A Three-Piece, All-Glass System
for Radial Chromatography Karl-Heinz A. Roslerl, Ralph N. Blomster, and Mark J. Schocken University of Maryland. School of Pharmacy. of Medicinal ChernlstrylPharmacognosy,20 N. Pine Street, Baltimore, . Department . MD 21201
For a number of . vears., in our courses we have been usine a simple system for the radial development of thin-layer plates. I t compares favorably with all other designs of radial chromatography systems that have heen described in the literature or are commercially available. Since the developing chamber is all-glass, the choice of developing solvents is virtuallv unlimited. Furthermore. as shown in Fiaure 1. the entire &face of the thin-layer plate is only millim&ersaway from the solvent surfa~e,ensuring total and uniform saturation. The design allows the solvent flow to start simply when downward pressure is applied to the center of the hack of the TLC plate. This leaves the system undisturbed between presaturation and actual development. Commercially available amaratus such as the U-Chamber hv Camae2 .. - trv.to achieve saturation and possibly presaturation by pumping solvent vavors into the void under the adsorbent laver. The design ch&acteristics of other systems such as the one by Analtech3 or Blome ( I ) do not allow presaturation of the plates. The speed of the system is dependent on the viscosity of the solvent. Kaiser ( 2 ) ,using the U-Chamber, found the solvent reached a diameter of 40 mm in 199 s with toluene as the solvent. In our system the 40-mm-diameter solvent front was reached in 123 s after 5 min preequilibration and 106 s after 10 min of preequilihration time using E. Merck HPTLC-Silicagel plates. Beyond 10 min of equilibration, the developing time remained constant. For other solvents of different viscosities such as n-hexane, benzene, and cyclohexane, we obtained values of 88 s, 109 s, and 225 s, respectively. Deslgn and Material Our system basically consists of three pieces:
Figwe 1. Side view of Petri dish,
10x10
m nc CUTE
10 CY
ccnn o m
5 CY OUSS CUTE 15 W W3'M D S H 10CYW3'MDgn 20.20 CY
nc CUTE
Figure 2. Circled area of Figure 1
n c CUTE
1. A glass Petri dish, whose rim had been ground to uniform
smoothness,which serves as solvent reservoir. 2. A short piece of 2-mm-o.d./l-mm-i.d. glass tubing which feeds the solvent to the TLC plate. 3. A rectangular slab of 5-mm glass with a 2-mm bore at its center which holds the feeder capillary in an upright position and also centers it. Figure 1shows a side view of the Petri dish assembly. Figure 2 is the enlarged view of the circled area in Figure 1 and shows the details of the capillary in the center and its placement within the svstem. Both ends of the ca~illarvare smoothly ground a i d at the bottom end the edgeihavibeen bevelled to accommodate a better fit into the little recess a t the center of the Petri dish. T o allow for access of solvent to the capillary, a small hole has been ground into its side 6 mm from the bottom end. The TLC plate is placed on top of the Petri dish with the
-
' Cleveland Twist Drill Co., Cleveland. OH.
Camag. Postfach 34. Muneny, Switzerland: US. Agents: Applied Analytical Industries. Wilmington. NC. Anaitech, Inc.. Newark. DE.
sa,VRIT
WW 2 U1O.D. IU)IWO
(YIan CUTE PETRI DISH
Figure 3. Tap view the complete assembly is shown only fw the 10 X 10-cm n c oiates.
