US3346486A - Apparatus and method for introduction of amino acid samples into a chromatography column - Google Patents

Apparatus and method for introduction of amino acid samples into a chromatography column Download PDF

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Publication number
US3346486A
US3346486A US358198A US35819864A US3346486A US 3346486 A US3346486 A US 3346486A US 358198 A US358198 A US 358198A US 35819864 A US35819864 A US 35819864A US 3346486 A US3346486 A US 3346486A
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United States
Prior art keywords
column
sample
liquid
buffer
resin
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Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Lifetime
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US358198A
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English (en)
Inventor
George D Winter
George N Catravas
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Technicon Corp
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Technicon Corp
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Filing date
Publication date
Application filed by Technicon Corp filed Critical Technicon Corp
Priority to US358198A priority Critical patent/US3346486A/en
Priority to GB13776/65A priority patent/GB1087301A/en
Priority to BE662089D priority patent/BE662089A/xx
Priority to SE04389/65A priority patent/SE335628B/xx
Priority to CH479865A priority patent/CH472903A/de
Priority to DE1673101A priority patent/DE1673101C3/de
Application granted granted Critical
Publication of US3346486A publication Critical patent/US3346486A/en
Anticipated expiration legal-status Critical
Expired - Lifetime legal-status Critical Current

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
    • G01N30/02—Column chromatography
    • G01N30/04—Preparation or injection of sample to be analysed
    • G01N30/16—Injection
    • G01N30/18—Injection using a septum or microsyringe
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S210/00—Liquid purification or separation
    • Y10S210/918—Miscellaneous specific techniques

