PROCESS AND APPARATUS FOR CHEMICAL ANALYSIS BY LIQUID CROMATOGRAPHY

Abstract: A process and an apparatus for performing analyses in liquid chromatography, by means of a pump capable of drawing a fluid and injecting it at high pressure, through at least a control valve, into a separation column. To obtain maximum precision as required by the analysis, before each analysis or each group of analyses, or with respect to each fluid used, a pump stroke is performed with the pump delivery side shut off in order to record and memorize the characteristics of compressibility of the fluid. These charac­ teristics are kept into consideration in the subsequent operative stages to obtain an exactly constant delivery of the feed pump to the separation column.

Description: The present invention relates to a process and an apparatus for performing analyses by high-pressure chromatography, wherein a syringe pump is used for injecting a fluid together with the sample at high pressure into a separation column. More exactly, the invention relates to a process and apparatus allowing to rectify the plunger stroke of said syringe pump in order to adjust the compressibility of the fluid it contains, depending upon the operating pressure value during an analysis. The apparatus comprises a syringe pump actuated by a motor means and capable of drawing a preset quantity of fluid from a container of same, then sending it, through a control valve, to a sample drawing zone and then to a column of chromatographic separation and finally to a detector. As it is well known to those skilled in the art, the chromatography analysis precision and reliability are connected to several factors, the most important of which is the need of exactly controlling the quantity of fluid introduced into the column and consequently the constancy of the fluid flow to the column. Up to now said delivery constancy was ensured by a motor means continuously acting on the piston pump, for example in the form of a step-by-step motor capable of ensuring a constant piston forward stroke and therefore, theoretically, a constant delivery feeding. US-E-(Re)37586 relates to a high pressure pump for liquid chromatography comprising a piston reciprocating in a chamber which is alternatively filled and emptied, in conjunction with the action of valve means. A valve placed downstream the pump and connecting the same with the column is maintained in its closed position until the analysis operating pressure is reached. Moreover, during compression, a signal is produced, proportional to the linear displacement of the piston, and a second signal is produced which indicates the pressure. These signals are used to control the piston speed. This pump allows to avoid chamber compliance compressibility of the fluid and obtain a certain constancy of the fluid flow to the chromatographic column, but its precision is not sufficient for particular high pressures and particular solvents, as will be explained later on. US-A-4347131 discloses a high pressure pump for liquid chromatography, comprising a piston reciprocating in a chamber under the control of two valves. A pressure sensor in foreseen for safety purposes only. It must be noticed that pressures involved during the above stated analyses are now very high, for example up to values of 50 MPa (500 kg/cm²),i.e. values at which the fluid compressibility, in particular that of certain fluids which are under supercritical temperature and pressure conditions, cannot be disregarded. Amongst those fluids creating heavy problems of compressibility at said high values of pressure, the following can be considered: carbon dioxide, pentane, ammonia, when used under supercritical temperature and pressure conditions, and moreover alcohols, chlorinated, aromatic and aliphatic fluids. In these cases precision on the amount pumped can be lost, in that a constancy in the forward movement of the piston does not correspond to a constancy of delivered flow, as at high pressures said flow rate is reduced due to the fluid compressibility. An object of the present invention is to provide a solution to this problem therefore providing a process and an apparatus which allow to obtain more precise and accurate as well as more reliable analyses in comparison with those performed up to now. According to the invention, said problem is solved by means of a process for performing sample analyses using a separation column to which fluid together with said sample is delivered at a high pressure by pump means having a mobile element, trough a fluidically connecting valve, said process comprising the steps of: drawing said fluid into said pump; sealingly closing said fluidical connecting valve to close off said pump means; compressing said fluid by said pump means up to a first predetermined pressure higher than the foreseen analysis pressure;detecting the fluid pressure values generated in said pump means during said fluid compression step, and the corresponding displacements of the pump mobile element; memorizing said values in a memory of a microprocessor together with said displacements of the said pump mobile element; determining the compressibility values of said fluid according to said detected pressures: opening said fluidically connecting valve to connect said pump means to said separation column; starting the sample analysis and the actual delivery of said fluid together with said sample to said separation