JP4456382B2 - Mass spectrometry reagent for measuring heavy metal ions, mass spectrometry method for heavy metal ions and mass spectrometry system for heavy metal ions analysis using the same - Google Patents

Description

The present ^{invention, Pb 2+, Cd 2+, Hg} 2+, Zn 2+, Ni 2+, is used to measure by mass spectrometry divalent heavy metal ions such as Cu ²⁺ and Co ^2+, 2 The present invention relates to a mass spectrometry reagent for measuring heavy metal ions.

  Causes of soil and groundwater contamination by heavy metals include leakage of raw materials including target substances and chemicals, leakage during storage and manufacturing processes, smoke drop, inadequate drainage of underground water or landfill disposal of waste. The behavior of heavy metals, etc. in the soil varies depending on their physicochemical properties and the nature of the soil that is the medium, but in general, heavy metals are difficult to dissolve in water and are easily adsorbed by the soil, It exists in the soil near the surface of the earth and often does not diffuse deep. However, when a load exceeding the adsorption capacity of the soil occurs or a substance having high water solubility and high mobility such as hexavalent chromium, it may diffuse to the deep underground with underground penetration such as rainwater.

  Against this background, there is a need for an analysis method that can easily and accurately grasp the state of heavy metal contamination. Conventional methods include ICP and atomic absorption methods as high-sensitivity measurement methods, but they are dangerous and unfavorable at the site because of the use of frames, and problems such as complicated operation and high running costs have been pointed out. Yes. A colorimetric analysis method can be cited as a simple analysis method, but the problem of low sensitivity has been pointed out.

  On the other hand, it has been reported that the complex formation ability of crown ether skeletons (including sulfur, nitrogen or oxygen atoms as heteroatoms) with heavy metal ions is measured by electrospray ionization mass spectrometry (ESI-MS) (non-patented) Reference 1). However, this method has a problem that the 1: 1 complex peak cannot be detected, the counter ion is added and detected, and the sensitivity is poor.

Sheldon M. et al., Anal. Chem. 2002, 74, 4423-4433

An object of the present invention is to provide a means for measuring divalent heavy metal ions easily and with high sensitivity without using a frame.

As a result of earnest research, the inventors of the present application have a structure that forms a complex with a divalent heavy metal ion to be measured in a test sample and an ionic functional group that becomes an anion in a solution in one molecule. The present invention was completed by finding that divalent heavy metal ions can be measured easily and with high sensitivity by using a compound containing the compound as a probe for mass spectrometry.

That is, the present invention provides the following general formula [I]
R ₁ −A−R ₂ [I]
(In the formula, R ₁ is a structure that forms a complex with a divalent heavy metal ion to be measured in a test sample , R ₂ is an ionic functional group that becomes a monovalent negative ion in solution, and A is any (Represents the spacer part)
Providing divalent heavy metal ions for measurement mass analysis reagent having a structure represented by the. Moreover, this invention provides the mass spectrometry method of a bivalent heavy metal ion including reacting the reagent of the said invention and the bivalent heavy metal ion in a test sample, and mass-analyzing a product. . Furthermore, the present invention aspirates the reagent container for storing the reagent of the present invention, the sample container for storing the test sample, the reagent stored in the reagent container and the test sample stored in the sample container. A suction means for suctioning the reagent housed in the reagent container and the test sample housed in the sample container via the tip of the suction means, and the middle of the suction pipe A first valve provided on the first valve, a liquid feed pump connected to the first valve via a liquid extrusion pipe, and a liquid mixing means connected to the first valve via a liquid feed pipe And a mass spectrometry system for analyzing divalent heavy metal ions, comprising a mass spectrometer connected downstream of the liquid mixing means.

When the reagent of the present invention is used, the charge of the divalent heavy metal ion is reduced by one due to the negative charge caused by the ionic functional group, and the complex formed by the complex formation is observed as an almost single peak. Therefore, divalent heavy metal ions can be measured with high sensitivity by simple mass spectrometry without using a frame. Further, the present invention, a reagent of the present invention, high-sensitivity divalent mass spectrometry and mass spectrometry system heavy metal ions is provided.

As described above, the divalent heavy metal ion measurement mass spectrometry reagent of the present invention has a structure represented by the general formula [I].

In the general formula [I], R ₁ represents a structure that forms a complex with a divalent heavy metal ion to be measured in the test sample . Preferred examples of such a structure include structures represented by the following formulas 1 to 5.

(In the Formula 1 to _{5, R 3, R 4,} R 5, R 6, R 7, R 8, R 9, R 10, R 11, R 12, R 13 and R _14, independently of one another, hydrogen An atom, a linear or branched alkyl group having 1 to 10 carbon atoms (preferably 1 to 4 carbon atoms), an amino group, a nitro group, or a halogen, and k, l, m, and n are independently of each other Represents an integer of 1 to 4.)
In general formula 2, it is a nitrogen atom that is bonded to the spacer moiety.

