JP2888092B2 - Pesticide analysis method - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for analyzing various kinds of pesticides by high performance liquid chromatography (hereinafter abbreviated as HPLC).

[0002]

BACKGROUND OF THE INVENTION At a golf course, pesticides (ashram, oxine copper, mecoprop, thiuram,
Gas chromatography and HPLC methods are used in water testing methods to check for residual water contaminants such as sidulone, iprodione, chlorothalonil, pencyclone, benzulide, etc.) in the water supply system. And thiuram are analyzed by the HPLC method.

[0003] The above-mentioned 3 by the HPLC method using a reversed-phase C18 filler.
Although it is possible to analyze components, for example, simultaneous analysis of highly polar ashram, oxine copper, and the like, and low-polarity pencyclon, benzulide, and the like must be performed by gradient elution, and conditions for that must be set. There is a limit to the simplicity and speed of the analysis due to the complexity and the like, and at present, a simple and quick method for analyzing the nine components of agricultural chemicals as described above is in great demand.

[0004]

SUMMARY OF THE INVENTION The present invention has been made in view of the above-mentioned circumstances, and an object of the present invention is to provide a method for simultaneous analysis of agrochemicals which can easily and quickly analyze nine components of agrochemicals by an HPLC method.

[0005]

The present invention relates to a method for analyzing pesticides by high performance liquid chromatography, wherein the analysis is carried out by isocratic elution using a cyanopropyl group-bonded filler as a filler. , Copper oxine, mecoprop, thiuram, sidurone, iprodione, chlorothalonil, pencyclon and benzulide.

[0006] That is, the present inventors have prepared a simple and convenient HPLC method for nine kinds of pesticides having a large polarity difference, specifically, ashram, oxine copper, mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide. As a result of intensive research to develop a method for rapid simultaneous analysis, if a filler with a cyanopropyl group is used as the filler, isocratic elution is easier,
It has been found that HPLC analysis can be performed quickly, and the present invention has been completed.

As the cyanopropyl group-bonding type filler used in the present invention, those in which a cyanopropyl group is bonded to a base material such as silica gel, porous polymer gel (alkylated polyvinyl alcohol type, methacrylic acid ester type, etc.), etc. Is mentioned. The preferable physical properties of the base material used as the raw material of the filler according to the present invention vary depending on the type of the base material. For example, when silica gel is used as the raw material base material, the physical property value of the raw silica gel has a particle diameter of 3 ~ 10μm,
Preferably 4 to 7 μm, specific surface area: 300 to 700 m ² / gr, preferably 450 to 600 m ² / gr, pore volume: 0.6 to 1.2 ml / gr,
Preferably, the filler has a pore diameter of 0.8 to 1.0 ml / gr and a pore diameter of 50 to 200 angstroms, preferably around 100 angstroms. The filler according to the present invention has, for example, such physical properties. A particularly preferred filler is a cyanopropyl-bonded silica gel having a carbon ratio of 7 to 14%, which is synthesized from a high-purity product having an extremely small metal content.

The method for producing a cyanopropyl group-bonded filler will be described below with reference to a case where silica gel is used as a raw material base material. That is, a pore diameter of 100 Å-
Before and after heating, spherical silica gel having an average particle size of 4 to 7 μm is heated, refluxed and reacted with toluene, dry pyridine and 3-cyanopropyldimethylchlorosilane for 6 to 10 hours with stirring. After cooling, the mixture is filtered, and the residue is washed with toluene, methanol, water, methanol, and acetone in this order, and then dried. Thereafter, the dried gel is subjected to a conventional method,
If the end capping treatment is performed by heating and reacting for ~ 10 hours, the intended cyanopropyl group-bonded filler can be easily obtained.

As a mobile phase in the isocratic elution according to the present invention, a solution comprising an organic solvent and a buffer is used. As the organic solvent used for this, usually, there is no UV absorption in a low wavelength region used under reversed phase conditions, and a highly polar solvent such as methanol, acetonitrile and the like. Usually, acetonitrile is preferably used because acetonitrile is used for extracting a sample for analysis.

[0010] The amount of the organic solvent used in the mobile phase is not particularly limited as long as the concentration in the buffer is 20 to 80 V / V%.

