ROCKVILLE, MD, United States
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Chen C.,Cc Biotech, Llc | Folk W.R.,University of Missouri | Lazo-Portugal R.,Cc Biotech, Llc | Finn T.M.,Cc Biotech, Llc | Knight M.,Cc Biotech, Llc
Journal of Chromatography A | Year: 2016

Spiral countercurrent-chromatography has great potential for improving the capacity and efficiency of purification of secondary metabolites, and here we describe applications useful for the isolation of flavonoids from the widely used South African medicinal plant, Sutherlandia frutescens (L.) R. Br. In the spiral tubing support rotor, STS-4 for high-speed counter-current chromatography, several polar butanol aqueous solvent systems were selected using a log. K plot, and the novel flavonol glycosides (sutherlandins A-D) were well separated by the optimized solvent system (ethyl acetate:n-butanol:acetic acid:water; 5:1:0.3:6 by vol.). The yield of purified flavonoids from 0.9. g extract varied from 8.6. mg to 54. mg of the sutherlandins for a total of 85.3. mg. The same extract was fractionated in the new STS-12 rotor of the same outside dimensions but with more radial channels forming 12 loops of the tubing instead of 4. The rotor holds more layers and increased length of tubing. From 0.9. g extract the STS-12 rotor yielded more recovery of 110.4. mg total with amounts varying from 11.2. mg to 64. mg of the sutherlandins and apparent increased separation efficiency as noted by less volume of each fraction peak. Thus from 1-g amounts of extract, good recovery of the flavonoids was achieved in the butanol aqueous solvent system. © 2017 Elsevier B.V.


Knight M.,Cc Biotech, Llc | Finn T.M.,Cc Biotech, Llc | Zehmer J.,APC Biotechnology Services Inc. | Clayton A.,APC Biotechnology Services Inc. | Pilon A.,APC Biotechnology Services Inc.
Journal of Chromatography A | Year: 2011

An important advance in countercurrent chromatography (CCC) carried out in open flow-tubing coils, rotated in planetary centrifuges, is the new design to spread out the tubing in spirals. More spacing between the tubing was found to significantly increase the stationary phase retention, such that now all types of two-phase solvent systems can be used for liquid-liquid partition chromatography in the J-type planetary centrifuges. A spiral tubing support (STS) frame with circular channels was constructed by laser sintering technology into which FEP tubing was placed in 4 spiral loops per layer from the bottom to the top and a cover affixed allowing the tubing to connect to flow-tubing of the planetary centrifuge. The rotor was mounted and run in a P.C. Inc. type instrument. Examples of compounds of molecular weights ranging from <300 to approximately 15,000 were chromatographed in appropriate two-phase solvent systems to assess the capability for separation and purification. A mixture of small molecules including aspirin was completely separated in hexane-ethyl acetate-methanol-water. Synthetic peptides including a very hydrophobic peptide were each purified to a very high purity level in a sec-butanol solvent system. In the STS rotor high stationary phase retention was possible with the aqueous sec-butanol solvent system at a normal flow rate. Finally, the two-phase aqueous polyethylene glycol-potassium phosphate solvent system was applied to separate a protein from a lysate of an Escherichia coli expression system. These experiments demonstrate the versatility of spiral CCC using the STS rotor. © 2011 Elsevier B.V.


Ito Y.,U.S. National Institutes of Health | Knight M.,Cc Biotech, Llc | Finn T.M.,Cc Biotech, Llc
Journal of Chromatographic Science | Year: 2013

For many years, high-speed countercurrent chromatography conducted in open tubing coils has been widely used for the separation of natural and synthetic compounds. In this method, the retention of the stationary phase is solely provided by the Archimedean screw effect by rotating the coiled column in the centrifugal force field. However, the system fails to retain enough of the stationary phase for polar solvent systems such as the aqueous-aqueous polymer phase systems. To address this problem, the geometry of the coiled channel was modified to a spiral configuration so that the system could utilize the radially acting centrifugal force. This successfully improved the retention of the stationary phase. Two different types of spiral columns were fabricated: the spiral disk assembly, made by stacking multiple plastic disks with single or four interwoven spiral channels connected in series, and the spiral tube assembly, made by inserting the tetrafluoroethylene tubing into a spiral frame (spiral tube support). The capabilities of these column assemblies were successfully demonstrated by separations of peptides and proteins with polar two-phase solvent systems whose stationary phases had not been well retained in the earlier multilayer coil separation column for high-speed countercurrent chromatography. © [2013] The Author.


Trademark
Cc Biotech, Llc | Date: 2012-11-18

Chromatography apparatus for laboratory use.


Trademark
Cc Biotech, Llc | Date: 2012-11-18

Liquid chromatography apparatus.


PubMed | Fuzbien Technology Institute Inc. and Cc Biotech, Llc
Type: | Journal: Journal of chromatography. A | Year: 2017

Over the last decade man-made carbon nanostructures have shown great promise in electronic applications, but they are produced as very heterogeneous mixtures with different properties so the achievement of a significant commercial application has been elusive. The dimensions of single-wall carbon nanotubes are generally a nanometer wide, up to hundreds of microns long and the carbon nanotubes have anisotropic structures. They are processed to have shorter lengths but they need to be sorted by diameter and chirality. Thus counter-current chromatography methods developed for large molecules are applied to separate these compounds. A modified mixer-settler spiral CCC rotor made with 3 D printed disks was used with a polyethylene glycol-dextran 2-phase solvent system and a surfactant gradient to purify the major species in a commercial preparation. We isolated the semi-conducting single walled carbon nanotube chiral species identified by UV spectral analysis. The further development of spiral counter-current chromatography instrumentation and methods will enable the scalable purification of carbon nanotubes useful for the next generation electronics.


