density gradient at the point of pseudoequilibrium of the protein solutionthat is, a t the point where the injected protein solution floats. This is due to the instability of the initial narrow, sharply defined zone formed by introduction of the protein solution, which is caused by the following chain of events ( 1 1 ) . In the protein zone the solution has a lon-er sucrose concentration than the solution above and below. Sucrose diffuses into this zone faster than protein diffuses out. The density of the solution a t the lower (and upper) surface becomes greater than that inmediately below, and droplets begin to settle out. Brakke ( 2 ) explains in detail the means for controlling this droplet sedimentation. The protein zone becomes stable as soon as a smooth sucrose gradient is re-established, and a protein gradient is formed which is of opposite sign and of no greater magnitude than that of the sucrose gradient. The depth of this stabilized protein zone can be minimized by the use of high density columns, in which the density increment due to the protein is relatively smaller, but the high viscosity of such a column makes this undesirable, A second method is to use a column m-hich has a steep density gradient; the steeper the density gradient of the sucrose, the steeper the gradient of the protein, and thus the narrower the protein zone. This narrower protein zone, L,, niight be expected t o give greater resolution of the electrophoresis procedure, for it is a ratio of the height of t h e original protein zone t o the total height of the column which defines the resolution that can be expectedthat is, the broader the initial protein zone, the broader will be the bell-shaped curve of any component after electrophoresis. The protein gradient formed within
a poorly stabilized protein zone, of opposite sign to the sucrose gradient, may give rise to asymmetry of the peaks of the individual components. This will result in asymmetry of the type in which the leading edge is blunted and the trailing edge is extended. I n the experiment reported in Figure 4, to stabilize the layered gradients, a long stirring rod was inserted from the top to the bottom of the column, gently rotated, and removed. Fifteen milligrams each of human serum albumin and globulin were dissolved in 0.5 ml. of buffer and dialyzed overnight against 5% sucrose solution in 0.05;ll Tris buffer a t pH 8.6. This sample was inserted a t a point about 10% of the distance from the top of the gradient to the bottom, and descending electrophoresis vas used. Although the width of the individual components is not noticeably greater in the gradients formed by layering, there is a perceptibly greater asymmetry than where a nonlinear gradient has been used, (Figure 4,first curve) as produced automatically by the apparatus shown in Figure 1,A. Thus use of the steeper gradient at the point where the sample is layered in probably results in a narrower stabilized protein zone. Figure 5 presents the results of the electrophoresis of human serum, human plasma, and the same sample of human serum analyzed by the usual Tiselius analytic electrophoresis. Protein determinations were carried out b y the Buffalo Black dye method of precipitating proteins reported by Plum, Hermansen, and Petersen (8). This extremely sensitive method avoids the difficulties involved in exchanging very viscous samples in cuvettes used for ultraviolet spectrophotometry. It is well adapted to the analysis of protein, except that plasma albumin combines in a different stoichiometric proportion than the plasma globulins. Thus the
peak of albumin in these diagrams is somewhat exaggerated. The p-globulins in plasma do not exceed the a-2-globulins, as might be expected, and this may be due t o the presence of some albumin in the a-2 peak. The correspondence of these peaks with known fractions of plasma proteins has been established by paper electrophoresis. It is advantageous t o use as small a volume as possible for the sample; however, good separation can be achieved with 1-nil. samples containing from 50 to 300 mg. of protein. ACKNOWLEDGMENT
The authors are indebted to John Gregory for helpful advice. LITERATURE CITED
Bock, R. M.,Ling, X.-S., ANAL. CHEW 26, 1843 (1954). Brakke, 11.K., Arch. Biochem. and Biophys. 55, 175 (1955). Brakke, RI. K., Phytopathology 43, 467 (1953). Brakke, RI. K., Vatter, A. E., Black, L. RI. “Abnormal and Pathological Plant Growth,” pp. 137-56, Brookhaven Sational Laboratory, Upton, N. Y., 1954. Cherkin, il., Martinez, F. E., Dunn, RI. S..J . Am. Chem. SOC.75, 1244 (i953j. Mitchell, H. K., Gordon, M.,Haskins, F.’B.,J . Biol. Chenz. 180, 1071 (1949). Philpot, J. St. L., Trans. Faraday SOC.36, 38 (1940). Plum, C. L., Hermansen, L., Petersen, I., Scand. J . Clin. Lab. Invest. 7, Suppl. 18 (1954). Sorof, S., Ott, h’. G., Young, E. ?VI., Arch. Biochem. and Bzophys. 57,
140 (1955). Svensson, H., 1 . V . d . 25, 252 (1954). Svensson, H., Hagdahl, L., Lerner, K. D., Science Tools 4, 1 (1957). RECEIVED for review Sovember 1, 1985. Accepted September 6, 1955. Kork supported in part by a research grant, G-4253, from the U. S. Public Health Service.
Process Control Methods in the Chlorination of Benzene RALPH A. TROUPE and JEROLD J. GOLNER Northeastern University, Boston, Mass.
