Table XI.
Compositions of Two Synthetic Mixtures Analyzed Six Months after Formation of the Analytical System
F
Known 37 4
Isobutane Isobutene 18 6 I-Butene 11 0 n-Butane 14 6 trans-2-Butene 11 0 7 4 cis-2-Butene a Average of two determinations. * One determination.
Found“ 37 4 18 6
Known 34 7
14 7
15 4 12 9 7 4
11 0
11 0
7 3
tures cited in Table I1 were analyzed conventionally, with typical precision (9). The marked improvement in precision resulting from differential rather than conventional photometry is evident from the comparison of standard deviations presented in Table XII. The precision of the analysis could he improved if necessary. Error could be decreased by use of higher analytical pressures, reference absorbers having higher absorbance values, and perhaps longer absorption paths. However, with determinations of error less than 0.05 or 0.107, absolute, other limiting factors come into play, such as the specification that the sample being
(3 ___
1-olume yo Error Expressed as Standard __ - ~- Deviation C’onventional Differential Isobutane 0 58 0.10 Isobutene 0 30 0.12 1-Butene n 52 0.12 n-Butane 0.73 0.29 frans-2-Buteiic. 0 40 0 19 cis-2-Butene 0 22 0 0;i
1 OUlldb
34 18 11 15 12
18 5 11 1
Table XII. Comparison of Results from Conventional and Differential Measurements
7 6 0
5
9 7 3
analyzed contains only the substances included in the calibration.
tion Spectroscopy,” D D . !5OT-13. JTiley, Kew York, 1950: Illichal, A . D., “Matrix and Tensor Calculus,” pp. 1-15, Wiley, New YorL
‘L,
LITERATURE CITED
( I ) Beroza, l\Iorton,
ASAL.
(?HEM.
25,
112-15 (1953). (2) Dwyer, Paul, “Linear Computations,” Chap. 3, Wiley, New York, 1951. (3) Ford, R. C., McDonald, R . S., hliller, F. A,, J. Opt. Soc .4??ter. 42, 149 (1952). (4) ~, Hiskev. C. F.. ASAL.C t r E l f . 21, 1440 (1919). (5) Jones, J. H., Clark, C;. It.. IIarrow, I,. S., J . Assoc. Oflc. Agi.. Chemists 34, 135-79 (1951). (6) llellon, M. G., “Analytical Absorp-
1Qd7 A ”
A,.
Milne, W.E., “Numerical Calculub,” pp. 17-35, Princeton Univ. Press, Princeton, 1949. Perrv, J. A , , ANAL.CHEM.23, 495 (1951). (10) Perry, J. 9.,Sutherland, R. G., Hadden. N.. ASAL. CHEM. 22, 1122 (1950). ’ (11) Smith, D. C., hliller, E. C., J . O p t . Soc. Amer. 34, 130 (1944). (12) Sutherland, G. B. B. XI., Willis, H. A,. Trans. Faradall SOC.41, 181 (1945). RECEIVED for revieTv November 16, lq.56. Accepted PIIRrch 30, 1957.
Inf ra red S pect ro pho to metric An a lysis of Fracti o na I Milligram Quantities of Solids J. J. KIRKLAND Grasselli Chemicals Departmenf, Experimenfal Station, E. 1. du Pont de Nemours & Co., Inc., Wilmington, Del.
,An evacuable, rectangular die has been designed for pressing small potassium bromide pellets used in the infrared analysis of solids. The die is particularly convenient, as it serves as the pellet holder in the infrared instrument, and no additional optical equipment, beam attenuation, or special instrumental adjustment is required. Qualitative and quantitative analysis of fractional milligram-size samples can b e carried out with this unit. Both freeze-drying and vibrator-grinding procedures are satisfactory for preparing the potassium bromide mixtures.
I
XFRARED
ABSORPTIOK
CURVES
Of
samples weighing less than 0.5 mg. are frequently needed for the identification and analysis of small samples isolated by column and paper
chromatography, and various extraction procedures. The potassium bromide-pellet technique (6-8) in many instances is the most convenient solution to the problem of analyzing such small quantities of material. Many commercially available dies produce disks which are approximately 0.5 inch in diameter and generally require 1 to 2 mg. of sample in order to produce satisfactory absorption curves. As the sample beam of the Perkin-Elmer Xodel 21 infrared spectrophotometer is a 1.5 x 11.5 mm. rectangle a t the optimum position in the microsanipling compartment, only about 157, of the sample incorporated in a 0.5-inch diameter disk is actually “seen” by the monochromator. Therefore, the amount of sample can be significantly decreased by pressing rectangular pellets in the shape of the sample beam of the spectrophotometer. Anderson and
Woodall (1) and Browning, Wiberley, and Nachod (2) have described nonevacuable rectangular dies which have proved the advantages of this design for studying very small quantities of solids. Described herein are the design and operation of an evacuable die n~hicli forms rectangular pellets just slightl! larger than the minimum external sample beam area of a Perkin-Elmer Model 21 infrared spectrophotometer. This equipment has several important advantages over other systems which have been proposed for obtaining spectral curves on fractional milligram quantities of sample: (1) No pellet holder is needed in conjunction with the die, as the body of the die containing the formed pellet is inserted directly into the sample beam without any instrumental adjustment; ( 2 ) no handling problems are encountered with the VOL. 29, NO. 8, AUGUST 1957
1127
very small, brittle pellets because of the direct mounting feature; (3) the die body does not interfere with the incident radiation of the spectrophotometer, hence, the beam is not attenuated and the instrument is operated under standard scanning conditions; (4) very small quantities of sample (0.25 to 0.4 pmole) can be scanned without auxiliary beam-condensing optical equipment; and (5) the unit can be employed for both qualitative and quantitatire studies.
