chromatogram of the sample showing a good breakdown of C4--Cecomponents along with small amounts of dissolved CI-CB hydrocarbons. For samples containing appreciable amounts of CSand higher aromatics the modified flow pattern (Figure 3) has been used with excellent results. The use of a silicone-modified alumina column for the analysis of light hydrocarbons has been previously reported (6). However, the present method combines this column with a pre-column in such a way as to provide a rapid, accurate method for a more complete characterization of pyrolysis off-gas. Previous reports also have not used the column to its full resolving potential. During the development of the method it was found that the programming rate or elution temperature had a pronounced effect on the resolution of two important, but minor, components-allene and acetylene. Programming rates greater that 6 “C per minute caused co-elution of these com-
ponents and, if very fast, caused both to elute with n-butane. It was further found that the elution of these components could occur even prior to propylene depending on the condition of the column. Acetylene in particular eluted prior to propylene after the column had been left at room temperature for several days. A graduai shift to longer elution times was observed as the column was temperature cycled several times until it eluted as shown on Figure 2. A standard practice of heating the column to the maximum temperature for one hour before use eliminated this variability. Another column combination which can be used in place of the seven-ring polyphenyl ether column is 6 ft X in. 6 ft X in. 15% SE-52, both on 20% Carbowax 20M Anakrom ABS. The Carbowax provides resolution of the aromatics and the SE-52 column provides the delay necessary to elute benzene after propane.
(6) C . G. Scott,J. Znst. Petrol., 45, 118 (1959).
RECEIVED for review June 4,1971.
+
Accepted August 26, 971.
Rapid Photochemical Determination of Uranium(V1) at Trace Levels William M. Riggs Central Research Department, Experimental Station, E. I . du Pont de Nemours and Company, Wilmington, Del. 19898 PHOTOCHEMICAL REACTIONS have great potential utility in analytical chemistry (1). The increased availability of convenient light sources has made possible the development of simple photochemical procedures for analysis using equipment which is readily available to most laboratories. The method described illustrates the high speed, sensitivity, and accuracy which can be achieved with simple techniques involving photochemical reactions. The new method is based upon photoreduction of uranium(V1) to uranium(1V) with a primary or secondary aliphatic alcohol. The U(1V) is then treated with excess iron (111), and the iron(I1) produced is colorimetrically determined with orthophenanthroline. This method has all the advantages of the best earlier photochemical methods (2, 3) and is significantly more convenient and sensitive. It has the further advantages of not being critically dependent on sulfuric acid concentration (2) or on close control of irradiation time, geometry, or intensity. EXPERIMENTAL
Apparatus. The light source was an 85-watt G.E. AH3 medium-pressure mercury vapor lamp. Its full spectrum was utilized for photolysis. Absorbance measurements were made with a Bausch & Lomb Spectronic 20 colorimeter. Two kinds of sample vessel were used. For larger amounts of solution (10-20 ml) the vessel was a Kimax 40-ml Erlenmeyer flask with a piece of capillary tubing attached near the bottom for de-aeration with nitrogen. Solutions in these vessels were stirred with polyfluorocarbon-covered magnetic (1) J. M. Fitzgerald, Ed., “Analytical Photochemistry and Photochemical Analysis,” Marcel Dekker, New York, N. Y., 1971. (2) A. A. Nemodruk and E. V . Bezrogova, Zh. Anal. Kliim., 21, 881 (1966), Eng. trans., p 744. (3) Zbid., 23, 388 (1968), Cheni. Abstr., 69, 1 5 9 1 7 ~(1968).
390
stirring bars. When small amounts of solution (-2.0 ml) were analyzed, the photolysis was carried out directly in a Kimax Spectronic 20 test tube-cell. Nitrogen bubbled into the solution through a tube inserted at the mouth of the test tube served to de-aerate and agitate the solution. Reagents. Standard uranyl solutions were prepared from Mallinckrodt analytical reagent grade ut any1 acetate dihydrate shown to be 99.3 pure by thermogravimetric was analysis before use. 0-Phenanthroline solution (0.1 prepared by dissolving the solid in hot distilled water. All other reagents were analytical reagent grade. Solutions were de-aerated when necessary with tank nitrogen which was saturated with water and scrubbed for traces of oxygen by bubbling through a basic sodium sulfite solution. Procedures. A standard curve was obtained by dissolving 0.0684 gram of ferrous ethylenediammonium sulfate tetrahydrate in water and diluting to 1 liter with 1 N HzS04. This solution contained 0.010 mg/ml of iron(I1). Further dilutions were made by pipetting from 1.00 to 50.00 ml into 100-ml volumetric flasks, Ten milliliters of 0.1 ophenanthroline was added, the pH was adjusted to about 5 with sodium acetate, and the solutions were diluted to volume. The absorbance of each solution was measured at 508 mp and plotted L’S. micrograms of iron present. All of the values fall directly on a straight line through the origin. By using the standard uranium(V1) solution in 1N &Sod, aliquots were transferred to the reaction vessel such that the final concentrations of UOzz+ would be in the range 0-20 pg/ml. Then 2.00 ml of 5M isopropyl alcohol in 1N HzS04 was added, followed by enough 1N H2S04to bring the volume to 18.00 ml. Oxygen-free nitrogen was bubbled through the solution for a few minutes before irradiation was begun and throughout the irradiation to exclude atmospheric oxygen. After the initial de-aeration period the solution was irradiated for 30 minutes, with the lamp mounted about 2 centimeters from the side of the reaction flask. Cold tap water (about 20 “C)was circulated around the base of the
ANALYTICAL CHEMISTRY, VOL. 44, NO. 2, FEBRUARY 1972
z
z)
Table I. Uranium Determination UraniumWI) concentration. d m l Added Found
a
20.00 16.00 12.00 8.00 4.00 2.00 Average of four determinations.
