Robert V. Dilts Vanderbilf University Nashville, Tennessee 37203
Methods of Separation The sophomore analytical course
O n e of the major problems involved in retaining the identity of the course in quantitative analysis is the updating of the lecture material and the selection of suitable experiments to illustrate the new topics. There is a current trend in the teaching of analytical chemistry toward incorporating some of the techniques of volumetric and gravimetric analysis into the second semester of the beginning chemistry course and then delaying a thorough discussion of the principles of analytical chemistry until the senior year, when the student has had both physical and organic chemistry. The ACS has recognized this trend and strengthened it in its 1962 revision of the "Minimum Standards Used as Criteria in Evaluating Undergraduate Professional Education in Chemistry." There is definite merit in this approach and it is unfortunate that one of the results has been a growing tendency to drop completely the analytical chemistry course which was usually taught to sophomores. Unless the laboratory of the beginning course is closely supervised by adequately trained personnel and standards are raised to those normally found in a sophomore analytical chemistry course, the quality of the work will be low and experience of little value. Since only a relatively few techniques are taught anyway, the student is deprived of many techniques and a great deal of analytical methPresented at the Joint Symposium on The Teaching of Analytical Chemistry at the 152nd Meeting of the American Chemied Society, New York City, September, 1966.
odology that he could he using during his undergraduate years. Moreover, while the graduating senior knows about the physical principles of acid-base, oxidationreduction, complex ion formation, and precipitation titrations, the use of modern instruments, and the theory of gravimetric analysis, he often is not particularly well versed in those important separation techniques that are of extreme value in modern analytical chemistry. His exposure to extraction, ion exchange, all forms of chromatography, and other separation techniques is minimal. These contacts, in an organic or physical chemistry course, have not been explored thoroughly except in terms of the specific organic separation desired or of the physical property to be studied. General Considerations
During a revision of the curriculum of the chemistry department at Vanderbilt University in 1962, it was decided to reorganize completely the two-semester Quantitative Analysis course that had been offered during the sophomore year. I t was felt that the classical quantitative analysis course no longer fulfilled our needs in training students in analytical chemistry. The subject matter barely touched upon the contemporary practice of analytical chemistry or upon those subjects in which there was active research. Separation methods and the laboratory techniques involved in their use had not been taught a t all. Important new reagents, such as EDTA, were not discussed at a
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depth commensurate with their significance. Consequently, the students received a false impression of the field of analytical chemistry and were not at all prepared to pursue graduate study in it. Since the students entered the course directly from the beginning chemistry course, they had not had any physical or organic chemistry. This meant that large amounts of the theory of analytical chemistry could not be taught properly, if a t all, and that organic analysis could not be included. Attempts were made to modernize the beginning analytical chemistry course so as to include modern analytical methods and to defer a thorough study of the physical basis for classical methods and instrumental analysis until the student had learned physical chemistry. The year course was separated into two onesemester portions; the first to be taught during the sophomore year and the second to be taught during the senior year. It was then decided to make the sophomore course a quantitative course in separation methods. The separated components of the mixtures would be analyzed by whatever methods were most suitable for them. I n this manner the basic laboratory skills of volumetric, gravimetric, and colorimetric analysis could be taught, as well as the separation techniques. All of these could be capitalized upon in subsequent organic, physical, and inorganic laboratory courses. For example, the use of the analytical balance is taught during the experiment on separation by precipitation. Nickel is determined by precipitation and subsequent weighing of its dimethylglyoxime chelate. Titrations and the use of pipets are covered in the ion exchange experiments. Standard solutions of sodium hydroxide are prepared and standardized as in a normal course, but instead of using these solutions to determine soda ash or an unknown potassium acid phthalate, the capacity of a cation ion exchange resin is determined. A second ion exchange experiment in which cadmium and zinc are separated illustrates the applications of standard EDTA solutions for the determination of metal ions. Pipetting technique, volumetric equipment, and colorimetry are covered in conjunction with the separation of fluorescein and methylene blue by adsorption chromatography and subsequent determination of each of the dyes spectrophotometrically. The second semester of analytical chemistry is a course taught during the senior year, with organic and physical chemistry prerequisites. It covers, in an intensive fashion, using instrumental techniques where applicable, those topics often covered in the classical quantitative analysis course, and is extended to include more modern and complex analytical instrumental techniques such as nuclear magnetic resonance, X-ray fluorescence, and infrared spectroscopy. Development of the Course in Separation Methods
