Abstract:
A liquid chromatography system includes a gradient proportioning valve in fluidic communication with sources of solvent. From the solvent sources, the gradient proportioning valve produces a low-pressure gradient stream. A first pump is in fluidic communication with the gradient proportioning valve to receive, pressurize, and move the pressurized low-pressure gradient stream to a flow-combining device. A second pump operates in parallel with the first pump and moves a pressurized solvent stream to the flow-combining device where the pressurized solvent stream combines with the low-pressure gradient stream to produce a high-pressure gradient stream. A second gradient proportioning valve can produce, from a second plurality of sources of solvent, a second low-pressure gradient stream, wherein the solvent stream moved by the second pump to the flow-combining device and combined with the other low-pressure gradient stream comprises the second low-pressure gradient stream.

Description:
RELATED APPLICATION 
       [0001]    This application claims the benefit of and priority to co-pending U.S. provisional application No. 61/570,446, filed Dec. 14, 2011, titled “Hybrid Gradient Delivery System and Operation,” the entirety of which is incorporated by reference herein. 
     
    
     FIELD OF THE INVENTION 
       [0002]    The invention relates generally to liquid chromatography systems. More specifically, the invention relates to hybrid gradient delivery systems that combine aspects of high-pressure gradient systems and low-pressure gradient systems. 
       BACKGROUND 
       [0003]    Chromatography is a set of techniques for separating a mixture into its constituents. In liquid chromatography systems, generally, one or more high-pressure pumps take in solvents and deliver a liquid solvent composition to a sample manager, where a sample awaits injection into the mixture. The sample is the material under analysis, examples of which include complex mixtures of proteins, protein precursors, protein fragments, reaction products, and other compounds, to list but a few. From the sample manager, the resulting liquid composition, comprised of the mixture of solvents and injected sample, moves to a point of use, such as a column of particulate matter. By passing the composition through the column, the various constituents in the sample separate from each other at different rates and thus elute from the column at different times. A detector receives the elution from the column and produces an output from which the identity and quantity of the analytes may be determined. 
         [0004]    High-performance liquid chromatography (HPLC) uses two basic elution modes: isocratic elution and gradient elution. In the isocratic elution mode, the mobile phase, comprised of either a pure solvent or a mixture of solvents, remains the same throughout the chromatography run. In the gradient elution mode, the composition of the mobile phase changes during the separation. Creation of the gradient (i.e., changing mobile phase composition) entails the mixing of multiple solvents, the proportions of which change over time in accordance with a predetermined timetable. Some HPLC systems create the gradient under high pressure, by mixing the solvents downstream of the pumps. Such HPLC systems are also referred to herein as high-pressure gradient systems. Other HPLC systems create the gradient under low pressure, using a gradient proportioning valve to select from up to four solvents, mixing the multiple solvents in front of a single aspirating pump, and changing the proportions of the solvents over time. Such HPLC systems are also referred to herein as low-pressure gradient systems. 
         [0005]    The decision between a high-pressure and a low-pressure gradient system involves a variety of tradeoffs, only a few of which are mentioned here. For one, high-pressure gradient systems have lesser dwell volumes than low-pressure gradient systems because the solvent mixing occurs after the pumps instead of before the intake side of the pump. However, because of the location of mixing, low-pressure gradient systems can produce a gradient with just one pump, whereas high-pressure gradient systems generally require one pump for each solvent. Hence, low-pressure gradient systems are more amenable than high-pressure gradient systems to tertiary and quaternary gradients, and are thus predominantly used for such chromatography applications, whereas high-pressure gradient systems are generally used for binary gradients. 
         [0006]    Often, however, it is desirable to blend more than two solvents in a gradient, with a third solvent being a modifier, such as TFA (triflouroacetic acid), introduced at a constant percentage. Furthermore, it is easier to blend in a more concentrated mixture of the modifier to the total composition than to add the modifier to each of the other solvents at the desired lower concentration. For example, if 0.1% TFA is the desired concentration, it is much easier to produce a 1% concentration of TFA, and introduce it in 10% proportion to the other two solvents, than to mix a 0.1% concentration of TFA in each of the other two solvents. Hence, a low-pressure gradient system is generally used for chromatography runs to introduce the third modifier solvent instead of a high-pressure gradient system. Use of the low-pressure gradient system for this purpose, though, has disadvantages of an increased dwell volume (in comparison to a high-pressure gradient system) and limiting the maximum percentage of one of the two other solvents to 90%. 
       SUMMARY 
       [0007]    In one aspect, the invention features a solvent delivery system for use in a liquid chromatography system. The solvent delivery system comprises a gradient proportioning valve in fluidic communication with a plurality of sources of solvent and producing therefrom a low-pressure gradient stream. A first pump is in fluidic communication with the gradient proportioning valve to receive and pressurize the low-pressure gradient stream and to move the pressurized low-pressure gradient stream to a flow-combining device. A second pump operates in parallel with the first pump. The second pump moves a pressurized solvent stream to the flow-combining device where the pressurized solvent stream combines with the pressurized low-pressure gradient stream to produce a high-pressure gradient stream. 