Of the structures represented by the general formulas 1 to 5, those represented by the formula 3 are particularly preferable, and R ₇ , R ₈ , R ₉ , R ₁₀ , R ₁₁ , R ₁₂ , R ₁₃ and R _{14 in the} formula 3 are particularly preferable. Is preferably a hydrogen atom.

In the general formula [I], R ₂ is an ionic functional group that becomes an anion, preferably a monovalent anion, in a solution, preferably an aqueous solution. Preferred examples include a carboxyl group or a salt thereof, a sulfone group. Alternatively, a salt thereof, a hydroxyl group or a salt thereof can be given. Among these, a carboxyl group or a salt thereof is particularly preferable. These preferred ionic functional groups, in particular carboxyl groups or salts thereof, not only reduce the charge of the divalent heavy metal ion monovalently by the charge of the anion, but are also stable and selective with the divalent heavy metal ion. This contributes to complex formation, and in turn contributes to highly sensitive measurement of divalent heavy metal ions.

In the general formula [I], A is an optional spacer portion. Reagents of the invention, form a divalent heavy ions and complexing by R ₁ to be measured in a test sample, since it is intended to monovalent from divalent to charge of the complex by R _2, and R ₁ A located between R ₂ can take any structure. Preferred examples of A include a linear or branched alkylene group having 1 to 10 carbon atoms (more preferably 1 to 4 carbon atoms), and a part of the alkylene group is an amino group, a halogen, an ether and / or a carbonyl group. And a substituted phenylene group in which a part of the phenylene group is substituted with an amino group, a halogen and / or a nitro group. The alkylene group substituted with ether means one in which a carbon atom in the alkylene chain is replaced with —O—. Similarly, an alkylene group substituted with a carbonyl group means one in which a carbon atom in the alkylene chain is replaced with —CO—. The number of substituents in the substituted alkylene group or substituted phenylene group may be one or more. Among these substituents, an alkylene group having 1 to 10 carbon atoms, more preferably 1 to 4 carbon atoms is preferable.

The divalent heavy metal ion that can be measured by the reagent of the present invention is not particularly limited as long as it is a divalent heavy metal ion capable of complexing with R ₁ , but a divalent cation is preferred. In recent years, time-of-flight mass spectrometers have been widely used as detectors for mass spectrometers. A time-of-flight mass spectrometer can detect monovalent ions more sensitively than multivalent ions such as divalent heavy metal ions. When a divalent cation is complex-bonded with the reagent of the present invention having a monovalent anionic functional group which is a preferable R ₂ , the charge of the divalent heavy metal ion is reduced by one valence, and the total number of complexes formed The charge is monovalent and can be measured with high sensitivity by a time-of-flight mass spectrometer. Preferred examples of the divalent heavy metal ions reagent is measured using the present ^{invention, Pb 2+, Cd 2+, Hg} 2+, Zn 2+, it is selected from Ni ^2+, Cu ²⁺ and Co ²⁺ At least one can be exemplified. As specifically described in the following examples, when the reagent of the present invention is used, a mixture of a plurality of divalent heavy metal ions can be separately measured. In the present specification, “measurement” includes both detection (qualitative analysis) and quantitative analysis.

The reagent of the present invention has a known structure of R ₁ , A and R ₂ , and can be obtained by combining them. Therefore, it can be synthesized based on a conventional method in the field of organic synthesis, and is preferable. Specific examples are detailed in the examples below.

When measuring a divalent heavy metal ion using the reagent of the present invention, first, a sample containing a divalent heavy metal ion is mixed to form a complex of the divalent heavy metal ion and the reagent, and then generated. The complex is subjected to mass spectrometry. Reaction of the reagent with a divalent heavy metal ions of the present invention, 1: Since proceeds well at 1 stoichiometric amount, the molar concentration of the reagent used is not particularly limited, the expected divalent heavy metal ions It may be about the same as the molar concentration. Therefore, an upper limit of the expected range of the molar concentration of divalent heavy metal ions or a reagent having a molar concentration slightly higher than that may be used. Alternatively, when investigating whether or not the divalent heavy metal ion concentration is below the allowable reference value, a reagent having a molar concentration that is the same as or slightly higher than the molar concentration of the reference value can be used. The temperature of the complex-forming reaction between the reagent and the divalent heavy metal is not particularly limited, but it reacts well at room temperature. In addition, since the reaction takes place quickly, the reaction time is not particularly limited, and is usually about 1 to 30 seconds, and in many cases about 2 to 3 seconds. The reaction solvent is preferably water or an organic solvent mixed with water at an arbitrary ratio, such as methanol or ethanol, and most preferably methanol. The mixing is preferably performed with stirring.

  The sample after the complex formation reaction can be directly subjected to mass spectrometry. Mass spectrometry itself can be performed by a conventional method using a commercially available mass analyzer. As the mass spectrometry, any known method may be used, but ESI-MS is preferable, and the detector is preferably a time-of-flight mass spectrometer.

