Selectivity. Electrochemical detectors are inherently selective. When operated in an amperometric mode, they will respond only to those species oxidizable at or below the set potential (or reduced at or below the set potential). However, when operated in a differential pulse mode, the detector response can be limited to those species whose oxidation potentials lie near to or between the initial and final potential (i.e., initial potential pulse modulation amplitude). The current-potential curves shown in Figure 5 were constructed from the respective chromatographic peak currents obtained by the injection of a solution containing 0.350,0.406, and 0.304 nmol of p -aminophenol, 3,4-dihydroxyphenylalanine,and sulfanilamide a t each applied potential. (These same curves could be determined a t a rotating disk electrode far more conveniently.) Amperometric and differential pulse chromatograms of this test solution are shown in Figures 6 and 7, respectively. The potentials (and modulation amplitudes AE,for the differential pulse mode) were chosen with the aid of the current-potential curves, so that each chromatogram was optimized for the selective detection of a single component. These results clearly illustrate the increased selectivity of the differential pulse mode. The small peaks from ~~-3,4-dihydroxyphenylalanine
and p-aminophenol in the lower trace of Figure 7 may be due to differential capacitive effects but are more likely the result of an imbalance in the differential circuits. A further increase in selectivity can be achieved by lowering the modulation amplitude. This does, however, lead to decreased sensitivity. In practice, the optimum setting will be determined by the analysis under development.
LITERATURE CITED (1) D. C. Johnson and J. Larochelie, Talanta, 20, 959 (1973) 12) P. T. Kissinaer. C. Refshauoe. R. Dreilina, and R . N. Adams, Anal. Lett., 6,
465(1973)(3) P. T. Kissinner,L. J. Felice, R. M. Riggin, _.L. A. Pachla, and D. C. Wenke, Clin. Chem., 20; 992 (1974). (4) Y. Takata and G. Muto, Anal. Chem., 45, 1864 (1973). (5) V. G. Levich, "Physicochemical Hydrodynamics" Prentice-Hall, Englewood Cliffs, N.J., 1962. (6) 2. Feher, G. Nagy, K. Toth, and E. Pangor, Analyst (London),' 99, 699 '
(1974). (7) D. J. Myers, R. A. Osteryoung, and J. Osteryoung, Anal. Chern., 46, 2089 (8)
(1974). W. J. Biaedel, C. L. Olsen, and L. R. Sharma, Anal. Chem., 35, 2100 (1963).
RECEIVEDfor review June 15, 1976. Accepted August 27, 1976.
Determination of Rosin in Shellac by High Performance Liquid Chromatography and by Gel Permeation Chromatography Alan F. Cunningham, Geoffrey C. Furneaux, and Donald E. Hillman" Materials Quality Assurance Directorate, Royal Arsenal East, Woolwich SE 18 6TD, England
Rosin in shellac has been quantitatively determined using either reverse phase high pressure liquid chromatography (HPLC) or gel permeation chromatography.The HPLC method uses methanol/water/ammonium carbonate as the eluting solvent at 60 OC. Quantlties down to 0.005% are determined with good accuracy after a prior solvent separation to remove the bulk of the shellac. A direct gel permeation chromatography method is used for the analysls of samples contalnlng rosin added at the 10-20% level.
Shellac is the refined form of lac, the resinous secretion of the parasitic insect, Laccifir Lacca. This secretion, deposited on branches of trees, is a solid solution of condensation products of mono- and dibasic hydroxy acids, the main component (up to 50%) ( I ) being aleuritic acid (trihydroxypalmitic acid). Rosin is a complex mixture of abietic and pimaric type acids which may contaminate shellac accidentally during production or may be deliberately added a t high levels (10-20%) to meet certain requirements (2).The majority of specifications for commercial grades of shellac include a clause requiring the absence of rosin, e.g., when tested by the Halphen-Hicks test which is applied after solvent extraction of the shellac ( 3 ) .The test will give semiquantitative results in experienced hands by comparison of color intensities of samples and standards and will detect 0.01% rosin in shellac. The only generally recognized quantitative methods are those of Wijs-Langmuir and of McIlhiney ( 4 ) , both of which are unreliable below 5%. This report describes two methods for the quantitative determination of low levels of rosin in shellac. Both use an 2192
initial solvent separation of rosin from the bulk of the shellac. The first method uses a reverse phase separation by high performance liquid chromatography (HPLC). The second method uses a gel permeation chromatography (GPC) separation. Chang ( 5 )used a Bio Beads SX2 SX8 GPC column system to separate rosin acids, fatty acids, and their dimers in tall oil. Zinkel and Zank (6) used GPC on a Styragel column for the separation of rosin from fatty acids after the conversion to methyl esters. The GPC separation has the advantage of eluting rosin as a single peak whereas gas chromatography requires formation of the methyl esters which give a complex pattern of peaks due to separation of the individual components. Quantitation by gas chromatography is therefore difficult. The GPC method was also used without the solvent separation step for the estimation of high levels of rosin in commercial grades of shellac where 10-20% of rosin had been deliberately added.
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EXPERIMENTAL Apparatus. Liquid Chromatography, A Du Pont model 820 high pressure liquid chromatograph was used with a 1 m X 2.1 mm i.d. stainless steel column packed with Permaphase ODS (reversed phase). The mobile phase was 40% v/v methanol/water containing 1.0 g ammonium carbonate per liter. The column oven was maintained a t 60 "C when a pressure of 800 psi produced a flow rate of 1.1 ml/min. Samples were introduced onto the column by syringe and were detected by their ultraviolet absorbance a t 254 nm with detector sensitivity 0.02 absorbance unit (AU) full scale deflection. Gel Permeation Chromatography, A Waters Associates GPC 200 chromatograph was used with an ultraviolet detector (Applied Research Laboratories Ltd) operating a t 254 nm. The column system was composed of four 4 f t X 3/~in columns packed with Styragel of exclusion limits 500 300 200 60 A. Tetrahydrofuran (THF)
ANALYTICAL CHEMISTRY, VOL. 48, NO. 14, DECEMBER 1976
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Table I. Quantitative Determination of Rosin in Shellac
Rosin added, %
Sample
A B C D E F G H I J K
Hand made shellac Hand made shellac rosin Hand made shellac rosin Hand made shellac + rosin Hand made shellac + rosin Hand made shellac + rosin Hand made shellac rosin Machine made orange flake shellac Orange shellac, dewaxed Garnet shellac, dewaxed Garnet shellac, dewaxed Limit of detection
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Rosin found, % HPLC system GPC
