US3881892A

US3881892A - Chromatograph system and method

Info

Publication number: US3881892A
Application number: US248384A
Authority: US
Inventors: Charles W Gehrke; Robert Zumwalt; Kenneth Kuo; David L Stalling
Original assignee: Individual
Current assignee: Individual
Priority date: 1972-04-28
Filing date: 1972-04-28
Publication date: 1975-05-06
Anticipated expiration: 1992-05-06

Abstract

A chromatograph system for a gas-liquid chromatographic analyses of complex samples of matter having a flow regulated carrier gas supply and sample injection means for vaporization and translation through a pre-column and a main analytical column to a detection unit, solvent venting means connected between a precolumn and analytical column for venting a substantial portion of the solvent from the chromatograph during conversion of the sample into detectable form and prior to translation thereof through the analytical column for detection, and means for bypassing a portion of the carrier gas supply into the analytical column downstream of the solvent venting means to facilitate the solvent venting operation. The invention also embodies the method of carrying out analysis of complex samples of matter in gas chromatograph-mass spectrometer combinations utilizing the step of solvent venting subsequent to the sample injection.

Description

United States Patent [1 1 Gehrke et al.

[ May 6,1975

[ CHROM ATOGRAPH SYSTEM AND METHOD [22] Filed: Apr. 28, I972 [21] Appl. No.: 248,384

Primary Examiner.lohn Adee Attorney, Agent, or Firm-Richard G. Heywood [57] ABSTRACT A chromatograph system for a gas-liquid chromatographic analyses of complex samples of matter having a flow regulated carrier gas supply and sample injection means for vaporization and translation through a pre-column and a main analytical column to a detection unit, solvent venting means connected between a pre-column and analytical column for venting a substantial portion of the solvent from the chromatograph during conversion of the sample into detectable form and prior to translation thereof through the analytical column for detection, and means for by-passing a portion of the carrier gas supply into the analytical column downstream of the solvent venting means to facilitate the solvent venting operation. The invention also embodies the method of carrying out analysis of complex samples of matter in gas chromatograph-mass spectrometer combinations utilizing the step of solvent venting subsequent to the sample injection.

9 Claims, 2 Drawing Figures CARRIER Q25 10 SUPPLY FLOW CONTROLLER ROTOMETER] [52] U.S. Cl... 55/67; 55/]97 [51] ..B01d 15/08 [58] Field of Search 55/67, 197

[56] References Cited UNITED STATES PATENTS 3,l 12,028 1/1973 Deans 55/l97 X 3,498,027 3/1970 Buchtel, 55/197 3,550,429 12/1970 MacMuataie et al. 551])? X 5 CLOSED cnnmen ans 0.! Aura oven 4/ /O CLOSED OFF AUTO p srnRr PATEHIEUMM sums 3.881.892

sum 20$ 2 DERIVATIZATION AND GLC ANALYSIS OF ng OF EACH AMINO ACID I ORN LYS TYR

(ILA

U LU GL Q VAL Q ASP D Z.

EU a: an L PHE ADJ HyPRO TRY lLE 0 I SER o J L Lu '7 E I I I I I 100 130 I60 :50 220 "c ESTERFICATION GLC ANALYSIS SUM-l -Butanob 3NHCL Sample Injected: 50 1 100C-30 Min. Solvent Vent Time: SEC.

Injection Port Temo: 180C ACYLATION Proqram Rate: 6/Min.

Final Temn. 230C 42 /41 CH Cl -IFAA (2 r 1 v/v) Attenuation: 32 x 10 AFS 100C-10 Min.

PRE-COLUI N: COLUMN:

1. n w/w% 0V-17 on 8 /1 mesh 0. w/w% EGA on /100 mesh II.P. Chromnscrb G 4" x 4 mm. a.w. Chromosorb W, l. 5 m x 4 mm LD.

Glass.

CHROMATOGRAPH SYSTEM AND METHOD BACKGROUND OF THE INVENTION The present invention relates to apparatus and methods of analyzing volatile compounds and more particularly to a sensitive and accurate system for analysis of separated constituents of mierosized samples.