         [0008]    In another aspect, the invention features a method for blending solvents in a liquid chromatography system. A low-pressure gradient stream is produced from a plurality of sources of solvent, pressurized, and moved to a flow-combining device while a pressurized solvent stream is moved to the flow-combining device. At the flow-combining device, the pressurized solvent stream mixes with the pressurized low-pressure gradient stream to produce a high-pressure gradient stream. 
         [0009]    In still another aspect, the invention features a liquid chromatography system comprising a solvent delivery system. The solvent delivery system includes a gradient proportioning valve in fluidic communication with a plurality of sources of solvent, producing therefrom a low-pressure gradient stream. A first pump is in fluidic communication with the gradient proportioning valve to receive and pressurize the low-pressure gradient stream and to move the pressurized low-pressure gradient stream to a flow-combining device. A second pump operates in parallel with the first pump. The second pump moves a pressurized solvent stream to the flow-combining device where the pressurized solvent stream combines with the pressurized low-pressure gradient stream to produce a high-pressure gradient stream. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0010]    The above and further advantages of this invention may be better understood by referring to the following description in conjunction with the accompanying drawings, in which like numerals indicate like structural elements and features in various figures. The drawings are not necessarily to scale, emphasis instead being placed upon illustrating the principles of the invention. 
           [0011]      FIG. 1  is a functional block diagram of an embodiment of a hybrid solvent delivery system for a liquid chromatography system, the hybrid solvent delivery system combining features of a high-pressure gradient system and a low-pressure gradient system. 
           [0012]      FIG. 2  is a functional block diagram of another embodiment of a hybrid solvent delivery system. 
           [0013]      FIG. 3  is a functional block diagram of another embodiment of a hybrid solvent delivery system. 
       
    
    
     DETAILED DESCRIPTION 
       [0014]    Solvent delivery systems described herein can be deemed hybrid systems because they combine features of a high-pressure gradient system with those of a low-pressure gradient system. As a hybrid, the solvent delivery systems function primarily as binary gradient systems having minimized dwell volume and flexibility for solvent selection. 
         [0015]      FIG. 1  shows an embodiment of a hybrid solvent delivery system  10  for producing and moving a solvent composition to a sample manager (not shown), also known as an autosampler, where a sample is introduced to the solvent composition. The features of a high-pressure gradient system adapted for use by the hybrid solvent delivery system  10 , outlined by dashed box  16 , include two pumps  14 - 1 ,  14 - 2  (generally,  14 ) operating in parallel. In brief overview, each pump  14 - 1 ,  14 - 2  includes a primary pumping actuator and an accumulator pumping actuator coupled in series. The pumps  14  can be of the type used in the 2545 Binary Gradient Module manufactured by Waters Corporation of Milford, Mass. 
         [0016]    The outlets of the pump  14 - 1 ,  14 - 2  are connected at the same or substantially the same mechanical location, here represented as a flow-combining device (FCD)  20 . Example implementations of the flow-combining device  20  include, but are not limited to, a T-section and a mixer. Each pump  14 - 1 ,  14 - 2  moves a solvent stream  18 - 1 ,  18 - 2 , respectively, at high pressure, to this flow-combining device  20 , where the pressurized solvent streams  18 - 1 ,  18 - 2  combine to produce a pressurized solvent composition  22  that is delivered over time to the sample manager. A gradient controller  24  is in communication with the pumps  14  to manage the speed of each pump  14  in order to deliver more or less of each solvent stream  18 - 1 ,  18 - 2  over the course of the separation. 
         [0017]    The features of the hybrid solvent delivery system  10  adapted from a low-pressure gradient system are shown within dashed box  26  and include two gradient proportioning valves (GPV)  28 - 1 ,  28 - 2  (generally,  28 ) operating in parallel. Each GPV  28  is in fluidic communication with up to four solvent reservoirs  30  (here, only two reservoirs per GPV are shown; the GPV  28 - 1  is in fluidic communication with solvent reservoirs  30 - 1 ,  30 - 2 ; and the GPV  28 - 2  is in fluidic communication with solvent reservoirs  30 - 3 ,  30 - 4 ). In addition, each GPV  28  is in fluidic communication with one of the pumps  14 ; the pump  14 - 1  acquires the low-pressure gradient stream  32 - 1  from the GPV  28 - 1 , and the pump  14 - 2  acquires the low-pressure gradient stream  32 - 2  from GPV  28 - 2 . 
