Patent Application
20080179252
The present invention relates to a method for analyzing organic compounds utilizing a SPE (Solid Phase Extraction) cartridge and an apparatus therefor by adsorbing analytes contained in a subject sample preliminarily to a SPE cartridge, eluting the analytes with a solvent, injecting the obtained eluate via a needle into a storage chamber in a gas chromatograph, whereby performing a quantitative analysis, in an organic compound analysis.
A liquid chromatographic analysis has been employed to analyze organic compounds, especially pesticides (for example, see Patent Reference 1). The liquid chromatographic analysis has also been applied to an analysis of pesticides employed in a golf course (for example, see Patent Reference 2). Meanwhile, as the safety of residual pesticides on agricultural produce became controversial, gas chromatography began to be employed in addition to liquid chromatography. It was also proposed to use a method employing gas chromatography in combination with infrared absorption spectrums or a method employing a microtrap in a pretreatment for a gas chromatographic analysis (for example, see Patent Reference 3 and Patent Reference 4). On the other hand, in response to an increasing concern on the environment, a method for analyzing dioxins and the like has also been studied (for example, see Patent Reference 5). A study was also made to connect a SPE cartridge to a gas chromatograph, and a method involving automation was also proposed utilizing a valve instead of packing for attenuating the analytes was also proposed (for example, see Non-Patent Reference 1).
Patent Reference 1: Japanese Unexamined Patent Publication No. 6-331618
Patent Reference 2: Japanese Unexamined Patent Publication No. 5-306998
Patent Reference 3: Japanese Unexamined Patent Publication No. 8-170941
Patent Reference 4: Japanese Unexamined Patent Publication No. 2002-328121
Patent Reference 5: Japanese Unexamined Patent Publication No. 2002-48688
Non-Patent Reference 1: Ryoichi Sasano et al., “Journal of Chromatography A” Elsevier Science (United States), 896 (2000) p. 41-49
It has been proposed, in response to an increasing concern on the environment, to increase the accuracy and the workability of the analysis of organic compounds such as residual pesticides or endocrine disturbing chemicals by a gas chromatographic analysis utilizing a SPE cartridge onto which analytes in a subject sample are adsorbed. In non-Patent Reference 1, the analytes adsorbed to the subject sample are eluted with a solvent, after which the eluate is injected to a storage chamber in the gas chromatograph through a piping that brings the SPE cartridge and the storage chamber in the gas chromatograph into communication, whereby the analytes are analyzed. After the completion of the analysis, the piping is washed. In such a case, a first valve is provided between one end of the piping and the SPE cartridge and a second valve is provided between the other end of the piping and the gas chromatograph, and it is enabled by switching these two valves to each other that the washing liquid (solvent) is supplied to the piping through the first valve, and said washing liquid is returned to the piping through the second valve and then drained via a drainage piping connected to the first valve whereby effecting the washing procedure, after which the abovementioned two valves are switched prior to the analysis of analytes adsorbed on the next SPE cartridge, but the analytical results may reflect the influence of the eluate because it is difficult to drain the eluate as completely as emptying the gap between the valve and the piping or a dead space.
An object of the invention is to know how to wash the inside of the piping and the needle to eliminate the influence of the eluate on the analysis and how to conduct these operations automatically.
The inventors made an effort to overcome the problems mentioned above, and finally have discovered the invention proposed here. Thus, the invention is a method for analyzing organic compounds utilizing a SPE cartridge comprising eluting analytes previously adsorbed onto the SPE cartridge with a solvent from a liquid feed pump, injecting an eluate obtained into a storage chamber in a gas chromatograph through a needle inserted into an injection port in said storage chamber, pulling said needle up from said injection port, washing the inside of said needle with a solvent fed from said liquid feed pump, discarding the wash liquid, followed by analyzing the organic compounds as analytes in the subject sample, and it is preferred to convey the eluate containing the analytes into a separate SPE cartridge to remove contaminants.
