Abstract:
Guard columns for high-pressure liquid chromatography (HPLC) apparatuses. These guard columns can be removable and/or metallic guard columns. Further, all of the outlet of the guard column can substantially engage the inlet of an HPLC column in the HPLC apparatus. Also, an HPLC apparatus that includes a guard column, a hand-tightened top end fitting at a first end of the HPLC apparatus and a hand-tightened bottom end fitting at a second end of the HPLC apparatus.

Description:
BACKGROUND OF THE INVENTION  
         [0001]    1. Field of the Invention  
           [0002]    The present application relates generally to the field of guard columns for high-pressure liquid chromatography (HPLC) devices. Guard columns are generally used to protect HPLC columns from physical or chemical contamination.  
           [0003]    2. Description of the Related Art  
           [0004]    High-pressure liquid chromatography (HPLC) is a process used for separating one or more compounds from a chemical mixture. The HPLC process consists of passing the mixture through a stationary packing material, under the influence of a high-pressure transport liquid, and separating the compounds by selective affinity, sieving, absorption or partition. The packing is typically housed within a thru-bored section of a body of a column and is typically held in place by frits at either end of the body.  
           [0005]    [0005]FIG. 1 illustrates a typical HPLC apparatus  10  according to the related art wherein a chemical mixture is injected into the HPLC apparatus  10  through an injection apparatus  300 . The mixture first passes through the frits  120 ,  140  and packing  115  of a guard column  30 . Then, the mixture is “fineled” through a narrow opening of a capillary connector  40  that is located between the guard column  30  and an HPLC column  20 . From that point, the mixture flows through the frits  180 ,  200  and packing  170  of the HPLC column  20  and the components of the mixture can then travel through an exit apparatus  340  after which they can be measured, collected, redirected or disposed of.  
           [0006]    The typical HPLC apparatus  10  illustrated in FIG. 1 shows the guard column  30  contained within a housing  45  and held in place by a threaded top end fitting  310 . The top end fitting  310  is screwed onto an end of the housing  45 . Because of the extreme pressures sometimes used to conduct HPLC processes (e.g., pressures up to and above 6,000 pounds per square inch (psi)), the top end fitting  310  is typically screwed to the housing  45  using wrenches or other methods of supplying high torque.  
           [0007]    The injection apparatus  300  through which the mixture enters the HPLC apparatus  10  is sheathed within the top end fitting  310 . A small region of the top end fitting  310  can form a top end fitting pathway  12  through which the mixture travels before flowing into the guard column  30 .  
           [0008]    The guard column  30  is positioned between the top end fitting  310  and the housing  45 . To prevent leaks, a guard column top seal  90  is placed between the guard column  30  and the top end fitting  310  and a guard column bottom seal  70  is placed between the guard column  30  and the housing  45 .  
           [0009]    The guard column  30  is removable and replaceable. To remove the guard column  30 , the top end fitting  310  is unscrewed from the housing  45  and the guard column  30  is pulled out. The guard column  30  can then be inspected and, if necessary, replaced. An advantage of a replaceable guard column  30  is that the HPLC column  20 , which is substantially more expensive than the guard column  30 , does not have to be replaced as often, if ever. The guard column  30  traps impurities or particulates in the sample mixture or in the transport liquid before the impurities or particulates can reach the HPLC column  20  and thereby extends the operable lifetime of the HPLC column  20 .  
           [0010]    The capillary connector  40  can be a piece of tubing or other device that connects the guard column  30  to the HPLC column  20 . It is, according to FIG. 1, held in place by a top capillary fitting  22  and a bottom capillary fitting  23 .  
           [0011]    The capillary connector  40  abruptly changes the cross-sectional area of the path of the mixture by forcing the mixture to flow from the thru-bore section of the guard column  30  to the relatively narrow opening of the capillary connector  40 . Then, the mixture passes through a relatively large thru-bore section of the HPLC column  20 . The sudden changes in the cross-sectional area of the flow path disrupt the flow of the mixture, cause unwanted mixing and blending of the mixture, and lower the capability of the HPLC column  20  to separate compounds from the mixture (i.e., causes band-spreading).  
           [0012]    An improvement to the HPLC apparatus  10  design depicted in FIG. 1 exists in the related art. This improvement eliminates the capillary connector  40  by making the guard column  30  “integral” to the HPLC column  20 .  
