Whatman filter paper (46 X 75 cm) No. 1for chromatography. The chromatogram was run in a mixture of n-butanokacetic acid:water (4:1:5) by the descending chromatographic technique. The chromatogram was dried and sugar spots were revealed by immersing the paper in aniline hydrogen phthalate reagent solution ( 3 ) .The chromatogram was then dried, first at room temperature and then at 110 "C (10 min). The colored fucose spots were cut off from the chromatogram and extracted with 10 ml of different solvents given in Table I. The intensity of the clear extract was measured on a Hilger spectrophotometer (Model No. H.700.304/54542) at 360 mp and the percent fucose extracted was calculated from the standard curve (straight-linewith slope = 0.44) obtained by plotting the concentration of L-fucose vs. optical density of the extract.
RESULTS AND DISCUSSION It will be seen from Table I that increase in the acidity of the solvent from 0.7 to 1.0 N has improved the extraction very little. Addition of ether along with the change in the acidity has decreased the extraction. B u t addition of 10%acetone t o the solvent quickened the extraction and all the sugar spots could be extracted within 6 h. The extraction could be carried out still faster by composition No. 9 (Table I) in which 20% acetone was added. How-
ever, increase in acidity of solvent No. 9 has no beneficial effect. Solvent mixture No. 9 was also used for the extraction of other hexose sugars from chromatogram and is found to work with equal efficiency.
ACKNOWLEDGMENT My sincere thanks are due to E. R. R. Iyengar, Head of the seawater algae and irriculture division, and D. J. Mehta, Director of the Institute, for their keen interest in this work. I am also thankful t o Y . A. Doshi and D. R. Baxi for their suggestions during the course of this work.
LITERATURE CITED (1) R. J. Block, E. L.
Durrum, and G. Zweig, "A Manual of Paper Chromatography and Paper Electrophoresis" Academic Press, New York, 1958, p 189. (2) G. Zweig and J. R. Whitaker, "Paper Chromatography And Electrophoresis", Vol. 1, Academic Press, New York and London, 1967, p 271. (3) C. M. Wilson, Anal. Chem., 31, 1199 (1959).
RECEIVEDfor review May 27, 1976. Accepted July 14, 1976.
Mercury Thin-Layer Electrode Permitting Gas Chromatographic Determination of Organic Electrolysis Products Joe L. Hanley' and Dennis G. Peters* Department of Chemistry, Indiana University, Bloomington, lnd. 4740 1
Electrodes for use in thin-layer electrochemistry have been developed for a variety of purposes (1-12). However, no report of a convenient mercury thin-layer electrode suitable for product analysis by means of gas chromatography, particularly for studies of the electrochemical behavior of organic compounds, has appeared in the literature. Several characteristics are desirable of such an electrode-it should allow for the rapid chromatographic analysis of electrolysis products, be applicable to nonaqueous as well as aqueous media, require only a small amount of sample, and be simple in design. In this communication are described a mercury thinlayer electrode and an electrolysis cell t h a t meet these requirements. To assess the usefulness of this system, we have followed the course of the chronopotentiometric reduction of 1-phenyl-1-hexyne in anhydrous dimethylformamide containing tetra-n -butylammonium perchlorate by interrupting the electrolysis a t various times and by determining the products with the aid of gas chromatography.