Since the molecular weight of the reagent used is known and the atomic weight of the divalent heavy metal ion is also known, the divalent heavy metal ion can be identified and measured by measuring the m / z of the complex by mass spectrometry. it can. In addition, as specifically shown in the following examples, even when a plurality of types of divalent heavy metal ions are included in the sample, since the atomic weight of each divalent heavy metal ion is different, a plurality of observations are made at different positions. Each divalent heavy metal ion can be separately measured from the m / z peak.

Using the reagent of the present invention, the anionic functional group R ₂ to cancel the charge of divalent heavy metal ions, since the charge of the complex can be monovalent, high using a time-of-flight mass spectrometer Mass spectrometry can be performed with sensitivity. Further, as the anionic functional group R _2, when using the preferred functional group described above, anionic functional groups, also contributes to the complex formation, a stable selective (i.e., one of the divalent Complex formation is achieved (only one type of complex is formed for heavy metal ions). For this reason, it is possible to accurately measure divalent heavy metal ions in a sample, and even when a plurality of types of divalent heavy metal ions are contained in one sample, each divalent heavy metal ion is fractionated. Can be measured accurately.

The present invention also provides a method for mass spectrometry of divalent heavy metal ions, which comprises reacting the reagent of the present invention with a divalent heavy metal ion in a test sample and mass analyzing the product. This can be done as described above.

The mass spectrometry method of the present invention can be performed manually using a general-purpose mass spectrometer using the reagent of the present invention, but the present invention is also suitable for automation using the reagent of the present invention. A mass spectrometry system for analyzing divalent heavy metal ions is also provided.

  This mass spectrometry system will be described with reference to FIGS. The system for mass spectrometry includes a reagent container 10 for storing a reagent and sample containers 12 and 12 'for storing a test sample. There may be one or a plurality of sample containers for storing the test samples. If there are a plurality of sample containers, a plurality of test samples can be analyzed. The mass spectrometric system includes suction means 14 for sucking the reagent contained in the reagent container and the test sample contained in the sample container. The suction means 14 is, for example, a syringe. A suction tube 16 is connected to the suction means 14, and the suction tube 16 sucks the reagent stored in the reagent container and the test sample stored in the sample container through its tip. A needle may be attached to the distal end portion of the suction tube 16, and in that case, a sample container or reagent container film in which the top of the container is covered with a film is pierced with the needle to suck the liquid inside. Can do. A first valve 18 is provided in the middle of the suction pipe 16. A liquid feed pump 22 is connected to the first valve 18 via a liquid extrusion pipe 20. A liquid mixing means 26 is connected to the first valve 18 via a liquid feed pipe 24. The liquid mixing unit 26 is not limited as long as it is a unit that can mix the sample solution and the reagent solution. For example, a simple and preferable example can be a mixing column. In the mixing column, glass beads are packed inside the column, and mixing of the two liquids takes place while the two liquids pass through them. A mass spectrometer is connected downstream of the liquid mixing means 26, which may be general purpose. Furthermore, in the preferred specific example shown in FIGS. 21 and 22, a second valve 28 is provided in the middle of the liquid extruding pipe 20 between the liquid feed pump 22 and the first valve 18, In the middle of the suction pipe 16, a third valve 30 is provided between the suction means 14 and the first valve 18, and these are connected by a second liquid extrusion pipe 32. Yes.

  In operation, first, the aspirating means 14 is operated to aspirate the reagent solution of the present invention from the reagent container 10. At this time, the first valve 18 and the third valve 30 are set on the side where the liquid is sucked by the suction means 14. The liquid feed pump 22 is stopped. Next, in this state, the tip of the suction tube 16 (or a needle if a needle is attached) is moved to suck the sample liquid from the sample container 12 or 12 '. The flow of fluid during suction is shown in FIG. 21 as a black triangle.

  When extruding the aspirated reagent liquid and test sample liquid into the mass spectrometer, the aspirating means 14 is stopped and the liquid feed pump 22 is operated to send out the solvent. As the solvent, like the reaction solvent, water or an organic solvent mixed with water at an arbitrary ratio such as methanol or ethanol is preferable, and methanol is most preferable. Further, the first, second and third valves are switched so that the fluid moves in the direction indicated by the black triangle in FIG. Then, the reagent solution and the test sample solution that are in the middle of the suction tube 16 are sent to the mass spectrometer via the liquid mixing means as shown by the black triangles in FIG. As described above, the reagent solution and the test sample solution are mixed when passing through the liquid mixing means. When the reagent solution and the test sample solution are mixed, the above-described binding reaction occurs, and the binding product is subjected to mass spectrometry as described above.

EXAMPLES Hereinafter, the present invention will be described more specifically based on examples. However, the present invention is not limited to the following examples.

  Prior to the description of each example, the reagent, separation generation method, and compound identification method employed in each example will be described.

