rate of proe x p e r i m e n t is collected. duction in a chamber and The result for the carbon at a certain rate of ventiladioxide production is calcution, the concentration of lated according to the equathe product (for example, tion: c a r b o n d i o x i d e ) is t h e s m a l l e r , the l a r g e r the R = V(C, - Co) chamber. The knowledge LXtXC, of this general r e l a t i o n is, where however, i n a d e q u a t e for R = amount of carbon estimating the influenceof a dioxide produced certain error in gas anaIysis in the chamber upon the error in the result in liters V = volume of chamber of an experiment. For that in liters purpose the functions must L = intensity of ventilabe expressed in mathematition (liters per cal terms. The equations hour) t = time in hours for these relations seem not 0 Co = concentrationof to have been w o r k e d out. V O umc ~ of chamber in W m . carbon dioxide at For a p a r t of t h e p r o b thestart (momenFIGURE 1. INFLUENCE OF VOLUMEOF CHAMBER ON CONCENl e m ( c o n c e n t r a t i o n of tary sample) TRATION OF CARBONDIOXIDEAND RESULTING ERROR
+