19.80 16.14‘= 12.07 8.02
4.46 2.11
flask during irradiation to prevent excessive heating. After irradiation 2.00 ml of 0.002M Fea+solution in 1N HzSOl was added to the flask, Then 20.00 ml of a 1 : 2 : 2 mixture of 0.1 o-phenanthroline solution, saturated sodium formate solution, and water was added. A sample of the resulting ferrous o-phenanthroline solution was taken for measurement of the absorbance at 508 mp us. a water blank. For measurements on smaller amounts of solution, the same technique was used except that a Bausch & Lomb Spectronic 20 test tube-cell was utilized as the reaction vessel and all the reagent volumes were reduced tenfold. This is advantageous since the final absorbance measurement is made directly in the reaction vessel, thereby eliminating one manipulation.
though IN HzS04 is a practical minimum (4, 5). The uranium(1V) produced in the photolysis is quite sensitive to reoxidation under the conditions of the photolysis; oxygen must be carefully excluded during the photolysis procedure by bubbling oxygen-free nitrogen through the solution. If the temperature of the solution rises significantly during photolysis, it becomes extremely difficult to prevent some reoxidation of U(1V) to U(V1). The temperature has therefore been kept at or slightly below room temperature by circulation of tap water around the base of the reaction vessel. The influence on the photoreduction of U0z2+ of many species which might interfere with an analytical determination is known (2, 4, 5). Sodium, potassium, ammonium, calcium, strontium, and aluminum ions can be tolerated in large excess as can fluoride, phosphate, and perchlorate ions. A great many other species can be tolerated in amounts at least equal to the amount of uranyl ion being determined. These include titanium(IV), molybdate, tungstate, arsenate, copper, chloride, nitrate, manganese(II1, and nickel(I1) ions. Iron will, of course, interfere strongly when present at a concentration of more than 3 or 4 parts per million. Below this level, however, account may be taken of its presence by use of a blank, allowing a uranium determination to be made. CONCLUSIONS
RESULTS AND DISCUSSION
Results of the analysis of uranium at several concentrations are given in Table I. As an example of the attainable precision, the standard deviation of a measurement at the 16.00 pg/ ml level is 0.81 based on four determinations. A calibration curve for UOz2+concentration can be prepared directly from the ferrous o-phenanthrolene curve since 1 .OO pg/ml of U0z2+ produces 0.414pg/ml Fez+in this procedure. A concentration of 0.10 pg/ml of UOz2+will produce an absorbance value of 0.009, and is therefore readily detectable. This may be extended to 0.01 pg/ml by utilizing IO-cm absorbance cells with an appropriate spectrophotometer. Thus, approximately 0.02 pg can be readily detected using a 2.0-ml sample, compared to claims of about 1 pg in earlier work (2, 3). The optimum alcohol concentration is between 0.4 and 4.0M. The sulfuric acid concentration is not critical, al-
The photochemical technique described makes it possible to analyze rapidly and with comparative ease uranium(V1) solutions at concentrations below 0.1 part per million. Other photochemical methods have been reported for analysis of uranium(V1) at trace levels (2, 3) but none can match the combination of high sensitivity, convenience, and accuracy of the method described here. RECEIVED for review June 22, 1971. Accepted September 7, 1971. (4) G . G . Rao, V. P. Rao, and N. C. Venkatamma, 2. Anal. Chem., 150, 178 (1956). ( 5 ) A. A. Nemodruk and E. V. Bezrogova, Zh. Anal. Khim., 21, 1210 (1966), Eng. trans., p 1074.
Use of a Substrate Gradient in the Automated Determination of Ornithine Transcarbamylase Albert Himoe, G . L. Catledge, and W. E. Kurtinl Department of Biochemistry, Bay for College of Medicine, Houston, Texas 77025
THE USE OF AUTOMATED determinations, including several enzyme assays, has become routine practice in clinical laboratories (1). In addition to the greater speed of analysis, automated enzyme assays are normally more accurate and reliable than manual assays because of the greater degree of Present address, Department of Chemistry, Trinity University, San Antonio, Texas 78212. (1) M. K. Schwartz and 0. Bodansky, “Methods of Biochemical Analysis,” Vol. 11, D. Glick, Ed., Interscience, New York, N.Y., 1963, p 211.
control over experimental variables. The same advantages of automated assays apply to enzyme kinetic studies (2). In clinical use, the same apalysis is normally made on a large number of discrete samples. For an enzyme kinetic study, the velocity of the reaction as a function of the concentration of substrates, cofactors, inhibitors, and activators must be determined. Therefore, instead of determining the velocity of the reaction at each substrate concentration, an experimental design in which the concentration of one of the substrates is continuously varied becomes feasible. (2) Zbid.,Vol. 16, 1968, p 489.
ANALYTICAL CHEMISTRY, VOL. 44, NO. 2, FEBRUARY 1972
391
