The spiral disks developed for separating biopolymers by type-J high-speed countercurrent chromatography (HSCCC) showed low resolution in protein samples probably due to a laminar flow effect of the viscous polymer phase system. The spiral disk has an advantage over the conventional multilayer coil separation column in that the channel geometry is easily modified in many ways. It also permits various inserts into the channel to improve its performance. We have found that introducing glass beads or fine Teflon threads into the channel produced intensive carryover of the stationary phase resulting in detrimental loss of the partition efficiency. However, inserting short segments of Teflon tubing into the channel at regular intervals produced positive results. It is assumed that such inserts served as a stream breaker to interrupt the laminar flow of the mobile phase especially for viscous polymer phase systems. A series of experiments was carried out by inserting short segments of Teflon tubing, 3 mm long and 2 mm ID, into the spiral channel of both single and four-spiral disks. The results indicated that in the single spiral disk the partition efficiency was improved by the number of inserts up to 300 which show similar separation with 500 inserts. A four spiral disk with 600 inserts produced the best separation for both dipeptide and protein samples, mainly by improved retention of the stationary phase due to its greater pitch of the sprial. Based on these findings, we have designed """"""""bead-chain spiral disk"""""""" (its appearance resembles a bead chain), each consisting of ca 4600 pits (2.7 mm diameter and 2 mm deep) connected with short ducts (0.5 mm long, 2 mm deep and 0.8 to 1.2 mm wide), and examined their performance with a set of test samples by a type-J HSCCC centrifuge. As expected, this new column design substantially improved the separation of both dipeptide and protein samples in terms of stationary phase retention and partition efficiency. Among those, the disk with four spiral channel and wider connection ducts (1.2 mm) produced the best results. Our studies demonstrated the following unique features of the spiral disk when mounted on the type-J CPC: 1. The spiral disk can retain a sufficient amount of polymer phase systems for the separation of biopolymers. 2. The spiral disk with an enhanced pitch permits high flow rates of the mobile phase to shorten the separation times. 3. The performance of the spiral disk can be improved by modifying the channel geometry such as spiral pitch and configuration of the channel. Fabrication of a multiple assembly of the bead-chain sprial disk is currently underway in the NIH Machine shop.
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