column; and regulating said fluid and sample delivery during said analysis according to said previously determined fluid compressibility values at the current analysis pressure, in order to maintain a controlled constant delivery of said fluid and sample to said separation column. In practice, during said fluid compression stroke a pressure-piston movements curve typical of the fluid treated is memorized in a microprocessor. During the subsequent analysis, the microprocessor is set on the delivery value involved and is then capable of controlling the speed of the pump piston taking into consideration the compressibility of the fluid at the operating pressure each time present during the compression stroke, as well as the fluid volume present at each moment in the pump chamber and other possible factors. The invention further comprises an apparatus for performing high pressure sample analysis using a separation column comprising, upstream of said separation column, a valve to fluidically connect pump means provided with a mobile element and control means, alternatively to a fluid container, to the separation column or to an exhaust, means for sealingly closing and disconnecting said pump means from said container, separation column and exhaust during a fluid compression step a pressure transducer to detect the fluid pressure values in the pump during said compression step characterized in that it further comprises: a microprocessor for memorizing, during said fluid compression step of the pump, the movements of said pump mobile element and the fluid pressure values recorded by the transducer, and for determining and memorizing fluid compressibility values; and means for connecting said microprocessor to pump control means in order to control the constancy of fluid and sample delivery during operative strokes sending the fluid to separation column according to the compressibility values of said fluid as recorded during said previous compression step. In order to sealingly close the pump delivery side, a valve is advantageously used, having a position in which the duct connecting the same to the pump is closed. As above stated, the pump is preferably a piston pump actuated by a step-by-step motor which is connected to the pump piston by a gear reduction unit and a screw-screw nut connection, a ball circulation connection, or the like, acting on the piston rod. Said valve typically comprises a head having a series of ducts leading to different points of an internal plane surface, as well as a counter-surface provided with at least a groove to connect the duct coming from the pump with the other ducts or with a closed space, said counter-surface being rotating on an axis perpendicular to the same, for the orientation of the connecting groove towards the operative positions, and being moreover elastically pressed against the first surface. The invention will be now described more in detail with reference to a preferred embodiment of same, illustrated as an example in the accompanying drawings, wherein: Figure 1 is a diagrammatic view of an apparatus according to the invention for samplings in chromatography. Figure 2 is an example of a curve: piston movements-pressure, as recorded by a pressure transducer in the apparatus of figure 1 and memorized in a microprocessor of the same apparatus, for an idle stroke of the pump. Figure 3 is a partially cutaway side view of the switch valve of figure 1. Figure 4 is a cross section of figure 3 Figure 5 is a partially cutaway side view of the piston pump and relevant control means. Figure 6 is an enlarged cross view illustrating the connection between the rod and piston in the pump of figure 5. With reference first of all to figure 1, an apparatus for performing chromatographic analyses is diagrammatically shown. The apparatus comprises, a pump 10 for example consisting of a cylinder 11 and a piston 12, actuated by an actuating means which in the shown example is constituted by a step-by-step motor 13 and by a gear unit 14 reducing the rotation speed of the motor shaft 15 and transforming the rotatory movement into a translation of rod 16 of piston 12. The pump 10 is connected downstream, through a duct 17, with a switch valve 18 capable of connecting said duct 17 alternatively to a duct 19 for drawing a fluid from a container 20, to an exhaust duct 21 and to a duct 22 connecting the separation column 23, through a device 24, capable of introducing the sample 25. Downstream of the column 23, the arrow 26 shows the connection to a detector. As it is known, in liquid or SFC chromatography the analyses are performed at pressures of the fluid introduced through the separation column 23 which reach very high values, for example up to 50 MPa (500 kg/cm²). In order to keep into consideration the compressibility of the fluids used, the apparatus comprises, on the duct 17 connecting the pump 10 to the valve 18, a pressure transducer 27 which is able to detect at any moment the pressure values present in the duct 17 and to communicate them, through the line 28, to a microprocessor 29, which also receives the data concerning the movements of piston 12, through the line 30 connecting the motor 13. By performing a plurality of detections it is possible to memorize in the microprocessor 29 a curve s-p such as the one shown in figure 2, in which pressure values are correlated with the movements of piston 12 as obtained with a given fluid, the