Analysis of complex samples of matter is carried out by vaporizing a liquid sample in the heated environment of a gas chromatograph. which separates the components of the sample into sequential analytical components and. in a conventional gas chromatograph or like separation apparatus. the gas or vaporized samplc to be analyzed is transported through the chromatograph to the detection unit for analysis by a stream of inert carrier gasv A principal problem in the past has been that microsized sample compounds are diluted in an organic solvent solution and consequently such small volume samples may be difficult to detect. or the large volume of volatile solvent is immediately swept through the analytical or separation column and frequently swamps the detector or obscures the sample peaks. Another major obstacle associated with the gasliquid chromatographic analysis of biological samples. or samples containing very low concentrations of amino acids or other molecules. has been that the sensitivity ofthe detection means has been minimized by the limited sample volume that could be injected into a packed analytical chromatographic column. In many cases. due to problems of maintaining intact the derivatives of the biological compounds at extremely low concentrations. problems are encountered. On removal of the solvent and excess derivatization reagents. many derivatized compounds become susceptible to hydrolysis by atmospheric moisture. Also. on the reduction of the sample volume to a few microliters. the various derivatives of the compounds to be analyzed in the solution may exhibit different solubility characteristics and separate as oils or crystals. thereby making analyses very difficult or impossible.

OBJECTS AND SUMMARY OF THE INVENTION It is an object ofthe present invention to improve the sensitivity and reliability of all kinds ofgas-liquid chromatographic analyses. such as steroids. carbohydrates. amino acids. nucleic acids. peptides. drugs. metabolics. volatile metals. petroleums and the like; and having limited applicability to liquid-liquid and ion-exchange chromatography.

Another object of the invention is the provision of a gas analysis system capable of accurately analyzing small quantities of sample. but permitting the introduction of larger volumes of sample into the chromatography column for better sensitivity of detection.

Another object is to provide a chromatographic sys tem that obviates the need for removal of halogenated solvents from the derivatized sample in electron cap ture and alkali-flame detection analyses Another object is to provide a chromatographic system that eliminates sul\enl and evtraneous peaks from interferring with early eluted compounds A further object of the invention is the provision of a highly scnsilnc chromatgrapliic gas-liquid analysis apparatus capable of flu and temperature program ming and yet providing accurate and reliable quantitatne data.

These and other objects and advantages of the invention will become more apparent hereinafter.

BRIEF DESCRIPTION OF THE DRAWINGS For purposes ofillustration and disclosure. the inven tion is embodied in the parts and the combination of parts hereinafter described and claimed. In the accompanying drawings which form a part of the specifica tion:

FIG. I is a diagrammatic view of a gas chromatographic system embodying the invention. and

FIG. 2 is a chromatogram resulting from a typical analysis of an amino acid sample taken in the chromatographic system of FIG. 1.

DESCRIPTION OF THE PREFERRED EMBODIMENT Referring to FIG. I. the present chromatographic system comprises the usual carrier gas supply I0. which maintains an inert gas such as nitrogen. helium or the like under pressure and discharges that gas to a flow controller I] and a rotameter l2 for regulating and reading the rate of gas flow to the system. The system also includes a main chromatograph device l3 having a chromatograph column I4 including a pre-column l5 and analytical column l6 modified according to the present invention. but which chromatograph I3 has the conventional controls (not shown] for varying the oven temperature between 50 to 300C or other usual ranges. The analytical column I6 is connected to a detector unit 17. such as a flame ionization detector or a mass spectrometer. for analysis of the eluted compounds. The system also includes an auxiliary chromatograph venting apparatus indicated diagrammab ically at 18. which includes a control panel I).

The carrier gas line from the rotameter 12 connects to a T-connection 2 l. and the main gas flow conduit 22 therefrom is connected into the pre-column IS. at 23. adjacent to the sample injection port 24. The injection port 24 is conventional in construction and includes an injector nut 25 sealably connecting a selfsealing rubber disc 26 to the inlet end of the precolumn 15. In practice. the pre-column I5 is lengthened so that it can be adapted to fit a number of different basic chromatograph devices 13. Preferably. the chromatographic column I4 is made of glass (to be inert and responsive to exposed temperatures) and. intcrmediate to the pre-column l5 and analytical column I6, a venting T-connection 30 is provided. This T- connection 30 is packed with a silanized glass wool plug 3! or the like. and the pro-column l5 and analytical column I6 are packed in a usual manner with a typi cal granular inert material 32. such as a high molecular weight polymer. above and below the plug 3]. The packing 32 in the pre-column 15 may be I"? O\'-l7 on (hromosorb O marketed by Ohio Valley Specialties and the packing 32 in the analytical column It: may be 0.65 EGA on (hromosorb W marketed by Johns- Manville. which are selected to retain N-trilluoroacetyl n-butyl esters of amino acids v. hile permitting the mating ofthc solvent and derivati/ation byproducts. as ill be described,