           [0013]    [0013]FIG. 2 illustrates such an alternative HPLC apparatus  10 , according to the related art, wherein a chemical mixture is injected into the HPLC apparatus  10  through an injection apparatus  300 . The mixture first passes through the frits  120 ,  140  and packing  115  of a guard column  30  that sits in a guard column housing  45 . Then, the mixture is “funneled” through a narrow opening that is located on the exit end of the guard column housing  45 . From that point, the mixture flows through the frits  180 ,  200  and packing  170  of the HPLC column  20  and the components of the mixture can then travel through an exit apparatus  340 , after which they can be measured, collected, redirected or disposed of.  
           [0014]    The components of the HPLC apparatus  10  illustrated in FIG. 2 are contained within an assembly comprised of an externally threaded bottom end fitting  360  and an internally threaded top end fitting  310 . This assembly is held in place by engaging two upper grommets  350  that fit into an upper groove  380  that is machined into the outer wall of the HPLC column  20 . Because of the extreme pressures sometimes used to conduct HPLC processes (e.g., pressures up to and above 6,000 psi), the top end fitting  310  and the bottom end fitting  360  are typically screwed together using wrenches or other methods of supplying high torque.  
           [0015]    The injection apparatus  300  through which the mixture enters the HPLC apparatus  10  is sheathed within the top end fitting  310 . A small region of the top end fitting  310  can form a pathway  12  through which the mixture travels before flowing into the guard column  30 .  
           [0016]    The guard column  30  is enclosed in a guard column housing  45  and positioned between the top end fitting  310  and the HPLC column  20 . To prevent leaks, a guard column top seal  90  is placed in the guard column housing  45  above the top end of the guard column  30 . This guard column top seal  90  mates with the exit end of the top end fitting  310  to form a leak-tight seal. An HPLC column top seal  100  is placed inside the HPLC column  20  to mate with the exit end of the guard column housing  45  to form a leak-tight seal.  
           [0017]    The guard column  30  and the guard column housing  45  are removable and replaceable. To remove the guard column  30  and the guard column housing  45 , the internally threaded top end fitting  310  is unscrewed from the externally threaded bottom end fitting  360  and the guard column  30  and the guard column housing  45  are pulled out. The guard column  30  can then be inspected and, if necessary, the guard column  30  and the guard column housing  45  can be replaced.  
           [0018]    The guard column housing  45  abruptly changes the cross-sectional area of the path of the mixture by forcing the mixture to flow from the thru-bore section of the guard column  30  to the relatively narrow exit opening  42  of the guard column housing  45 . Then, the mixture passes through a relatively large thru-bore section of the HPLC column  20 . The sudden changes in the cross-sectional area of the flow path disrupt the flow of the mixture, cause unwanted mixing and blending of the mixture, and lower the capability of the HPLC column  20  to separate compounds from the mixture (i.e., causes band-spreading).  
           [0019]    Below the guard column housing  45  is the HPLC column top seal  100  that is placed inside the top of the HPLC column  20 . An HPLC column bottom seal  110  is placed between the HPLC column  20  and an exit fitting  375  that sheathes the exit apparatus  340 . The exit fitting  375 , when screwed onto a lower HPLC column fitting  365 , holds the exit fitting  375  and the HPLC column  20  together with the use of two lower grommets  385  that fit into a lower groove  395 . The end of the exit fitting  375  contains a pathway through which the components of the mixture travel before flowing into the exit apparatus  340 , after which they can be measured, collected, redirected or disposed of. The HPLC column bottom seal  1   10 , the HPLC column top seal  1   00 , and the guard column top seal  90  are typically formed from polymeric materials that are specifically designed and manufactured to withstand the high-pressure and potentially corrosive conditions necessary for HPLC processes. Hence, the seal materials are extremely expensive and each seal increases the overall cost of the HPLC apparatus illustrated in FIG. 2. In addition, the HPLC apparatus in FIG. 1 and in FIG. 2 both require the sample mixture to “funnel” through narrow channels between the guard columns and the HPLC columns. The sudden changes in the cross-sectional area of the flow path disrupt the flow of the mixture, cause unwanted mixing and blending of the mixture, and lower the capability of the HPLC column  20  to separate compounds from the mixture (i.e., causes band-spreading). Accordingly, a need exists for an improved HPLC apparatus  10 .  