curve being recorded when the fluid is submitted to a compression stage within the pump 10 and the duct 17, up to pressure values at least corresponding to the ones involved in the sample analyses, by a pump stroke as performed after closure of the duct 17 by means of the valve 18. In practice, before each analysis or before each group of analyses performed with the same fluid, a step is foreseen during which the desired fluid is drawn from the container 20, the valve 18 is closed and the piston 12 of the pump is moved forward, then the values of pressure and displacement of the same piston 12 at any moment are recorded and memorized. After this stage, the analysis goes on normally and the movement of piston 12 is controlled by a direct control of the microprocessor 29 on the step-by-step motor 13 in a way as to maintain a constant and preset value of delivery through the column 23, keeping into consideration the operating pressure, the volume of fluid present at any instant in the pump chamber and finally the compressibility of the fluid which has been memorized in the microprocessor 29. It is possible to foresee a thermoregulation of the syringe in which the mobile phase or fluid is contained, which allows to obtain programmed deliveries with the maximum accuracy even if the environmental temperature varies. The connection of duct 17, coming from the pump, respectively with duct 19 for the fluid drawing, with exhaust duct 21 and with duct 22 leading to the separation column, are controlled by a motorized valve 18, as shown more in detail in figures 3 and 4. Said valve comprises a body 31 which carries a head 33 fixed by means of bolts 32, and wherein four ducts are inserted, 34, 35, (figure 4), which are connected to the ducts 17, 19, 21 and 22. The ducts 34-37 all lead to a plane surface 38, as diagrammatically indicated by references 39-42 in figure 4, the outlet 39 of duct 34, connected to the pump 10, being positioned centrally with respect to said surface. For the alternative connection of duct 34 with one of the other ducts 35, 36, 37, a counter-surface 43 bearing an elongated groove 44 is resting on the surface 38 (figure 4), said counter-surface being capable of fluidically connecting the passage 39 with one of passages 40, 41 and 42 or, if rotated to a position opposite to the one illustrated in figure 4, of closing said duct 34 from the external environment. As shown in figure 3, said counter-surface 43 is carried by a circular plate 54 mounted on a support 45 which on its turn is connected, by means of a slit 46 and a pin 47, to an extension 56 of the shaft 48 of a motor means 49, capable of being rotated for a preset angle. The rotations of counter-surface 43 are controlled by a tubular appendix 50 as shown by the support 45 and provided with fins 51 which pass through a known position detecting device 52 to ensure the utmost precision in the positioning of support 45 and therefore of groove 44. The plate 54 and support 45 are mounted in a rotatory way with respect to the valve body 31 by means of a self-centering thrust bearing 53 which rests on one side on an adjusting nut 55 and is stressed on the other side by a compression spring 57 which bears, on the opposite side, against the support 45, to press the surface 43 against the surface 38 with the desired necessary strength, in order to ensure that no leakage occurs even at very high pressures. Sealing between the two surfaces is ensured, even in long time periods, by a self-adaptation as determined by a "floating" assembling of support 45 on the shaft 48 of the motor 49. The side sealing to the counter-surface 43 is in any case ensured by gaskets 58. By rotating the shaft 48, the support 45 and the plate 54, it is possible to selectively place the groove 44 in a way as to connect the outlet 39 with outlets 40, 41 and 42 respectively, or in a position in which pump connecting duct 17 is completely closed. Figures 5 and 6 illustrate some details of pump 10 and of the relevant control motor. The pump 10 essentially consists of a cylinder 60 and piston 62 pump, whose cylinder 60 is closed at one end by a head 61 with a passage 63 to connect the duct 17. On the opposite side, the cylinder 60 is fixedly connected, by a nut 64 screwed on a thread 65, to a bell body 66 which ensures a connection with the step-by-step motor and with a movement reducing and transforming unit. More exactly, the motor 13 has a shaft 67 bearing a pinion 68 which actuates two reducing gears 69 and a ring gear 70 which controls a rectilinear feeding device 71 of the screw-screw nut type, preferably of the ball circulation type, mounted on support 66 by means of a bearing 72. The piston rod 73 has the configuration of a but screw and moves inside the housing 74, said rod 73 being connected to the piston 62 in the way more specifically illustrated in figure 6. With reference to said figure 6, the piston is formed by a block 75 having two heads 76 and 77 connected for instance by means of screws 78 and 79 in such a way as to form seats 80 and 81 for gaskets 82 and 83 (figure 5). The head 77 has a head having an oval shape in section, which houses a shaped head 84 fixed to the rod 73 at 85, the connection between the two heads 77 and 84 being in this case too of the "floating" type and such as to ensure a self-adaptation of the piston 62 in its seat inside the cylinder 60, in order to reduce wear of the gaskets 82 and 83.