1. Reagents The reagents and solvents used were special grades of commercial products manufactured by Tokyo Kasei, Kanto Chemical, Wako, Aldrich, Fulka and Merck. Dry methylene chloride is obtained by distilling industrial grade water and adding dry molecular sieves to remove moisture.

2 Separation and purification
2.1 Column Chromatography Aluminum oxide 60 aktiv neutral made by Merck was used for the stationary phase. For the mobile phase, primary equivalent n-hexane, ethyl acetate and chloroform were used.

2.2 Preparative TLC
Merck TLC plates Aluminum oxide 60 F254 and Merck TLC plates silica gel 60 F245 with concentrating zone were used for the stationary phase. As the mobile phase, primary equivalent n-hexane and ethyl acetate were used.

2.3 Thin layer chromatography (TLC)
Merck TLC plates Aluminum oxide 60 F254 and Merck TLC plates silica gel 60 F245 were used for the stationary phase. For the mobile phase, primary equivalent n-hexane, ethyl acetate and chloroform were used. The compound was detected using iodine and UV absorption (254 nm).

3 Compound identification
3.1 Nuclear magnetic resonance spectrometer (NMR)
As the NMR apparatus, LNM-LA300FT-NMR and JNM-GSX270FT-NMR manufactured by JEOL Ltd. were used. As a heavy solvent, chloroform-d and acetone-d6 manufactured by Merck were used, and tetramethylsilane (TMS) was used as an internal standard. Signal abbreviations are as follows.
s: singlet, d: doublet, t: triplet, q: quartet, m: multiplet

3.2 Mass spectrometer The ionization method of the mass spectrometer is the ESI method, and the mass spectrometer is TOFMS. A Mariner Biospectrometry Workstation from Applied Biosystems was used. TOF is a reflector type.

Synthesis of KHM-1 KHM-1 which is a reagent of the present invention having a structure represented by the following formula was synthesized according to the following scheme.

Synthesis scheme

(1) Synthesis of 3- (bis-pyridin-2-ylmethyl-amino) -propionic acid methyl ester

3-Bromo-propionic acid methyl ester (2.0 ml, 12.0 mmol, 1 eq.) Was placed in an Ar-substituted 300 ml three-necked eggplant flask, and acetonitrile (100 ml) was added. 2,2′-Dipicolylamine (1; 7.20 ml, 35.9 mmol, 3 eq.) And potassium carbonate (3.32 g, 24.0 mmol, 2 eq.) Were added and refluxed for 19 hours. After the reaction was stopped by adding H ₂ O, the solvent was distilled off under reduced pressure. The obtained solid was dissolved in ethyl acetate, washed with H ₂ O, and the organic layer was dried with sodium sulfate. The solvent was distilled off under reduced pressure, and the residue was purified by column chromatography (Al ₂ O ₃ , n-hexane: ethyl acetate = 1: 1 v / v) to obtain a brown oily compound (2; 2.83 g, 82.9%) It was.

TLC (Al ₂ O ₃ ); R _f = 0.8 (n-hexane: ethyl acetate = 1: 1 v / v)
¹ H-NMR (300 MHz, CD ₃ OD, TMS, rt, δ / ppm) 2.57 (t, J = 7.2 Hz, 2H, -CH _2- ), 2.94 (t, J = 7.2 Hz, 2H, -CH _2- ), 3.64 (s, 3H, -CH ₃ ), 3.83 (s, 4H, -CH _2- ), 7.20 (t, J = 6.3 Hz, 2H, ArH), 7.49 (d, J = 4.1 Hz, 2H, ArH), 7.66 (d, J = 5.5 Hz, 2H, ArH), 8.52 (d, J = 4.9 Hz, 2H, ArH)

(2) Synthesis of 3- (bis-pyridin-2-ylmethyl-amino) -propionic acid

In a 50 ml two-necked eggplant flask, put 3- (bis-pyridin-2-ylmethyl-amino) -propionic acid methyl ester (2; 2.00 ml, 7.01 mmol), methanol (50 ml), 10w% aq. NaOH ( 3 ml) was added and refluxed for 24 hours. After the reaction was stopped by adding H ₂ O, most of the solvent was distilled off under reduced pressure. 1M aq. HCl (15 ml) was added to adjust the pH to 5, and then the solvent was distilled off under reduced pressure. Ethanol was added and the deposited precipitate was filtered off, and the filtrate was concentrated under reduced pressure to obtain a white crystalline compound (3; 1.82 g, 95.9%).

TLC (Al ₂ O ₃ ); R _f = 0.0 (n-hexane: ethyl acetate = 1: 1 v / v)
ESI-TOFMS (+)
[M + H] ⁺ = 272.2
¹ H-NMR (300 MHz, CD ₃ OD, TMS, rt, δ / ppm) 2.71 (t, J = 6.3 Hz, 2H, -CH _2- ), 3.12 (t, J = 6.2 Hz, 2H, -CH _2- ), 4.05 (s, 4H, -CH _2- ), 7.26 (t, J = 6.2 Hz, 2H, ArH), 7.44 (d, J = 7.8 Hz, 2H, ArH), 7.71 (t, J = 6.0 Hz, 2H, ArH), 8.58 (d, J = 4.1 Hz, 2H, ArH)

Comparative Example 1
Comparative compounds were synthesized according to the following synthesis scheme. The comparative compound has a structure obtained by removing the carboxyl group from KHM-1 synthesized in Example 1.

