METHOD FOR DECOLORIZING CHROMATOGRAPHY SOLUTIONS JACK RADELL and P. D. HUNT Wright-Patterson Air Force Base, Ohio D E C O L ~ R I Z I N Gof a solution of an organic compound prior to chromatography requires heating the prepared solution with carbon black or carbon black and filter aid. The heated solution is filtered and then added to the chromatographic column. The collected fractions are concentrated to recover the organic compound. Some of the shortcomings of this method result from the number of transfers required and the difficulty of completely filtering off carbon. Consequently significant quantitative information is more difficult to obtain. In this laboratory a dry packing procedure has been employed which considerably eliminated the ahove difficulties. The adsorbent ( A ) is added with tapping and periodic tamping until the desired length of column is obtained. The adsorbent is covered by a layer of absorbent cotton ( B ) followed by a layer of intimately mixed activated carbon: filter aid1 (3: 1 by vol.) (E) another layer of cotton (C) and finally a
layer of purified sand (D). The sand is used to prevent the cotton from floating up when liquid is added. The decolorizing layer (E) should not he more than onetenth the length of adsorbent. The effluent from the column is fed directly into a continuous concentration apparatus through a cork stopper (F). This consists of a 3-way connecting tube (G) leading to the tared concentration flask (H). The third connection carries the distilled developer to the combined condenser and analytical still head (I)from which it is collected in a graduated cylinder (J). The conrentration flask was heated by a constant level hot ~vaterbath (K) on a hot plate (L). When a fraction is ready for removal, the bottom end of the column is corked for 10 minutes to assure complete removal of the developer. The developer may be fed into the column by a constant level device (M) prepared from a 5-mm. i.d. tube sealed on to the neck of a single necked round bottom flask filled with developer and supported so that the end of the tube is a t the desired liquid level in the column ahove the sand. This procedure may be modified so that developers boiling above 100°C. can be removed by an appropriate constant temperature hath. If it were desirable the adsorhent could be added to the column by one of the wet-pack methods. The dry-pack method using Florisi12 as an adsorbent has been found most convenient in this laboratory. The proposed method has a number of advantages over some more common practices. After the solution to be decolorized and chromatographed is added to the column, no transfers are required. Thus reliable quantitative data are easily obtained. In fact, traces of material can be isolated easily from a large volnme of developer with no losses. The apparatus once in operation requires little care. Overheating of the residue obtained by evaporation is minimized by the hot water hath. When a fraction is removed, the volume of developer distilled can he read directly in the graduated cylinder and the weight of residue can he obtained promptly by weighing the tared concentration flask.
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A suitable filter aid is Celite Analytical Filter-Aid, manufactured by the Johns-Manville Co., New York, N. Y. 1 A product of the Floridin Company of Tallahassee, Florida.
JOURNAL O F CHEMICAL EDUCATION
croanalysis under the name of "purple of Cassius" test. This reaction is hased upon the formation of a suspension of colloidal gold and tin hydroxide by treatment of gold(II1) chloride with tin(I1) chloride. Shortly after this, in 1666, Tachenins discovered that a drop of a mixture of gold solution with tincture of nutgall, when placed upon paper, produced a metallic spot. Microscopic analysis appears to be the oldest of the various branches of microanalysis. So far as is known, the first man to use the microscope in chemistry was Sigismund Margraf (1709-82) during a study on cane sugar in beet juice. There is some question as to who first applied microscopic methods in analytical chemistry, hut according to Chamot and Mason, it was Raspail in 1833. The latter first presented a system of microscopic tests for chemists, and also proposed the identification of compounds by observing crystals under the microscope. On the other hand, Deniges and Feigl give credit for the development of microchemistry to Tichman, whose work in 1853-57 described the microscopic identification of small traces of blood through the formation of characteristic hemin crystals. Still other investigators, including Helwig, Harting, Wormley, and Boricky, are credited with originating microanalysis. In a book published in 1865, Helwig discussed the use of the microscope in toxicology. Harting prepared a text in 1865 on the theory and use of the microscope, and in the following year prepared a second volume in which the forms of some precipitates were described. In 1867, Wormley published a book on the microchemistry of poisons. In this book he described methods whereby a drop of the solution under examination was treated with a suitable chemical reagent, and the reaction product was viewed under the microscope. Ten years later Boricky prepared a dissertation on the chemical microscopic analysis of minerals and offered the first rational system of microchemical reactions. The method which he described was based upon the decomposition of minerals by means of fluosilicic acid, and the identification of the fluosilicates of various metals by means of their crystalline forms. In 1869, Sorhy carried out some studies on the use of the microscope in blowpipe analysis, and later Streng published a paper