Claims: A process for performing sample analyses using a separation column (23) to which fluid together with said sample is delivered at a high pressure by pump means (10) having a mobile element (12), through a fluidically connecting valve (18), said process comprising the steps of: drawing said fluid into said pump (10); sealingly closing said fluidical connecting valve (18) to close off said pump means (10); compressing said fluid by said pump means (10) up to a first predetermined pressure higher than the foreseen analysis pressure; detecting the fluid pressure values generated in said pump means (10) during said fluid compression step, and the corresponding displacements of the pump mobile element (12); memorizing said values in a memory of a microprocessor (29), together with said displacements of the said pump mobile element (12); determining the compressibility values of said fluid according to said detected pressures; opening said fluidically connecting valve (18) to connect said pump means (10) to said separation column (23); starting the sample analysis and the actual delivery of said fluid together with said sample to said separation column (23); and regulating said fluid and sample delivery during said analysis according to said previously determined fluid compressibility values at the current analysis pressure, in order to maintain a controlled constant delivery of said fluid and sample to said separation column (23). A process according to claim 1, characterized in detecting the current operating pressures and the current volume of fluid present in said pump (10) and in using these values together with the said memorized fluid compressibility values to maintain said controlled constant fluid and sample delivery. A process according to claim 2, characterized in detecting the current fluid temperature values and in using this further values to maintain said controlled constant fluid and sample delivery. A process according to claim 1, characterized in that said fluid compression step is performed before each analysis or each cycle of analyses with the same fluid. A process according to claim 1, characterized in that said fluid compression step is performed only once for each fluid and the relevant values are indefinitely memorized. An apparatus for performing high pressure sample analysis using a separation column (23) comprising, upstream of said separation column (23), a valve (18) to fluidically connect pump means (10), provided with a mobile element (12) and control means (13-15), alternatively to a fluid container (20), to the separation column (23) or to an exhaust, said valve (18) being able to sealingly close off said pump means (10) from said container (20), separation column (23) and exhaust during a fluid compression step, a pressure transducer (27) being provided for to detect the fluid pressure values in the pump during said compression step characterized in that it further comprises: a microprocessor (29) for memorizing, during said fluid compression step of the pump, the movements of said pump mobile element (12) and the fluid pressure values recorded by the transducer, and for determining and memorizing fluid compressibility values; and means (30) for connecting said microprocessor (29) to pump control means (13-15) in order to control the constancy of fluid and sample delivery during operative strokes sending the fluid to separation column (23), according to the compressibility values of said fluid as recorded during said previous compression step. An apparatus according to claim 6, wherein said valve for sealingly closing off the pump (10) consists of a valve (18) having a position for closing a duct (17) connecting said valve (18) to the pump (10). An apparatus according to claim 6, wherein said pump means (10) is a piston pump (62) and said pump control means (13-15) is a stepping motor (49), the latter being connected to the pump piston by means of a reducing gear (69) and a screw-screw nut connection (71), connected to the rod (73) of said piston (62). An apparatus according to claim 7, wherein said valve (18) comprises a head (33) having a series of ducts (19, 21, 22; 34-37) leading to different points of a first internal plane surface (38), as well as a counter-surface (43) provided with at least a connecting groove (44) for selectively connecting the duct (17; 34) coming out from the pump (10) to the other ducts (19, 21, 22; 34-37) or to a closed space, said counter-surface (43) being rotatively mounted on an axis (56) perpendicular to the same, for the orientation of said connecting groove (44) and being elastically pressed against said first plane surface (38). An apparatus according to claim 9, wherein the counter-surface (43) is flotatingly mounted on said axis (56) by means of a tubular appendix (50).

Inventors: CARERA, ROSOLINO, TRISCIANI, ADRIANO
Applicants: CARLO ERBA STRUMENTAZIONE S.P.A.
Application year: 1985
Main classification: G01N30
IPC codes: G01N30