EXPERIMENTAL Construction and Use of Thin-Layer Electrode and Electrolysis Cell. As shown in Figure 1,the thin-layer electrode is basically an immersible electrode similar to the design described by Tom and Hubbard ( 2 1 ) employing a boundary made from Corning 7930 porous Vycor glass. Vycor glass was obtained as thick-walled capillary tubing (7.4-mm o.d., 2.1-mm i.d.), but was ground down with a wetwheel saw to produce tubing having a wall thickness of approximately 0.45 mm; the decreased wall thickness lowers the ohmic resistance of the porous boundary significantly, thereby improving the apparent reversibility of electrode processes that occur in the thin-layer cavity. Using E-63 Epoxy Adhesive purchased from Techkits (Demarest, N.J.), we cemented a 6- to 10-mm length of the thinned Vycor tubing at the top to constricted Pyrex tubing (8-mm 0.d.) and at the bottom to a glass cup fabricated from Pyrex tubing (12-mm 0.d.) which served as the auxiliary-electrode compartment. Teflon (Slic-Seal, made by Permanent address, Department of Chemistry, Lake Land College, Mattoon, Ill. 61938. 2036
Hoke, Inc., Cresskill, N. J.) was applied to each joint to prevent contact of the adhesive with the supporting electrolyte-solvent system. A length of platinum wire (0.080-in.diameter), coated with mercury according to the method of Ramaley, Brubaker, and Enke ( I 3 ) ,served as the working electrode. For the design depicted in Figure 1, the positioning of the working electrode within the porous Vycor tubing as well as the non-precision bore of the tubing itself precludes the use of the electrode for kinetics studies because the thin-layer cavity is not necessarily uniform. However, as discussed by Hubbard (12),the thin-layer cavity can easily be made uniform, if desired. A platinum wire (0.025-in. diameter) was connected to the mercury-coated electrode and was extended upward through the entire length of the Pyrex tube. This platinum wire was held in place at the top with a fitting that allowed the mercury-coated working electrode to be raised or lowered in order to achieve proper positioning within the Vycor tubing; the working electrode was positioned visually so that it protruded just beyond the bottom of the Vycor tubing. Fine platinum gauze was wrapped around the outside of the Vycor tubing to form the auxiliary electrode; as an electrical contact, a length of platinum wire was heat-welded to the gauze and then sealed through and passed inside the Pyrex tubing. As reference electrode, we used a saturated cadmium amalgam in contact with anhydrous dimethylformamide saturated with both cadmium chloride and sodium chloride ( 1 4 ) ;the potential of this electrode is -0.750 V vs. SCE (aqueous saturated calomel electrode). This reference electrode terminated in a cracked glass tip ( 1 5 )positioned a few millimeters below the thin-layer electrode. All potentials in this paper are quoted with respect t o the cadmium amalgam-dimethylformamide electrode. To the inner wall of the cell at the level of the supporting electrolyte-solvent therein was sealed a circular glass platform upon which a small open beaker containing the desired sample solution was placed. A stiff platinum wire inserted through a port in the top of the cell and wrapped around the beaker allowed it to be moved to and from the platform as well as to a position beneath the working electrode. To deaerate the sample solution, the working electrode was raised above the level of supporting electrolyte-solvent in the cell, and the beaker containing the sample solution was swung under the working electrode so that the tip of the electrode was immersed in the solution; pre-purified nitrogen saturated with solvent vapor was then allowed
ANALYTICAL CHEMISTRY, VOL. 48, NO. 13, NOVEMBER 1976
to flow past the working electrode and to bubble through the sample solution. An ordinary sintered-glass nitrogen inlet permitted deaeration of the supporting electrolyte-solvent in the main compartment of the cell. Screwing the thin-layer electrode assembly downward allowed the auxiliary-electrode compartment to become filled with deaerated supporting electrolyte-solvent from the main compartment; then the thin-layer electrode assembly was screwed upward to its normal position. To fill the thin-layer cavity,the beaker of deaerated sample solution was positioned beneath the working electrode, and the Teflon fitting supporting the working electrode was screwed downward until the tip of the electrodejust touched the sample solution and the thin-layer cavity filled by capillary action. Then the Teflon fitting was screwed upward, the beaker was returned to the platform, and the Teflon fitting was screwed downward until the mercury-coated working electrode made contact with the supporting electrolyte-solvent, Although a small beaker containing approximately 2 ml of solution was employed as the sample reservoir, there is no reason why as little as a total of 100 p1 of sample solution cannot be tolerated; it is only necessary that the container be large