The students taking the sophomore course consist of professional chemistry majors, premedical students, chemical engineers, and liberal arts students. Consequently, attempts were made to include topics that would be of interest and also useful in the student's chosen career. Insofar as possible, attempts were made to build upon whatever previous exposure to 314 / Journal of Chemical Educofion
analytical chemistry the students had had in the beginning chemistry course. It was decided to use only inorganic chemistry in the laboratory and lecture to illustrate the separation techniques. Many of the separation methods included in this course are most useful and powerful when they are applied to organic or biochemical systems. I t was felt, however, that our primary purpose should be to explain the underlying principles of the technique in the lecture and to teach the general experimental manipulations in the laboratory. The specific chemistry of the laboratory experiment is less important since each teacher will have favorite systems that he feels most challenging and illuminating. The probability that a student will apply the separation technique to the exact system used in an undergraduate laboratory is relatively small, so that regardless of the particular system chosen, later applications will be different ones. If the basic information has been learned, then modifications of this for other applications should prove to be no serious obstacle for the student. Consequently, it was felt that it would be less confusing, remove one complication factor, and lead to a stronger emphasis on the technique per se and less on the unfamiliar chemicals involved if the laboratory experiments involved primarily inorganic chemicals. Since this decision was reached, organic chemistry has been moved to the sophomore year. Beginning this year, the students will have had one semester of organic chemistry and take the second one concurrently, so that organic systems will be included in the course in the future. An equilibrium approach to the theory of the separations provides an underlying connecting theme and is the simplest approach for the students, who have been exposed to the concept in the beginning chemistry course. In almost all of the methods covered in the course, the equilibrium approach is the major one used commonly, so that there is no distortion of the method in order to fit it into this general framework. Where other theoretical approaches-such as the theoretical plate concept or the Donnan membraneare alternatively used, this fact is merely pointed out to the student. Selection of textbooks has proved a problem that has not yet been solved satisfactorily. No sophomore level text that treats equilibrium, applies the concepts to methods of separation, and then gives illustrative problems and laboratory experiments is available. E. W. Berg's "Physical and Chemical Methods of Separation" is used as the descriptive lecture text; either J. N. Butler's "Solubility and pH Calculations" or A. J. Bard's "Chemical Equilibrium" as a text for treating equilibrium concepts and providing numerical problems; and J. Waser's "Quantitative Chemistry" as part of the laboratory manual. For eleven experiments mimeographed instructions are provided for the students. A. I. Vogel's "A Textbook of Quantitative Inorganic Analysis" is recommended to the teacher as a source for laboratory experiments. The Lecture Portion of the Course
The course consists of two hours of lecture each week for a fourteen week semester. About the first half of the material in the lecture treats general equi-
librium and its application to the problems of separations. A unified equilibrium concept is taught. Each kind of equilibrium is merely considered as a specific application of the general case. Not only are aqueous acid-base and polyprotic equilibria treated, but some discussion of non-aqueous concepts is included. After this, the emphasis is upon the combination of equilibrium conditions, so that the use of precipitation as a means of separation is treated and the effect of pH on solubility is well covered. Complex ion equilibria are discussed through the means of stepwise formation constants, the distribution of species in solutions of complexing agents, and the average ligand number. This is followed by the treatment of the effect of pH on complex ion equilibria, the effect of complex ion form* tion on solubility equilibria, ending finally with a discussion of et.hylenediminetetraacetic acid, its chelates, and the effect of pH on their stability. Partition equilibria have not been encountered by the students before, so a general discussion of the distribution law and the factors that affect the distribution of a substance are treated in considerable detail. The distribution ratio is defined and theu the effect of dissociation, association, and complex ion formation upon it and the distribution of solutes is explained. Finally, the