The type of buffer used in the mobile phase is as follows:
There is no particular limitation as long as there is no UV absorption in the low wavelength region. For example, potassium phosphate, sodium phosphate and the like are usually preferably used. The salt concentration of the buffer is usually in the range of 10 to 500 mM, but the concentration of 10 to 100 mM is desirable in consideration of maintenance of the HPLC apparatus, column life, and the like.

The pH range of the mobile phase is not particularly limited as long as the target pesticide can be analyzed, but pH 2.0 to 7.0 is preferable in consideration of column life and the like.

When the above nine agricultural chemicals are analyzed simultaneously, it is desirable to set the pH within the range of 3.6 to 3.9 in order to maintain the analysis accuracy of ashram, oxine copper and mecoprop.

It is needless to say that the mobile phase may contain a chelating agent such as EDTA, if necessary, in addition to the organic solvent and the buffer.

In the present invention, the flow rate at the time of elution is usually 0.1.
The column temperature is usually 3 to 2.0 ml / min, preferably 0.7 to 1.3 ml / min, and the column temperature is usually 20 to 60C, preferably 30 to 40C.

The analysis method of the present invention can be carried out, for example, as follows. That is, an analytical column filled with a cyanopropyl group-bonded packing material was attached to an HPLC device, equilibrated with a mobile phase, and a pretreated sample was injected.
By performing isocratic elution under conditions of about 0.3 to 2.0 ml / min and a column temperature of 20 to 60 ° C., ashram, oxine copper, mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide can be easily and rapidly alloed. Can be analyzed.

Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited thereto.

[0018]

【Example】

Example 1 (filler) silica gel (pore size 100 Å,
Spherical silica gel with an average particle diameter of 5 μm)
Add 20 ml, stir and heat, toluene-water azeotrope 70 m
l was distilled off. 10 ml of dry pyridine and 6.0 g of 3-cyanopropyldimethylchlorosilane were added thereto, and the mixture was refluxed for 8 hours while stirring. After cooling, the reaction solution was filtered and the residue was washed three times with 20 ml of toluene. Followed by methanol,
Washing was repeated in the order of water, methanol and acetone, followed by drying to obtain 10.8 g of cyanopropyl silica gel. To 8 g of the above silica gel, 40 ml of dry toluene and 9 ml of hexamethyldisilazane were added, and reacted by heating for 8 hours. After cooling, the mixture was filtered, and the residue was washed with toluene, methanol, water, methanol, and acetone in this order, and then dried to obtain 8 g of a cyanopropyl group-bonded filler.

(Sample) (A) Standard solution 1 Using an agricultural chemical standard, ashram: 5 μg / ml, oxine copper: 5 μg / ml, mecoprop: 10 μg / ml, thiuram: 10
μg / ml, Siduron: 10 μg / ml, Iprodione: 10 μg /
ml :, chlorothalonil: 5 μg / ml, pencyclone: 10 μg
/ ml and benzulide: prepared by dissolving in acetonitrile to 10 μg / ml. (B) River water solid phase extract 0.1N nitric acid was added to 500 ml of river water to adjust the pH to 4.0.
This was applied to a solid phase column (SPE-GLF manufactured by Yokogawa Analytical Systems Co., Ltd., washed with 5 ml of acetonitrile and 10 ml of distilled water) at a flow rate of 10 to 20 ml / min by suction under reduced pressure. Subsequently, extraction was performed from the solid phase column using 5 ml of acetonitrile to obtain a river water solid phase extract. (C) Standard water-added river water solid phase extract 500 ml of river water adjusted to pH 4.0 using 0.1 N nitric acid, and 5 ml of standard solution 1 of (A) above added to the same as (B) above Extraction was performed to obtain river water solid phase extract with standard solution added.