In the second aspect, the invention is a method for analyzing organic compounds utilizing a SPE cartridge comprising eluting analytes previously adsorbed onto the SPE cartridge with a solvent from a liquid feed pump, injecting an eluate obtained into a storage chamber in a gas chromatograph through a needle inserted into an injection port in said storage chamber while dissolving in said eluate a derivatizing reagent which is then injected together with the analytes into the storage chamber in the gas chromatograph, pulling said needle up from said injection port, washing the inside of said needle with a solvent fed from said liquid feed pump, discarding the wash liquid, followed by derivatizing in said storage chamber the organic compounds as analytes in a subject sample prior to the analysis of said derivatives, and it is preferred to convey the eluate containing the analytes into a separate SPE cartridge to remove contaminants.
In the third aspect, the invention is a method for analyzing organic compounds utilizing a SPE cartridge comprising eluting analytes previously adsorbed onto the SPE cartridge with a solvent from a liquid feed pump, injecting an eluate obtained into a storage chamber in a gas chromatograph through a needle inserted into an injection port in said storage chamber, pulling said needle up from said injection port, washing the inside of said needle with a solvent fed from said liquid feed pump, discarding the wash liquid, followed by analyzing the organic compounds as analytes in a subject sample, further followed by eluting analytes previously adsorbed onto a second SPE cartridge with a solvent from a liquid feed pump, injecting an eluate obtained into a storage chamber in a gas chromatograph through a needle inserted into an injection port in said storage chamber, pulling said needle up from said injection port, washing the inside of said needle with a solvent fed from said liquid feed pump, discarding the wash liquid, followed by analyzing the organic compounds as analytes in the subject sample again consecutively, and it is preferred to convey the eluate containing the analytes into a separate SPE cartridge to remove contaminants.
In the fourth aspect, the invention is a method for analyzing organic compounds utilizing a SPE cartridge comprising eluting analytes previously adsorbed onto the SPE cartridge with a solvent from a liquid feed pump, injecting an eluate obtained into a storage chamber in a gas chromatograph through a needle inserted into an injection port in said storage chamber while dissolving in said eluate a derivatizing reagent which is then injected together with the analytes into the storage chamber, pulling said needle up from said injection port, washing the inside of said needle with a solvent fed from said liquid feed pump, discarding the wash liquid, followed by derivatizing in said storage chamber the organic compounds as analytes in the subject sample prior to the analysis of said derivatives, further followed by eluting analytes previously adsorbed onto a second SPE cartridge with a solvent from a liquid feed pump, injecting the eluate containing the analytes into a storage chamber in a gas chromatograph through a needle inserted into an injection port in said storage chamber while dissolving in said eluate a derivatizing reagent which is then injected together with the analytes into the storage chamber, pulling said needle up from said injection port in the storage chamber, washing the inside of said needle with a solvent fed from said liquid feed pump, discarding the wash liquid, followed by derivatizing in said storage chamber the organic compounds as analytes in the subject sample prior to the analysis of said derivatives again consecutively, and it is preferred to convey the eluate containing the analytes into a separate SPE cartridge to remove contaminants.
In the fifth aspect, the invention is a method for analyzing organic compounds in a SPE cartridge comprising a readily unclampable clamp component for clamping a certain cartridge among a plural of SPE cartridges onto which analytes contained in a subject sample have previously been adsorbed in a pretreatment step and a washing cartridge, a liquid feed pump for supplying a solvent to a cartridge clamped by said clamp, a first piping for supplying the solvent from this liquid feed pump to said SPE cartridge, a second piping for transferring an eluate containing the analytes which were supplied to said SPE cartridge and then eluted to a needle communicating to a storage chamber in a gas chromatograph, a liquid draining part movable from a receiving position through a non-receiving position, said receiving position serving to receive as a waste fluid a washing liquid which is the solvent, supplied here as a washing liquid to the needle via said second piping from said washing cartridge, from said liquid feed pump upon clamping said washing cartridge by said clamp component after the analyte-containing eluate eluted from said SPE cartridge was supplied to the storage chamber of the gas chromatograph, characterized in that an insertion state switching means is provided for switching, in the case of said SPE cartridge, to the first insertion state in which the tip of said needle is inserted into the storage chamber in said gas chromatograph, and for switching, in the case of said washing cartridge, to the second insertion state in which the tip of said needle is inserted into an inlet in said liquid draining part.
