Spiral Disk Assembly For High-speed Countercurrent Chrom

开发用于分离生物聚合物的J型高速逆流色谱（HSCCC）的螺旋盘在蛋白质样品中显示出低分辨率，这可能是由于粘性聚合物相系统的层流效应。螺旋盘相对于传统的多层盘管分离柱的优点在于，通道几何形状容易以多种方式修改。它还允许各种插入到通道中以改善其性能。 我们已经发现，引入玻璃珠或精细的聚四氟乙烯线到通道中产生的固定相的强烈携带，导致分配效率的有害损失。然而，将短段的特氟隆管定期插入通道产生了积极的结果。据推测，这种插入物作为一个流断路器中断的层流的移动的相，特别是粘性聚合物相系统。通过将3 mm长和2 mm ID的特氟隆管的短段插入单螺旋盘和四螺旋盘的螺旋通道中来进行一系列实验。 结果表明，在单螺旋盘中，当插入数达到300个时，分离效率得到提高，与500个插入物的分离效果相似。具有600个插入物的四螺旋盘产生了对二肽和蛋白质样品的最佳分离，主要是由于其较大的螺旋节距而改善了固定相的保留。 基于这些发现，我们设计了“珠链螺旋盘”（其外观类似珠链），每个由约4600个凹坑（直径2.7 mm，深2 mm）与短管道（0.5 mm长，2 mm深，0.8 - 1.2 mm宽）连接组成，并通过J型HSCCC离心机用一组测试样品检查其性能。正如预期的那样，这种新的柱设计在固定相保留和分配效率方面大大改善了二肽和蛋白质样品的分离。其中，具有四个螺旋通道和更宽的连接管道（1.2mm）的盘产生最好的结果。 我们的研究证明了安装在J型CPC上时螺旋盘的以下独特功能： 1.螺旋盘可以保留足够量的聚合物相系统用于分离生物聚合物。 2.具有增强节距的螺旋盘允许移动的相的高流速以缩短分离时间。 3.螺旋盘的性能可以通过修改通道的几何形状（诸如螺旋节距和通道的配置）来改善。 珠链螺旋盘的多重组装的制造目前正在NIH机械车间进行。

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.