         [0018]    Each GPV  28  includes an inlet for each reservoir  30 , an inlet valve (not shown) for controlling each flow of fluid being drawn into one of the inlets, and a common outlet through which fluid flows from the GPV  28  to one of the pumps  14 . A conduit for transporting fluid, for example, a tube, extends from each reservoir  30  to one of the inlets of the GPV  28  and from the outlet of the GPV  28  to the intake side of the pump  14 . An example implementation of a gradient proportioning valve is described in U.S. Pat. No. 5,862,832, issued Jan. 26, 1999, the entirety of which patent is incorporated by reference herein. 
         [0019]    The gradient controller  24  is in communication with the GPVs  28  to actuate their individual valves sequentially at the appropriate times, thereby managing the intake of fluid from the reservoirs  30  for mixing in desired proportions and producing low-pressure gradient streams  32  over time. From solvent reservoirs  30 - 1 ,  30 - 2 , the GPV  28 - 1  produces low-pressure gradient stream  32 - 1 , and from solvent reservoirs  30 - 3 ,  30 - 4 , the GPV  28 - 2  produces low-pressure gradient stream  32 - 2 . These gradient streams  32  are produced ahead of the pumps  14 , and thus under low pressure. 
         [0020]    During operation, the pump  14 - 1  draws and pressurizes the low-pressure gradient stream  32 - 1  produced by the GPV  28 - 1 , and moves the resulting pressurized low-pressure gradient stream  18 - 1  to the flow-combining device  20 , while the pump  14 - 2  draws and pressurizes the low-pressure gradient stream  32 - 2  produced by the GPV  28 - 2 , and moves the resulting pressurized low-pressure gradient stream  18 - 2  to the flow-combining device  20 , where the two pressurized low-pressure gradient streams  18 - 1 ,  18 - 2  combine to produce the pressurized solvent stream  22 . Because the flow-combining device  20  is downstream of the pumps  14  the solvent stream  22  is produced at high pressure (e.g., in the range between 5000-15000 psi). As used herein, the phrase “pressurized low-pressure gradient stream” refers to a low-pressure gradient stream that is produced by a GPV and subsequently pressurized to a high pressure by a pump  14 . 
         [0021]    As an illustration of the operation, consider for example that solvent reservoir  30 - 1  contains water, solvent reservoir  30 - 2  contains 1% TFA in water, solvent reservoir  30 - 3  contains solvent B, and solvent reservoir  30 - 4  contains 1% TFA in solvent B. To achieve a solvent composition with 0.1% TFA modifier, the gradient controller  24  can control the GPV  28 - 1  to take a 90% proportion of solvent reservoir  30 - 1  (water) and a 10% proportion of solvent reservoir  30 - 2  (1% TFA in water) to produce a low-pressure gradient stream  32 - 1  of 0.1% TFA in water. In addition, the gradient controller  24  can control the GPV  28 - 2  to take a 90% proportion of solvent reservoir  30 - 3  (solvent B) and a 10% proportion of solvent reservoir  30 - 4  (1% TFA in solvent B) to produce a low-pressure gradient stream  32 - 2  of 0.1% TFA in solvent B. The pumps  14  draw and pressurize the low-pressure gradient streams  32 - 1 ,  32 - 2 , and combine the resulting pressurized low-pressure gradient streams  18 - 1 ,  18 - 2  to produce a pressurized solvent stream  22  comprised of water, solvent B, and 0.1% TFA. The maximum achievable proportion of solvent B is 100%. 
         [0022]    In an alternative embodiment, the hybrid solvent delivery system can have one GPV  28  only. For example, consider that GPV  28 - 2  and the solvents  30 - 3 ,  30 - 4  are not part of the hybrid solvent delivery system  10  shown in  FIG. 1 , and that the pump  14 - 2  is instead in direct fluidic communication with a solvent reservoir  30 - 5 . During operation of this embodiment, the pump  14 - 1  draws and pressurizes the low-pressure gradient stream  32 - 1  produced by the GPV  28 - 1 , and moves the resulting pressurized low-pressure gradient stream  32 - 1  to the flow-combining device  20 , while the pump  14 - 2  draws, pressurizes, and moves solvent from reservoir  30 - 5  to the flow-combining device  20 , where the pressurized low-pressure gradient stream  18 - 1  combines with the solvent  30 - 5 . If the pump  14 - 2  is turned off, any of the embodiments of the hybrid solvent delivery system  10  can be adapted to operate like a conventional low-pressure gradient system. 
