Indicators for determining the pH of a solution

Abstract

The present invention relates to an indicator (I) for determining the pH value of a solution, wherein the indicator (I) comprises a dye molecule (F) and at least one first amino acid (A1), wherein the dye molecule (F) and the at least one first amino acid (A1) are bonded by covalent bonding, wherein when the at least one first amino acid (A1) is bonded to the dye molecule (F), the at least one first amino acid (A1) has at least one unbonded pH sensitive group. The present invention also relates to a sensor membrane (1) for an optical sensor (2), a sensor cap (4) for being placed on the optical sensor (2), and an optical sensor (2).

Description

Technical Field

[0001] The invention relates to an indicator for determining the pH value of a solution. The invention also relates to a sensor film for an optical sensor, a sensor cap for placement on an optical sensor and an optical sensor. Background Art

[0002] Indicators suitable for determining the pH value of a solution generally have at least one pH sensitive group. The pH sensitive group is sensitive to the pH value of the solution, that is to say that the pH sensitive group undergoes protonation or deprotonation depending on the pH value of the solution. Hydroxyl groups and amino groups are, for example, pH sensitive groups. Indicators are also generally dye molecules. The state of the pH sensitive group of the indicator and thus the optical properties of the indicator change according to the pH value of the solution. The pH range in which, for example, the color of the indicator changes is also referred to as a color change range. Therefore, when the indicator is directly present in the solution, the pH range of the solution can be read out in a simple manner using the eyes or a photometer.

[0003] The indicator can be an alternative component of an optical sensor, which is at least partially immersed in the solution. The optical sensor can also be placed on a flow measurement unit, for example. In the optical sensor, the indicator is preferably fixedly present, for example, the indicator is combined in a matrix. The optical sensor usually has at least one light source for emitting an excitation signal that varies with the indicator and a receiving unit for receiving a reception signal from the indicator. In the scope of the optical sensor, for example, beads are used, which have a defined concentration of the indicator and are dissolved in the sample solution.

[0004] In order to determine the pH value of a solution, there are different methods to choose from, such as measuring the transmittance, luminescence or Raman signal of an indicator. Thus, the fluorescence quenching of the indicator can be determined or a so-called double life cycle reference (DLR) can be performed, in which the phase difference between a steady-state long-term fluorescence signal and a short-term pH-sensitive fluorescence signal is determined. In addition, a short-term pH-sensitive fluorescence signal is described in US6602716B1. In addition, the measuring principle of ratiometric measurement can also be applied, in which the pH value is evaluated based on a comparison of at least two fluorescence maxima of the indicator.

[0005] Indicators are often used in titrations. In titrations, the unknown concentration of a sample (also referred to as analyte) in a sample solution is determined by chemical reaction with the aid of a titrant solution, which contains a titrant with a defined concentration. Indicators can be used to display the endpoint of the titration.

[0006] Many indicators have only one color change range and therefore can only be used for a limited pH range. And universal indicators should cover as large a pH range as possible, so that they can be used in many different applications with different pH values. Yet typically, this universal indicator is also composed of a mixture of multiple indicators that have separate color change ranges. Therefore, in order to make this universal indicator, each required indicator must be weighed individually and mixed with other indicators in the correct ratio. Summary of the invention

[0007] It is therefore an object of the present invention to specify an indicator, a sensor membrane, a sensor cap and an optical sensor which can be used in the desired pH range.

[0008] According to the invention, this object is achieved by an indicator for determining the pH value of a solution, wherein the indicator comprises a dye molecule and at least one first amino acid, wherein the dye molecule and the at least one first amino acid are bonded by means of a covalent bond, wherein the at least one first amino acid is bonded to the dye molecule such that the at least one first amino acid has at least one unbonded pH-sensitive group.

[0009] Amino acids form a class of chemical compounds which have at least one hydroxyl group and at least one amino group and thus already have two pH-sensitive groups. In addition, amino acids usually have at least one further functional group, which is sometimes also pH-sensitive. Of particular interest for the present invention are proteinogenic α-amino acids, in which the amino group occurs directly adjacent to the carboxyl group and which serve as substrates for proteins. Such proteinogenic α-amino acids are usually available at low cost and can be supplied quickly.