The 'l'-connection 30 between the pre-column I5 and analytical column If) is connected by a tle\ible stainless steel tube conduit 33 or the like to a solenoid valve 34 in the .iu\iliary unit 18. and an outlet conduit 35 from this tube 34 wins to atmosphere. The solenoid valve 34 is operated by a timer mechanism 36 set by a manual control 36' on the control panel 19.

An important feature of the present system is the carrier gas hy-pass arrangement designed to facilitate the venting operation. A conduit 37 is connected to the T- connection 2] in the main carrier gas line 20.22 and is connected through a gas metering needle valve 37' in the auxiliary unit 18 to a conduit 38 connected to another T-connection 39 downstream of the venting connection 30. both intermediate of the pre-column l5 and analytical column 16. The carrier gas hy-pass lines 37.38 are regulated by the needle valve 37' through a control knob 40 on the control panel I9 to bypass approximately 20% of the carrier gas into the chromatographic column 14 to assist in the venting operation. as will he described. The control panel 19 of the auxiliary unit 18 also includes a master on-off" switch 41. a venting switch 42 for conditioning the opening and closing of the timer 36 in conjunction with the timer control 36. and a timer start button 43 (together with

indicator lamps

44 and 45 showing the venting and non-venting conditions of the auxiliary unit 18).

In operation. the flow controller H is set in the normal manner to regulate the total volumetric flow rate of carrier gas from the carrier gas supply it] through the rotameter l2 and conduit 20 to the T-connection 2t. and the gas metering valve 37' in the auxiliary unit 18 is adjusted by the control 40 so that approximately It) milliliters per minute of carrier gas (approsimately 20%) pass through the by-pass lines 37.38 to the T- connection 39. The timer control 36' is set for the desired venting time. such as 30 seconds (0.5 as shown in FIG. I l. and appropriate time adjustments may he made depending upon whether there is solvent remaining or the detector peaks of early eluted compounds are lost or their quantities are diminished. Of course. other parameters in the chromatograph operation that affect solvent and sample separation are column and injection port temperatures. polarity of the column packing and the percent loading which are characteristic of gas-liquid chromatographic principles.

The venting switch 42 is turned to open and the sample solution to be analyzed is then injected into the injection port 24. usually by using a hypodermic sy ringe (not shown) penetrating the disc 26. but any liquid injection devices may be utilized. Upon injection of the sample. the timer button 43 is immediately depressed and released to activate the timer 36 and open the solenoid valve 34. The venting switch 42 is then turned to auto to control the closing of the solenoid valve 34 at the end of the pre-set venting cycle of the timer control 36'.

L'pon injection of the sample into the heated precolumn 15. the sample solution is immediately vapori/ed and separates into the constituent components of the sample by reason of their \olatilities and polarities which proceeds slovvly through the pre-column 15 in a normal manner its a function of the packing and temperaturc. whereas the highly volatile solvent and estraneous volatile substances from the sample solution rapidly traverse the pre column 15. With the venting valve 34 in the open position. these volatile substances and solvent as v\ ell a the main carrier gas tream are di- \erted through the venting conduit following the path of least resistance relative to the packed analytical column and are vented to atmosphere the bypass carrier gas also flows through the conduits 37.38 into fill the column l4 below the venting line and. similarly. is diverted upwardly and vented to atmosphere. The sample compounds of interest proceed slowly through the pre-colurnn t5 and are separated from the solvent and other volatile substances. When the timer control 36 times out and the timer 36 closes the solenoid valve 34 to terminate the venting operation. normal separation occurs as the carrier gas sweeps the constituent compounds of the sample on through the analytical column If: to the detector 17. Thus. the venting of solvent and volatile reaction byproducts of the sample from the column l4 prevents interference with the normal separation and detection of the sample compounds of interest. and the column and detector life are prolonged.

