JP2008164498A - Sample injection device, and liquid chromatography apparatus - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a sample injection device and liquid chromatography apparatus that can suppress the occurrence of carry-over and increase the number of theoretical stages. <P>SOLUTION: The sample injection device comprises a needle 21A for a sample whose tip for delivering the sample has a tapered shape, and a pipe 15 through which the sample delivered from the needle 21A for the sample is fed out. The pipe 15 has a bowl-like sample injecting section 15a to which the needle 21A for the sample can be mounted and the sample delivered from the needle 21A for the sample is injected, and a pipe section 15b through which the sample injected from the sample injecting section 15a is fed out. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to a sample injection device and a liquid chromatography device.

  In general, a liquid chromatography device is a storage tank that stores a mobile phase (elution solvent), a degassing device that degass the mobile phase, a pump that supplies the mobile phase from the storage tank, and a sample that is injected together with the mobile phase into a pipe toward the column. A sample injection device, a column packed with a packing for separating components in the sample, a thermostat for maintaining the column at a substantially constant temperature, a detector for detecting the components in the separated sample, and the like. Among these, as disclosed in Patent Documents 1 to 3, for example, the sample injection device attaches a sample injection needle that sucks a sample to a sample injection port, and injects the sample into a pipe together with the mobile phase.

  By the way, in recent years, the detection sensitivity of a liquid chromatography apparatus has been improved, and accordingly, a phenomenon called carry-over has become a problem. Carryover is a phenomenon that shows the detection results as if the sample previously measured in time series remains in the liquid chromatography device and the substance is present in the sample currently being measured, It reduces the reliability of the analysis results. In carry-over, when a sample is injected into a pipe together with a mobile phase, the sample adheres to the metal and / or resin in the sample injection device and remains, and the remaining sample is mixed when the next sample is injected. Caused by.

  On the other hand, in liquid chromatography / mass spectrometry (LC / MS) and the like, since high-speed liquid chromatography (HPLC) is used because measurement in a short time is possible, further improvement in the number of theoretical plates is achieved. It has been demanded.

FIG. 13 shows a part of a conventional sample injection apparatus. FIG. 13A is a perspective view, and FIG. 13B is a cross-sectional view. The sample injection device shown in FIG. 13 is connected to the pipe 65 using the male screw 64 in a state where the sample injection port 61 and the needle seal 62 are fixed by the holder 63. The sample tends to adhere and a spiral flow tends to occur. As a result, carryover is likely to occur, and the number of theoretical plates is likely to decrease.
Japanese Patent Laid-Open No. 10-10103 WO reissue 2004/102182 JP 2006-201121 A

  An object of the present invention is to provide a sample injection device and a liquid chromatography device capable of suppressing the occurrence of carryover and improving the number of theoretical plates in view of the problems of the above-described conventional technology.

  The invention according to claim 1 includes a first needle having a tapered tip end for discharging the sample, and a pipe through which the sample discharged from the first needle is sent out in the sample injection device, The pipe can be fitted with the first needle, and a mortar-shaped sample injection part into which a sample discharged from the first needle is injected, and a sample injected from the sample injection part It has the piping part which is sent out.

  According to a second aspect of the present invention, in the sample injection device according to the first aspect, the inclination angle of the tip of the first needle is substantially the same as the inclination angle of the sample injection part. .

  The invention according to claim 3 is the sample injection device according to claim 1 or 2, wherein the inner diameter of the first needle is equal to or larger than the inner diameter of the pipe section.

  A fourth aspect of the present invention is the sample injection device according to any one of the first to third aspects, further comprising a pressure-bonding member that pressure-bonds the first needle to the sample injection portion. The needle has a pressed part that is pressed by the pressure-bonding member.

  According to a fifth aspect of the present invention, in the sample injection device according to any one of the first to fourth aspects, the second needle that can be attached to the sample injection portion and the first needle A sample suction means for sucking a sample into the first needle while being connected to the first needle, and the first needle is connected to the sample suction means. And a switching valve capable of switching between a state in which the second needle is connected to the mobile phase supply means for supplying the mobile phase and a state in which the first needle is connected to the mobile phase supply means. Furthermore, it is characterized by having.

  A sixth aspect of the present invention is a liquid chromatography apparatus comprising the sample injection device according to any one of the first to fifth aspects.