(Operation method) A cyanopropyl group-bonded filler was packed in a slurry into a 4.6 mm x 150 mm stainless steel column (manufactured by Sugiyama Shoji), set in an HPLC apparatus, and analyzed under the following analysis conditions. Analysis conditions Mobile phase: 50 mM mono-potassium phosphate and 50 mM phosphoric acid 1000: 22
(V / V) and a mixture of buffer (pH 3.8) and acetonitrile at a ratio of 59:41 (V / V) were added to EDT.
The mobile phase was prepared by adding A · 2Na to 100 mg / l. Flow rate: 1.0 ml / min (pump; LC-6A Shimadzu Corporation)
Column temperature: 35 ° C (Column oven; CTO-6A, manufactured by Shimadzu Corporation) Detection: UV absorption wavelength of each component Maximum absorption wavelength in 230 to 270 nm (Detector: MCPD-3600 manufactured by Otsuka Electronics Co., Ltd.) ) Sample injection volume: 10μl

(Results) The results obtained are shown in FIG. still,
Each peak number in FIG. 1 indicates the following nine agricultural chemical components. 1: ashram, 2: copper oxine, 3: mecoprop, 4: thiuram, 5: siduron, 6: iprodione, 7: chlorothalonil, 8: pencyclon, 9: bensulide. From the results of FIG. 1 (A), when standard solution 1 was used as a sample, complete separation and analysis of nine components of ashram, oxine copper, mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide were possible within 25 minutes. It can be seen from the results of FIGS. 1B and 1C that the above-mentioned nine components contained in the river water can be completely separated and analyzed.

Example 2 (Filler) Same as that obtained in Example 1.

(Sample) (A) Standard solution 1 Same as that used in Example 1. (B) Standard solution 2 Using an agrochemical standard, ashram: 5 μg / ml, oxine copper: 5 μg / ml, mecoprop: 10 μg / ml, thiuram: 10
μg / ml, iprodione: 10 μg / ml, pencyclon: 10 μ
g / ml and benzulide: prepared by dissolving with acetonitrile so as to be 10 μg / ml.

(Procedure) The pH of the buffer solution was adjusted to 2.0 to 2.0 by changing the mixing ratio of 50 mM monopotassium phosphate and 50 mM phosphoric acid.
A solution which was changed by 0.5 in the range of 7.0 was used as a buffer solution for a mobile phase, and otherwise, under exactly the same conditions as in Example 1,
Simultaneous analysis of standard solution 1 (9 agricultural chemical components) was performed in the same manner as in Example 1. Standard solution 2 (7 agricultural chemical components) was simultaneously analyzed in the same manner as above using a mobile phase in which the pH was changed by 0.1 in the range of 3.4 to 4.4.

(Results) The obtained results are shown in FIGS. In addition, each symbol in FIG. 2 and FIG. 3 shows the following nine components of agricultural chemicals, respectively. Ben. : Benthulide, Pen. : Pencyclone, Chl. : Chlorothalonil, Ipr. : Iprodione, Si-1: Siduron-1, Si-2: Siduron-2 (isomer of Siduron-1), Thi. :
Thiuram, 8-qu: copper oxine, MCPP: mecoprop, Asu. : Ashram. As is evident from the results in FIG. 2, oxine copper and mecoprop are easily affected by the pH of the mobile phase, and when the pH is low, the retention of oxine copper tends to be small and the retention of mecoprop tends to be large. When it is higher than 5, it is understood that the retention of the ashram is small and it is not sufficiently separated from the mecoprop. Further, as is apparent from the results of FIG. 3, it is preferable to carry out the analysis in the pH range of 3.7 to 3.8 in order to accurately separate and analyze ashram, oxine copper, and mecoprop which are easily affected by pH.

Example 3 (Filler) Same as that obtained in Example 1.

(Sample) The standard solution 1 prepared in Example 1 was used.

(Operation method) The detection wavelengths for determining the peak area and the peak height were as follows: ashram: UV 270 nm, oxine copper: UV 240 nm, mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide: UV 230 nm. Using the same column and the same apparatus as in Example 1 except that the analysis was performed, the same operation as in Example 1 was repeated and analyzed repeatedly (n = 11), and the elution time, peak area, and reproducibility of the peak height were examined. Was.

(Results) The results obtained are shown in Table 1.

[0030]

[Table 1]

As is clear from the results in Table 1, all the obtained coefficients of variation (CV) are 0.13% or less in the retention time, 1.9% or less in the peak area, and 2.6% or less in the peak height. It can be seen that the reproducibility of the analysis method is good.