enough and the depth of sample solution sufficient to permit the tip of the thin-layer electrode to be immersed into the sample. After an electrolysis, the Teflon fitting holding the working electrode was removed from the cell. With the electrode assembly held horizontally, careful withdrawal of the mercury-coated platinum wire left almost all the sample solution inside the porous Vycor tubing. A syringe was utilized to remove the sample from the Vycor tubing for purposes of analysis. Because dimethylformamide interfered with the gas chromatographic analysis of electrolysis products, the sample could not be injected directly into a gas chromatograph, although this might be done if the retention time for the solvent differs significantly from the retention times of products. For the studies mentioned in this paper, the electrolyzed sample was syringed from the Vycor tubing and injected into a melting-point capillary. A small volume (2 to 3 pl) of diethyl ether was syringed into the capillary tube, followed by water (10 pl), and the open end of the capillary was sealed with the aid of a propane torch. Repeated inversion and centrifugation of the capillary caused the electrolysis products to be extracted into the ether, whereas most of the dimethylformamide entered the aqueous phase. Then the capillary was broken, and the ether layer was injected directly into a gas chromatograph. This procedure reduced the size of the chromatographic peak for the solvent so that it no longer obscured or overlapped the peaks for the products. With practice, the entire procedure, starting with the actual electrolysis, followed by removal of the sample from the thin-layer cavity and by the microextraction, and ending with injection of the ether extract into the chromatograph, requires no more than 5 min. Reagents, Instrumentation, and Procedures. Dimethylformamide (Fisher Spectranalyzed material) employed as solvent in all the studies was dried over Linde 4A molecular sieves before use. Tetra-n-butylammonium perchlorate, purchased from the G. Frederick Smith Chemical Company and dried in a vacuum desiccator, served as the supporting electrolyte. Except for 1-phenyl-1-hexyne, obtained as described previously ( I 6 ) ,all other chemicals were reagent grade. Chronopotentiometric apparatus utilized in this work was of a conventional design ( 1 5 ) , and transition times were measured according to a procedure described elsewhere (17).A Hewlett-Packard Model 700 gas chromatograph equipped with a flame ionization detector and a 14-ft X Ya-in. column packed with 15%UCON Polar on 80-100 Chromosorb W was used for product analyses. Using different lengths of Vycor tubing, we constructed thin-layer electrodes with capacities ranging from 1 to 2 p1 of solution; the thickness of the solution layer was approximately 3 pm. Chronopotentiometric measurements (10) with standard solutions of anthracene in dimethylformamide containing 0.1 M tetra-n-butylammonium perchlorate were employed to determine the exact volume of the thin-layer cavity of each electrode. A thin-layer chronopotentiogram for reduction of anthracene exhibits two waves with half-wave potentials of -1.17 and -1.73 V corresponding, respectively, to formation of the radical-anion and dianion of anthracene; the first wave is more sharply defined and, therefore, better for calibration of thin-layer electrodes. T o begin a series of experiments, a 0.1 M solution of tetra-n-butylammonium perchlorate in dimethylformamide was transferred to the main compartment of the cell. Approximately 2 ml of a solution of 0.1 M tetra-n-butylammonium perchlorate in dimethylformamide containing a known concentration of 1-phenyl-1-hexyne was added to the small beaker on the platform. Then the cell was closed, the solutions were deaerated, and the auxiliary-electrode compartment was filled with air-free supporting electrolyte-solvent. Next, the
f
WORKING ELECTRODE
AUXILIARY ELECTROD
*>TEFLON
.'
AUXILIARY ELECTRODE (FINE Pt GAUZE AROUND VYCOR TUBE1
AUXILIARY-ELECTRODE
-
POROUS VYCOR
TUBE
r'
Hg-COATED P I WIRE
THIN-LAYER CAVITY
Flgure 1. In the upper part of the drawing is the completely assembled electrolysis cell with the thin-layer electrode in position for an electrolysis. In the lower portion is a detailed view of the tip of the thin-layer electrode
thin-layer cavity was filled and a complete chronopotentiogram was recorded; then the electrolyzed solution was expelled from the thinlayer cavity into the main compartment by passage of nitrogen through the tube supporting the mercury-coated working electrode. Finally, a series of individual electrolyseswas performed in which the thin-layer cavity was refilled and the potential of the mercury-plated cathode was allowed to reach various preselected values along the chronopotentiometric wave, after which the solution in the thin-layer cavity was analyzed by means of gas chromatography. For the majority of experiments, the constant current was chosen so that complete reduction of the starting material was achieved in 20 to 50 s.