  According to the present invention, it is possible to provide a sample injection device and a liquid chromatography device capable of suppressing the occurrence of carryover and improving the number of theoretical plates.

  Next, the best mode for carrying out the present invention will be described with reference to the drawings.

  FIG. 1 shows an example of the liquid chromatography apparatus of the present invention. The configuration of the liquid chromatography apparatus will be described. The liquid chromatography apparatus shown in FIG. 1 has a storage tank 11 that stores a mobile phase (elution solvent), a degassing apparatus 12 that degass the mobile phase, a pump 13 that supplies the mobile phase from the storage tank 11, and a sample. A sample injection device 14 that injects into the pipe 15 toward the column 16 together with the phase, a column 16 that is filled with a filler for separating components in the sample, a thermostatic chamber 17 that maintains the column 16 at a constant temperature, The detector 18 detects components in the separated sample. In order to stabilize the measurement, it is desirable that the mobile phase is always supplied from the storage tank 11 to the column 16 via the sample injection device 14 by the pump 13.

  FIG. 2 shows an example of the sample injection apparatus of the present invention. 2 includes a pipe 15, a sample needle 21A, a mobile phase needle 21B, a syringe 22, a cleaning liquid pump 23, a valve 24, a sample container 25, an injection valve 26, a cleaning liquid container 27, and a cleaning apparatus. 28 and a needle moving means (not shown).

  The sample needle 21A can be connected to the syringe 22 via an injection valve 26 and a valve 24. It is also possible to connect to the pump 13 via the injection valve 26.

  As shown in FIG. 3, the piping 15 through which the sample discharged from the sample needle 21A together with the mobile phase is sent is a mortar-shaped sample injection portion 15a into which the sample is injected, and the sample injected from the sample injection portion 15a. The sample injection unit 15 a is fixed by a fixing member 31. The pipe 15 is preferably directly connected to the column 16. In addition, as a material of the piping 15, metals, such as PEEK (polyetheretherketone), stainless steel, and titanium, are mentioned.

  The tip of the sample needle 21A has a tapered shape, and the inclination angle is preferably 10 to 45 °, and more preferably 10 to 30 °. At this time, it is preferable that the inclination angle of the distal end portion of the sample needle 21A and the inclination angle of the sample injection portion 15a are substantially the same. Thereby, the junction area of 21 A of sample needles and the sample injection part 15a can be enlarged, and the leakage of a sample and a mobile phase can be suppressed.

  The inner diameter of the sample needle 21A is preferably 0.1 to 0.8 mm, and more preferably 0.1 to 0.3 mm. At this time, the inner diameter of the sample needle 21A is preferably equal to or larger than the inner diameter of the pipe portion 15b, and more preferably substantially the same as the inner diameter of the pipe portion 15b. Thereby, when a sample and a mobile phase are inject | poured from the sample injection | pouring part 15a, generation | occurrence | production of a spiral flow can be suppressed. In addition, it is preferable that the internal diameter of the piping part 15b is 0.05-0.3 mm, and 0.065-0.25 mm is further more preferable.

  Furthermore, as shown in FIG. 4, the sample needle 21 </ b> A may have a pressed portion 41 on both sides, preferably on both sides near the tip. At this time, when the sample and the mobile phase are injected from the sample needle 21A into the sample injection portion 15a by pressing the pressed portion 41 so as to sandwich the sample needle 21A using the crimping member 42, The deformation of the needle 21A can be suppressed.

  In addition, the mobile phase needle 21 </ b> B can be connected to the pump 13 via the injection valve 26. Although the mobile phase needle 21B is not particularly limited, it is preferable to use the same needle as the sample needle 21A.

  Next, the sample injection apparatus of the present invention will be described with reference to FIGS. 5 to 10 show the connection state of each component of the sample injection device of the present invention, and each of the standby state, the sample collection time, the preliminary cleaning of the sample needle, the ultrasonic cleaning of the sample needle, The states at the time of sample analysis (part 1) and at the time of sample analysis (part 2) are shown.

  When the sample needle 21A is connected to the syringe 22 by switching the valve 24 and the injection valve 26, the sample needle 21A can be sucked and discharged from the sample needle 21A by pushing and pulling the syringe 22. it can. When the sample needle 21A is connected to the pump 13 by switching the injection valve 26, the mobile phase is supplied from the pump 13 to the sample needle 21A.