1-Bromo-propane (1.0 ml, 8.13 mmol, 1 eq.) Was placed in an Ar-substituted 100 ml neck flask, and acetonitrile (50 ml) was added. 2,2′-Dipicolylamine (4; 4.90 ml, 24.4 mmol, 3 eq.) And potassium carbonate (2.26 g, 1.63 mmol, 2 eq.) Were added, and the mixture was refluxed for 24 hours. After suction filtration, the solvent was distilled off under reduced pressure. Purification was performed by column chromatography (Al ₂ O ₃ , n-hexane: ethyl acetate = 1: 1 v / v) to obtain a reddish brown crystalline compound (5; 1.67 g, 85.2%).

TLC (Al ₂ O ₃ ); R _f = 0.8 (n-hexane: ethyl acetate = 1: 1 v / v)
ESI-TOFMS (+)
[M + H] ⁺ = 242.3
^{1 H-NMR (300 MHz,} CD 3 OD, TMS, rt, δ / ppm) 0.87 (t, J = 7.4 Hz, 3H, -CH 3), 1.55 (m, 2H, -CH 2 -), 2.51 ( t, J = 7.4 Hz, 2H, -CH _2- ), 3.81 (s, 4H, -CH _2- ), 7.14 (t, J = 6.7 Hz, 2H, ArH), 7.56 (d, J = 7.8 Hz, 2H, ArH), 7.66 (t, J = 7.6 Hz, 2H, ArH), 8.52 (d, J = 5.6 Hz, 2H, ArH)

Using KHM-1 synthesized in Example 1, Cu ²⁺ ions were subjected to mass spectrometry as follows.

(1) Measurement conditions of mass spectrometer Fig. 1 shows the configuration of the apparatus. Using a syringe pump, the mobile solvent (MeOH) is constantly flowed at a flow rate of 10.0 μL / min. 20 μL of the sample solution is introduced from the injector using a microsyringe. The sample solution is directed to the mass spectrometer along the flow of the mobile solvent. The setting conditions of the mass spectrometer are as follows.
Model: Mariner manufactured by Applied Biosystems
Ionization method: Electrospray ionization detection method: Time-of-flight detection method Mobile solvent: Methanol spray tip potential: 3500 V
Nozzle potential: 90 V
Detector voltage: 2280 V
Quad RF voltage: 700 V
Flow rate of Nebulizer gas (N ₂ ): 0.25 L / min Flow rate of Auxiliary gas (N ₂ ): 1.0 L / min Temperature of the counter stream: 160 ℃

(2) Operation method Add 6.0 × 10 ^-10 M CuCl ₂ · 2H ₂ O and 6.0 × 10 ^-10 M KHM-1 to the screw tube (solvent is methanol), and stir at room temperature for 10 seconds. Measurement was performed with the analyzer under the above measurement conditions.

(3) Results The results are shown in FIG. FIG. 2 is a plot of m / z on the horizontal axis and relative intensity (%) on the vertical axis. In FIG. 2, M means a reagent (the same applies to FIG. 3 and subsequent figures). As shown in FIG. 2, Cu2 ⁺ , which is a divalent heavy metal ion, was detected with high sensitivity as a monovalent metal complex peak using KHM-1 which is the reagent of the present invention.

Comparative Example 2
The same operation as in Example 2 was performed except that the comparative compound synthesized in Comparative Example 1 was used as a reagent. The results are shown in FIG. As shown in FIG. 3, three types of chemical species containing Cu ions were generated, and the Cu complex could not be detected with high sensitivity as a monovalent metal complex.

The same operation as in Example 2 was performed except that ZnCl ₂ was used instead of cupric chloride dihydrate, and Zn ²⁺ ions were ^subjected to mass spectrometry. The results are shown in FIG. As shown in FIG. 4, Zn ²⁺ , which is a divalent heavy metal ion, could be detected with high sensitivity as a monovalent metal complex peak.

Comparative Example 3
The same operation as in Example 3 was performed except that the comparative compound synthesized in Comparative Example 1 was used as a reagent. The results are shown in FIG. As shown in FIG. 5, complex formation with Zn ions is less than in Example 3, and a plurality of chemical species containing Zn ions are generated, and the Zn complex is detected as a monovalent metal complex with high sensitivity. I couldn't.