         [0023]      FIG. 2  shows another embodiment of a hybrid solvent delivery system  50 , which adds a second stage of GPVs  52  to the hybrid solvent delivery system  10  of  FIG. 1 . More specifically, the hybrid solvent delivery system  50  includes a GPV  52 - 1  in fluidic communication with a plurality of solvent reservoirs  54 - 1 ,  54 - 2  and a GPV  52 - 2  in fluidic communication with a plurality of solvent reservoirs  54 - 3 ,  54 - 4 . The outlet of GPV  52 - 1  is in fluidic communication with one of the inlets of GPV  28 - 1 , while the outlet of GPV  52 - 2  is in fluidic communication with one of the inlets of GPV  28 - 2 . The gradient controller  24  is in communication with the GPVs  52  to manage the intake of fluid from the reservoirs  54  for mixing in desired proportions and producing low-pressure gradient streams  56 - 1 ,  56 - 2  (generally,  56 ) over time. From solvent reservoirs  54 - 1 ,  54 - 2 , the GPV  52 - 1  produces low-pressure gradient stream  56 - 1 , and from solvent reservoirs  54 - 3 ,  54 - 4 , the GPV  52 - 2  produces low-pressure gradient stream  56 - 2 . 
         [0024]    During operation, the low-pressure gradient stream  32 - 1  produced by the GPV  28 - 1  includes a proportion of the low-pressure gradient stream  56 - 1  produced by the GPV  52 - 1 . The pump  14 - 1  draws and pressurizes the low-pressure gradient stream  32 - 1 , and moves the resulting pressurized low-pressure gradient stream  18 - 1  to the flow-combining device  20 . Concurrently, the GPV  28 - 2  produces the low-pressure gradient stream  32 - 2 , which includes a proportion of the low-pressure gradient stream  56 - 2  produced by the GPV  56 - 2 . The pump  14 - 2  draws and pressurizes the low-pressure gradient stream  32 - 2 , and moves the resulting pressurized low-pressure gradient stream  18 - 2  to the flow-combining device  20 , where the two pressurized low-pressure gradient streams  18 - 1 ,  18 - 2  combine to produce the pressurized solvent stream  22 . In one embodiment, the gradient controller  24  centrally controls the various compositions of each low-pressure gradient stream  32 ,  56  and the resulting high-pressure gradient stream (i.e., solvent stream  22 ). In other embodiments, the gradient controller  24  includes a plurality of decentralized controllers that intercommunicate and manage the various compositions in fashion. 
         [0025]    In alternative embodiments, the hybrid solvent delivery system  50  can have one second-stage GPV  52  only or one second-stage GPV  52  and one first-stage GPV  28 . For example, for one alternative the GPV  52 - 2  and the solvents  54 - 3 ,  54 - 4  are not part of the hybrid solvent delivery system  50  shown in  FIG. 2 ; in another example alternative, both the GPV  52 - 2  and GPV  28 - 2  are not part of the hybrid solvent delivery system  50 . Again, if the pump  14 - 2  is turned off, any of the embodiments of the hybrid solvent delivery system  50  can be adapted to operate like a conventional low-pressure gradient system. 
         [0026]      FIG. 3  shows another embodiment of a hybrid solvent delivery system  60 , which fluidically connects a switch valve  62  to one of the GPVs  28  of the hybrid solvent delivery system  10  of  FIG. 1 . More specifically, the hybrid solvent delivery system  60  includes a switch valve  62  in fluidic communication with a plurality of solvent bottles (or reservoirs)  64 - 1 ,  64 - 2 ,  64 - 3 ,  64 - 4 ,  64 - 5 , and  64 - 6 . In this example, the outlet of the switch valve  62  is in fluidic communication with one of the inlets of the GPV  28 - 2 . The gradient controller  24  is in communication with the switch valve  62  to select one of the inlets of the switch valve  62 , and, thus, the particular solvent bottle  64  from which to draw solvent. 
         [0027]    From the solvent drawn through the switch valve  62  and from solvent reservoirs  30 - 3 ,  30 - 4 , the GPV  28 - 2  produces the low-pressure gradient stream  32 - 2 . The pump  14 - 1  draws and pressurizes the low-pressure gradient stream  32 - 1  produced by the GPV  28 - 1  and moves the resulting pressurized low-pressure gradient stream  18 - 1  to the flow-combining device  20 , while the pump  14 - 2  draws and pressurizes the low-pressure gradient stream  32 - 2 , and moves the resulting pressurized low-pressure gradient stream  18 - 2 , which includes the solvent from the selected solvent bottle  64 , to the flow-combining device  20 , where the pressurized low-pressure gradient streams  18 - 1 ,  18 - 2  combine. 
         [0028]    In like fashion, another switch valve and set of solvent bottles can be fluidically connected to an inlet of the GPV  28 - 1  instead of or in combination with the switch valve  62  and solvent bottles  64  connected to the GPV  28 - 2 . The embodiments of  FIG. 3  are merely illustrative examples of the varying complexity that can be built into a hybrid solvent delivery system. 
         [0029]    While the invention has been shown and described with reference to specific preferred embodiments, it should be understood by those skilled in the art that various changes in form and detail may be made therein without departing from the spirit and scope of the invention as defined by the following claims.