  When the mobile phase needle 21B is connected to the pump 13 by switching the injection valve 26, the mobile phase is supplied from the pump 13 to the mobile phase needle 21B.

  A three-way solenoid valve 29 is provided in the pipe connecting the mobile phase needle 21B and the injection valve 26 to prevent dripping. The three-way solenoid valve 29 is closed when the injection valve 26 is in the connection state indicated by B in the figure when the mobile phase needle 21B is not attached to the sample injection portion 15a such as during movement, and the mobile phase needle 21B. It is possible to prevent the mobile phase from flowing out.

  The cleaning liquid container 27 stores therein a cleaning liquid (usually a solvent used for a mobile phase) and is connected to the cleaning liquid pump 23. At this time, the cleaning liquid is sucked by the cleaning liquid pump 23 and is pumped to the valve 24.

  The valve 24 can selectively supply the cleaning liquid pumped by the cleaning liquid pump 23 to the cleaning device 28 or the sample needle 21A. Specifically, the valve 24 has three ports P1 to P3, and two of these ports can be selectively connected. The valve 24 can be connected to none of the ports P1 to P3.

  The syringe 22 and the injection valve 26 are both connected to the port P3 of the valve 24. That is, the syringe 22 and the injection valve 26 are always connected.

  The cleaning device 28 includes cleaning units 28a and 28b, an ultrasonic transducer 28c, a waste liquid port 28d, and a waste liquid pipe 28e. When the cleaning device 28 is connected to the cleaning liquid pump 23 by switching the valve 24, the cleaning liquid is supplied from the cleaning liquid container 27. In addition, an excessive amount of excess cleaning liquid flows into the waste liquid port 28d and is discharged to the outside as waste liquid from the waste liquid pipe 28e connected to the waste liquid port 28d. As will be described later, by inserting the sample needle 21A into the cleaning device 28, the sample needle 21A to which the sample has adhered is cleaned. Thereby, generation | occurrence | production of carry over can be suppressed. Further, the cleaning device 28 is provided with an ultrasonic transducer 28c, and the sample needle 21A can be ultrasonically cleaned. Thereby, the cleaning effect of the sample needle 21A can be enhanced, and the occurrence of carry-over can be more reliably suppressed.

  The pipe 15 is connected to the column 16. That is, the pipe 15 is not connected to the injection valve 26 and is separated and independent from the injection valve 26. As will be described later, the sample needle 21A that sucks the sample is attached to the sample injection part 15a of the pipe 15, and the sample and the mobile phase are discharged to the sample injection part 15a, so that the sample is moved along the pipe part 15b together with the mobile phase. Stream and delivered to column 16.

  The injection valve 26 has six ports, and the pump 13, the sample needle 21A, the mobile phase needle 21B, the valve 24, and the cleaning device 28 are connected to the five ports. The injection valve 26 can switch between a connection state indicated by a solid line A in the figure (hereinafter referred to as connection state A) and a connection state indicated by a broken line B in the figure (hereinafter referred to as connection state B).

  When the injection valve 26 is in the connection state A, the sample needle 21A is connected to the pump 13 via the injection valve 26, and the valve 24 is connected to the cleaning unit 28a of the cleaning device 28 via the injection valve 26. . At this time, the mobile phase needle 21B is not connected to any of them. When the injection valve 26 is in the connection state B, the sample needle 21A is connected to the valve 24 via the injection valve 26, and the mobile phase needle 21B is connected to the pump 13 via the injection valve 26. The

  Next, a sample injection method using the sample injection apparatus of the present invention will be described.

  As shown in FIG. 5, in a standby state before taking a sample, the injection valve 26 is in the connection state A, and the valve 24 is not connected to any of the ports P1 to P3. Furthermore, the sample needle 21 </ b> A is attached to the sample injection portion 15 a of the pipe 15 by a needle moving means (not shown). Further, the mobile phase needle 21B stands by in the vicinity of the sample injection portion 15a by a needle moving means (not shown) and is in a disconnected state. Further, the three-way solenoid valve 29 closes a pipe connecting the mobile phase needle 21B and the injection valve 26. Accordingly, during standby, the mobile phase supplied from the pump 13 is supplied to the column 16 via the sample needle 21A and the sample injection portion 15a.