equilibria involved in the distribution of a metal ion between an aqueous phase and an organic solution of a chelating agent are treated according to the method given by Irving and Williams.' The remainder of the lecture material deals with descriptive material, the theory and operations of separations themselves. The discussion of partition equilibrium leads naturally into the treatment of extraction, and counter-current distribution. Examples are given for the extraction of nonsolvated solutes such as osmium tetroxide, separation of metal ions by extraction as their chelates, and extraction of ion pairs with the ferric chloride system being discussed in detail. A treatment of some of the theory of adsorption is necessary prior to a discussion of adsorption chromatography. This is followed by a description of the methods of column operation and then specific systems are mentioned. Partition chromatography is explained and the effect of operation variables is pointed out insofar as possible. The techniques and terminology of paper chromatography are treated and the meaning and uses of R, values given. Only about two lectures are devoted to gas chromatography, these being used to point out the similarities and differences of apparatus and technique between this and the other types of column chromatography. The last topic covered is ion exchange chromatography, the application of the equilibrium principle to exchangers, a description of resins and their properties, and finally some brief treatment of the applications of ion exchange. The Laboratory Portion of the Course
The laboratory meets for two three-hour periods each week during the semester. The experiments attempt to illustrate, insofar as it is possible under the 'IRVING, H. M., (1949).
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WILCIAMS, R. J. P.,J . Chem. Sac., 1841
limitation given above, those techniques discussed in lecture for which equipment is available. I t also presents standard analytical techniques within the framework of separation techniques. Precipitation. The separation of nickel from iron by precipitation with dimethylglyoxime illustrates the effect of pH on chelate formation, the use of masking agents-tartrate in this c a s e t h e nature of uncharged chelates, as well as teaching the standard laboratory techniques of gravimetric analysis. Student results on standard nickel samples has been quite satisfactory. Ion Exchange. The preparation and standardiaation of standard sodium hydroxide introduces the concept of standard solutions and their properties, as well as teaching titration technique. This solution is theu used to determine the capacity of an unknown ion exchange resin. A known weight of air-dried cation exchange resin in the hydrogen cycle is placed in a 25-ml buret and eluted with sodium sulfate. From a titration of the sulfuric acid produced during the complete exchange on the column, the student calculates the capacity of the dry resin. Student results on this experiment have been excellent, the experiment is simple and straightforward to perform. EDTA is prepared and standardized against pure zinc using Eriochrome Black T as the metallochromic indicator, demonstrating to the student the nature of chelating agents, more about indicator action, and chelatometric titrations. The anion exchange separation of zinc and cadmium as their chloro complexes on Dowex-1, illustrates the effects of chloro- and hydroxycomplex ion formation on selectivity coefficients, as well as giving the student practice in the actual technique of ion exchange. The separated metal ions are titrated with EDTA and the number of milligrams of each in the initial sample reported. Since EDTA will titrate both metal ions under the conditions used, no direct indication of the cleanness of the separation is obtained. The only clues the students have are their grades and the posted metal ion contents for the uuknown solutions. Since students have difficulties in observing the Eriochrome Black T end point, results on this experiment are only moderately good. Extraction. A study of the effect of pH on the extraction of copper into a chloroform solution of 8hydroxyquinoline illustrates the lecture material on the extraction of metal chelates and shows the students how one determines the pH,,,. Varying amounts of dilute acid are added to copper solutions; these solutions are shaken with a chloroform solution of oxine; the pH of the aqueous phases is measured; and the concentration of the copper oxinate in the chloroform phases is determined colorimetrically. A graph of the percent copper extracted vs pH is drawn and the pH,,, ascertained from this. The separation and determination of copper, lead, and calcium by extraction of their oxinates into chloroform shows how the pHLl,values and pH adjustment of the aqueous phase can be used to separate first copper, then lead, and finally calcium by selective extraction. The extracts are analyzed colorimetrically a t 425 mfi using calibration graphs that are provided. Care must be taken to keep the metal ion concentration low Volume 44, Number 6, June 1967