Definitions

  • the primary object of the present invention is to obviate the need for restoring to the abovementioned manipulations and facilitate the introduction of the amino acid samples into the chromatography column, and to avoid disturbing the resin in the column.
  • An important ancillary object is to provide a fitting which may be readily secured to and removed from the top of the column and which is well adapted to enable the introduction of the samples into the column with facility and without disturbing the resin in the column.
  • amino acid samples for example, protein and/or peptide hydrolysates
  • the amino acid sample is dissolved in a quantity of a water solution of sucrose, and an aliquot of the the resulting solution is layered on the upper surface of the resin in the column, after which the buffer when pumped into the column by a positive displacement pump in the usual way forces the sample into the resin.
  • FiG. 1 is a vertical sectional view of the upper part of a chromatography column having the fitting of the present invention mounted thereon;
  • FIG. 2 is a fragmentary sectional view showing part of the fitting in the condition thereof after the introduction of the sample.
  • the fitting housing 12 is preferably a molded plastic member, the preferred plastic being known ,as Kel-F. Said housing has a lower part 14 which fits snugly into the top of the column 10 and an O-ring 16 provides a fluid-tight seal between the inner surface of column 10 and the peripheral surface of part 14. The housing 12 is releasably held in position at the top of the column by a pair of clamps 18.
  • a vertically extending passage 20 in housing 12 communicates with the upper part of column 10 and with the transverse passages 22 and 24 provided in said housing.
  • a fitting 26 has a part 28 which is threaded into the opening 30 of housing 12 in fluid-tight relation in said opening. Fitting 26 provides a passage for the buffer supplied by a positive displacement pump (not shown) through a tube 32 connected to said fitting and to the outlet of said pump.
  • a Luer-Lok thimble or hollow plug 34 is screwed into an opening at the top of the housing in fluid-tight engagement with the housing as indicated at 36. Said thimble communicates with the upper end of ,a hypodermic needle or capillary tubing 38 which has a fluidtight connection with the lower end of said thimble.
  • Said tubing 33 extends from the lower end of thimble 34 through passage 2% into the upper part of column 10 below the housing part 14.
  • a length of polyethylene tubing 49 is fitted on tubing 38 and extends downwardly into the upper part of column 10 to a point adjacent the upper surface 43 of the resin with which the column is packed.
  • the polyethylene tubing can be cut to any required length and thereby provides means for adjusting the outlet of the hypodermic needle tubing in relation to the upper surface of the column resin.
  • a Luer-Lok plug 42 is screw-threaded into a housing opening 44 and communicates with passage 24 to provide an overflow opening when the Luer-Lok cap is removed. It will be understood that during the operation of the column for the stripping thereof, at which time the buffer liquid is being introduced into the column by way of fitting 26, the Luer-Lok plugs 34 and 42 are each tightly closed by a Luer-Lok cap, one of which is indicated at 46 in FIG. 3.
  • FIG. 1 shows the condition of the apparatus at the time of the introduction of the sample into the column, as hereinafter more particularly described. At this time, the caps 46 are removed from the plugs 34 and 42 and the positive displacement pump is shut down.
  • the sucrose-sample solution is introduced into the column by means of a syringe of a well known type which has a lower cylindrical end portion which is inserted into the thimble 34 in fluid-tight relation therewith, when the LuerLok cap 46 is removed.
  • a syringe of a well known type which has a lower cylindrical end portion which is inserted into the thimble 34 in fluid-tight relation therewith, when the LuerLok cap 46 is removed.
  • the sucrose-sample solution has a higher density than the buffer, the sucrosesample flows downwardly through the buffer and is thus layered onto the top surface 43 of the resin in the chromatography column.
  • the buffer which is in the column above the resin at the time of the interrupting of the operation of the positive displacement pump and which is displaced from the column by the introduction of the sucrose-sample solution, flows out of the fitting 3 through the passage 20 around the capillary tubing 40, 38, through the transverse passage 24 and out through the Luer-Lok thimble 42.
  • the syringe is removed from the Luer-Lok thimble 34. Then said thimbles are closed in fluid-tight condition by the attachment of the Luer-Lok caps 46, and the pump for introducing the butfer into the column is started, thereby resulting in the sample-sucrose solution being forced into the resin by the butter.
  • sucrosesample solution which has been found to be highly satisfactory in the practice of the present invention.
  • sucrose Fifty grams of sucrose are dissolved in sufficient water to provide 100 ml. of the sucrose solution.
  • the sucrosesample solution is prepared by adding three parts of the sample to one part of the sucrose solution. Ordinarily, 0.20 ml. of the sample-sucrose solution is introduced into the column in the manner described above. This quantity of the sucrose-sample solution is thus composed of 0.15 ml. of sample and 0.05 ml. of the sucrose solution.
  • a chromatography apparatus comprising a chromatography column containing an ion exchange resin, and means for the introduction of buffer and samples into the column, each sample being contained in a liquid which has a higher density than the butter, said introduction means including a member releasably secured to the top of the column and having a plurality of passages each independently in flow communication with the column, one of said passage having an inlet part accessible externally of said member for the introduction of the sample into the column when the flow of butter into the column is interrupted, another of said passage having an inlet for providing for the flow of buffer into the column during the eluting operation, said member having an outlet opening in communication with said buffer passage for outflow of such buflers as is displaced by the sample introduced through said one passage, and means for releasably closing said outlet and said inlet to said one passage.
  • a chromatography apparatus comprising a chromatography column containing an ion exchange resin, and means for the introduction of buffer and samples into the column, each sample being contained in a liquid which has a higher density than the butter, said introduction means including a member releasably secured to the top of the column and having a plurality of passages each independently in flow communication with the column, one of said passage comprising a capillary tube having an inlet part accessible externally of said member for the introduction of the sample into the column when the flow of buffer into the column is interrupted, said capillary tube being vertically disposed and having at its lower end a removable length of tubing to provide an outlet for the sample into the column at a predetermined level above the resin in the column, another of said passages having an inlet for providing for the flow of buffer into the column during the eluting operation, said member having an outlet opening in communication with said bufler passage for outflow of such bufler as is displaced by the sample introduced through said one passage, said lower end of said tubing being disposed below said
  • a chromatography apparatus comprising a chromatography column containing an ion exchange resin, and means for the introduction of buffer and samples into the column, each sample'being contained in a liquid which has a higher density than the buffer
  • said introduction means including a member releasably secured to the top of the column and having a plurality of passages each independently in flow communication with the column, one of said passages having an inlet part accessible extrenally of said member for the introduction of the sample into the column when the flow of butter into the column is interrupted, said inlet part having an inner wall with a fitting which is releasably engageable in fluid-tight relation by the discharge end of a syringe for the transmission of the sample through said one passage from said inlet into the column, another of said passages having an inlet for providing for the flow of buffer into the column during the eluting operation, said member having an outlet opening in communication with said buffer passage for outflow of such butter as is displaced by the sample introduced through said one passage, and means for relea
  • a chromatography apparatus comprising a chromatography column containing an ion exchange resin, and means for the introduction of buffer and samples into the column, each sample being contained in a liquid which has a higher density than the butter, said introducing means including a member releasably secured to the top of the column and having a plurality of passages each independently in flow communication with the column, one of said passages comprising a capillary tube having an inlet part accessible externally of said member for the introduction of the sample into the column when the flow of bufler into the column is interrupted, said capillary tube being vertically disposed and having at its lower end a removable length of tubing to provide an outlet for the sample into the column at a predetermined level above the resin in the column, said inlet part having an inner wall with a fitting which is releasably engageable in fluid-tight relation by the discharge end of a syringe for the transmission of the sample through said one passage from said inlet into the column, another of said passages having an inlet for providing for
  • a method for the chromatography analysis of an amino acid sample by apparatus having a chromatography column containing an ion exchange resin through which a butter liquid is pumped for forming the chromatogram and for the stripping operation comprising: incorporating the sample in a liquid having a higher density than said buffer liquid, flowing the buffer liquid into the column, interrupting the flow of said butter into the column, and introducing a quantity of said sample-containing liquid into the column above the resin whereby said sample-containing liquid flows downwardly through the butter liquid and is thereby layered on the top surface of the resin displacing an equal volume of the buffer liquid.
  • a method for the chromatography analysis of an amino acid sample by apparatus having a chromatography column containing an ion exchange resin through which a buffer liquid is pumped for forming the chromatogram and for the stripping operation comprising: incorporating the sample in a water solution of sucrose and thereby providing a liquid having a higher density than said buffer liquid, flowing the buffer liquid into the column, interrupting the flow of said buffer into the column, and introducing a quantity of said sample-containing liquid into the column above the resin whereby said sample-containing liquid flows downwardly through the butter liquid and is thereby layered on the top surface of the resin displacing an equal volume of the buffer liquid.
  • An inlet apparatus for a chromatography column containing a porous material, for the introduction of an eluting liquid and a sample liquid having a higher density than the eluting liquid, said apparatus comprising: means for the fluid-tight attachment of said apparatus to the top of the column; a first substantially vertical passageway having a lower end adapted to be disposed within the column above the level of the porous material contained therein, and an upper end with a fitting adapted to be releasably connected to a source of sample liquid; a second passageway independent of said first passageway having an inlet end adapted to be connected to a source of eluting liquid and disposed between said upper and lower ends of said first passageway, having an intermediate port adapted to be disposed in fluid flow communication with the interior of the column above the level of the lower end of said first passageway, and having an outlet end disposed between said upper and lower ends of said first passageway and adapted to discharge such eluting liquid as is displaced from the column by the sample liquid; and means for