  As shown in FIG. 6, when a sample is collected on the sample needle 21A, the sample needle 21A is inserted into the sample container 25 by a needle moving means (not shown), and the mobile phase needle 21B. Is attached to the sample injection portion 15a. The injection valve 26 is switched to the connection state B. Further, the three-way solenoid valve 29 is also opened, and the mobile phase needle 21B is communicated with the injection valve 26. Therefore, at the time of sample collection, the mobile phase supplied from the pump 13 is supplied to the column 16 via the mobile phase needle 21B and the sample injection unit 15a. For this reason, the mobile phase is always supplied to the column 16 and the measurement can be stabilized. The sample needle 21 </ b> A is connected to the syringe 22 via the injection valve 26 and the valve 24. For this reason, the sample in the sample container 25 is sucked into the sample needle 21 </ b> A by sucking using the syringe 22. At this time, the suction amount of the sample is set so that the sucked sample does not enter the injection valve 26. Thereby, it can prevent that a sample adheres in the injection valve | bulb 26, and can suppress generation | occurrence | production of carry over. In order to increase the amount of sample suction, a sample loop may be provided in the sample needle 21A.

  When the sample collection is completed, the sample adheres to the outer wall of the sample needle 21A, and thus the sample needle 21A to which the sample has adhered is washed. Specifically, preliminary cleaning and ultrasonic cleaning of the sample needle 21A are performed.

  As shown in FIG. 7, when the sample needle 21A is preliminarily washed, the sample needle 21A is washed by the washing device 28 by the needle moving means (not shown) while maintaining the injection valve 26 in the connected state B. Insert into part 28b. Further, the valve 24 is switched so that the ports P1 and P2 are connected, and the cleaning liquid in the cleaning liquid container 27 is supplied to the cleaning unit 28b via the cleaning liquid pump 23. Therefore, the cleaning liquid is ejected into the cleaning unit 28b, and the outer wall of the sample needle 21A is preliminarily cleaned. The cleaning liquid overflowing from the cleaning unit 28b is discharged through the waste liquid port 28d and the waste liquid pipe 28e.

  As shown in FIG. 8, when ultrasonically cleaning the sample needle 21A, the sample needle 21A is removed from the cleaning device 28 by a needle moving means (not shown) while maintaining the injection valve 26 in the connected state B. Insert into the cleaning section 28a. Further, the valve 24 is switched again to a state where none of the ports P1 to P3 is connected. In this state, by driving the ultrasonic transducer 28c to generate ultrasonic waves, the cleaning liquid loaded in the cleaning unit 28a is ultrasonically vibrated, and the outer wall of the sample needle 21A is ultrasonically cleaned.

  Note that the mobile phase is supplied from the pump 13 to the column 16 via the injection valve 26, the mobile phase needle 21B, and the sample injection unit 15a during the preliminary cleaning and the ultrasonic cleaning.

  When the ultrasonic cleaning is completed, the sample collected by the sample needle 21A is supplied to the column 16 to analyze the sample.

  As shown in FIG. 9, when the sample is analyzed, the injection valve 26 is switched again from the connection state B to the connection state A, and the sample needle 21A is connected to the sample injection portion 15a by the needle moving means (not shown). It is attached to. Further, the mobile phase needle 21B is also moved to a standby position near the sample injection portion 15a by a needle moving means (not shown). When the mobile phase needle 21B starts moving, the three-way solenoid valve 29 closes the pipe connecting the mobile phase needle 21B and the injection valve 26. When analyzing the sample, the sample needle 21A that sucks the sample is attached to the sample injection portion 15a, and the sample needle 21A is connected to the pump 13 by the injection valve 26. Thereby, the sample in the sample needle 21 </ b> A is supplied to the column 16 without passing through the injection valve 26. The sample supplied to the column 16 is subjected to predetermined separation in the column 16 and then sent to the detector 18 for analysis. Therefore, it is possible to prevent the sample from remaining in the injection valve 26 and to suppress the occurrence of carryover. In addition, the number of theoretical plates can be improved by suppressing the occurrence of carryover in the sample injection apparatus.