It is also possible that a cartridge holder for holding said plural of SPE cartridge s and a washing cartridge is provided, said cartridge holder is constituted from a holding component capable of holding a plural of cartridges at a certain interval on a horizontal straight line and an actuator for moving said cartridge holder in a horizontal direction is provided.
It is also possible that said insertion state switching means is provided with an ascending-descending driving means for moving the clamp component clamping said cartridge upward or downward and a moving operation means for moving said liquid draining part toward a receiving position which is located inside of a moving path of the cartridge moved by said ascending-descending moving means and which allows the tip of said needle connected to said clamp component to be inserted into the inlet in said liquid draining part and also toward a non-receiving position which is located outside of said moving path and which allows the tip of said needle to be inserted into the storage chamber (injection port) of said gas chromatograph.
It is also possible that said insertion state switching means is provided with an ascending-descending driving means for moving said needle connected via a hose to said clamp component in an upward or downward direction and a moving operation means for moving said liquid draining part toward a receiving position which is located inside of a moving path of the cartridge moved by said ascending-descending moving means and which allows the tip of said needle connected to said clamp component to be inserted into the inlet in said liquid draining part and also toward a non-receiving position which is located outside of said moving path and which allows the tip of said needle to be inserted into the storage chamber (injection port) of said gas chromatograph.
In the sixth aspect, the invention is an apparatus for analyzing organic compounds in a SPE cartridge comprising a grasp means for grasping a certain cartridge among a plural of SPE cartridges onto which analytes contained in a subject sample have previously been adsorbed in a pretreatment step and a washing cartridge to move it to a certain position, a readily unclampable clamp component for clamping the certain cartridge moved by said grasp means, a liquid feed pump for supplying a solvent to a cartridge clamped by said clamp, a first piping for supplying the solvent from this liquid feed pump to said SPE cartridge, a second piping for transferring an eluate containing the analytes which were supplied to said SPE cartridge and then eluted to a needle communicating to a storage chamber (injection port) in a gas chromatograph, a liquid draining part movable from a receiving position through a non-receiving position, said receiving position serving to receive as a waste fluid a washing liquid which is the solvent, supplied here as a washing liquid to the needle via said second piping from said washing cartridge, from said liquid feed pump upon clamping said washing cartridge by said clamp component after the analyte-containing eluate eluted from said SPE cartridge was supplied to the storage chamber of the gas chromatograph, characterized in that an insertion state switching means is provided for switching, in the case of said SPE cartridge, to the first insertion state in which the tip of said needle is inserted into the storage chamber (injection port) in said gas chromatograph, and for switching, in the case of said washing cartridge, to the second insertion state in which the tip of said needle is inserted into an inlet in said liquid draining part.
It is also possible that a contaminant removing means for removing contaminants is provided between said second piping and said needle.
It is also possible that a cartridge holder for holding a plural of solid phased cartridges onto which contaminants are to be adsorbed is provided, and said cartridge holder is constituted from a holding component capable of holding a plural of cartridges at a certain interval on a horizontal straight line and an actuator for moving said cartridge holder in a horizontal direction is provided, whereby constituting said contaminant removing means.