The same operation as in Example 2 was performed except that Ni (NO ₃ ) ₂ .6H ₂ O was used in place of cupric chloride dihydrate, and Ni ²⁺ ions were ^subjected to mass spectrometry. The results are shown in FIG. As shown in FIG. 6, Ni ²⁺ , which is a divalent heavy metal ion, could be detected with high sensitivity as a monovalent metal complex peak.

Comparative Example 4
The same operation as in Example 4 was performed except that the comparative compound synthesized in Comparative Example 1 was used as a reagent. The results are shown in FIG. As shown in FIG. 7, a plurality of chemical species containing Ni ions were generated, and the Ni complex could not be detected with high sensitivity as a monovalent metal complex.

The same operation as in Example 2 was performed except that CoCl ₂ .6H ₂ O was used instead of cupric chloride dihydrate, and Co ²⁺ ions were ^subjected to mass spectrometry. The results are shown in FIG. As shown in FIG. 8, Co ²⁺ , which is a divalent heavy metal ion, could be detected with high sensitivity as a monovalent metal complex peak.

Comparative Example 5
The same operation as in Example 5 was performed except that the comparative compound synthesized in Comparative Example 1 was used as a reagent. The results are shown in FIG. As shown in FIG. 9, a plurality of chemical species containing Co ions were generated, and the Co complex could not be detected with high sensitivity as a monovalent metal complex.

A Pb ²⁺ ion was ^subjected to mass spectrometry analysis in the same manner as in Example 2 except that (CH ₃ COO) ₂ Pb · 3H ₂ O was used instead of cupric chloride dihydrate. The results are shown in FIG. As shown in FIG. 10, Pb ²⁺ , which is a divalent heavy metal ion, could be detected with high sensitivity as a monovalent metal complex peak.

The same operation as in Example 2 was performed except that CdCl ₂ was used in place of cupric chloride dihydrate, and Cd ²⁺ ions were ^subjected to mass spectrometry. The results are shown in FIG. As shown in FIG. 11, Cd ²⁺ , which is a divalent heavy metal ion, could be detected with high sensitivity as a monovalent metal complex peak.

The same operation as in Example 2 was performed except that (CH ₃ COO) ₂ Hg was used in place of cupric chloride dihydrate, and Hg ²⁺ ions were subjected to mass spectrometry. The results are shown in FIG. As shown in FIG. 12, Hg ²⁺ , which is a divalent heavy metal ion, could be detected with high sensitivity as a monovalent metal complex peak.

After adding 4.0 × 10 ⁻¹¹ M to 1.6 × 10 ⁻⁹ M CdCl ₂ and 8.0 × 10 ⁻¹⁰ M KHM-1 to the screw tube (solvent is methanol), stirring at room temperature for 10 seconds, mass spectrometer The measurement was carried out under the above measurement conditions. As an internal standard, 1-methyl-pyridinium was used at a concentration of 1.1 × 10 ⁻⁹ M. In preparing the calibration curve, the peak intensity of the complex in the mass spectrum was calculated as a relative ratio to the peak intensity of the internal standard substance (relative intensity = peak intensity of the complex ÷ peak intensity of the internal standard substance).

The results are shown in FIG. As shown in FIG. 13, the relative intensity increases depending on the Cd ²⁺ ion concentration in the range of 1.6 × 10 ⁻¹⁰ M to 8.0 × 10 ⁻¹⁰ M, and at least Cd ²⁺ is increased in this range. It was confirmed that quantification was possible. This concentration range is smaller than the upper limit of the environmental standard value, and it is possible to check whether the Cd ²⁺ concentration is within the environmental standard range by this method. In this example, since the concentration of KHM-1 was 8.0 × 10 ⁻¹⁰ M, the relative strength did not increase even if the heavy metal ion concentration exceeded this, but if a higher concentration of KHM-1 was used, High concentrations of heavy metal ions can also be quantified (same in the following examples).

The same operation as in Example 9 was performed, except that 1.6 × 10 ^{−11 to} 1.6 × 10 ⁻⁹ M (CH ₃ COO) ₂ Pb · 3H ₂ O was used. The results are shown in FIG. As shown in FIG. 14, the relative intensity increases depending on the Pb ²⁺ ion concentration in the range of 8.0 × 10 ⁻¹⁰ M to 1.6 × 10 ⁻⁹ M, and at least Pb ²⁺ is increased in this range. It was confirmed that quantification was possible. This concentration range is smaller than the upper limit of the environmental standard value, and it is possible to check whether or not the Pb ²⁺ concentration is within the environmental standard range by this method.

The same operation as in Example 9 was performed except that 4.0 × 10 ^{−12 to} 1.6 × 10 ⁻⁹ M CuCl ₂ .2H ₂ O was used. The results are shown in FIG. As shown in FIG. 15, the relative intensity increases depending on the Cu ²⁺ ion concentration in the range of 1.6 × 10 ⁻¹¹ M to 8.0 × 10 ⁻¹⁰ M, and at least Cu ²⁺ is absorbed in this range. It was confirmed that quantification was possible. This concentration range is smaller than the upper limit of the environmental standard value, and it is possible to check whether or not the Cu ²⁺ concentration is within the environmental standard range by this method.

