VERSATILITY
The instrument described offers a number of advantages. Construction is simple and inexpensive. As opposed to some instruments based on electrolytic conduction, it causes no electric current flow in the system monitored and, of course, it works equally well for nonconducting solvents. It is designed for
continuous duty for long periods of time, even in a cold room. The instrument is versatile. It can be set either to turn on or off automatically, and operation with any desired degree of sensitivity can be achieved with a variable delay system. I n addition to its application to the control of solvent flow in column chro-
matographic systems for which i t was intended, the instrument can be used in a number of other designs in which a light path can be interrupted by the system to be controlled. Thus, meniscus detectors, solvent transfer controls, and overflow devices based on the same principle have been developed.
Practical Double-Prism Cell for Differential Refractometry Andreas Rosenberg, Rufus Lumry, and Kirk Aune, Department of Chemistry, University of Minnesota, Minneapolis, Minn. and Lee Gropper, Spinco Division, Beckman Instruments, Inc., Palo Alto, Calif.
with studies of the Isolutions refractive index of dilute protein containing organic additives, N CONNECTION
we designed a suitable cell for the measurement of differential refractive indices using the interference optics of the Model H Beckman/Spinco electrophoresis apparatus. To be useful in a wide range of protein studies, such a cell must be easy to clean and fill while positioned in the bath, and it must be free of grease, since the smallest amounts of added organic solvents are enough to transport grease from joints to cell walls. The new cell is extremely simple to use
Figure 2. Front view of refractorneter cell Figure 1. Side view of cell assembly
1 168
ANALYTICAL CHEMISTRY
and yields precision considerably greater than that of commercially avaiIabIe differential refractometers. The cell is a modification of the prismatic double cell of Svensson and Odengrim (1). Figure 1 shows a side view of the cell assembly, and Figure 2 shows a front view of the refractometer cell alone. The three borosilicate glass parts are held together by clamps positioned at the ball joints. Silicone rubber gaskets are used for a water tight seal between the ball and the socket. The cell fits into a modified rack of the electrophoresis apparatus and can be used without change or readjustment of the standard interference optics. Solutions or solvent in the upper compartments come in contact only with glass. We have found that the rubber gasketing material, ahich is in contact with the contents of the lower compartment, is impervious to dilute solution of ordinary watersoluble organic solvents over long periods of time. As shown in Figure 1, the height of the fringe-forming triangular compartments is reduced by filling the sharp corners at the bottom of these compartments. This modification slightly reduces the number of fringes which can be observed and thus the accuracy, but is essential if the cell is to be easily cleaned or cleaned in situ. One of the upper compartments and the lower compartment are generally filled with solvent, and the unknown sample is placed in the remaining upper compartment. The accuracy was determined from readings of fringe pictures with a Bausch and Lomb optical comparator. The picture is read across the diagonal fringe pattern at any vertical setting. Individual fringes vary in width by as much as 1 part in 30, but we read the fringes in groups of 5 to 25 fringes, depending on the total number of fringes in the pattern. In a typical experiment the plate distances for 14 fringes is read for fringes 1 to 15, 2 to 16, 3 to 17, and so on, across the plate.
The average fringe density was determined from the average of these readings, making corrections for the magnification errors of the apparatus. Differences in densities obtained along several horizontal measuring paths from top to bottom of the picture were within a standard deviation. Hence, only one set of readings across the plate is necessary. For fringe densities from 14 to a t least 28 fringes per cm. the standard error was 0.015 fringes per cm. If a standard error of 0.02 fringe per cm. is accepted as the reading sensitivity, the real minimum change in measurable refractive index is given by An =
A8 v
h tan
in which An is this change, X is the wave length employed, s is the reading sensitivity, h is the height of the cell (or the liquid column, if not completely filled) and Y is the angle of the triangular cell. For this cell: h = 3.30 cm., tan Y = 1.003, and X = 5.461 X cm. The quantity A n i s then 3.3 X lo-' refractive index unit and provides a reliable measure of the accuracy of refractive index measurement. The precision in replicate experiments has been found to be about the same as the accuracy. The sensitivity depends on the fringe density; the accuracy given here is valid for the range from 10 to 50 fringes per cm. The cell is designed for differences in refractive index up to about 1.5 X 10-3 unit or slightly over 1% protein in water. For slightly higher concentrations the equations of Svensson and Odengrim ( I ) may be utilized, but for appreciably more concentrated solutions, a cell with a smaller value of tan v must be used. LITERATURE CITED
(1) Svenfiaon, H., Odengrim, K., Acta. Chem. Scand. 6 , 720 (1952).
THISwork was supported by the U. S. Public Health Service, NIH Grant No. AM 05853.