[0010] Since the pKa values ​​of amino acids are generally known or can be determined in a simple manner, a list of amino acids is provided to the user, from which he can select at least one first amino acid according to the desired pKa value, which has at least one pH-sensitive group with the desired pKa value. The user can thus select at least one first amino acid in a targeted manner that is compatible with the reaction, for example a titration, in which the indicator is to be used. After the addition of the indicator, the pH range of the solution to be analyzed can be determined, for example, by means of a photometer, based on the transmittance of the solution.

[0011] The amino acid itself is usually non-optically active in the visible range of light and therefore covalently combines with the dye molecule in the process of the present invention. The dye molecule is especially active in the visible range of light and in the near infrared range, i.e. between 380nm and 3000nm. The dye molecule typically has an extended delocalized π-electron system. The dye molecule itself may already be pH-sensitive or not pH-sensitive. The bonding of the dye molecule to at least one first amino acid is carried out in the following manner, i.e. when covalent bonding has been achieved between the dye molecule and at least one first amino acid, at least one pH-sensitive group of at least one first amino acid exists in a free form.

[0012] In a possible design, at least one second amino acid is covalently bonded to at least one first amino acid, wherein the at least one first amino acid and the at least one second amino acid are identical or different amino acids, wherein the at least one second amino acid is bonded to the at least one first amino acid so that the at least one second amino acid has an unbonded pH sensitive group. In order to expand the pH range covered by the indicator and / or to increase the number of pH sensitive groups, at least one second amino acid may be provided on at least one first amino acid.

[0013] Advantageously, at least one second amino acid is bonded on at least one first amino acid by peptide bond. Peptide bond can form in simple and known manner between amino acid and is extremely stable with respect to acid and alkali. At least one first amino acid is bonded to the situation on the dye molecule by its amino group or carboxyl group, and also at least one first amino acid whose unbonded amino group or carboxyl group are provided are used to form with at least one second amino acid whose peptide bond.

[0014] In an improved design, at least one first amino acid and / or at least one second amino acid has at least three pH sensitive groups in an unbonded state, wherein at least one first amino acid and / or at least one second amino acid is especially histidine. If histidine (which has three pH sensitive groups with three different pKa values) is selected as at least one first amino acid, then a wide pH range can be covered using only histidine. Additionally, at least one second amino acid that also has at least three pH sensitive groups can be combined with histidine or at least one other first amino acid that has at least three pH sensitive groups. Therefore, the pH range that can be covered by the indicator is significantly increased.

[0015] Preferably, at least one first amino acid is bonded to the dye molecule via a peptide bond or a CC bond.

[0016] In another possible design, at least two first amino acids are combined with the dye molecule. In order to improve the pH sensitivity of the indicator, for example, at least a plurality of first amino acids are combined with the dye molecule.

[0017] Possible design scheme provides: at least two identical second amino acids are respectively combined with at least two first amino acids. In this case, the at least two first amino acids are respectively extended with at least one second amino acid.

[0018] In an alternative design, at least two different second amino acids are combined with at least two first amino acids. In this case, the at least two first amino acids are extended with different second amino acids respectively and potentially more different pH sensitive groups are introduced into the indicator than in the previous case.

[0019] In an improved design, the indicator has at least two pH-sensitive groups, wherein the pH-sensitive groups each have a pKa value, wherein the pH-sensitive groups of the indicator are selected such that the respective pKa values ​​of the pH-sensitive groups differ from one another by at least one pKa unit. This results in a good resolution of the pH range with as little overlap of the respective color change ranges of the pH-sensitive groups as possible. For a desired indicator having a pH range of pH=4 to pH=10, pH-sensitive groups having pKa values ​​of approximately 1 to 5.5 and pKa values ​​of approximately 2 to 8.5 can be selected.

[0020] In a preferred embodiment, the indicator has at least three pH sensitive groups, wherein the pH sensitive groups of the indicator are selected so that the indicator has a pH sensitive range including at least three pH units. Such an indicator, for example, covers a pH range between at least three decimal numbers (e.g., from pH=4 to pH=7).

[0021] Advantageously, the dye molecule is selected from the class of BODIPY dyes, ADOTA dyes or DAOTA dyes.

[0022] Preferably, the dye molecule is pH insensitive. Therefore, the pKa value of the dye molecule need not be considered when constructing the indicator, but the desired pH range is achieved by arranging the pKa values ​​of the pH sensitive groups of at least one first amino acid and / or at least one second amino acid.