         BRIEF SUMMARY OF THE INVENTION  
         [0020]    According to one embodiment, a high-pressure liquid chromatography (HPLC) apparatus that includes a top end fitting, a removable, metallic guard column in direct contact with the top end fitting, wherein the guard column includes a guard column inlet and a guard column outlet, and an HPLC column, wherein the HPLC column includes an HPLC column inlet and an HPLC column outlet, and wherein substantially all of the guard column outlet engages the HPLC column inlet.  
           [0021]    According to another embodiment, an An HPLC apparatus that includes a hand-tightened top end fitting at a first end of the HPLC apparatus and a hand-tightened bottom end fitting at a second end of the HPLC apparatus wherein the top end fitting and the bottom end fitting provide seals sufficient for operation of the HPLC apparatus, and an HPLC column positioned between the top end fitting and the bottom end fitting, the HPLC column having an HPLC column inlet and an HPLC column outlet, a removable guard column having a guard column inlet and a guard column outlet wherein substantially all of the guard column outlet engages the HPLC column inlet.  
           [0022]    According to yet another embodiment, a method of gathering HPLC data that includes hand-tightening a top fitting and a bottom fitting around a removable guard column and an HPLC column engaged with the guard column to provide a seal, injecting an experimental sample into the guard column, and allowing the experimental sample to travel through a path of substantially constant cross-sectional geometry as the experimental sample travels through a guard column outlet, to an HPLC column inlet, and through the HPLC column inlet. 
       
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0023]    The invention will be described by way of example, in the description of exemplary embodiments, with particular reference to the accompanying drawings in which:  
         [0024]    [0024]FIG. 1 illustrates a cross-sectional view of a high-pressure liquid chromatography (HPLC) apparatus incorporating a non-integral guard column according to the related art.  
         [0025]    [0025]FIG. 2 illustrates a cross-sectional view of an alternative liquid chromatography (HPLC) apparatus incorporating an integral guard column according to the related art.  
         [0026]    [0026]FIG. 3 illustrates a cross-sectional view of a guard column used in HPLC columns within the scope of the present invention.  
         [0027]    [0027]FIG. 4 illustrates a cross-sectional view of a guard column and of an HPLC column connected to it, wherein the HPLC column top frit and the guard column bottom frit have substantially similar diametral cross-sectional geometries.  
         [0028]    [0028]FIG. 5 illustrates a cross-sectional view of a first embodiment of an HPLC apparatus according to the present invention.  
         [0029]    [0029]FIG. 6 illustrates a cross-sectional view of a guard column and of an HPLC column connected to it, wherein the HPLC column top frit and the guard column bottom frit have different diametral cross-sectional geometries. 
     
    
     DETAILED DESCRIPTION OF THE INVENTION  
       [0030]    [0030]FIG. 3 illustrates a guard column  30  for use with an HPLC apparatus  10 . The guard column  30  includes a thru-bored guard column grooved body  105  with a diametral groove capable of accommodating the presence of a guard column bottom seal  70 .  
         [0031]    The guard column bottom seal  70 , and all other seals of the present invention, can be made of any material(s) capable of preventing leaking under the conditions necessary to conduct the high-pressure liquid chromatography (HPLC) process. Preferred materials include perfluoroelastomers such as, for example, CHEMRAZ and KALREZ perfluoroelastomers. All seals of the present invention are also preferably, though not necessarily, substantially square cross-sectional quad-ring seals.  
         [0032]    Inside the thru-bored section of the guard column grooved body  105  is the guard column packing  115 . The guard column packing  115  is kept in place between a porous guard column top frit  120  at a guard column inlet  130  and a porous guard column bottom frit  140  at a guard column outlet  150 . The materials from which all of the packings  115 ,  170  and porous frits  120 ,  140 ,  180 ,  200  of the present invention are made are those materials that allow for HPLC processes to be carried out. They include, but are not limited to, silane-derivatized silica particles for the packings  115 ,  170  and to porous stainless steel for the frits  120 ,  140 ,  180 ,  200 .  
         [0033]    [0033]FIG. 4 illustrates a portion of an HPLC column  20  and how it is arranged relative to the guard column  30  according to one embodiment of the present invention. According to the embodiment shown, the HPLC column  20  includes an HPLC column body  160  with HPLC column packing  170  contained within it. The HPLC column packing  170  is kept in place between a porous HPLC column top frit  180  at the HPLC column inlet  190  and a porous HPLC column bottom frit  200  at the HPLC column outlet  210 .  