F A rapid, simple, and accurate method of analysis of reaction mixtures of chlorinated benzenes is based on density measurements. It can be used for either process control or product analysis, if a density-product distribution chart is prepared for the specific chlorination system under the conditions used in its operation. Vapor phase chromatography was used as a primary standard and was found
to be an accurate qualitative and quantitative method of analyzing mixtures of chlorobenzenes.
I
N THE AI4KUFbCTURE O f most O f the
chlorobenzenes, such as the commercially important p-dichlorobenzene, a complex reaction mixture results @), a-hich contains not only the desired product and hydrogen chloride but
also other chlorobenzenes. For example, in the manufacture of p-dichlorobenzene, monochlorobenzene, odichlorobenzene, and some trichlorobenzenes may be produced in addition to the principal product. As the economics of the process depend upon being able t o produce the maximum yield of the desired product or combination of desired products and upon knowing the concentration of the by-products, VOL. 30, N O . 1, JANUARY 1958
129
a rapid, simple, accurate method of process control is needed. The flowsheet (Figure 1) illustrates a typical commercial batch chlorination (in this case for p-dichlorobenzene). Process control is carried out by analysis of samples taken from the reactor at periodic intervals. Methods currently employed consist of stopping the reaction a t a certain density. established through experience (Z), and analyzing the product by a series of distillations and freezing point determinations (?). As the ratio of isomers or the product distribution may be affected to varying extents b y reaction temperature, rate of chlorination, type of catalyst, and catalyst concentration, and by whether the chlorination is batch or continuous (Y), the fragmentary density values available in the literature may be misleading. These references do not disclose the technique employed to obtain the density values nor the operating conditions for the chlorination from which the samples were taken, and no attempt has been made to use accurate density determinations in analyzing the composition of the reaction mixture. The difficulties with analysis by distillation and freezing point are the length of time needed for analysis, evcessive handling, and the need for skilled analysts. iilternative methods of analysis involve infrared anaIyzers and mass spectrometers. Cost of these installations is somewhat prohibitive for small manufacturers, and professional skills are needed to operate these devices and to interpret the results. I n developing the method presented here, samples were taken at intervals from a commercial batch chlorinating unit similar to that shown in Figure 1. The process was operated using ferric chloride catalyst a t 50' C. t o produce p-dichlorobenzene. Twelve 250-ml. samples mere withdraivn over a period of 63 hours. The samples were neutralized with a 10% sodium hydroxide solution to remove any dissolved hydrogen chloride gas which might hinder subsequent analysis. Samples which were partially solidified, because of a high concentration of p-dichlorobenzene, were first heated until homogeneous to ensure coniplete neutralization.
Figure 1.
Flowsheet of typical process for chlorination of benzene
I 9-
MONOCHLOROBENZENE I
-
I
1
P - DICHLOROBENZENE
/I
0 5 W
3
L
2
1
It-
4
-
0
I
I
60
50
I
I
40
I
1
I
20
30
10
0
T I M E , MINUTES
Figure 2. Sample recorder chart from Vapor Fractometer showing distribution of products ATOMS C I / M O L E
0.0 100 90
0.2
0.4
0.6
0.8
BENZENE 1.2
1.0
1.4
1.61.8
2.02.1
DICHLORS
d
W V
C
e
W
w
0 i
I 2-
0 t rn 0 e I V 0
PRIMARY ANALYSIS
The primary analysis of the neutralized samples mas performed on a Perkin-Elmer Model 154 Vapor Fractometer (gas chromatography equipment) equipped with a column containing a n 80-20 mixture of diatomaceous silica and dinonyl phthalate. The carrier gas Tvas helium. The detector unit, which measured the thermal conductivity of the exit gases, was equipped with a strip-chart recorder. I n performing the analysis, a neutralized sample of the reaction mixture
130
ANALYTICAL CHEMISTRY
D E N S I T Y A T 50'C.,
Figure 3.
G. IML.
Variation of product composition with density in chlorination of benzene
mas injected into the sampling section of the Vapor Fractometer with a graduated microsyringe; this allowed the components present to be separated in and eluted from the column. Samples containing polychlorinated ben-
zenes required approximately 2.5 hours for the last component to appear a t the column exit. T o prevent mixtures which contained undissolved p-dichlorobenzene from clogging the small aperture of the in-
REACTION
Figure 4.