Patent
Cc Biotech, Llc | Date: 2013-12-01

A method for constructing a spiral tube support apparatus used in countercurrent chromatography, improvements to countercurrent chromatography tube support design, and methods of using the improved countercurrent chromatography apparatus are described. The spiral tube support apparatus may be constructed by a shape forming process such as a three dimensional printing process that in turn uses a laser sintering technique, and can be made out of any easily formed material. Shape changes on both the tube support and the top improve the performance of the tube support and ease of manufacturing. The improved tube support and new method of creation permit use in both micro or macro scale preparations and use in small or large molecule preparations. In particular, specific solvent systems are described that permit purification of proteins.


Grant
Agency: Department of Health and Human Services | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 217.39K | Year: 2014

DESCRIPTION (provided by applicant): Recent advances in countercurrent chromatography (CCC) have created novel spiral-design separation columns that hold a very high stationary phase of all the solvent systems, such that now all molecules of any size and water solubility can be successfully separated in high-speed countercurrent chromatography. With spiral CCC there is inherent versatility, more than any type of chromatography. Preliminary studies with the new spiral tubing support rotor succeeded in purifying Sutherlandioside B, a major water soluble cycloartane glycoside from Lessertia frutescent (Sutherlandia) is using an n- butanol solvent system. With the small volume of the coil (~100 ml) and a stationary phase at 50% (~50 ml), it is impressive that amass of 1.2 g of crude plant extract could be efficiently fractionated in one run with high recovery of the target compound. In addition, the active iridoid glycoside, harpagoside was isolated from an extract of Harpagophytum procumbens (Devil's Claw


Grant
Agency: NSF | Branch: Standard Grant | Program: | Phase: | Award Amount: 150.00K | Year: 2016

The broader impact/commercial potential of this project lies in the availability of single-wall carbon nanotubes in their most highly purified form in laboratory scale quantities. This would be enabled by the use of a spiral countercurrent chromatograph, invented and made in the USA. Single-wall carbon nanotube is one of the most promising semi-conductor materials and is being actively considered for making state-of-the-art high performance electronic circuits. The associated know-how will be of great value to the high volume semi-conductor manufacturing industry. The availability of discrete chiral types of the single-wall carbon nanotubes separated by the proposed system will have additional impact on the research market. Research labs in the semi-conductor industry and academic labs may be able to develop the carbon nanotube based nanomaterials, components and final products for point of care diagnostics, genomics devices and sensors for environmental and security identification. The societal benefits of these potential applications are significant. The benefits extend more broadly to the life science research industry also where the technology works for protein and particle separations in the development of novel vaccines and drug delivery.



This Small Business Innovation Research (SBIR) Phase I project focuses on developing a novel laboratory scale system for carbon nanotube purification. Carbon nanotubes are challenging to manufacture and isolate in their pure form for use as robust logic and sensor elements in advanced electronics. They are hard crystal carbon polymers that are heterogeneous tubes of various lengths and diameters and occur as single-walled or multi-walled with other structures. The existing processes to solubilize and purify the various single-walled semi-conductor species are quite difficult and have low mass yields. The methods mostly used to date are column chromatography and density gradient ultracentrifugation. To meet the need of higher mass loading and increased selectivity, this proposal focuses on developing spiral countercurrent chromatography. A new spiral design rotor used with a polymer solvent mix has shown improved nanotube fractionation over static extraction in preliminary experiments. Recent innovative research on dextran polymer liquid stationary phase makes it possible to improve the fractionation of carbon nanotubes even further. It also opens up many exciting options for large molecule separation. This development has a very significant intellectual merit for both chromatography and semi-conductor research.


Grant
Agency: National Science Foundation | Branch: | Program: SBIR | Phase: Phase I | Award Amount: 150.00K | Year: 2016

The broader impact/commercial potential of this project lies in the availability of single-wall carbon nanotubes in their most highly purified form in laboratory scale quantities. This would be enabled by the use of a spiral countercurrent chromatograph, invented and made in the USA. Single-wall carbon nanotube is one of the most promising semi-conductor materials and is being actively considered for making state-of-the-art high performance electronic circuits. The associated know-how will be of great value to the high volume semi-conductor manufacturing industry. The availability of discrete chiral types of the single-wall carbon nanotubes separated by the proposed system will have additional impact on the research market. Research labs in the semi-conductor industry and academic labs may be able to develop the carbon nanotube based nanomaterials, components and final products for point of care diagnostics, genomics devices and sensors for environmental and security identification. The societal benefits of these potential applications are significant. The benefits extend more broadly to the life science research industry also where the technology works for protein and particle separations in the development of novel vaccines and drug delivery. This Small Business Innovation Research (SBIR) Phase I project focuses on developing a novel laboratory scale system for carbon nanotube purification. Carbon nanotubes are challenging to manufacture and isolate in their pure form for use as robust logic and sensor elements in advanced electronics. They are hard crystal carbon polymers that are heterogeneous tubes of various lengths and diameters and occur as single-walled or multi-walled with other structures. The existing processes to solubilize and purify the various single-walled semi-conductor species are quite difficult and have low mass yields. The methods mostly used to date are column chromatography and density gradient ultracentrifugation. To meet the need of higher mass loading and increased selectivity, this proposal focuses on developing spiral countercurrent chromatography. A new spiral design rotor used with a polymer solvent mix has shown improved nanotube fractionation over static extraction in preliminary experiments. Recent innovative research on dextran polymer liquid stationary phase makes it possible to improve the fractionation of carbon nanotubes even further. It also opens up many exciting options for large molecule separation. This development has a very significant intellectual merit for both chromatography and semi-conductor research.

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