Measuring and testing – Gas analysis – Gas chromatography
Reexamination Certificate
2000-11-07
2002-11-26
Williams, Hezron (Department: 2856)
Measuring and testing
Gas analysis
Gas chromatography
C073S023420, C073S864810, C073S864210, C095S086000, C096S105000
Reexamination Certificate
active
06484560
ABSTRACT:
TECHNICAL FIELD
The present invention relates to the field of chromatography and more particularly toward an apparatus and method for concentrating an analyte prior to an analytical detector such as a mass spectrometer.
BACKGROUND
Although analytical instrumentation is becoming increasingly sensitive and analyte detection continues to improve, many chemical analytes require concentration prior to chemical analysis. Typically this is done using bench-top chemical processes specifically developed or tailored to the analytical problem. Representative of these approaches are solvent condensation or evaporation techniques that eliminate the solvent while retaining the analyte (or analytes—in all cases analyte could just as well as be analytes) by exploiting differences in physical properties such as volatility. Nitrogen or inert gas “blown-downs”, rotary evaporation, Kuderna-Danish condensers, distillation (steam, etc.), tube heaters, vacuum evaporation, (freeze drying), and related techniques are typical of approaches to pre-concentrate an analyte by removal of a solvent. A number of commercial machines exist specifically for this purpose such as RapidVap™, CentriVap™, SafetyVap™, as examples. Physical techniques such as centrifugation or selective adsorption (solid-phase extraction, column chromatography), selectively separate analytes from a bulk phase prior to redesolving in another solvent prior to injection. It is after these bench-chemistry steps that an analyte has been concentrated to an appropriate degree that makes detection and quantitation possible with an existing detector. These steps are time-consuming, subject to loss, usually require specialized equipment and technical expertise all of which lead to increased expense.
In gas chromatography, large volume injection techniques have been developed with special hardware (pre-column inlets) to allow more sensitive detection by evaporation of solvent while attempting to retain analyte inside the pre-column inlets prior to the analyte being delivered to the analytical column for chromatographic separation and analysis/detection. Typical volumes are less than or equal to 100-&mgr;l (by single injection) and the most frequent approach is to inject the solution, either in portion or in entirety, then evaporate the solvent, which is vented from the chromatograph, and transfer the analyte to the analytical column. This approach suffers from a limitation on volume that can be contained inside the pre-column inlet and, therefore, any increases in volume must be obtained by consecutive injections and evaporation cycles that can result in sample losses. These pre-column inlets that thermally program the vaporization of the solvent are called PTVs.
Another similar approach is the cool-on-column solvent venting arrangements that use a long, large diameter of capillary tubing (approximately 1 ml volume) to retain the injected volume. The operation is again the same as the PTV in that the temperature is programmed to vaporize the solvent and retain the analyte. Again the injection volume is fixed by the mechanical configuration of the assorted tubing. In both these approaches the ability to retain analytes is determined by the difference in the boiling point between the solvent and the analyte and the ability of the pre-column inlet and /or analytical column (phase) to selectively capture and retain the analyte.
However, these large-volume pre-column inlets have been considered attractive relative to the standard pre-column inlet such as split/splitless or on-column that only allow less than 4-&mgr;l injections and provide no capability for in-situ concentration.
Both the standard volume pre-column inlet arrangements and the existing large volume port technologies are limited in the volume that can be concentrated by the fact that liquid injections vaporize inside the port and mechanical arrangements (namely, the liquid or vapor volume that can be contained) place an upper bound on the concentration factors that can be achieved. It would be desirable to obtain in-situ concentration that is flexible and less constrained in the concentration factors that can be obtained.
In the biochemical and organic chemistry fields there is often a need to concentrate an analyte before it is loaded into an analytical instrument. For instance, some products may need to be concentrated, because they are expensive to derivatize, difficult to extract or synthesize. Small amounts of product or intermediates are produced and need to be characterized and identified accurately before proceeding to future research steps. However, low concentrations of product can be a problem for a researcher, because they may challenge the limits of instrument sensitivities or increase the possibility of inaccurate abundance measurements. In addition, the transfer of these analytes from instrument to instrument or from instrument to storage container can result in significant additional loss of product. For these reasons, simple techniques and instruments for both concentrating and analyzing analytes would be of interest to researchers.
A number of instruments already exist for concentrating analytes. For instance, in the biochemical fields analytes may be concentrated using centrifuges, ultra-centrifuges and filtering. A number of commercial products exist for this purpose. For instance, Amicon™ produces a number of filters that allow for desalting of samples, removal of solvent and concentration of small amounts of analyte. However, these devices often require access to a centrifuge or micro-centrifuge for spinning the analytes and are effective only when concentrating small volumes.
On a larger scale, typical analyte concentration is performed on the bench top using rotary evaporation, K-D, dry nitrogen blow down or in a PTV injection port by depositing the analyte in a liner and then evaporating the solvent by heating the liner. Each of these methods provides for concentration of analyte by removal of solvent. However, most of these methods and instruments suffer from the limitation of possible loss of analyte. In addition, since the concentration of analyte is performed separately, these methods can be time intensive and laborious. It would, therefore, be of particular interest to be able to concentrate an analyte without having to transfer, or resolvate the analyte.
Recent trends in analytical instrumentation include components for concentrating analytes in situ (i.e. directly in the instrument or more preferably before the analytes are introduced onto the pre-column). For instance, quartenary pumps are being used in HPLC instruments to pre-mix analytes before the analyte is injected and separated. Present chromatographic technology, however, requires additional hardware, components or complex instrumentation design to appropriately concentrate analytes. In addition, most of the chromatography instrumentation is designed to concentrate the analyte inside the pre-column just before it is introduced onto the column. A problem with this type of device is that the analyte is too dilute or contains a volatile solvent that significantly lowers the amount of sample concentration. It would, therefore, be desirable to be able to adapt existing chromatography hardware to serve the purpose of concentrating analytes in situ (i.e. on the syringe needle tip). A brief review of pre-column inlets, therefore, is in order to clarify existing technology that may be adapted to serve these purposes.
A number of different pre-column inlets or injectors exist to provide accurate, reproducible and predictable introduction of analytes into gas chromatography columns. Usually the analyte is a liquid and can be injected using a syringe, but other devices are available. For instance, analytes can be introduced onto columns by automatic analyzers or valves. Pre-column inlets can be divided into two major categories including packed-column inlets and capillary-column inlets. In gas chromatography the packed column inlets are fairly popular. A second type of column called a capillary-column inlet i
Agilent Technologie,s Inc.
Cygan Michael
Joyce Timothy H.
Williams Hezron
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