Since a method for analyzing organic compounds utilizing a SPE cartridge and an apparatus therefor makes a valve unnecessary, during the organic chemical analysis, by means of switching between the state where a needle is inserted into an injection port of a storage chamber in a gas chromatograph and the state where the needle is pulled out of the injection port to discard the washing liquid, the eluate is never retained in a piping or the needle whereby accomplishing a sure washing step. Accordingly, it is excellent in analyzing analytes accurately, and exerts its ability especially in analyzing organic compounds in atmosphere or organic compounds in water, and thus can analyze a trace substance such as residual pesticides or endocrine disturbing chemicals rapidly and accurately. Also by means of an automation, an analysis requires reduced labor, time, cost and the like.
Also by providing a contaminant removing means for removing contaminants between a second piping and a needle described above, the contaminant, if any, which was escaped from removal in a previous processing step can be removed whereby increasing the reliability regarding to the contaminant removal, and such a removal of the contaminant during the analysis makes the step for removing the contaminant in the previous step unnecessary whereby increasing the arbitrariness of designing advantageously.
While organic compounds, as used herein, are not limited particularly, preferred analytes in conducting the invention may for example be a trace component such as residual pesticides or endocrine disturbing chemicals. Residual pesticides may for example be asulam, oxine-Cu, mecoprop, thiuram, siduron, iprodion, chlorothalonil, pencycuron, bensulide and the like. Endocrine disturbing chemicals may for example be phenols such as nonylphenol, bisphenol A and the like, phthalates, PCBs and dioxins, organic tin compounds such as tributyltin, triphenyl tin and the like, synthetic estrogens such as ethylestradiol, estriol and the like.
As used herein, a subject sample refers to an atmospheric air and waters such as drinking water, waste water and the like, as well as vegetable and animal food product, whose organic compounds contained therein and whose organic compounds deposited thereon are to be analyzed. Examples include vegetable pesticides or endocrine disturbing chemicals deposited on whose surface is to be analyzed.
As used herein, an analyte refers to an organic compound which is contained in the subject sample and whose content is to be analyzed. For example, it refers to an organic compound intended to be analyzed including organic compounds contained in atmospheric air such as endocrine disturbing chemicals and organic compounds contained in water such as endocrine disturbing chemicals and residual pesticides as well as organic compounds contained as a constituent of a food product or organic compounds deposited on the surface of a food product.
While the subject sample may itself be used as a analyzable sample in the case of atmospheric air and various waters such as a drinking water and a waste water, a vegetable or animal food product is usually prepared into a analyzable sample by extracting organic compounds to be analyzed with a solvent followed by diluting to a certain volume. For example, a vegetable or a fruit which was cut into small pieces, or a cereal or a pea which was grained, is combined with water to effect a full swelling, and then homogenized together with acetonitrile as a solvent, filtered, subjected to an extraction of organic compounds, which is then made to a certain volume. An analyzable sample is adsorbed onto the column of a liquid chromatograph, and then eluted with a mobile phase liquid different from the mobile phase liquid employed upon introduction to the liquid chromatograph, while the retention time of the analytes is set preliminarily to transfer the target substance exclusively to the SPE cartridge. For example, a mobile phase other than acetonitrile is employed to transfer the target organic compounds to the SPE cartridge, to which then a syringe needle is attached, via which the eluate from the SPE cartridge is injected directly to a gas chromatograph to perform an analysis.
As used herein, a SPE cartridge onto which analytes have previously been adsorbed refers to a SPE cartridge employing as a packing a styrdivinylbenzene polymer (SDB), an octadecylsilane silica gel (C18), an ethyldiamine-N-propylsilane silica gel (PSA) and the like, and the packing may be selected depending on the intended analytes to obtain a satisfactory separation. While a SPE cartridge for removing contaminants may contain a packing which may vary depending on the types of the contaminants, those which may be exemplified are silica gel, florsil, PSA, ion exchange resin-based solid phase and the like. A washing cartridge has no packing filled therein, and used for washing the needle and the injection port of a storage chamber of the gas chromatograph.
