# US 6,458,273 B1 · Sample separation apparatus and method for multiple channel high throughput purification > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 6,458,273 B1 - Document ID: US6458273B1 - Status: Granted - Kind: B1 - Application number: 09/569,378 - Filed: 2000-05-11 - Granted: 2002-10-01 - Earliest claimed: 1999-10-29 - Assignees: Ontogen Corporation - Inventors: Jonathan D. Krakover; Romaine Maiefski - Web page: https://patentgrep.com/patent/US6458273B1 ## CPC - B01D 15/08: Selective adsorption, e.g. chromatography - B01D 15/1871: {placed in series} - B01D 15/14: relating to the introduction of the feed to the apparatus - B01D 15/1885: {placed in parallel} - B01D 15/40: using supercritical fluid as mobile phase or eluent - B01J 2219/00759: Purification of compounds synthesised - G01N 30/02: Column chromatography - G01N 30/24: Automatic injection systems - G01N 30/466: {with separation columns in parallel} - G01N 30/6004: {end pieces} - G01N 30/6052: {body} - G01N 30/82: Automatic means therefor - B01D: SEPARATION (separating solids from solids by wet methods B03B, B03D; by pneumatic jigs or tables B03B; by other dry methods B07; magnetic or electrostatic separation of solid materials from solid materials or fluids, separation by high-voltage electric fields B03C; centrifuges, vortex apparatus B04; presses per se for squeezing-out liquid from liquid-containing material B30B9/02; treatment of water C02F, e.g. softening by ion-exchange C02F1/42; {arrangements of air intake cleaners in gas turbine plants F02C7/05} ; arrangements or mounting of filters in air-conditioning, air-humidification or ventilation F24F13/28) - B01J: CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) ## Abstract A high throughput liquid chromotagraphy column assembly for separating a large sample with a selected mass weight and fluid volume. The assembly includes a loading column with a loading chamber with a first inner diameter and a first length. The loading chamber contains a packing material on to which sample is loaded and spatially distributed. The length of the loading chamber's volume is sufficient to fully load the sample therein but the length of the loading chamber is insufficient to achieve the selected chromotagraphic separation. A separation column is in fluid communication with the loading column. The separation column has a separation chamber with a diameter smaller than the loading column's inner diameter and a length greater than the loading column's length. In one embodiment, the loading column has a diameter two or more times greater than the separation chamber's diameter, and the loading column has a length one-half or less than the length of the separation chamber. The separation chamber's diameter is such that the separation chamber has a volume over the same length as the loading column's length that is insufficient to act as a loading area for the entire selected sample for the given mass and volume of the sample. ## Claims ### Claim 1 A high through liquid chromatography column assembly configured to receive first and second packing materials and a selected sample for flow therethrough to achieve a selected chromatographic separation of the sample, the sample having a mass weight and a fluid volume, comprising: a loading column having a first packing material and a loading chamber having a first inner diameter and first length, the loading chamber being sized to contain a volume of a first packing material with a vertical absorptive profile, the loading chamber being sized to retain a selected volume of the first packing material to spatially distribute the sample within the loading chamber to load the sample prior to chromatographic separation of the sample, the length of the loading chamber being insufficient to achieve the selected chromatographic separation of the sample as the sample passes through the loading chamber; and a separation column having a second packing material and a separation chamber therein in fluid connection with the loading chamber and being positioned to receive the sample from the loading column, the separation chamber having a second diameter smaller than the first diameter and having a second length greater that the first length, the separation chamber being sized to retain the second packing material, the second length of the separation chamber being sufficient to achieve the selected chromatographic separation of the sample as the sample passes therethrough, the separation chamber having a volume of the second packing material over a same length as the first length that is insufficient to act as a loading region for the entire selected sample. ### Claim 2 (depends on 1) The chromatography column assembly of claim 1 wherein the first diameter is two or more times greater than the second diameter. ### Claim 3 (depends on 1) The chromatography column assembly of claim 1 wherein the first length is one half or less than the second length. ### Claim 4 (depends on 1) The chromatography column assembly of claim 1 wherein the loading column has an outlet and the separation column has an inlet, the loading column and the separation column are spaced apart from each other and interconnected by a carrier tube, connected to the loading column's outlet and the separation column's inlet. ### Claim 5 (depends on 1) The chromatography column assembly of claim 1, further comprising a dilution column having a dilution chamber in fluid connection with the loading chamber. ### Claim 6 (depends on 5) The chromatography column assembly of claim 5, wherein the dilution chamber has a first inlet and outlet, the loading column has a second inlet and outlet and the separation column has a third inlet, the dilution column, the loading column, and the separation column are spaced apart from each other, the dilution column being in fluid communication with the loading chamber through a first carrier tube connected to the distribution column's first outlet and the loading column's second inlet, the loading column