[0023] The task based on the present invention is also solved by a sensor film for an optical sensor, which comprises an indicator according to the present invention and a matrix for embedding the indicator. The indicator is covalently or physically bonded to the matrix, which can be composed of, for example, silica gel, organically modified silicates or sols, or can also be composed of polystyrene derivatives, methacrylate derivatives, polyurethane derivatives, acrylate derivatives or cellulose derivatives. The three-dimensional network and the base layer are understood as a matrix, on which the so-called SAM structure (self-assembled monolayer) can be applied. The sensor film is in contact with the solution and the indicator incorporated in the sensor film can react to the pH value of the solution.

[0024] The object underlying the invention is also achieved by a sensor cap for placement on an optical sensor, wherein the sensor cap has a sensor film according to the invention with an indicator according to the invention, or wherein the sensor cap has a coating with an indicator according to the invention. The region of the sensor cap to which the coating with the indicator is applied can be made, for example, of glass.

[0025] The object underlying the invention is also achieved by an optical sensor comprising a light source for emitting an indicator-dependent excitation signal, a receiving unit for receiving a reception signal from the indicator, and a sensor cap according to the invention. The reception signal can be, for example, luminescence or transmittance. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Next, the present invention will be described in detail with reference to the following Figures 1 to 6, wherein:

[0027] Figure 1a , 1b Showing the first and second designs of the indicator according to the present invention;

[0028] Figure 2a An exemplary schematic diagram of the synthesis of a first design of an indicator according to the present invention is shown;

[0029] Figure 2b An exemplary schematic diagram of the synthesis of a third design of the indicator according to the present invention is shown;

[0030] Figure 3 An exemplary schematic diagram of the synthesis of a fourth design of the indicator according to the present invention is shown;

[0031] Figure 4 The design of the sensor membrane according to the present invention is shown;

[0032] Figure 5 The design of the sensor cap according to the present invention is shown;

[0033] Figure 6 A configuration of an optical sensor according to the present invention is shown. DETAILED DESCRIPTION

[0034] exist Figure 1a , a first design of the indicator I according to the present invention is shown in FIG. 1 , wherein the indicator comprises a dye molecule F and at least one first amino acid A1. Figure 1a In the first embodiment of the indicator I shown, the three first amino acids A1 are covalently bonded to the dye molecule F via peptide bonds. Alternatively, a CC bond is also possible, for example. Figure 1b , an optional second design of the indicator I according to the present invention is shown, wherein the first amino acid A1, exemplified here by histidine, is combined with the dye molecule F via a CC bond. At least one first amino acid A1 should be covalently bonded to the dye molecule F, so that when at least one first amino acid A1 is bonded to the dye molecule F, the at least one first amino acid has at least one unbonded pH-sensitive group. Figure 1a , 1b In the embodiment, three first amino acids A1 are bonded to the dye molecule F, wherein other numbers of first amino acids A1 may be bonded to the dye molecule F, such as at least one or at least two first amino acids A1.

[0035] For the sake of overview, FIG. 2 shows an exemplary schematic diagram of a chemical reaction in which a dye molecule F and at least one first amino acid A1 are converted into Figure 1a The chemical synthesis of the indicator is carried out according to known synthesis strategies and is not further explained here. It is pointed out that when synthesizing the indicator I, protective groups P1, P2 are preferably used on the pH-sensitive groups of at least one first amino acid and / or at least one second amino acid A1, A2, such as Figure 2a , 2b and Figure 3 As shown in Figure 2 to 3. The first and / or second amino acid A1, A2 protected by protecting groups P1, P2 are abbreviated as A, P in Figures 2 to 3. Here, amino acids can also have different protecting groups. After dye molecule F is combined with at least one first amino acid A1 or after at least one first amino acid A1 is combined with at least one second amino acid A2, the protecting groups P1, P2 present are removed to release the pH sensitive group. The respective protecting groups of the first and / or second amino acid A1, A2 can be the same or different from each other. For the use of synthetic strategies and protecting groups in the case of amino acids, for example, reference is made to the following literature:

[0036] Brückner, R. Reaktionsmechanismen-Organische Reaktionen, Stereochemie, Moderne Synthesemethoden; 3.Auflage; Springer-Verlag Berlin Heidelberg; 2004 (Brückner, R.; "Reaction Mechanism-Organische Reaktionen, Stereochemie, Moderne Synthesemethoden"; 3rd edition; Springer-Verlag Berlin Heidelberg; 2004); and

[0037] · Isidro-Llobet, A.; M.; Albericio, F. Chem Rev. 2009, 109, 2455-2504 (Isidro-Llobet, A.; M.; Albericio, F.; Chemical Reviews; 2009; page 109, lines 2455-2504).

