JP5024540B2 - Visual density determination method and reaction vessel - Google Patents

Description

  The present invention relates to a simple analysis method for visually determining the component concentration of a reagent and a reaction vessel used therefor.

  In the past, development of simple analysis methods that are quick, inexpensive, and do not require special techniques has been actively conducted in environmental analysis. In particular, a colorimetric method for converting the concentration of sample water into color shading by adding a coloring reagent to the sample water as a simple analysis of the components contained in moisture and determining the shading of the color (for example, the following patent document) 3) was the mainstream.

JP 2001-333797 A JP 2004-340759 A JP 2007-183104 A

  However, since the human eye cannot clearly quantify the color shading, the analysis results of these measurement methods vary depending on the field environment, including individual differences depending on the measurer and the state of light rays. There is a problem. On the other hand, the calculation method using a microplate consisting of a large number of storage units that also serve as reaction vessels and colorimetric cells is mainly used in the clinical field because it can simultaneously measure absorbance of multiple samples. . However, since the microplate reader as a measuring device is expensive and sufficient sensitivity cannot be obtained, there have been few reports on applications in other fields. Moreover, many spectrophotometers used in other colorimetric methods are also expensive, and there is a problem that it takes time to determine the concentration of a large amount of reagents.

  In order to solve such a conventional problem, an object of the present invention is to provide a visual concentration quantification method capable of obtaining a constant measurement result regardless of a measurer or a physical condition, and a reaction container used therefor.

  The visual concentration quantification method according to the present invention made to solve the above-mentioned problem is to contain a plurality of reaction reagents that change color when the concentration of the sample exceeds a certain concentration ratio with respect to the reaction reagent, each at a different specified concentration. A concentration scale is set, and the component concentration of the sample is determined by the number of discolored reaction reagents.

  In the concentration scale, the concentration of the reaction reagent is monotonously increased (a (i) ≦ a (i + 1) (i ≧ 1)) or monotonically decreased (a (i) ≧ a (i + 1) ( i ≧ 1)), and the density scale has a configuration in which a plurality of the gray scale rows are arranged side by side in the vertical direction or the horizontal direction. Sometimes it is done.

  As the reaction container used for the visual concentration determination method, the concentration scale including a plurality of storage portions constituting a grayscale gradation row in which the concentration of the reaction reagent is monotonously increased or monotonously decreased stepwise is formed. Features. The density scale may be formed by juxtaposing a plurality of the shade gradation rows vertically or horizontally.

  The present invention enables inexpensive visual analysis with improved accuracy of analysis results that do not depend on the individuality and qualities of the judge as compared to commercially available conventional colorimetric methods. In addition, since the present invention can be applied to various reactions in which a color change reaction has a concentration threshold, such as a titration reaction, visual quantification of many types of analytes is possible.

Hereinafter, embodiments of a visual density determination method according to the present invention and a reaction vessel used therefor will be described with reference to the drawings.
FIG. 1 shows an example of a reaction vessel in which a concentration scale 1 used in the visual concentration determination method according to the present invention is formed, and is an example in which a plurality of grayscale gradation rows 2 are configured on one concentration scale 1. is there.

  In this example, like the macro plate 3 or the like, the concentration of the reaction reagent is increased stepwise from the top in each column (may be in each row) of a container in which a plurality of receiving portions 4 that are recessed vertically and horizontally are formed. A plurality of grayscale gradation rows 2 are arranged side by side so as to be adjusted so as to decrease monotonously. The state of monotonic decrease is appropriately set according to the properties of the component to be detected and the reaction reagent, such as proportional decrease, logarithmic decrease, or irregular monotonous decrease.

  In this state, when the sample is added in units of rows, only the portion where the concentration of the sample exceeds a certain concentration ratio with respect to the reaction reagent is continuously discolored, and the sample concentration is determined by the number of the storage portions 4 that are continuously discolored. Is quantified (see FIG. 2).

  As shown in FIG. 2, the sulfite-hydrogen peroxide-based autocatalytic reaction has a characteristic of rapidly changing pH (color tone: in the presence of BTB solution) after a certain induction period, and there is a concentration threshold in the reaction. The threshold value can be adjusted by the concentration ratio of the reactants. That is, in this reaction, the occurrence of the reaction can be controlled like ON / OFF of the switch by the concentration ratio of the aqueous hydrogen peroxide solution to the concentration of the aqueous sulfite ion solution.

  Therefore, as the reaction reagent, for each row of microplates or the like, sodium sulfite is accommodated in the accommodation portion in a stepwise manner in which the concentration is monotonically decreased in a stepwise manner from top to bottom. (It may be singular.) A density scale is formed. When the concentration scale 1 has a configuration as a ready-made kit, the opening of the storage unit 4 in which the reagent is stored is sealed with rubber or the like into which an injection needle can be inserted, so that the reagent does not leak. Is required. The rubber or the like needs to be a material that has high water tightness and air tightness and is not denatured by the components of the reagent.