DIE DESIGN A N D OPERATION
The evacuable microdie shown in Figure 1 was constructed of Carpenter Vega tool steel drawn to Rockwell C59/60 hardness. This unit forms “rectangular” pellets which are actually a 1.8-mm. wide cross section of a 13.8mm. diameter circle. A slightly larger pellet area is required over the minimum beam dimensions (1.5 X 11.5 mm.) for mounting purposes. In order for the pellets to be produced with the minimum of aberration, the faces of the plunger, A, and the anvil, B , were held flat and parallel using close tolerances, and both surfaces were mirror-finished. All corners were rounded and angles filleted in order to produce a die design of maximum structural strength. After the die is loaded, air is withdrawn from the potassium bromide-sample mixture, C, before pressing, by means of a semicircular grdove, D, which is inside the body of the die, E, and juqt above the top of the potassium bromide mixture. This air evacuation channel is connected to a removable nipple, F .
via the mounting slots, I , into the micro-cell holder assembly of a PerkinElmer Model 21 infrared spectrophotometer. If a pellet is to be retained for reference it may be punched out into a ring using a punch similar to that of A , but longer. I t is desirable to coat the inside of the die and the plunger and anvil faces with a very thin coating of powdered graphite in order to ensure easy removal of the pellets. Potassium bromide pellets prepared in the manner described are visually transparent and transmit about 85% of the incident energy of the spectro-
photometer. To date about 250 potassium bromide pellets have been prepared in the microdie without noticeable deterioration of the unit or its operation, QUALITATIVE APPLICATION
In general, about 25 to 100 y of sample, molecular weight 100 to 250, are required to obtain an absorption spectrum in which the medium or weak intensity absorption bands are clearly visible. Qualitative scans have shown that about 10 y of urea and 20
A\
E-
Figure 1 . Evacuable die for solid phase submilligram infrared spectrosCOPY
F
D’
A. Plunger 8. Anvil C. KBr mixture D. Air evacuation groove E. Die body and holder F. Air evacuation nipple G. 0 ring H. Mounting pin I. Mounting slots
The pellet is formed in the following manner : The plunger containing an 0 ring, G, a t the base is positioned in the die body and the unit turned upside down. Approximately 50 mg. of potassium bromide-sample mixture is loaded into the chamber in this inverted position and tamped down. (The plunger rests just a t the bottom of the air evacuation groove in this position; therefore, no mixture enters the air evacuation groove during this operation.) The anvil and the 0 ring, G, are then carefully assembled in the die body, making sure that no mixture becomes trapped between the plunger or the anvil and the die wall. The assembled die containing the mixture is then placed in a laboratory hydraulic press, and sufficient force is applied to seal the 0 rings to the metal surfaces. The die is then evacuated to less than 5 mm. of mercury via the air evacuation nipple for about 2 minutes. About 2-ton total pressure is applied for 1 minute t o form the pellet. The die is then removed from the press, and the plunger, anvil, and air evacuation nipple disassembled. The mounting pin, H , is screwed into place and the die body containing the formed pellets is mounted
1128
ANALYTICAL CHEMISTRY
WAVE LENGTH, MICRONS Absorption spectrum of 44 y of 3-phenyl- 1,l -dimethylurea
Figure 2.
0.4 0 W
E ln 4 m
Yz
A. F R E E Z E - D R I E D F R O M WATER 0.30 -B.F R E E Z E - D R I E D FROM DIOXANE 0.20
4
m
a
2m
0.10
d
C
IO
20
30
40
so
p0 I N PELLET
Figure 3. Band intensity vs. sample quantity curves, freeze-drying technique
i
o'60t
"
cent Dental llanufacturing Co., Chicago, Ill.), in somewhat the same manner as described in a previous publication (5). Mixtures made by this procedure are satisfactorily homogeneous with many compounds.
BENZAMIDE
20
40
60
80
100
120
140
160
180
AG IN P E L L E T
Figure 4. Band intensity vs. sample quantity curves, vibrator-grinding technique y of 3-phenyl-1,l-dimethylureacan lie positively identified using the micro rectangular die equipment described. Figure 2 shows the infrared spectrum of 44 y of 3-phenyl-1,l-dimethylurea.
Toribara and DiStefano (9) was used. The freeze-drying process can usually t o 11/* hours. It is be completed in sometimes desirable to ribrate the fluffy mixture resulting from the freezedry process for about 20 to 30 seconds in the manner described below in order t o facilitate pressing of the pellets.