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  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Treatment Of Liquids With Adsorbents In General (AREA)
  • Sampling And Sample Adjustment (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
US358198A 1964-04-08 1964-04-08 Apparatus and method for introduction of amino acid samples into a chromatography column Expired - Lifetime US3346486A (en)

Priority Applications (6)

Application Number Priority Date Filing Date Title
US358198A US3346486A (en) 1964-04-08 1964-04-08 Apparatus and method for introduction of amino acid samples into a chromatography column
GB13776/65A GB1087301A (en) 1964-04-08 1965-04-01 Apparatus and method for introduction of amino acid samples into a chromatography column
BE662089D BE662089A (de) 1964-04-08 1965-04-05
SE04389/65A SE335628B (de) 1964-04-08 1965-04-05
CH479865A CH472903A (de) 1964-04-08 1965-04-06 Verfahren und Vorrichtung zur chromatographischen Analyse einer Aminosäureprobe
DE1673101A DE1673101C3 (de) 1964-04-08 1965-04-06 Verfahren und Vorrichtung zum Auf tragen von Proben bestimmter Aminosäuren auf eine chromatografische Säule

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US358198A US3346486A (en) 1964-04-08 1964-04-08 Apparatus and method for introduction of amino acid samples into a chromatography column

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US3346486A true US3346486A (en) 1967-10-10

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US (1) US3346486A (de)
BE (1) BE662089A (de)
CH (1) CH472903A (de)
DE (1) DE1673101C3 (de)
GB (1) GB1087301A (de)
SE (1) SE335628B (de)