[0038] exist Figure 1a in and Figure 2a , histidine is shown as the first amino acid A1, as an example, the first and / or second amino acid A1, A2 have at least three pH sensitive groups in the unbonded state. Therefore, other amino acids are not excluded. Based on the peptide bond, two pH sensitive groups are still unbonded after bonding to the dye molecule. Figure 1a and Figure 2a Indicator I in has six pH sensitive groups in total, is respectively three amino groups and three imidazole groups, and therefore has two different pKa values.Dye molecule F has had the pH sensitive amino group of itself at first, but this pH sensitive amino group is bonded with the first amino acid A1 with peptide bond when converting.But optionally, dye molecule F also can be pH insensitive.For example, dye molecule F can be selected from the category of BODIPY dye, ADOTA dye or DAOTA dye.

[0039] Figure 2b A schematic diagram of the synthesis of a third embodiment of the indicator I according to the invention is shown, wherein Figure 2a The indicator I obtained in exemplarily works with at least one second amino acid A2, which is covalently bonded to at least one first amino acid A1. Figure 2bAspartic acid is shown as the second amino acid A2, which, like histidine, has three pH-sensitive groups in the unbonded state. The second amino acid A2 can be different from the first amino acid A1 or the same as it. Aspartic acid as the second amino acid A2 is bonded to the first amino acid A1, i.e. histidine, by means of a peptide bond, wherein other possibilities of bonding are not excluded. It also applies to the second amino acid A2 that when at least one second amino acid A2 is bonded to at least one first amino acid A1, the second amino acid has at least one unbonded pH-sensitive group.

[0040] Optionally, the indicator I has at least three pH-sensitive groups, which are selected such that the indicator I has a pH-sensitive range that includes at least three pH units. Figure 2b The indicator I in the embodiment has three different pH-sensitive groups, i.e., an imidazole group (pKa-6), an ammonium group (pKa-9) and a carboxyl group (pKa-4), which respectively appear three times in the indicator I, and by means of which, for example, a pH range of pH=2.5 to pH=10.5 can be determined. For example, the indicator I also has at least two pH-sensitive groups, whose pKa values ​​differ from each other by at least one pKa unit, such as also from Figure 2b As obtained in indicator I.

[0041] Figure 2b An example for selection is shown, that is, at least two identical second amino acids are respectively combined with at least two first amino acids. Figure 3 That is, at least two different amino acids are combined with at least two first amino acids. Figure 3 In the fourth design scheme of the indicator I, the dye molecule F in FIG. 1 which is combined with the first amino acid A1 is converted with three different second amino acids A2 and thus obtains. By combining different second amino acids A2 with the first amino acid A1, a series of different pH-sensitive groups can be introduced into the indicator I. Therefore, the indicator I has a sulfonate group (pKa~1), a carboxyl group (pKa~4), an imidazole group (pKa~6), an ammonium group (pKa~9) and a guanidine group (pKa~12) as pH-sensitive groups. Therefore, the indicator I is almost regarded as a universal indicator capable of covering a pH range of about -0.5 to 13.5. By using different second amino acids A2, a large number of different indicators I (not shown) can be produced, but their relationship to each other generally corresponds to the relationship of the reactants in terms of stoichiometry. In the sense of the present invention, it is possible to abandon the use of traditional methods to distinguish different indicators I. However, purification is not excluded in the sense of the present invention.

[0042] Figure 4The sensor film 1 for an optical sensor 2 according to the present invention is shown, and the sensor film has a matrix 3 in which an indicator I is embedded and fixed. The matrix 3 can be composed of silica gel, organic modified silicate or sol, or can also be composed of polystyrene derivatives, methacrylate derivatives, polyurethane derivatives, acrylate derivatives, cellulose derivatives. The embedding of the indicator I in the matrix 3 can be achieved by chemical covalent bonds or physical bonds.