QUANTITATIVE EXPERIMENTS
Fractional milligram quantities of nimy solids may also be analyzed quantitatively using the equipment m d techniques described in this paper. -1s virtually the entire area of the rectangular pellet is used in the infrnred measurement, quantitative relationships depend only on the weight of the pellet and the concentration of the sample vontained in it. The pellets need not be of uniform or reproducible thickness, but the sample-potassium bromide mixture-must be homogeneous and the particle size of the sample sufficiently small. Freeze-drying techniques are often preferred in the preparation of potassium bromide-sample mixtures to be used for quantitative purposes, as homogeneity is more nearly assured than with a vibrator-grinding procedure.
A satisfactory procedure which was used for mixture preparation of fractional milligram quantities of sample by freeze-drying, is to contain the sample in about 2 t o 3 ml. of a suitable freeze-drying solvent, add 50 mg. of optical quality potassium bromide, and quick-freeze with liquid nitrogen or a dry iceacetone mixture. Some of the more useful solvents for the freezedrying preparation of potassium bromide mixtures appear to be water, dioxane, glacial acetic acid, tert-butanol. tert-amyl alcohol, and benzene. Finely divided potassium bromide (250 mesh) is added if organic solvents are to be used in this operation. The organic solution-potassium bromide mixture is first thoroughly slurried and then frozen in the usual manner. During this study, a micro freeze-dry apparatus similar to that previously described by
Figure 3 shows band intensity 21s. sample quantity curves which were obtained TI ith 3-phenyl-1,l-dimethylurea. Two series of pellets rrere prepared by the freeze-drying technique preyiously described, using dioxane as the solvent in one set, and water in the other. These pellets were then scanned in the 6- to 14-micron region and intensity us. sample quantity curves constructed for several peaks a t varioas wave lengths. From the curves shon n in Figure 3, the precision of the technique appears to be satisfactory for most purposes, even when working n-ith very small amounts (0.04 to 0.4 lmole) of substances. As many compounds have appreciable vapor pressures a t the temperature and pressure of a freeze-drying process, caution must be exercised when quantitative measurements in the microgram range are desired. For instance, significant amounts of sulfanilic acid, DL-serine, and benzamide were volatilized when attempts mere made to freeze-dry 25- to 110-7 quantities of these compounds from water. Losses can often be made negligible by making sure that a minimum freeze-drying period is used in removing the particular solvent employed. Quantitative data can also be obtained with a vibrator-grinding technique. This procedure is particularly useful for preparing salt mixtures with samples which cannot be freeze-dried because of volatility or solubility limitations. Satisfactory particle-size reduction and mixing can often be accomplished by grinding in a dental vibrator such as the Wig-L-Bug (Cres-
Benzamide, which is too volatile to be quantitatively determined by the freezedrying method, was incorporated into micropellets by a vibrator-grinding procedure and yielded very acceptable quantitative data. This was accomplished by weighing the submilligram sample into a stainless steel Wig-L-Bug vibrator cylinder. Next, 50 mg. of pure powdered potasssium bromide and four '/*-inch diameter steel balls were added, and the cylinder contents were vibrated for about 5 minutes. After removal of the steel balls, the mixture was pressed in the die without further treatment. Figure 4 s h o w the band intensity sample quantity curves for the 6.33and 6.90-micron bands of benzamide using 20 to 160 y of this compound. Although the 6.33-micron band does not show a linear relationship, both curves indicate the good reproducibility of the sampling technique. Based on the limited amount of data obtained during this and other similar studies, it would appear that solid-phase quantitative analyses can be carried out on fractional milligram quantities of sample with only a slight decrease in reproducibility from that which is normally expected for conventional circular disk operation (3-5). 23s.
ACKNOWLEDGMENT
The author is indebted to L. A. Wharry who assisted in the design and fabrication of the microdie and to G. S. 'VC'alser who obtained many of the data.
LITERATURE CITED
(1) Anderson, D. H., Woodall, N. B., ANAL.CHEM.25, 1906 (1953). (2) Browning, R. S., Wiberley, S. E., Yachod, F. C., Ibid., 2 7 , 7 (1955). ( 3 ) Hausdorff, H., A p p l . Spectroscopy 8, 131 (1954). (4) Jensen, J. B., Acta. C h e w Scand. 8, 393 (1954). (5) Kirkland, J. J., Ar.4~.CHEM.27, 1537 (1955). (6) Schiedt, U., Z. Saturforsch. 8 b , 66 (1953). ( 7 ) Schiedt, U., Rheinmein, H., Ibid., 7 b , 270 (1952). (8) Stimson, bl. M., O'Donnell, 11, J., J . Am. Chem. Soc. 74, 1805 (1952). ( 9 ) Toribara, T. Y., DiStefano, V., BSAL.CHEM.26, 1519 (1954).
RECEIVED for review October 22, 1956. Accepted March 28, 1957. VOL. 29, NO.
a,
AUGUST 1957
1129