The same operation as in Example 9 was performed, except that 4.0 × 10 ^{−13 to} 1.6 × 10 ⁻⁹ M ZnCl ₂ was used. The results are shown in FIG. As shown in FIG. 16, the relative intensity increases depending on the Zn ²⁺ ion concentration in the range of 1.6 × 10 ⁻¹⁰ M to 8.0 × 10 ⁻¹⁰ M, and at least Zn ²⁺ is within this range. It was confirmed that quantification was possible.

The same operation as in Example 9 was performed, except that 4.0 × 10 ^{−13 to} 1.6 × 10 ⁻⁹ M Ni (NO ₃ ) ₂ .6H ₂ O was used. The results are shown in FIG. As shown in FIG. 17, in the range of 8.0 x 10 ^-10 M from 1.6x 10 ^-11 M, and the relative intensity increases depending on the Ni ²⁺ ion concentration, the Ni ²⁺ least in this range It was confirmed that quantification was possible.

The same operation as in Example 9 was performed except that 1.6 × 10 ^{−12 to} 1.6 × 10 ⁻⁹ M CoCl ₂ .6H ₂ O was used. The results are shown in FIG. As shown in FIG. 18, in the range of 8.0 x 10 ^-10 M from 1.6x 10 ^-11 M, and the relative intensity increases depending on the Co ²⁺ ion concentration, the Co ²⁺ least in this range It was confirmed that quantification was possible.

With ^{1.6 x 10 -10 ~1.6 x 10 -9} M of (CH ₃ COO) ₂ Hg, except that the concentration of internal standard and 2.2 x 10 ^-9 M, The same procedure was followed as in Example 9 . The results are shown in FIG. As shown in FIG. 19, the relative intensity increases depending on the Hg ²⁺ ion concentration in the range of 4.0 × 10 ⁻¹⁰ M to 1.6 × 10 ⁻⁹ M, and at least in this range, Hg ²⁺ is increased. It was confirmed that quantification was possible. This concentration range is larger than the upper limit of the environmental standard value. However, if the sample is concentrated and analyzed, Hg ^{2+ having} a concentration below the upper limit of the environmental standard value can be measured.

Various ions with an ion concentration of 4.0 x 10 ^-10 M using 2.0 x 10 ^-9 M KHM-1 (compounds used are Cu ²⁺ , Cd ²⁺ , Ni ²⁺ , Co ²⁺ , Ni ²⁺ , Pb ²⁺ was the same as in Examples 2 to 8 and the internal standard was 2.0 × 10 ⁻⁹ M), and mass spectrometry was performed in the same manner as in Example 2. The results are shown in FIG. As shown in FIG. 20, each heavy metal ion was observed as a peak of a monovalent metal complex, and it was possible to measure each heavy metal ion separately. Table 1 below shows the error of each peak observed in this example, calculated based on the results of quantitative analysis in Examples 9 to 14. As shown in Table 1, even when a mixture of heavy metal ions was measured simultaneously, it could be measured with an error range of about 3%.

  By using the reagent of the present invention, various heavy metal ions can be easily measured with high sensitivity. Therefore, the reagent of the present invention is useful as a reagent for measuring various heavy metal ions by mass spectrometry.

It is a figure which shows typically the structure of the mass spectrometer used in the Example of this invention. In Example 2 of this invention, it is a figure which shows the result of having conducted mass spectrometry of Cu2 ⁺ . In the comparative example 2 of this invention, it is a figure which shows the result of having mass-analyzed Cu2 ⁺ . In Example 3 of this invention, it is a figure which shows the result of having mass-analyzed Zn2 ⁺ . In the comparative example 3 of this invention, it is a figure which shows the result of having mass-analyzed Zn2 ⁺ . In Example 4 of this invention, it is a figure which shows the result of having mass-analyzed Ni ^<2+> . In the comparative example 4 of this invention, it is a figure which shows the result of having mass-analyzed Ni2 ⁺ . In Example 5 of this invention, it is a figure which shows the result of having carried out mass spectrometry of Co2 ⁺ . In the comparative example 5 of this invention, it is a figure which shows the result of having carried out mass spectrometry of Co2 ⁺ . In Example 6 of this invention, it is a figure which shows the result of having conducted mass spectrometry of Pb2 ⁺ . In Example 7 of this invention, it is a figure which shows the result of having carried out mass spectrometry of Cd < ²⁺ >. In Example 8 of this invention, it is a figure which shows the result of having conducted mass spectrometry of Hg < ²⁺ >. In Example 9 of this invention, it is a figure which shows the result of having quantitatively analyzed Cd <^2+> by mass spectrometry. In Example 10 of this invention, it is a figure which shows the result of having quantitatively analyzed Pb <^2+> by mass spectrometry. In Example 11 of this invention, it is a figure which shows the result of having quantitatively analyzed Cu ^<2+> by mass spectrometry. In Example 12 of this invention, it is a figure which shows the result of having quantitatively analyzed Zn ^<2+> by mass spectrometry. In Example 13 of this invention, it is a figure which shows the result of having quantitatively analyzed Ni ^<2+> by mass spectrometry. In Example 14 of this invention, it is a figure which shows the result of having quantitatively analyzed Co ^<2+> by mass spectrometry. In Example 15 of this invention, it is a figure which shows the result of having quantitatively analyzed Hg <^2+> by mass spectrometry. In Example 16 of this invention, it is a figure which shows the result of having quantitatively analyzed the mixture of various heavy metal ions by mass spectrometry. It is a figure which shows typically the state at the time of the liquid suction of the preferable specific example of the mass spectrometry system for heavy metal ion analysis of this invention. It is a figure which shows typically the state at the time of the liquid extrusion of the preferable specific example of the mass spectrometry system for heavy metal ion analysis of this invention.