Specifically, those provided include a cartridge holder 3 having a number of holding parts for holding SPE cartridges 1 and a washing cartridge 2 at a certain interval over a horizontal straight line (vertical or angled line may also be employed), a pair of upper/lower unclampable clamp components 5,5 for clamping a certain SPE cartridge 1 held by the abovementioned cartridge holder 3 to supply the solvent from a liquid feed pump 4 without leakage, a needle 7 A of a syringe 7 for supplying an eluate containing analytes eluted from the certain SPE cartridge 1 with the solvent supplied as mentioned above to an injection port 6A in a storage chamber in a gas chromatograph 6, and a liquid draining part 8 which receives a waste water containing the washing liquid supplied to and then drained from said washing cartridge to which a solvent was supplied from the abovementioned liquid feed pump 4 while clamping the abovementioned washing cartridge 2 with clamping components 5, 5. The abovementioned liquid feed pump 4 is connected to one clamp 5 via a flexible hose 9 which is a first piping, and the other clamp 5 is connected to the syringe 7 via a flexible hose 10 which is a second piping.
More specifically, the abovementioned liquid chromatograph 11 is provided with a pump 17 for supplying a mobile phase liquid to an injection port 16 for introducing an analyzable sample into the column, a pre-column 18 for removing analyzable sample-carried contaminants which degrade an LC column and an LC column 19 for separating the contaminants from the analytes, and the analytes contained in the eluate from this LC column 19 will be detected by the abovementioned detector 12. The detector 12 may also be a constituent component of the liquid chromatograph 11.
While the abovementioned cartridge holder 3 consists of a first holding component 3A which is in a form of board having a plural of through holes formed therein whereby allowing a plural (may be 2 or more although 12 in Figure) of the cartridges 1 to be held at a certain interval on a horizontal straight line as shown in
As shown in
Accordingly, by driving the operation shaft 22A of the abovementioned electric motor 22 rotationally from the state shown in
24 in
Accordingly, as shown in
With regard to an automatic analysis by an analytical apparatus, the power of the apparatus is first turned on to effect an origin action of respective electric motors based on the detection signals from origin sensors 24, 36 (while the figure shows those only for the electric motor 22 and a needle driving electric motor 37, an origin sensor for a holder driving electric motor M or an origin sensor for a liquid feed pump 4 may also be provided). While the second insertion state in which the tip of the abovementioned needle 7A is inserted into an inlet 8A in the abovementioned liquid draining part 8 is established here because of the start from the washing step, an analytical step may first be conducted at a very beginning because of no solvent retained initially in a piping.
Then, the cartridge holder 3 is moved to a certain position allowing the washing cartridge 2 to be clamped by the clamp components 5,5. Thereafter, the washing cartridge 2 is clamped by the pair of upper/lower clamp components 5,5 and then the liquid feed pump 4 is driven to pass the solvent through the hose 9, the washing cartridge 2, the hose 10, the needle 7A, and then is discarded into the inlet 8A in the abovementioned liquid draining part 8, whereby washing the hose 9, the washing cartridge 2, the hose 10, the needle 7A.