  Note that the cleaning liquid in the cleaning unit 28a of the cleaning device 28 may be replaced during sample analysis. In this case, as shown in FIG. 10, the valve 24 is switched so that the ports P2 and P3 are connected, and the cleaning liquid in the cleaning liquid container 27 is passed through the cleaning liquid pump 23, the valve 24 and the injection valve 26. 28a. Thereby, the cleaning liquid contaminated by cleaning the sample needle 21A is discharged through the waste liquid port 28d and the waste liquid piping 28e, and the cleaning liquid not contaminated is supplied to the cleaning section 28a. Therefore, the outer wall of the sample needle 21A can be more reliably cleaned at the next ultrasonic cleaning.

  Note that the sample injection device of the present invention is not limited to the above-described configuration, and may be one in which the pipe 15 and the sample needle 21A shown in FIG. 3 or 4 are applied to a conventional sample injection device.

  FIG. 11 shows an example of such a sample injection apparatus. In FIG. 11, the same components as those in FIGS. 1 to 10 are denoted by the same reference numerals and description thereof is omitted. The sample injection device shown in FIG. 11 includes a pipe 15, a sample needle 21A, a syringe 22, a valve 24, a sample container 25, an injection valve 26, a cleaning device 51, and a needle moving means (not shown).

  The injection valve 26 has six ports, to which a pump 13, a pipe 15, a column 16, a sample needle 21A, a valve 24, and a cleaning device 51 are connected. The injection valve 26 can switch between a connection state indicated by a solid line A in the figure (hereinafter referred to as connection state A) and a connection state indicated by a broken line B in the figure (hereinafter referred to as connection state B).

  In a standby state before collecting a sample, the injection valve 26 is in a connection state A, and the sample needle 21A is attached to the sample injection portion 15a of the pipe 15. At this time, the sample needle 21 </ b> A is connected to the pump 13 via the injection valve 26, and the pipe 15 is connected to the column 16 via the injection valve 26. Therefore, the mobile phase supplied from the pump 13 is supplied to the column 16 via the sample needle 21 </ b> A, the pipe 15, and the injection valve 26.

  On the other hand, when the sample is collected, the injection valve 26 is switched to the connection state B, and the sample needle 21A is inserted into the sample container 25. At this time, the sample needle 21 </ b> A is connected to the syringe 22 via the injection valve 26 and the valve 24. For this reason, by operating the syringe 22, the sample in the sample container 25 is sucked into the sample needle 21A. In addition, since the pump 13 is connected to the column 16 via the injection valve 26, the mobile phase is supplied to the column 16 even during sample collection by the sample needle 21A.

  When the sample collection is completed, the sample adheres to the outer wall of the sample needle 21A, and thus the sample needle 21A to which the sample has adhered is washed. Specifically, after the sample needle 21A is inserted into the cleaning unit 51b of the cleaning device 51, the sample needle 21A is inserted into the cleaning unit 51a to clean the outer wall of the sample needle 21A. The cleaning units 51a and 51b of the cleaning device 51 are always supplied with a new cleaning agent by connecting the valve 24 to the cleaning liquid pump 23 at a predetermined timing. The surplus cleaning liquid is discharged from the waste liquid port 51c and the waste liquid pipe 51d.

  When the cleaning is completed, the sample collected by the sample needle 21A is supplied to the column 16 and the sample is analyzed. Specifically, the sample needle 21A is attached to the sample injection portion 15a, and the injection valve 26 is set to the connection state A again. Thus, the sample is sent from the sample needle 21A to the sample injection unit 15a by the mobile phase supplied from the pump 13 to the sample needle 21A, and then rides on the flow of the mobile phase and then the piping unit 15b and the injection valve 26. To be supplied to the column 16. The sample supplied to the column 16 is subjected to predetermined separation in the column 16 and then sent to the detector 18 for analysis.

  Using the liquid chromatography apparatus shown in FIG. 1, 2 μl of a standard sample (reserpine 10 ng / ml) was injected, and the number of theoretical plates was evaluated. In Example 1, the sample injection apparatus shown in FIG. 2 has the configuration shown in FIG. 4, and in Comparative Example 1, the sample injection apparatus shown in FIG. 11 has the configuration shown in FIG. Using.