         [0034]    Because the guard column bottom seal  70  is positioned within the guard column grooved body  105  instead of below the guard column  30 , the guard column bottom frit  140  can be placed in direct contact with the HPLC column top frit  180 . Hence, the guard column  30  and HPLC column  20  are directly connected or engaged to each other, eliminating the need for a capillary connector  40  or guard column housing  45 .  
         [0035]    According to other embodiments of the present invention, the guard column bottom frit  140  and the HPLC column top frit  180  are not quite in direct contact. Instead, any or all of the frits  120 ,  140 ,  180 ,  200  are proximate (e.g., within  0 . 1  millimeters) of the column inlets and outlets to which they are closest to, but are not quite flush with the inlet or outlet. These embodiments allow for greater manufacturing tolerances.  
         [0036]    In other embodiments of the present invention, items such as, but not limited to, washers and additional seals can be placed between the guard column  30  and the HPLC column  20 , causing the guard column bottom frit  140  and the HPLC column top frit  180  to be slightly separated. However, such items do not prevent the guard column outlet  150  from engaging with the HPLC column inlet  190 . Even with, for example, the presence of a washer, the guard column outlet  150  and the HPLC column inlet  190  can still come together and/or interlock to an extent sufficient to maintain the capability of the HPLC apparatus to separate compounds from the mixture with minimum band-spreading.  
         [0037]    The HPLC column  20  and guard column  30  illustrated in FIG. 4 are illustrated in FIG. 5 within a simpler design for an HPLC apparatus  10 . The HPLC apparatus  10  in FIG. 5 requires fewer seals than the designs in FIG. 1 or FIG. 2 and therefore reduces the overall cost of the HPLC apparatus  10 . Also, without the constrictive openings of the capillary connector  40  or guard column housing  45 , the mixture does not have to experience a sudden widening of the flow path as it enters the HPLC column  20 . Instead, according to the embodiments of the present invention illustrated in FIG. 5, the guard column bottom frit  140  and the HPLC column top frit  180  have identical diametral cross-sectional dimensions and geometries with relation to the flow path of the mixture.  
         [0038]    According to the embodiment of the present invention illustrated in FIG. 5, only two seals, the guard column bottom seal  70  and an HPLC column bottom seal  110 , are needed to prevent leaking. Hence, the cost of the HPLC apparatus  10  shown in FIG. 5, and the cost of the subsequent maintenance for it, is substantially reduced when compared to an HPLC apparatus  10  where three or more seals are used.  
         [0039]    The HPLC apparatus  10  illustrated in FIG. 5, along with other embodiments of the present invention, has the advantage that the top end fitting  330  and the bottom end fitting  360  can usually be screwed onto a spacer  370  by hand-tightening. Even under 6,000 psi internal operating conditions, no wrenches are typically needed to tighten the fittings  330 ,  360  sufficiently to prevent leaking in the HPLC apparatus  10 . In fact, for one embodiment of the present invention wherein the HPLC column  20  had a 0.250-inch outer diameter, the amount of force needed to prevent leaking was approximately 0.5 foot-pounds. For another embodiment of the present invention wherein an HPLC column  20  with a 1.0-inch outer diameter, 2 foot-pounds of force were required, an amount of force well within the capability of human hands.  
         [0040]    [0040]FIG. 6 illustrates another embodiment of the present invention wherein a guard column  30  and an HPLC column  20  are in direct contact with each other and where only one seal, a guard column bottom seal  70 , is placed between them. Although a guard column grooved body  105  is illustrated in FIG. 6, other embodiments of the present invention make use of an HPLC column grooved body  150  and use an HPLC column top seal as the only seal between the guard column  30  and the HPLC column  20 .  
         [0041]    According to the apparatus illustrated in FIG. 6, the guard column bottom frit  140  is smaller than the HPLC column top frit  180 . This embodiment of the present invention minimizes the costs of the HPLC apparatus  10  because it utilizes only a single seal between the HPLC column  20  and the guard column  30 .  
         [0042]    The embodiment illustrated in FIG. 6 also shows that the spacer  370  illustrated in FIG. 5 is not always required. According to the embodiment illustrated in FIG. 6, the top end fitting  330  and the bottom end fitting  360  are both screwed directly onto a threaded HPLC column  20 .  
         [0043]    The foregoing detailed description has been given for understanding exemplary implementations of the invention only and no unnecessary limitations should be understood therefrom as modifications will be obvious to those skilled in the art without departing from the scope of the appended claims and their equivalents.