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or precipitate formation will occur. When this is done student results are satisfactory. Chromatography. Partition chromatography is illustrated by the experiment in which chlorocomplexes of copper and nickel are separated on a cellulose column using HC1-acetone as the eluting agent. The separated metal ions are then titrated with standard EDTA and the number of milligrams of each present in the initial sample reported. Students enjoy this experiment and the results obtained are good. The separation of methylene blue from fluorescein on an alumina column is used as an example of adsorption chromatography. Ethanol is the eluate for the methylene blue and water is used for the fluorescein. The fractions are collected in volumetric flasks and analyzed calorimetrically by comparison against standard dye solutions. The color of the dyes against the white column indicates clearly that the separation is clean, for the fluorescein does not move from the top of the column with ethanol. This experiment is also a good test of the students' ability to use volumetric equipment, and the grades reflect this. The experiment in paper chromatography is only qualitative in nature. A mixture of copper(II), cobalt(II), iron(III), and nickel(I1) ions is separated on a piece of Whatman #1 circular filter paper in a Petri dish. HC1 in acetone is used as the eluting agent. An unknown containing any or all of these ions is run simultaneously with a standard containing all of them and from a comparison of the R, values obtained and the visual evidence, the ions present in the unknowns are identified. The students like the experiment greatly since almost everyone gets his unknown correct. The potency of thin-layer chromatography is illustrated by the separation of the cis- and trans-isomers of dichlorobis(ethylenediamine)cobalt(II) chloride on thin-layer chromatographic sheets. Methanolie solutions are used as the eluting agent and rubeanic acid is the developer. The experiment is only qualitative in nature, but serves to illustrate how two very similar compounds with only slight differences in polarity and solubility can be resolved by this technique. The separation is not complete and careful technique is essential to avoid obscuring the spots. Instmcmental Techniques. Students who will take no additional analytical chemistry should be introduced t o two of the most widely used instrumental techniques. Therefore, experiments in potentiometric pH determinations and colorimetry are also included in the course. The equivalent weight and acid dissociation constant of an unknown weak acid are determined by means of titration with the standard sodium hydroxide using the glass electrode and a direct reading p H meter. This not only introduces the students to the use of the pH meter, but also shows them how equivalent weights and ionization constants are determined. The results obtained compare very favorably with tabulated values.
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The colorimetric determination of iron with 1,lOphenanthroline is a classic experiment designed to introduce the student to the technique of obtaining spectra, preparation of Beer's Law plots, and analysis of unknown iron samples. The results are quite good in general. Grading
The grading system was designed so that 50% of the student's final grade was determined by his performance in the lecture portion of the course and 50% by his results on the laboratory experiments. Results among the students that have completed the course in the past four years show that 80% of all those students receiving A's in the course became chemistry majors and only 20% majored in science departments other than chemistry. Fifty-eight percent of all of the A students were premedical students, 4% were chemical engineers, and 31y0 were to go to graduate school in chemistry. Attempts to use this course as the Advanced Placement course have met with varied success. Most students did not perform as well as was expected of them. There appears to be a level of maturity and scientific sophistication, a greater drive, and a degree of laboratory skill required that is greater than that found in the usual Advanced Standing student. It should be pointed out that several of these students were at the top of the class, so that much more seems to depend upon the individual student than performance on any tests. Conclusions
I n general, student performance in the laboratory exceeded that in the lecture. Results 011 the experiments were felt to be more than compensating for a low test score, so that greater time, effort, and interest was spent on the laboratory material. Some of the experiments require considerable attention to detail and meticulous work that a great many students seemed unwilling to put forth, but the majority of the students enjoyed these challenges. I n designing and developing new experiments, a great care must be taken to ensure that the system is foolproof in the hands of these relative beginners, for experiments reported that are successful when performed by skilled technicians can be complete failures when run by students inexperienced in the manipulative techniques. The course has been taught for four years now and it is felt to be successful. Experimental results, in general, are good. Student response has been enthusiastic. Students are pleasantly surprised when they discover that they actually use in summer jobs; in graduate or medical school; or in other chemistry, biology, or engineering courses the techniques that they have learned in this course. It is felt that the course has achieved its purpose of presenting modern analytical techniques without abandoning the sophomore year course in analytical chemistry.