Referring to FIG. 2. from this chromatograph and the accompanying data setting out the conditions under which it was obtained. it will be readily apparent to those skilled in the art of chromatography that the analysis of such a low concentration as a 10 nanogram solution of amino acids indicates the high degree of detection sensitivity provided by the present invention. It will be understood that in a conventional. unvented gas chromatograph. the detector would be completely swamped by the solvent and no worthwhile reading could be obtained.

It will be readily apparent to all skilled in the art that the applicability of the present invention to the principles of gas-liquid chromatography may be equally applicable to certain liquid-liquid or ion-exchange chro matography situations and in which essentially equivalent rcsults can be obtained.

The invention encompases all changes and modifications of the embodiment disclosed as will be readily apparent to all skilled in the art. and is only limited in scope by the claims which follow.

What is claimed is;

I. In combination. a gas chromatographic system having a pre-column with a flow regulated carrier gas supply and injection means through which a sample so lution is injected for separation into solvent and sample compounds and movement thereof through said precolumn. and an analytical column adapted to receive sample compounds from said pre-column for analysis in a detection unit; a chromatograph accessory device comprising solvent venting means connected interme diate of the pre-column and analytical column. said solvent venting means being adapted to vent from the precolumn a substantial portion of the carrier gas and the solvent from said sample solution during conversion of the sample compounds into a detectable form in the precolumn. said solvent venting means including means for controlling said venting operation prior to movement of the sample compounds from the pre-column and past said ttcccssor) dev ice to the analytical column.

2. The chromatographic system according to claim I in which said solvent venting means comprises a con duit and valve means for selectively closing said conduit and opening it to atmosphere 3. The chromatographic system according to claim 2 in which said last-mentioned means comprises timer means for controlling the valve means for opening and closing of said Cttlttltltl.

4. The chromatographic sy stem according to claim l in which said pic-column and analytical column are packed with granular chromatographlc column material. and means for separating the packing material in the respectiv c columns.

5. The chromatograplnc s \\lcm according to claim I including other means lot I passing a portion of the carrier gas from said carrier gas supply past said prc column to a point downstream of said sohent venting means.

6. The chromatographic system according to claim 5 in which said solvent venting means and said other bypass means are connected between said pre-column and analytical column in closely adjacent relationship.

7. The chromatographic system according to claim 5 including adjustable valve means in said hy-pass means for regulating the amount of h \-pass carrier gas.

8. The method of analyzing complex samples of matter in a gas chromatograph having a pre-eolumn and an analytical column. comprising the steps of flowing a carrier gas into the pre-column at a predetermined flow rate. injecting a predetermined amount of the sample matter into the pre-column. introducing a minor portion of the carrier gas intermediate to the pre-column and analytical column, venting the solvent and carrier gas to atmosphere from a point intermediate the precolumn and analytical column for a predetermined time interval, and moving the vaporized sample constituents through the analytical column to a detector subsequent to said venting operation.

9. In combination. a chromatographic system comprising first column means having an inlet. a flow regulated carry gas supply for introducing a major portion of carrier gas into the inlet of said first column means. said first column means also having an injection port adapted to recene a sample solution. means in said first column means for separating said sample solution into its \olatile solvent and component fractions during movement of said carrier gas and sample solution through said first column means. and second column means sequentially arranged with said first column means and being adapted to receive the component fractions of said sample solution from said first column means. and detection means for sensing and analyzing said component fractions of said sample solution moving through said second column means; a chromatograph accessor) device comprising solvent venting means including an outlet conduit connected intermediate of said first and second column means and adapted to vent from said first column means a substantial amount of said volatile solvent and carrier gas during separation and movement of said component fractions of said sample through said first column means. timer operated valve means in said outlet conduit for controlling the venting operation therethrough. it -pass conduit means connected intermediate of said first and second column means downstream of said outlet conduit means and upstream of said second column means for hy-passing a minor portion of carrier gas past said first column means and outlet conduit means. and adjustable valve means in said hy-pass conduit means for regulating the amount of hy-pass carrier gas therethrough.