RESULTS AND DISCUSSION To test the usefulness of the thin-layer electrode for studies of organic compounds, we examined the chronopotentiometric reduction of 1-phenyl-1-hexyne in dimethylformamide containing 0.1 M tetra-n-butylammonium perchlorate as supporting electrolyte. In accord with the polarographic behavior of 1-phenyl-1-hexyne (16),a thin-layer chronopotentiogram exhibits a single well-defined wave for the four-electron reduction of starting material t o 1-phenylhexane. In one experiment, a 0.0088 M solution of 1-phenyl-1-hexyne was electrolyzed at a constant current of 193 FA to a point on the chronopotentiometric wave equivalent to one-half the transition time. Gas chromatographic analysis of t h e solution revealed t h e presence of 1-phenylhexane (46%), trans-l-phenyl-1-hexene (ll%), cis-1-phenyl-1-hexene (6%), and trans1-phenyl-2-hexene (5%) along with unreduced l-phenyl-lhexyne (32%). These results agree remarkably well with those found previously (16) for the large-scale controlled-potential M 1-phenyl-1-hexyne electrolytic reduction of 2.5 X under conditions corresponding t o the transfer of two electrons per molecule of starting material. However, if a solution containing 0.0088 M l-phenyl-l-
ANALYTICAL CHEMISTRY, VOL. 48, NO. 13, NOVEMBER 1976
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hexyne is subjected to a large-scale electrolysis at controlled potential, the distribution of products after addition of two electrons per molecule of alkyne-trans- 1-phenyl-1-hexene (72%), trans -1-phenyl-2-hexene (17%), 1-phenylhexane (8%), and cis-1-phenyl-1-hexene (3%)-differs substantially from that seen in the thin-layer chronopotentiometric experiment. This difference in product distributions is attributable to the fact that electrolytically induced isomerization of l-phenyl1-hexyne to l-phenyl-1,2-hexadiene occurs to a much greater extent during the large-scale electrolysis of a well stirred solution than during the electrolysis of an immobile thin layer of solution; it is reduction of the allene isomer which yields relatively little 1-phenylhexane but large amounts of trans1-phenyl-1-hexene and trans-1-phenyl-2-hexene (16). We believe that the mechanism of isomerization involves an interaction between 1-phenyl-1-hexyne and its electrogenerated radical-anion and that the longer time these two species have to react during a large-scale electrolysis is responsible for the abundance of allene-derived products. Presumably, for systems kinetically less complicated than the reduction of 1phenyl-1-hexyne, the product distributions observed for thin-layer and large-scale experiments with any given concentration of electroactive substance should be comparable. On the other hand, for systems whose behavior is governed by kinetically controlled processes, it might be possible to gain valuable mechanistic information through a comparison of the product distributions found in thin-layer and large-scale electrolyses. Repeated cathodic polarization of the working electrode causes deterioration of the mercury coating on the platinum wire; the more negative the potential to which the electrode is polarized, the shorter the lifetime of the mercury coating. In experiments with 1-phepyl-1-hexyne, as many as ten electrolyses could be performed before renewal of the mercury coating was necessary. Besides our work with 1-phenyl-1-hexyne, the thin-layer chronopotentiometric reduction of 6-chloro-l-phenyl-lhexyne has been examined briefly. As reported previously (IS),an exhaustive large-scale controlled-potential reduction at -1.75 V of a 2.5 X M solution of 6-chloro-1-phenyl1-hexyne a t a mercury pool in dimethylformamide containing 0.1 M tetra-n-butylammonium perchlorate yielded benzylidenecyclopentane (81%),benzylcyclopentane (6%), 1 phenyl-1-hexyne (4%), trans - 1-phenyl-1-hexene (3-) , cis -1phenyl-1-hexene (2%), trans-1-phenyl-2-hexene (2%), 1phenylhexane (l%), and 1-benzylcyclopentene (