After completion of the washing, an analytical step is initiated, and the pair of clamp components 5,5 is moved to an unclamping position, and the cartridge holder 3 is moved to a certain position where a SPE cartridge 1 to be clamped can be clamped. Thereafter, the SPE cartridge 1 is clamped by the pair of upper/lower clamp components 5, 5, and is allowed to wait until the gas chromatograph 6 becomes in an operating condition. At the same time, a solvent suction signal is output to the liquid feed pump 4 and the needle 7A is raised to the origin position indicated in
Thus, a bracket 41 of the abovementioned syringe 7 in which the syringe 7 is attached as being capable of moving upward and downward without restriction to a vertically longitudinal slide rail 40 via the bracket 41 is connected to a timing belt 42 driven by the abovementioned needle driving electric motor 37 (ascending-descending driving means), and a basal end of a swinging bracket constituting the abovementioned liquid draining part 8 is attached as being capable of rotating integrally to a driving rotation shaft 38A of the abovementioned rotary solenoid 38. Accordingly, upon detection by a cartridge location detecting means 34 that a certain SPE cartridge 1 is positioned, the clamp driving electric motor 22 is driven to clamp the cartridge 1 by a pair of clamp components 5,5, which state, when detected by a clamp detection sensor 32, allows the rotary solenoid 38 to be driven whereby operating the swinging bracket 8 in a swinging manner from the receiving position indicated by a solid line in
While, in
The grasping means shown in
The abovementioned first grasping components 52, 52 is protruded downward from a grasping unit 53 consisting of a casing and capable of moving proximally or distally in a horizontal direction (right and left directions) to the condition of grasping the cartridge 1 and to the condition releasing the grasp, and the abovementioned grasping unit 53 is capable of being moved upward and downward by the ascending-descending unit 54 and also capable of moving forward and backward as a result of connection via a bracket 67 to an endless belt 66 driven by the electric motor 55. The abovementioned ascending-descending unit 54 is supported via a movably engaged slide component 65 on a slide rail 64 elongated from front to back as being fixed on the wall of the vertical wall component 63 built vertically. The abovementioned box 56 is mounted on a moving table 57, and this moving table 57 is fixed on a slide component 60 movably engaged with a slide rail 59 elongated from right to left placed on a fixation platform 58, and the moving table 57 is connected via a bracket (not shown) to the endless belt 62 driven by the electric motor 61, and thus by driving the electric motor 61, the moving table 57 can be moved in the right and left directions.
Accordingly, as shown in
As shown in
Residual pesticides in water were analyzed by a fortification recovery test.
10 ml of water was taken and supplemented with 2 ng of chlorpyrfos, and then loaded onto a SPE cartridge, which was flushed with a nitrogen gas to remove any moisture.
Gas chromatograph (GC/MS)
Storage chamber: For voluminous injection (using gastric bag-shaped insert)
Storage chamber temperature: 70° C.-120° C./min-250° C. (15 min)
Column oven temperature: 60° C. (3 min)-20° C./min-280° C. (5 min)
SPE cartridge: Solid phase C18
A SPE cartridge in which a sample was concentrated was placed in an injector part in a main unit. The eluate hexane was used to effect a direct elution into the gas chromatograph storage chamber, and a measurement was conducted using a GC/MS. As a result, chlorpyrfos exhibited a recovery of 90% or higher and a satisfactory chromatogram as shown in
Residual pesticides in water were analyzed by a fortification recovery test.
10 ml of water was taken and supplemented with 2 ng of bisphenol A, and then loaded onto a SPE cartridge, which was flushed with a nitrogen gas to remove any moisture. An eluate derivatizing reagent employed was N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) diluted with acetone.
Gas chromatograph (GC/MS)
Storage chamber: For voluminous injection (using gastric bag-shaped insert)
Storage chamber temperature: 70° C.-120° C./min-250° C. (15 min)
Column oven temperature: 60° C. (3 min)-20° C./min-280° C. (5 min)
SPE cartridge: Solid phase C18
Eluate: 10% BSTFA-containing acetone solution 40 μl
A SPE cartridge in which a sample was concentrated was placed in an injector part in a main unit. The eluate containing the derivatizing reagent (BSTFA) was used to effect a direct elution into the gas chromatograph storage chamber, and a measurement was conducted using a GC/MS. As a result, a derivatized bisphenol A-TMS was observed, showing that the derivatization was accomplished actually.
The method for analyzing organic compounds utilizing an inventive SPE cartridge is capable of measuring 5 to 20 certain residual pesticides and endocrine disturbing chemicals rapidly and accurately, and suitable for evaluating the safety of a target such as a food product rapidly. The constitution regarding to the ascending and descending of the needle 7A in an inventive analytical apparatus and the entire constitution for controlling are not limited to those shown in figures, and may vary as desired.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2004-250205 | Aug 2004 | JP | national |
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/JP05/10880 | 6/14/2005 | WO | 00 | 9/5/2007 |