[0043] exist Figure 5 The design of the sensor cap 4 according to the present invention is shown in FIG. Figure 4 Alternatively, the indicator I can be designed as a coating 5 on the sensor cap 4.

[0044] exist Figure 6 2 shows a design of an optical sensor 2 according to the present invention. The optical sensor 2 has a light source 6 for exciting the indicator I, and a receiving unit 7 for receiving a reception signal from the indicator I. In addition, the optical sensor 2 includes a sensor cap 4, wherein the sensor cap 4 is optionally provided with a coating 5 with the indicator I.

[0045] Reference numerals list

[0046] 1 Sensor membrane

[0047] 2 Optical Sensors

[0048] 3 Matrix

[0049] 4 Sensor cap

[0050] 5. Coating

[0051] 6 Light Source

[0052] 7 Receiving unit

[0053] F Dye molecule

[0054] A1 First amino acid

[0055] A2 Second amino acid

[0056] P1 Protecting group of the first amino acid

[0057] P2 Protecting group of the second amino acid

[0058] I Indicator.

Claims

1. An indicator (I) for determining the pH value of a solution, wherein: The indicator (I) comprises a dye molecule (F) and at least one first amino acid (A1), wherein the dye molecule (F) and the at least one first amino acid (A1) are bonded to the dye molecule (F) by means of a covalent bond, wherein the at least one first amino acid (A1) is bonded to the dye molecule (F) so that the at least one first amino acid (A1) has at least one unbonded pH-sensitive group, wherein the dye molecule (F) is selected from the category of ADOTA dye or DAOTA dye, wherein the at least one first amino acid (A1) is bonded to the dye molecule (F) by means of a peptide bond or a CC bond.

2. The indicator according to claim 1, wherein At least one second amino acid (A2) is covalently bonded to the at least one first amino acid (A1), wherein the at least one first amino acid (A1) and the at least one second amino acid (A2) are the same or different amino acids, and wherein the at least one second amino acid (A2) is bonded to the at least one first amino acid (A1) so that the at least one second amino acid (A2) has at least one unbonded pH-sensitive group.

3. The indicator according to claim 2, wherein The at least one second amino acid (A2) is bonded to the at least one first amino acid (A1) via a peptide bond.

4. The indicator according to claim 2 or 3, wherein The at least one first amino acid (A1) and / or the at least one second amino acid (A2) have at least three pH-sensitive groups in an unbound state.

5. The indicator according to claim 4, wherein The at least one first amino acid (A1) and / or the at least one second amino acid (A2) is histidine.

6. The indicator according to any one of claims 1 to 3, wherein There are at least two first amino acids (A1) bonded to the dye molecule (F).

7. The indicator according to claim 6, wherein At least two identical second amino acids (A2) are bonded to the at least two first amino acids (A1), respectively.

8. The indicator according to claim 6, wherein There are at least two different second amino acids (A2) bonded to the at least two first amino acids (A1).

9. The indicator according to any one of claims 1 to 3, wherein The indicator (I) has at least two pH sensitive groups, wherein the pH sensitive groups each have a pKa value, wherein the pH sensitive groups of the indicator (I) are selected such that the respective pKa values ​​of the pH sensitive groups differ from each other by at least one pKa unit.

10. The indicator according to any one of claims 1 to 3, wherein The indicator (I) has at least three pH sensitive groups, wherein the pH sensitive groups of the indicator (I) are selected such that the indicator (I) has a pH sensitive range comprising at least three pH units.

11. The indicator according to any one of claims 1 to 3, wherein The dye molecule (F) is pH insensitive.

12. A sensor film (1) for an optical sensor (2), comprising an indicator (I) according to any one of the preceding claims and a matrix (3) for embedding the indicator (I).

13. A sensor cap (4) for placement on an optical sensor (2), wherein: The sensor cap (4) comprises a sensor membrane (1) according to claim 12, or the sensor cap (4) comprises a coating (5) comprising an indicator (I) according to any one of claims 1 to 11.

14. An optical sensor (2), comprising a light source (6) for emitting an excitation signal that varies depending on the indicator, a receiving unit (7) for receiving a reception signal from the indicator (I), and a sensor cap (4) according to claim 13.

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