in which he recommended methods similar to those described by Sorby. Still later in 1884 Steng proposed several new reactions and descrihed new techniques to aid in microscopic studies. Reinsch also published a paper in 1881 which dealt with the identification of compounds by means of their crystal forms. In 1885 Haushofer published a rather extensive paper in which microscopic tests were described as an aid to qualitative analysis. He listed reactions for most of the elements and thereby laid the foundation for general microscopic analysis. Haushofer and also Klement and Renard suggest that microchemical methods could he applied to all chemical elements. Of the many chemists who have contributed to the development of qualitative microanalysis, Behrens perhaps most properly might be called the founder of microscopic chemical analysis. During the last 20 years of the nineteenth century he published a series of papers in which he proposed a large number of microscopic tests and also descrihed methods for the separa-
tion of compounds from each other so as to give greater accuracy t o the confirmatory tests. Between the years 1907 and 1909, Schoorl published a series of papers in which he described a scheme of microchemical qualitative analysis based upon the classical hydrogen sulfide system. Emich, who died in 1940, made many very valuable contributions to qualitative microanalysis and was responsible for the development of many of the techniques used in microanalysis a t the present time. DEVELOPMENT OF SPOT TESTS
One of the more important branches of microanalysis is that known as spot analysis. Spot tests, in the modern sense, date from about the middle of the nineteenth century, hut methods which would now be described as spot tests were applied even during the time of Christ. One of the most common of all spot tests, namely the testing for acidity or basicity with filter paper impregnated with plant extracts, was first used by Boyle. For this purpose he used a litmus paper and paper impregnated with the extract of violets and cornflowers. I t has not been definitely established who made the first use of spot reactions for analytical purposes. One of the earliest published records appeared in a paper by Schiff in 1859. He used filter paper impregnated with silver carbonate to detect uric acid in urine. A drop of the specimen produced a brown spot of free silver. The fundamental work for that division of spot test analysis in which filter paper is used as the reaction medium is found in a study by Schonbein in 1861. He showed t,hat when aqueous solutions rise in strips of filter paper, the water precedes the dissolved material, and the distance travelled by different solutes can differ enough to make it possible to detect the cosolutes in separate zones. These observations led to the classic studies by Goppelsroeder from 1861-1907, which were compiled in his Kapillaranalyse, published in Dresden in 1910. Bayley in 1878-86 described a method of analysis in which a drop of solution was placed upon filter paper, whereby concentric rings of varying concentrations of substances were formed. Using this terhnique, he described a method for the detection of cadmium in the presence of copper. In 1896 Trey found that with an ammoniacal solution containing copper and cadmium, cadmium occurs in the outer ring and can he detected with ammonium sulfide, which results in the formation of a yellow ring of cadmium sulfide about a black circle of copper sulfide. In 1809, Krulla proposed an interesting method of analysis hased upon capillarity. He found that when solutions of salt mixt,ures containing different ions mere allowed to rise in filter paper, each component was adsorbed to the same extent as if alone in the solution. The addition of similar ions, however, caused a repression of the capillarity of the ion in question. This fact was used in the following method: to the solution to be tested, different ions were added as long as a distinct decrease in capillarity occurred. Each ion which caused this decrease was, therefore, shown to be present in the test solution. In 1918 Feigl and co-workers began a systematic study of the use of specific and selective reactions for JOURNAL OF CHEMICAL EDUCATION
the analysis of most of the cations and anions, as well as organic compounds, and also the special techniques of spot tests. The result has been numerous publications in the literature, and a number of important books devoted to this subject. In 1921, Hauser first suggested the use of spot reaction techniques for the systematic analysis of the ions. Using the tests described by Feigl and Stern he devised a method for the analysis of the ions of Group I11 of the hydrogen sulfide system. Since that time, many schemes of qualitative microanalysis have been reported. Some of these apply spot and microscopic tests as supplementary tests in the usual hydrogen sulfide system and some use other group reagents, while
VOLUME 34, NO. 8, AUGUST, 1957
others simply carry out the analysis by means of specific and selective spot tests. For these methods we are greatly indebted to Tananaeff, Gutzeit, Heller, Kmmholz, Engelder, and Schiller, Gerstenzang, Emschwiller and Charlot, van Nieuenburg, Davies, BenedettiPichler, Wilson, and Emich. In conclusion, it is interesting to note that many of the so-called "newer" methods are not actually new at all. It is only their development that is new. Thus it might seem that much of value may be derived from a study of the historical records of chemistry, and the publications of the early chemists. Some chance observation might contain the germ of an idea, which if more fully developed, could lead to important results.