  The conditions for liquid chromatography are shown below. As the column 16, a column having a length of 5 cm and an inner diameter of 2 mm packed with octadecylsilyl silica gel having an average particle diameter of 3 μm was used, and the temperature of the column 16 was kept at 40 ° C. in a constant temperature bath 17. In addition, a 0.1 wt% aqueous solution of acetic acid / acetonitrile (volume ratio 50:50) was used as the mobile phase, and the mobile phase flow rate was 200 μl / min. Note that water was used as the cleaning liquid. Further, as the detector 18, a mass spectrometer using an electrospray ionization method was used. The parent ion was 609 (m / z), and 195 (m / z) cations were detected.

  FIG. 12 shows the evaluation result of the number of theoretical plates. From FIG. 12, the theoretical plate number of Comparative Example 1 was 1400, and the theoretical plate number of Example 1 was 5600. From this, Example 1 has 4200 theoretical plates higher than Comparative Example 1. Further, since the peak appears sharper in Example 1 than in Comparative Example 1, it can be seen that the separation of the peak is performed cleanly, and the analysis accuracy of the liquid chromatography apparatus of Example 1 is compared. It can be seen that it is higher than Example 1.

It is a block diagram which shows an example of the liquid chromatography apparatus of this invention. It is a perspective view which shows an example of the sample injection apparatus of this invention. It is a figure which shows an example of piping used by this invention. It is a figure which shows an example of the needle used by this invention. It is a figure which shows the state at the time of the standby of the sample injection apparatus of this invention. It is a figure which shows the state at the time of sample collection of the sample injection apparatus of this invention. It is a figure which shows the state at the time of the preliminary washing of the sample needle of the sample injection device of the present invention. It is a figure which shows the state at the time of ultrasonic cleaning of the needle for samples of the sample injection apparatus of this invention. It is a figure which shows the state at the time of the sample analysis of the sample injection apparatus of this invention (the 1). It is a figure which shows the state at the time of the sample analysis of the sample injection apparatus of this invention (the 2). It is a figure which shows the other example of the sample injection apparatus of this invention. It is a figure which shows the evaluation result of Example 1 and Comparative Example 1. It is a figure which shows a part of conventional sample injection apparatus.

Explanation of symbols

DESCRIPTION OF SYMBOLS 11 Storage tank 12 Deaeration apparatus 13 Pump 14 Sample injection apparatus 15 Piping 15a Sample injection part 15b Piping part 16 Column 17 Thermostatic chamber 18 Detector 21A Sample needle 21B Mobile phase needle 22 Syringe 23 Cleaning liquid pump 24 Valve 25 Sample container 26 Injection valve 27 Cleaning liquid container 28 Cleaning device 28a, 28b Cleaning unit 28c Ultrasonic vibrator 28d Waste liquid port 28e Waste liquid piping 29 Three-way solenoid valve 31 Fixed member 41 Pressed part 42 Pressure bonding member 51 Cleaning device 51a, 51b Cleaning unit 51c Waste liquid Port 51d Waste liquid piping

Claims (6)

A first needle having a tapered tip for discharging the sample;
A pipe through which the sample discharged from the first needle is delivered;
The pipe can be fitted with the first needle, and a mortar-shaped sample injection part into which a sample discharged from the first needle is injected, and a sample injected from the sample injection part A sample injection device having a piping section through which the water is delivered.   The sample injection apparatus according to claim 1, wherein an inclination angle of a tip portion of the first needle is substantially the same as an inclination angle of the sample injection part.   3. The sample injection device according to claim 1, wherein an inner diameter of the first needle is equal to or larger than an inner diameter of the pipe portion. A crimping member for crimping the first needle to the sample injection portion;
The sample injection device according to any one of claims 1 to 3, wherein the first needle has a pressed portion that is pressed by the pressure-bonding member. A second needle that can be attached to the sample injection portion;
A sample suction means that can be connected to the first needle and sucks the sample into the first needle in a state of being connected to the first needle;
The first needle is connected to the sample suction means, the second needle is connected to mobile phase supply means for supplying a mobile phase, and the first needle is connected to the mobile phase supply means. The sample injection device according to any one of claims 1 to 4, further comprising a switching valve capable of switching a state connected to.   A liquid chromatography apparatus comprising the sample injection device according to claim 1.

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