V O L U M E 2 5 , NO. 9, S E P T E M B E R 1 9 5 3
1413 _____
Table I. .\rialq-sis of Synthetic 7)Zixtures of Phthalic .4cid and Phthalic .4nhydride Synthetic Mixture Anhydride Acid
Error, % _ _ _ -4nhydride Acid
Found Anhydride Acid
~
.
extracts to pH 2 with approximately 3 N hydrochloric acid, transfer to a 100-ml. volumetric flask, and dilute to the mark with 0.1 N hydrochloric acid. Make appropriate dilutions of the solution with 0.1 iV hydrochloric acid and measure the absorbance a t 276 mp in 1.0-cm. cells against a blank of 0.1 N hydrochloric acid. For maximum accuracy, the absorbance should fall in the vicinity of 0.4 absorbance units. Calculations. Calculate the concentration of phthalic acid in grams per liter, C p , of the final diluted solution by the following equation : cp = A / u , where A is the measured absorbance and up is the absorptivity value found for pure phthalic acid in the calibration. Calculate the percentage of phthalic acid in the original sample from the following equation: C p X aliquot factor X 100 acid, % weight of
Table 11. Determination of Phthalic Acid i n Commercial Phthalic 4nhydride _
Acid, %
Sample Source A Lot 1 Lot 2 Lot 3 Source B Source C Source D Lot 1 Lot 2
0 20 0.17 0 16
Average.
0 26 0 24 0.12
0.23 0.21 0.14
0.17 0.13 0.21
0.18
0 . 9 5 0.86 0.33 0.27
0.91 0.30
0.18
cc
0.l i
Calculation of phthalic. aiihydride ma)- be made by the same formulas, substituting the valurs found for phthalic anhydride. Results obtained from analyses of both synthetic and commercial samples (Tables I and 11) show that the standard deviation of a single determination, as calculated from differences between duplicate determinations, was 0.045%. LITERATURE CITED
(1) S h r e v e , 0. D., a n d Heet,her, AI. R., . ~ N A L .CHEM.,23, 441 (1951). ( 2 ) Siggia, S., a n d F l o r a m o , PI'. A . Ibid., 25,797 (1953). (3) Vogel, -4. I., " P r a c t i c a l O r y a n i r C h e m i s t r y , " p. 491, Ken- York, L o n g m a n s . Green :nid Co.. 194R. R E c R i w n March 20, 19.53.
Arrepted .June A. 1953.
Preparation of Water Samples for Deuterium Analysis in the Mass Spectrometer FRANCIS P. CHINARD AND THEODORE ENNS Departments of Medicine and of Physiological Chemistry, The Johns Hopkins I riirersif>.School of Medicine, Baltimore. Md. of the distribution oi heavy water in animal organI isms, . the limiting factor in the number of deuterium analyses N STUDIES
is the reduction of the separate, water samples rather than the actual measurements in the mass spectrometer. Use of conventional zinc reduction trains (2) with transfer of the gas into a sample tube by means of a Toeppler pump is tedious and time-consuming [Other procedures involving reaction of water with methyl magnesium iodide (4)or n i t h dirthyl zinc. ( 1 ) arc not suited to routine analysrq 1
+-5cm.-
Zn DUST
PYREX WOOL
fore thc thickened constriction is made. Approximately 2 grams of zinc dust are added to each tube. The tubes are then heated in a drying oven a t about 120" C. overnight and allowed to cool in a desiccator over calcium chloride. The pipets to be used for introducing the samples into t'he tubes are similarly treated. A jig, such as is shown in Figure 2, is advisable. The st,op cock is initially in position 11, and a sample tube is connect'ed to the vacuum system by means of vacuum tubing. (The end of t,he sample tube is made to touch the end of the stopcock side arm so that the exposed surface of rubber is kept to a minimum.) The cock is turned to position 111; the calcium chloride tube is thus connected t o the sample tube. The sample tube is then gently heated with a small flame in order to drive off residual water. The cock is then t'urned to position I. The sample tube is allowed to cool in place while the sample (approximately 0.01 ml.) is taken up in a pipet. The stopcock is now turned to position 11, and the sample tube is disconnected.
Figure 1. Sample Tuhe Because of the large volume and surface of the apparatus, considerable care is required to avoid contamination of the samples and to ensure complete reduction of the water (2). I n addition, improper packing of the zinc and the progressive oxidation of the zinc may result in incomplete reduction of the water samples. The surface and volume of the apparatus may be considerably reduced, and the tedious inconvenience of renewing the zinc may be eliminated by heating the water samples in the presence of zinc in individual sample tubes. The number of samples which can be processed a t one time is limited only by the size of the available muffle furnace. EXPERI\IE\T&L
The sample tubes are shown 111 Figure 1. The seal of the guard tube of the capillary end need not be perfect if the entire tube is introduced into the mass spectrometer system. The borosilicate glass wool is used to prevent gross contamination of the mass spectrometer with zinc dust; it should be inserted be-
TO PUMP
-
,"-&-
rubber
I
IS AMPLE c A
h
U
SOLID C 0 2
PYREX
I
WOOL CaC$
II
m
POSITION
Figure 2. Diagram of Jig The sample is introdured while the tube is a t a temperature between 45" and 50" C. The sample tube is now quickly reconnected to the rubber tubing and the cock is turned to position 111. The pressure in the sample tube is thus reduced and most of the atmospheric moisture removed. The sample is then frozen by the application of solid carbon dioxide to the outqide
1414 -
ANALYTICAL CHEMISTRY ~
Table I. Found, Atom '% D 0.011 0.016
Sample4 14
Illustrative data of analyses with the mass spectrometer are shown in Table I.
