Waters Technologies Corporation

Micromass UK Limited

Waters Investments Limited

Robert Stephen Plumb

Chris Lee Stumpf

Marc Victor Gorenstein

Jennifer Harnisch Granger

Jose M Castro-Perez

Alan Millar

Christopher Clarkson Jones

Joseph D Michienzi

Geoff C Gerhardt

James P Murphy

Moon Chul Jung

Michael John Tomany

Paul D Rainville

Jose Castro-Perez

An apparatus for use in a chromatography system includes a microfluidic substrate having a fluidic channel configured as an analytical chromatographic column and a fluidic port on one side of the microfluidic substrate. The fluidic port opens at a head end of the analytical chromatographic column. A dried blood spot (DBS) collection device holds one or more dried biological samples. The DBS collection device is directly coupled to the microfluidic substrate whereby one of the biological samples is placed into fluidic communication with the fluidic channel of the microfluidic substrate and an extraction of that biological sample flows toward the head end of the analytical chromatographic column. A diluent source fluidically coupled to the fluidic port supplies a solvent to the head end of the analytical column to dilute the extracted biological sample before the biological sample flows into the analytical chromatographic column.

Analysis of dried blood spot samples in a microfluidic system with dilution of extracted samples

A method of searching for potentially unknown metabolites of pharmaceutical compounds is disclosed. The accurate mass of a pharmaceutical compound will generally be known and can be rendered in the form of an integer nominal mass or mass to charge ratio component and a decimal mass or mass to charge ratio component. Possible metabolites are searched for on the basis of having a decimal mass or mass to charge ratio component which is substantially very similar to the decimal mass or mass to charge ratio of the parent pharmaceutical compound.

Mass spectrometry

A separation device is formed of a first tube with a cylindrical wall enclosing a chamber having a first diameter, where the device has a first end for discharging a fluid and a second end for receiving a fluid. At least two stationary phase medias are packed into the chamber with separating frits between the medias. At least one end frit element, secured to an inner surface of the first tube contains the media in its section of the chamber and allows the beds formed of the media to be formed and packed. When the separation device so constructed forms a nanocolumn with a guard bed, the guard bed may be cleaved from the separation device extending the useful life of the nanocolumn. When the separation device so constructed comprises two analytical sections, and a plurality of guard beds, complex analysis may be performed on a column having an extended useful life.

Separation device with integral guard column

Mass spectrometer

Chromatograms and mass spectra produced by an LC/MS system are analyzed by creating a two-dimensional data matrix of the spectral and chromatographic data. The two-dimensional matrix can be created by placing the spectra generated by the mass spectrometer portion of the LC/MS system in successive columns of the data matrix. In this way, the rows of the data matrix correspond to chromatographic data and the columns of the data matrix correspond to the spectra. A two-dimensional filter is specified and applied to the data matrix to enhance the ability of the system to detect peaks associated with ions. The two-dimensional filter is specified according to desired criteria. Rank-1 and rank-2 filters can be specified to improve computational efficiency. One method of applying the two-dimensional filter is through convolution of the data matrix with the two-dimensional filter to produce an output data matrix. Peaks corresponding to detected ions are identified in the output data matrix. Parameters of the peaks are determined and stored for later processing including quantitation, or simplification of chromatograms or spectra by, for example, identifying peaks associating with ions having retention times falling within a specified retention time window or having mass-to-charge ratios falling within a specified mass-to-charge ratio window.

Apparatus and method for identifying peaks in liquid chromatography/mass spectrometry data and for forming spectra and chromatograms

Disclosed herein are an apparatus and method for effectuating increased chromatographic efficiency in a capillary column by employing a retaining frit disposed within an analytical capillary. By disposing the retaining frit within the analytic capillary, the void volume is significantly minimized. The columns and methods described herein produce a simplified analytical capillary and retaining frit apparatus that provides greater chromatographic efficiency. Additionally, the column of the instant invention maintains chromatographic fidelity by reducing the transfer diameter as well as facilitating fluidic connections in situ.

Capillary loop with a built-in retaining frit

Nanocolumn chromatography devices useful, e.g., in CEC and nanoLC are disclosed. An exemplary chromatography device of the invention includes a nanocolumn, e.g., a capillary, packed with a particulate stationary phase material and a solid support. The solid support, or in situ frit, is adjacent to and integral with the stationary phase material. An in situ frit of the invention may be a mixture of the stationary phase material and a polymeric network of cross-linked poly(diorganosiloxane), e.g., poly(dimethylsiloxane), which may optionally be sintered. The invention also provides methods of making and using such devices.

Polymeric solid supports for chromatography nanocolumns

Chromatograms and mass spectra produced by an LC/MS system are analyzed by creating a two-dimensional data matrix of the spectral and chromatographic data. The two-dimensional matrix can be created by placing the spectra generated by the mass spectrometer portion of the LC/MS system in successive columns of the data matrix. In this way, the rows of the data matrix correspond to chromatographic data and the columns of the data matrix correspond to the spectra. A two-dimensional filter is specified and applied to the data matrix to enhance the ability of the system to detect peaks associated with ions. The two-dimensional filter is specified according to desired criteria. Rank-1 and rank-2 filters can be specified to improve computational efficiency. One method of applying the two-dimensional filter is through convolution of the data matrix with the two-dimensional filter to produce an output data matrix. Peaks corresponding to detected ions are identified in the output data matrix. Parameters of the peaks are determined and stored for later processing including quantitation, or simplification of chromatograms or spectra by, for example, identifying peaks associating with ions having retention times falling within a specified retention time window or having mass-to charge ratios falling within a specified mass-to-charge ratio window.

Disclosed herein is a method of analyzing three-dimensional data obtained from a chromatography/spectrometry process, in particular an LC/MS process using a two-dimensional multi-variant statistical analysis. The LC portion permits separation of analytes within a sample. The information obtained from such a procedure typically depends on retention time (R). As the analytes progress through the system, they enter the MS region of the LC/MS system. There they are ionized and a mass detector then detects these ionized species. The information procured from this procedure is generally reported as intensity for a corresponding m/z value. Therefore, an LC/MS system provides at least three pieces of information. Principle component analysis (PCA) is a robust method of multi-variant analysis of this type of data between different samples. However, typically, PCA analysis is performed using only two-dimensional data. Therefore, it is expected that at least one parameter of data obtained from a LC/MS operation is lost, however, the methods disclosed herein preserves all three mathematical dimensions.

Method of using data binning in the analysis of chromatography/spectrometry data

Mass intensity values, isotopic ratios and exact mass differences between isotopes may be obtained and analyzed to determine if there are one or more masses of interest. In one implementation, a method will look among the mass intensity values for peaks of interest. The mass intensity values, representative of isotopic signatures, will be compared to a criteria in order to determine which of the masses are of interest. Although the invention is not so limited, examples of such criteria include mass intensity values above a specified threshold, within a certain ratio of another mass intensity value, and/or separation between the masses themselves. Optionally, a tolerance may be applied to the criteria. If masses are found to be of interest, MS/MS can automatically be triggered for one or all of the masses of interest, when an analyzing system such as an LC-MS-MS, GC-MS-MS or MALDI-MS-MS system is used.

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Milford, MA, United States of America

Robert Stephen Plumb