layer facing the developing solvent. In order to allow for reproducibility of matching the center of the TLC plate and the feeder capillary, a complementary base plate is used. This base plate has blocks attached to two of its sides, which serve as stops for the Petri dish during its alignment. Figure 3 is the top view to illustrate how the bottom plate assembly can accommodate Petri dishes of different diameVolume 63 Number 9
September 1966
813
ters. In operation, the Petri dish is placed onto the bottom h late, touching both of the 5-cm aligning hlocks. Solvent is Hdded until it kahoiit 2 mm l~rlowthr ul;;)rr rim of the Petri dish. A T1.C d a r e is lar red on tor, of the Petri dish and two edges of the dish are pressed against the aligning hlocks. In a properly made system, the plate should not make contact with the capillary a t this point. Contact, which is manifested as a wet spot on the TLC plate, should occur only after a slight finger pressure has been applied to the center of the glass plate. T o develop the TLC plate, it is placed on the Petri dish in the same orientation as in the center-marking procedure. Ultimate saturation of the TLC adsorbent with solvent vapors is achieved if a time interval of 10 minis allowed before contact between the capillary and the TLC layer is made by applying slight finger pressure to the center of the hack of the plate. The materials required to build the system are commercially available. The base plate is a standard 20 X 20-cm glass plate and the aligning hlocks can be of any suitahle, solvent resistant material, which are attached with douhlesided tape. The capillary is cut from standard glass tubing of 2-mm o.d. and 1-mm i.d. (5Ie4-in. o.d. and 5/12s-in. id.). A tungsten carbide drill hit of 2 mm' and a quarter-inch electric drill was used to hore the hole for the capillary and the recess a t the center of the Petri dish. All the glass-cutting and grinding that is necessary to match the capillary support with the Petri dish and to prepare the capillary can be done with a handheld grinding stone, a diamond pencil, and a file. The glass plate which holds the capillary is cut from a standard 20 X 5 X 0.5-cm TLC glass plate. Since it is difficult to cut this plate accurately enough so that it has minimal play inside the Petri dish, it is custom fitted by cutting i t slightly longer than needed and rounding off the corners by grinding. For accurate centering it is advisable to make permanent alignment marks on the plate near one of its corners and another one on the outside of the Petri dish. A third alignment mark should he placed on one of the two hlocks on the base plate. After the capillary holding slab had thus been prepared and aligned, the center of i t is determined by mark-
814
Journal of Chemical Education
ing the spot above the recess in the bottom of the Petri dish. At this mark, finally the 2-mm hore is drilled through the plate. The last steD is the design of the solvent feeding..ca~illarv. . First the butrim edgesar&moothed offso that the capillary iictunllv firs inside the recess in the bottom of the Petri dish. Then, the length of the capillary which initially was cut slightly too long is adjusted by grinding its top end. During this adjustment precedure, the Petri dish is filled with water and the shortening of the capillary is continued until it just fails to touch the layer of a TLC plate when it is placed on top of it. An optimal length has been achieved, when slight Dressure establishes contact and this contact remains after the pressure has been stopped. Finally, a small opening is filed into the side of the capillary 6 mm from the bottom. ..
Classes of compounds that are routinely sepnrnted in our la11I)? circular'I'I.C include: Plant ~henolicssuch as anthraquinones and flavonoids on Silica gel with organic solvents, Betalains on Polyamide with aqueous buffers, and Monosaccharides on Silicagel with polar organic solvents. In all cases, superior separations were achieved in comparison with ascending chromatography under corresponding conditions on a time-for-time basis or on a distance-for-distance basis. Experiments have heen carried out with various other adsorbents, including Silica gel G 60 (E. Merck), Cellulose (E. Merck), and a Polyamide which had been especially prepared for TLC by partial hydrolysis and precipitation (3).In some separation experiments 2-mm preparative plates (20 cm X 20 cm) were used to separate compounds which migrated very slowly. In this case, an overnight run, using continuous development gave excellent results. For this technique of continuous development radial chromatography is especially well suited. Literature Cited Ziatkir, A,, and Kaiser, R. E., Edr.: Eiaevier: New York. 1977:Vol9, p 51. (2) Kaiser. R. E. In '"HPTLC-High Performance Thin Layer Chromsfography": Zlaws, A.. and Kaiser, R.E., Edn.. Elsevier: NcuYork, 1977; Vol9, p 73. (31 Robler. H.: Heinrich, W.:Msbry,T. J. J. Chrornarogr. i973,78,432. (1) Blome, J. In "HPTLC-High Performance Thin Layer Chromatography";