Results .%tom % Excessb Found Calculated
0.017
2
0.080
0 Oil
0.063
3
0 ,>A 0 . %57
0 ,34 0 .ii
0 58 0 58
4
0.6.5 0 64 1 17 1.17
0 63 0 62
0 63 0 63
11.7
1 I6
5
113
1 Ih
6
2.83 2.74
;8:7 2 -.
2 90 2 90
7
2.90
2 8%
2 83
8
3.71 5.66 5.2:
3.70
3.61
5.64 5 23
a , 80
9
4 . 80
6.12 6 2.7 6.13 10 6.11 6.10 6.23 a Sample 1 is for Baltimore tap water. Samples 2 , 4, 7, and 8 were prepared b y dilution of deuterium oxide with t a p or once distilled water; samples 3, 5 , 6,9. and 10 were prepared b y dilution of deuterium oxide with whole blood. T h e actual values for calculated atom per cent excess may deviate by = t 2 % from the values given. 6 Atom % D minus average atom % D of tap water. C The average value, 0.015 atom % ' D is in agreement with the d a t a cited in ( 8 ) .
of the tube. As soon as the sample is frozen, the cock is turned to position I, and the sample tube is sealed a t the constriction. (The solid carbon dioxide need not be applied during the sealing.) Approximately 20 samples can be prepared per hour. (The vacuum pump should be protected from possible introduction of zinc dust by an appropriately placed plug of borosilicate glass Wool.)
The sample tubes are then placed in individual brass tubes (to prevent catastrophes in case one of the tubes explodes) and are heated in a suitable furnace a t 400" f 10" C. for 30 minutes. It is advisable to allow the furnace to cool to near room temperature before the door is opened.
DISCUSSION
The sample tubes after reduction of the mater are small bombs The samples of water used should be no greater than 0.01 ml. ; a pressure of approximately 24 atmospheres may be expected from 0.1 ml. of water in a 10-ml. sample tube. The sealed sample tubes should never be heated in an open flame. Data on samples containing greater enrichments of deuterium than 7 atom % excess are not included; in the experiments maximum sample enrichment is less than this value. Accurate analysis of highly enriched samples would have required conversion of the mass spectrometer to an instrument for deuterium gas. This was not considered justifiable in view of the purpose for which the mass spectrometer was intended: hydrogen isotope analysis. The "memory effect" is discussed in detail by Kirshenbaum (5). The procedure described here permits more numerous analyses to be made and reduces or eliminates certain of the sources of wror inherent in the conventional zinc reduction trains; it doe3 not affect in any n a v the memory effect arising from adsorbed water in the mass spectrometer itself. ACKNOWLEDGMENT
This work was supported in part by a grant from the Life Insurance Medical Research Fund and in part by contract No. \'-lo01 M-527 between the Veterans Administration and the Johns Hopkins University. LITERATURE CITED
(1) Friedman, L., and Irsa, -1. P., ANAL.CHEM.,24,876 (1952). (2) Graff, J.,and Rittenberg, D., Ibid., 24, 878 (1952). (3) Kirshenbaum, I., "Physical Properties and Analysis of Heavy Water," p. 130, New York, McGraw-Hill Book Co., 1951. (4)Orchin, M., mender, I., arid Friedel. R. .1.,.%s.\L. CHEY..21, 1072 (1919).
RECEIVED for review Sovember 3, 1932. Accepted April
1. 19S3
Colorimetric Determination of Calcium with Ammonium Purpurate MAX B. WILLIAMS
AND JAMES H. MOSER Oregon State College, Corcallis, Ore.
development of new colorimetric methods for the deterT mination of calcium has been curtailed by the limited availability of suitable reagents that change color in the presence of this HE
ion. The colorimetric method in the most general use a t this time consists of precipitating calcium oxalate with a known excess of oxalate reagent, then determining the residual oxalate concentration colorimetrically by measuring the color remaining after the addition of a known amount of standard permanganate or ceric solution. This method, while good and fairly free from interferences, is not applicable to very low concentrations of calcium (0.1 to 10 mg. per liter) for the possibi1it.v of precipitate loss is relatively great; also, it is often difficult to obtain complete precipitation of calcium ovalate when certain organic materials are present. Schwarzenbach and Gysling ( 4 ) investigated the formation of si colored complex of ammonium purpurate (murexide), ?;H*CsH,Os"., with calcium in aqueous solution. The structural forniula for the purpurate ion is
r
TH-C-0
/
O=C--SH
\
SH-CLO
They showed that the ratio of calcium ion to purpurate ion wm 1 to 1, with an equilibrium constant for the formation of the calcium complex of approximately 1380 a t a pH of 8.53. The value of the equilibrium constant varies from log IC = 2.6 a t pH 4.65 to 5.1 a t pH 12.50. The stability t,o decomposition of murexide decreases, however, with increasing pH of the solution. A possible structure of the calcium complex was suggest,ed, and! also, complexes of certain other cations wpre investigated. Ost,ertag and Rinck (2-3) described a method employing ammonium purpurate for the determination of macro amount,s of calcium. The aqueous solutions were compared at' pH 6 in a Lange photoelectric colorimeter equipped with a 500 to 550 mp filter. In their method, pH 6 was chosen, so that the dye would be more stable. I n this spectrophotometric study; it was found that the sen+ tivity of the colorimetric reaction is greatly increased by a considerahly higher pH, thus making possible its development into a micromethod. At the same time, errors caused by the decomposition of the dye in alkaline solut,ion v-ere minimized to the point where the method became reproducible and reliable.
1STABILITY O F A.MMONIUM PURPURATE SOLUTIONS
O=C--SH
The instability of ammonium purpurate in aqueous solution has been a drawback to the development of it3 w e in analytical