  Next, the hydrogen peroxide sample solution, which is a sample for obtaining the concentration of the component to be determined, is added to all the accommodating parts 4 constituting the predetermined grayscale gradation sequence 2. As a result, only the portion where the hydrogen peroxide concentration of the added hydrogen peroxide sample solution is equal to or higher than a certain concentration ratio with respect to the sulfite ion concentration of the reaction reagent is continuously discolored (transparent). The gradation row 2 presents an appearance of a kind of histogram on the concentration scale 1, and the hydrogen peroxide concentration can be quantified visually by the connection of the continuously discolored storage portions 4.

  If the concentration of the reaction reagent is known and the reaction concentration of the sample can be derived therefrom, the reaction concentration of the sample corresponding to the concentration of the reaction reagent is specified at the end of the concentration scale 1 as shown in FIG. By doing so, the component concentration of the sample can be quantified numerically from the description of the reaction concentration corresponding to the row of the containing portion where the highest concentration is indicated among the changed containing portions (see FIG. 3).

Moreover, 3 * 10 < ^-4 > -3.5 * 10 < ^-3 > mol containing EBT which is an indicator like FIG.
Using an EDTA aqueous solution of dm ³ (hardness equivalent to 30 to 350 mg / L, with a low factor) as a reaction reagent, the concentration of the component to be quantified is simply reduced stepwise from top to bottom in each row of the microplate 3 etc. For example, the hardness of the sample water can be measured by configuring the concentration scale 1 including the grayscale gradation row 2 accommodated by 100 μL each, and adding Mg ²⁺ sample water to the concentration scale in units of rows.

In this case, when the concentration of the Mg ²⁺ component in the added sample water is equal to or less than the concentration of EDTA in the reaction reagent (EDTA> Ca ²⁺ , Mg ²⁺ ), the blue color that is the EBT color is shown (see the following reaction formula (1)). Above the concentration of EDTA (EDTA <Ca ²⁺ , Mg ²⁺ ), purple which is an EBT chelate compound is shown (see the following reaction formula (2)).

EDTA, EBT + Ca ²⁺ , Mg ²⁺ → Ca ²⁺ -EDTA, Mg ²⁺ -EDTA + EBT (blue) (1)
EBT + Ca ²⁺ , Mg ²⁺ → Ca ²⁺ -EBT, Mg ²⁺ -EBT (purple)
... (2)

In this way, since the reaction reagent in the container 4 in which the concentration of the Mg ²⁺ component contained in the sample water exceeds the concentration of EDTA is discolored (chelate titration reaction), the container 4 that has been discolored singly or plurally. The grayscale gradation row 2 including “” indicates the hardness of the sample water added in the form of a histogram (see FIG. 5). At this time, when distilled water, tap water (hardness 70 mg / L), and commercially available deep sea water (hardness 310 mg / L) were added, the number of storage portions 4 corresponding to the respective concentrations changed color. (See the sixth to eighth columns in FIG. 5).

  The above-described invention can be expected to be useful as a simple analysis kit for a water environment with high accuracy of analysis results of various substances.

It is a conceptual diagram which shows the concept of the visual concentration determination method and reaction container by this invention. It is a principle figure which shows an example of the visual density determination method and the visual density determination method using the reaction container by this invention. It is a photograph which shows an example of the quantification result of the visual density | concentration quantification method by this invention and the visual density | concentration quantification method using the reaction container. It is a principle figure which shows an example (water hardness measurement) of the visual concentration determination method and reaction concentration determination method using reaction container by this invention. It is a photograph which shows an example (water hardness measurement) of the quantification result of the visual concentration quantification method by this invention and the visual concentration quantification method using the reaction container.

Explanation of symbols

1 concentration scale,
2 shades of gray,
3 microplates,
4 containment section,

Claims (5)

Concentrations in which reaction reagents that change color when the concentration of the sample exceeds a certain concentration ratio with respect to the reaction reagent of the sulfite-hydrogen peroxide autocatalytic reaction are stored in a plurality of accommodating portions at different specified concentrations. A visual concentration determination method that determines the component concentration of a sample by setting the scale and the number of reaction reagents that have changed color. Set the concentration scale to store the reaction reagent that changes its color when the concentration of the sample exceeds a certain concentration ratio with respect to the reaction reagent in multiple storage units. a visual concentration determination method determines component concentration of the sample drives out,
The visual concentration quantification method , wherein the reaction reagent is an EDTA aqueous solution containing an indicator . The visual concentration determination method according to claim 1 , wherein the concentration scale is configured as a grayscale sequence in which the concentration of the reaction reagent is monotonously increased or decreased monotonically. Each containing a reaction reagent comprising a sulfite-hydrogen peroxide autocatalytic reaction reagent or an EDTA aqueous solution containing an indicator as a reaction reagent that changes color when the concentration of the sample exceeds a certain concentration ratio with respect to the reaction reagent, In addition, a reaction container for a visual concentration quantification method in which a concentration scale is formed, which includes a plurality of storage units constituting a grayscale sequence in which the concentration of the reaction reagent is monotonously increased or monotonously decreased stepwise. 5. The visual concentration determination according to claim 4, wherein the opening of each storage unit storing the reaction reagent has a structure in which the opening is sealed with a material having water tightness and air tightness and capable of inserting an injection needle. Law reaction vessel.