Example 4 (Filler) Same as that obtained in Example 1.

(Sample) (A) Using an agrochemical standard, ashram, oxine copper, mecoprop, thiuram, ciduron, iprodione, chlorothalonil and penciclone are adjusted to 5 μg / ml to 100 μg / ml, and benzolide is adjusted to 10 μg. The solution was prepared by dissolving with acetonitrile so as to have a concentration of / ml to 100 µg / ml.

(Operation method) The detection wavelength for obtaining the peak area was as follows: Ashram: UV 270 nm, Oxin copper: UV240 n
m, mecoprop, thiuram, sidurone, iprodione, chlorothalonil, pencyclon and benzulide: UV
Using the same column and the same apparatus as in Example 1 except that the wavelength was set to 230 nm, the same operation as in Example 1 was performed to obtain the pesticide concentration (μ
g / ml) and a calibration curve representing the relationship between the peak area (absorbance · min) and their correlation coefficients.

(Results) The results obtained are shown in FIG. Each symbol in FIG. 4A is the following pesticide 4 respectively.
The components are shown. □: Ashram, +: Oxin copper, Δ: Thiuram, ×:
Chlorothalonil. In addition, each symbol in FIG. 4B indicates the following five pesticide components. □: Iprodione, +: Penciclone, Δ: Benthuride, Δ: Siduron, ×: Mecoprop.

[0036]

[Table 2]

As is clear from the results of Table 2 and FIG. 4, the calibration curves of ashram, oxine copper, mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide are all straight lines passing through the origin with a correlation coefficient of 0.999. It turns out that it is excellent in quantitative property.

[0038]

The present invention provides a simple and rapid method for simultaneous analysis of ashram, oxine copper, mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide, which has been difficult to carry out simultaneously by the conventional method. This is a great invention that contributes to the industry.

[0039]

[Brief description of the drawings]

FIG. 1 shows the chromatogram obtained in Example 1.

FIG. 2 shows the pH of the mobile phase (p (p) obtained in Example 2).
9 is a graph showing the relationship between (H2.0 to 7.0) and the retention time.

FIG. 3 shows the mobile phase pH (p) obtained in Example 2.
14 is a graph showing the relationship between H3.4 to 4.4) and the retention time.

FIG. 4 shows a calibration curve obtained in Example 4.

[0040]

[Explanation of symbols]

FIG. 1 shows 1: Ashram, 2: Oxin Copper, 3: Mecoprop, 4: Thiuram, 5: Siduron, 6: Iprodione, 7: Chlorothalonyl, 8: Pencyclon, 9: Benthuride.

FIG. 2. In FIG. : Benthuride, Pe
n. : Pencyclone, Chl. : Chlorothalonil, Ip
r. : Iprodione, Si-1: Siduron-1, Si
-2: Sidulon-2 (isomer of Sidulon-1),
Thi. : Thiuram, 8-qu: oxine copper, MCP
P: Mecoprop, Asu. : Indicates ashram.

FIG. 3 shows that Ben. : Benthuride, Pe
n. : Pencyclone, Ipr. : Iprodione, Th
i. : Thiuram, 8-qu: copper oxine, MCPP:
Mecoprop, Asu. : Indicates ashram.

FIG. 4 (A) shows □: ashram, +: copper oxine, Δ: thiuram, ×: chlorothalonil. In FIG. 4B, □: iprodione, +: penciclone, Δ: bensulide, Δ: siduron, ×: mecoprop.

Continued on the front page (58) Fields surveyed (Int.Cl. ⁶ , DB name) G01N 30/88 A01N 33/10 A01N 47/26 A01N 57/14 G01N 30/48 CA (STN)

Claims (1)

(57) [Claims] 1. A method for analyzing pesticides by high performance liquid chromatography, wherein the analysis is carried out by isocratic elution using a cyanopropyl group-bonded filler as a filler, wherein ashram, oxine copper, Analytical method of at least one of mecoprop, thiuram, siduron, iprodione, chlorothalonil, pencyclon and benzulide. [0001]