Claims

1. IN COMBINATION, A GAS CHROMATOGRAPHIC SYSTEM HAVING A PER-COLUMN WITH A FLOW REGULATED CARRIER GAS SUPPLY AND INJECTION MEANS THROUGH WHICH A SAMPLE SOLUTION IS INJECTED FOR SEPARATION INTO SOLVENT AND SAMPLE COMPOUNDS AND MOVEMENT THEREOF THROUGH SAID PRE-COLUMN, AND AN ANALYTICAL COLUMN ADAPTED TO RECEIVE SAMPLE COMPOUNDS FROM SAID PRE-COLUMN FOR ANALYSIS IN A DETECTION UNIT; A CHROMATOGRAPH ACCESSORY DEVICE COMPRISING SOLVENT VENTING MEANS CONNECTED INTERMEDIATE OF THE PRE-COLUMN AND ANALYTICAL COLUMN, SAID SOLVENT VENTING MEANS BEING ADAPTED TO VENT FROM THE PRE-COLUMN A SUBSTANTIAL PORTION OF THE CARRIER GAS AND THE SOLVENT FROM SAID SAMPLE SOLUTION DURING CONVERSION OF THE SAMPLE COMPOUNDS INTO A DETECTABLE FORM IN THE PRECOLUMN, SAID SOLVENT VENTING MEANS INCLUDING MEANS FOR CONTROLLING SAID VENTING OPERATION PRIOR TO MOVEMENT OF THE SAMPLE COMPOUNDS FROM THE PRECOLUMN AND PAST SAID ACCESSORY DEVICE TO THE ANALYTICAL COLUMN.

2. The chromatographic system according to claim 1 in which said solvent venting means comprises a conduit and valve means for selectively closing said conduit and opening it to atmosphere.

3. The chromatographic system according to claim 2 in which said last-mentioned means comprises timer means for controlling the valve means for opening and closing of said conduit.

4. The chromatographic system according to claim 1 in which said pre-column and analytical column are packed with granular chromatographic column material, and means for separating the packing material in the respective columns.

5. The chromatographic system according to claim 1 including other means for by-passing a portion of the carrier gas from said carrier gas supply past said pre-column to a point downstream of said solvent venting means.

6. The chromatographic system according to claim 5 in which said solvent venting means and said other by-pass means are connected between said pre-column and analytical column in closely adjacent relationship.

7. The chromatographic system according to claim 5 including adjustable valve means in said by-pass means for regulating the amount of by-pass carrier gas.

8. The method of analyzing complex samples of matter in a gas chromatograph having a pre-column and an analytical column, comprising the steps of flowing a carrier gas into the pre-column at a predetermined flow rate, injecting a predetermined amount of the sample matter into the pre-column, introducing a minor poRtion of the carrier gas intermediate to the pre-column and analytical column, venting the solvent and carrier gas to atmosphere from a point intermediate the pre-column and analytical column for a predetermined time interval, and moving the vaporized sample constituents through the analytical column to a detector subsequent to said venting operation.

9. In combination, a chromatographic system comprising first column means having an inlet, a flow regulated carry gas supply for introducing a major portion of carrier gas into the inlet of said first column means, said first column means also having an injection port adapted to receive a sample solution, means in said first column means for separating said sample solution into its volatile solvent and component fractions during movement of said carrier gas and sample solution through said first column means, and second column means sequentially arranged with said first column means and being adapted to receive the component fractions of said sample solution from said first column means, and detection means for sensing and analyzing said component fractions of said sample solution moving through said second column means; a chromatograph accessory device comprising solvent venting means including an outlet conduit connected intermediate of said first and second column means and adapted to vent from said first column means a substantial amount of said volatile solvent and carrier gas during separation and movement of said component fractions of said sample through said first column means, timer operated valve means in said outlet conduit for controlling the venting operation therethrough, by-pass conduit means connected intermediate of said first and second column means downstream of said outlet conduit means and upstream of said second column means for by-passing a minor portion of carrier gas past said first column means and outlet conduit means, and adjustable valve means in said by-pass conduit means for regulating the amount of by-pass carrier gas therethrough.