T I M E , HOURS
Variation of composition
of reaction mixture with time
jection needle, a sufficient amount of analytical grade toluene was added t o solubilize this ingredient. The resultr ing peak representing toluene appearing on the recorder chart was therefore ignored in computing the composition of the injected sample. With adjustment of two variables-sample quantity and detector sensitivity-it was possible t o avoid running the peak of the major component in the sample beyond the range of the recorder chart. The sample quantities employed ranged between 10 and 40 pl. and the detector sensitivities between and of its maximum output. For all runs the helium pressure and flow were kept constant a t 7 pounds per square inch gage and 250 ml. per minute, respectively. The operating temperature was maintained a t 150" C.
fitted into the neck of the flask and the flask immersed in a constant temperature bath until the volumetric reading remained at a constant value for 5 minutes. Volumetric readings were made a t 40', 45', and 50" C. A t lower temperatures some samples exhibited a solid phase; therefore, densities were not determined a t temperatures below 40' C. Corrections, based on the water calibrations, were made t o the apparent densities determined as outlined above in order to arrive a t the true densities. A specific gravity balance or a cassia flask of smaller volume could be used for the determination of these densities, but in the latter case some accuracy would be sacrificed.
Figure 2 shows the recorder chart for one of the samples. The components of the mixtures rrere identified qualitatively on inspection of the series of peaks scribed for each sample, as in this type of mixture the components are eluted in order of decreasing vapor pressure a t the temperature of operation (1). As the area under each peak is directly proportional to the molar concentration of the corresponding component ( S ) , the area relationship was usrd to calculate the relative concentration of the constituent? present.
T o check the accuracy of the procedure, a synthetic mixture of chlorinated benzenes was prepared corresponding in composition to each sample analyzed by the Vapor Fractometer. The reagents used to prepare these mixtures JTere Eastman organic chemicals:
DETERMINATION OF DENSITIES
The density of each neutralized sample mas determined by a modification (6) of the method of Leyea and Othmer (4).
A thin-walled cassia flask of 100-ml. capacity, with a graduated neck of 10-nil. capacity, was first calibrated with water. T o determine the densities of the samples, the flask was filled to the zero mark at 40' C. with a weighed quantity of the sample. The stopper n a s then
DENSlTY
OF SYNTHETIC MIXTURES
Eastman No. 777 benzene (thiophenefree), melting point 5-5.5' C. Eastman S o . 499 o-dichlorobenzene 99 +yo Eastman S o . 89 p-dichlorobenzene melting point 53-4' C . Eastman No. 70 chlorobenzene, boiling point 13032' C. Eastman No. P 1641 1,2,4-trichlorobenzene (practical) The density of each synthetic mixture was determined by the procedure described. Average deviation from the density of the corresponding reactor sample mas O.l5%, the maximum deviation being -0.4%. DISCUSSION OF RESULTS
The graphical presentation of the data (Figures 3 and 4) is based on densities measured at 50" C. Similar curves could be drawn for the data a t 40' and 45" C. or a density correction curve could be prepared by plotting
the average deviation of densitj from the value at 50" C. against the teniperature in degrees centigrade. Figure 3 shows the manner in which product distribution varies with density for the particular operation studied. (The density-product distribution curve is applicable only t o the specific chlorination unit used in its derivation and only when operated under the conditions employed during the collection of the data used in obtaining the curve.) It is apparent that the density of the reaction mixture will serve as a satisfactory method of analysis at a n y stage of the operation. For comparison of the product distribution with that reported by Machlullin (6) and by Wiegandt and Lantos ( 7 ) , a scale of chlorine atoms substituted per mole of benzene has been drawn at the top of the diagram. Figure 3 substantiates the findings of TS7iegandt and Lantos (Y), that o-dichlorobenzene converts more rapidly to trichlorobenzene than does the para isomer, and the percentage of para in the dichlorobenzene fraction increases further after the trichlorcbenzene begins to form. MacMullin ( 5 ) reports a somewhat higher maximum concentration of monochlorobenzene and a somewhat lower maximum concentration of dichlorobenzenes than Figure 3 indicates, although in both cases the peaks occur a t the same ratio of chlorine to benzene. If the change in density with time under the conditions studied is plotted, it will be found t h a t the density increases continuously a t a slightly decreasing rate as the reaction proceeds. This indicates that the correct point to terminate the reaction can be obtained easily on a "predicted time" basis. Rate of change of composition of the reaction mixture is presented in Figure 4. Although this is not needed as part of the development of the analytical procedure, it was felt that the kinetic data might be of interest. LITERATURE CITED
(1) Barefoot, R. R., Currah, 3 . E., Chem. in Canada 7,45-8, 50,52 (1955). ( 2 ) Groggins, P. H., "Unit Processes in Organic Synthesis," 4th ed., Mc-
Graw-Hill, Yew York, 1945. (3) Hausdorff, H. H., "Vapor Fractometry," Perkin-Elmer Corp., ?lorwalk, Conn., 1965. ( 4 ) Leyes, C. E., Othmer, D. F., Ind. Eng. Chem. 37, 968 (1945). ( 5 ) MacMullin, R. B., Chem. Eng. Progr. 44, 183-8 (1948). (6) Troupe, R. A., Kobe, K. A , , Ind. Eng. Chem. 42, 801 (1950). (7) Wiegandt, H. F., Lantos, P. R., Ibid., 43, 2167 (19513. RECEIVEDfor review June 17, 1957. Accepted September 6, 1957. VOL. 30, NO. 1, JANUARY 1958
131