Cited By (15)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3474908A (en) * 1965-12-23 1969-10-28 Technicon Corp Chromatography apparatus
US3826373A (en) * 1973-04-03 1974-07-30 Us Army Liquid chromatography apparatus
US3878099A (en) * 1974-04-08 1975-04-15 Beckman Instruments Inc Mounting for liquid chromatograph column
US3961534A (en) * 1975-09-02 1976-06-08 Richard Gundelfinger Two position rotary valve for injecting sample liquids into an analysis system
US4068528A (en) * 1976-01-19 1978-01-17 Rheodyne Incorporated Two position rotary valve for injecting sample liquids into an analysis system
USD254445S (en) 1977-08-25 1980-03-11 Warner-Lambert Company Chromatography column
US4457846A (en) * 1982-07-08 1984-07-03 Milton Roy Company Liquid chromatography methods and devices
US4752391A (en) * 1986-08-21 1988-06-21 Ceskoslovenska Akademie Ved Column for preparative liquid chromatography
US4755558A (en) * 1986-05-30 1988-07-05 Beckman Instruments, Inc. Using internal marker
US4855486A (en) * 1986-05-30 1989-08-08 Kalbag Suresh M Blocked, marked amino acids
US5607581A (en) * 1995-10-31 1997-03-04 Systec, Inc. Debubbling and priming system for chromotography
US5861125A (en) * 1986-08-15 1999-01-19 City Of Hope Protein or peptide sequencing method and apparatus
US6090190A (en) * 1997-12-12 2000-07-18 Hoechst Marion Roussel Deutschland Gmbh Closure systems for chromatographic columns
US20050104373A1 (en) * 2003-11-14 2005-05-19 Zarubaiko Ted M. Draining and/or stagnant-pocket-minimizing instrument tee
US10107407B2 (en) 2010-09-28 2018-10-23 Parker-Hannifin Corporation Modular valve manifold system

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP4291569B2 (ja) * 2000-11-21 2009-07-08 ウォーターズ・インヴェストメンツ・リミテッド 強化型クロマトグラフィ用移動相希釈スキーム

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US2769454A (en) * 1954-01-13 1956-11-06 Modern Faucet Mfg Co Pressure control fittings

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US2769454A (en) * 1954-01-13 1956-11-06 Modern Faucet Mfg Co Pressure control fittings

Cited By (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3474908A (en) * 1965-12-23 1969-10-28 Technicon Corp Chromatography apparatus
US3826373A (en) * 1973-04-03 1974-07-30 Us Army Liquid chromatography apparatus
US3878099A (en) * 1974-04-08 1975-04-15 Beckman Instruments Inc Mounting for liquid chromatograph column
US3961534A (en) * 1975-09-02 1976-06-08 Richard Gundelfinger Two position rotary valve for injecting sample liquids into an analysis system
US4068528A (en) * 1976-01-19 1978-01-17 Rheodyne Incorporated Two position rotary valve for injecting sample liquids into an analysis system
USD254445S (en) 1977-08-25 1980-03-11 Warner-Lambert Company Chromatography column
US4457846A (en) * 1982-07-08 1984-07-03 Milton Roy Company Liquid chromatography methods and devices
US4755558A (en) * 1986-05-30 1988-07-05 Beckman Instruments, Inc. Using internal marker
US4855486A (en) * 1986-05-30 1989-08-08 Kalbag Suresh M Blocked, marked amino acids
US5861125A (en) * 1986-08-15 1999-01-19 City Of Hope Protein or peptide sequencing method and apparatus
US4752391A (en) * 1986-08-21 1988-06-21 Ceskoslovenska Akademie Ved Column for preparative liquid chromatography
US5607581A (en) * 1995-10-31 1997-03-04 Systec, Inc. Debubbling and priming system for chromotography
US6090190A (en) * 1997-12-12 2000-07-18 Hoechst Marion Roussel Deutschland Gmbh Closure systems for chromatographic columns
US20050104373A1 (en) * 2003-11-14 2005-05-19 Zarubaiko Ted M. Draining and/or stagnant-pocket-minimizing instrument tee
US6991263B2 (en) 2003-11-14 2006-01-31 Itt Manufacturing Enterprises, Inc. Enhanced-draining and/or stagnant-pocket-minimizing instrument tee
US10107407B2 (en) 2010-09-28 2018-10-23 Parker-Hannifin Corporation Modular valve manifold system

Also Published As

Publication number Publication date
BE662089A (de) 1965-10-05
CH472903A (de) 1969-05-31
GB1087301A (en) 1967-10-18
DE1673101C3 (de) 1973-11-29
DE1673101A1 (de) 1970-01-02
DE1673101B2 (de) 1973-05-10
SE335628B (de) 1971-06-01

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