Explanation of symbols

DESCRIPTION OF SYMBOLS 10 Reagent container 12 Sample container 12 'Sample container 14 Suction means 16 Suction pipe 18 1st valve 20 Liquid extrusion pipe | tube 22 Liquid feed pump 24 Liquid feed pipe 26 Liquid mixing means 28 2nd valve 30 3rd valve 32 3rd 2 Liquid extrusion tube

Claims (15)

The following general formula [I]
R ₁ AR ₂ [I]
(In the formula, R ₁ is a structure that forms a complex with a divalent heavy metal ion to be measured in a test sample , R ₂ is an ionic functional group that becomes a monovalent negative ion in solution, and A is any (Represents the spacer part)
A mass spectrometric reagent for measuring divalent heavy metal ions having a structure represented by: The reagent according to claim _1, wherein R ₁ is represented by any one of the following formulas 1 to 5.

(In the general formulas 1 to 5, R ₃ , R ₄ , R ₅ , R ₆ , R ₇ , R ₈ , R ₉ , R ₁₀ , R ₁₁ , R ₁₂ , R ₁₃ and R ₁₄ are each independently hydrogen. An atom, a linear or branched alkyl group having 1 to 10 carbon atoms, an amino group, a nitro group or a halogen, and k, l, m and n each independently represent an integer of 1 to 4) The reagent according to claim 2, wherein R ₁ is represented by Formula 3. The reagent according to claim 3, wherein, in the formula 3, R ₇ , R ₈ , R ₉ , R ₁₀ , R ₁₁ , R ₁₂ , R ₁₃ and R ₁₄ are hydrogen atoms. The reagent according to any one of claims 1 to 4, wherein R ₂ is a carboxyl group or a salt thereof, a sulfone group or a salt thereof, or a hydroxyl group or a salt thereof. The reagent according to claim 5, wherein R ₂ is a carboxyl group or a salt thereof.   A is a linear or branched alkylene group having 1 to 10 carbon atoms, a substituted alkylene group in which part of the alkylene group is substituted with an amino group, a halogen, an ether and / or a carbonyl group, a phenylene group, or phenylene The reagent according to any one of claims 1 to 6, which is a substituted phenylene group in which a part of the group is substituted with an amino group, a halogen and / or a nitro group.   The reagent according to claim 7, wherein A is an alkylene group. The reagent according to claim 8, which is a carboxylic acid having the following structure or a salt thereof.

The reagent according to any one of claims 1 to 9, wherein the divalent heavy metal ion is a divalent cation. The reagent according to claim 10, wherein the divalent cation is at least one selected from Pb ²⁺ , Cd ²⁺ , Hg ²⁺ , Zn ²⁺ , Ni ²⁺ , Cu ²⁺ and Co ²⁺ . And reagent according to any one of claims 1 to 11, by reacting a divalent heavy metal ions in a test sample, the product comprises mass spectrometry, divalent mass spectrometric method heavy metal ions . A reagent container containing the reagent according to any one of claims 1 to 11, a sample container containing a test sample, a reagent contained in the reagent container, and a test sample contained in the sample container A suction means for aspirating the reagent, a suction tube connected to the suction means for sucking the reagent contained in the reagent container and the test sample contained in the sample container via the tip, and the suction A first valve provided in the middle of the pipe, a liquid feed pump connected to the first valve via a liquid extrusion pipe, and a liquid connected to the first valve via a liquid feed pipe A bivalent heavy metal ion analysis mass spectrometry system comprising a mixing means and a mass spectrometer connected downstream of the liquid mixing means.   A second valve is provided in the middle of the liquid extruding pipe and between the liquid feed pump and the first valve, and in the middle of the suction pipe, the suction means and the first valve The mass spectrometry system according to claim 13, wherein a third valve is provided between the second valve and the third valve, and the second valve and the third valve are connected by a second liquid extrusion tube. A mass spectrometry system using the mass spectrometry method for divalent heavy metals according to claim 12.

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