being in fluid communication with the separation column by a second carrier tube connected to the loading column's second outlet and the separation column's third inlet. ### Claim 7 (depends on 5) The chromatography column assembly of claim 5 wherein the dilution column, the loading column, and the separation column are spaced apart from each other, the dilution column being fluidly connected to the loading chamber by a first carrier tube, and the loading column being fluidly connected to the separation column by a second carrier tube. ### Claim 8 (depends on 7) The chromatography column assembly of claim 7 wherein the first and second carrier tubes are small bore high pressure liquid chromatography tubes. ### Claim 9 (depends on 5) The chromatography column assembly of claim 5 wherein the dilution chamber contains inert packing material therein. ### Claim 10 (depends on 1) The chromatography column assembly of claim 1 wherein the loading column and the separation column are spaced apart from each other and are in fluid communication through a carrier tube extending therebetween. ### Claim 11 (depends on 1) The chromatography column assembly of claim 1 wherein the loading column and the separation column are integrally connected to each other. ### Claim 12 (depends on 1) The chromatography column assembly of claim 1, Teflon coating portions of the first and second columns that come in contact with fluid. ### Claim 13 (depends on 1) The chromatography column assembly of claim 1, further comprising a dilution column having a dilution chamber in fluid communication with the loading chamber, a portion of the dilution column being positioned in the loading chamber, the dilution chamber and the loading chamber being separated by a frit. ### Claim 14 (depends on 13) The chromatography column assembly of claim 13 wherein the frit is positionable in the loading chamber and sandwiched between the dilution column and the first packing material in the loading chamber. ### Claim 15 (depends on 1) The chromatography column assembly of claim 1, further comprising a dilution column having a dilution chamber in fluid connection with the loading chamber, a bottom end of the dilution column being securely connected to a top end of the loading column. ### Claim 16 (depends on 15) The chromatography column assembly of claim 15 wherein a frit is sandwiched between the dilution column and the loading column. ### Claim 17 (depends on 1) The chromatography column assembly of claim 1 wherein the separation chamber has a substantially constant cross sectional area along the second length. ### Claim 18 (depends on 1) The chromatography column assembly of claim 1 wherein the separation chamber has first and second end portions, the first and portion being closest to the loading column, the second diameter being at the first end portion, and the separation chamber having a third diameter at the second end portion smaller than the second diameter. ### Claim 19 (depends on 1) The chromatography column assembly of claim 1 wherein the separation chamber has a truncated conical shape. ### Claim 20 A system for liquid chromatography, comprising a carrier tube with inlet and outlet portions, and two columns; the first column being coupled to the inlet portion of the carrier tube and being about ½ or less than the length of the second column and having internal diameter approximately two or more times the internal diameter of the second column, and the second column connected to the outlet portion of the carrier tube. ### Claim 21 (depends on 20) The system of claim 20, further including a dilution chamber adjacent to and prior of the first column, said dilution chamber having a selected geometric interior volume. ### Claim 22 (depends on 21) The system of claim 21 wherein the first and second columns are integrally connected to each other. ### Claim 23 (depends on 20) The system of claim 20 wherein the first and second columns are integrally connected to each other. ### Claim 24 (depends on 20) The system of claim 20, further including Teflon coating portions of the first and second columns that come in contact with fluid. ### Claim 25 A chromatographic column, comprising first and second column portions, the first column portion having a loading chamber with a first inner diameter and first length, the second column portion having a separating chamber with a second inner diameter and second length, the first inner diameter being approximately two times greater than the second inner diameter and the first length being approximately ½ or less than the second length. ### Claim 26 (depends on 25) The column of claim 25 wherein the first inner diameter is at least approximately two times greater than the second inner diameter. ### Claim 27 (depends on 26) The column of claim 26 wherein the first length is approximately equal to or less than one-half of the second length. ### Claim 28 (depends on 25) The column of claim 25 wherein the first length is approximately equal to or less than one-half of the second length. ### Claim 29 (depends on 25) The column of claim 25 wherein the first and second column portions contains a solid phase material. ### Claim 30 (depends on 25) The column of claim 25 further comprising a dilution chamber adjacent to the first chamber with the first chamber being between the dilution chamber and the second chamber. ### Claim 31 (depends on 25) The column of claim 25 wherein the first column portion has a funnel-shaped transition portion connected to the second column portion. ### Claim 32 (depends on 25) The column of claim 25 wherein the first column portion is releasably connected to the second column portion. ### Claim 33 (depends on 25) The column of claim 25 wherein the first column portion is integrally connected to the second column portion. ### Claim 34 (depends on 25) The column of claim 25 wherein the second chamber portion is a tapered chamber that tapers inwardly as the second column portion extends away from the first column portion.