WO2015013731A1

WO2015013731A1 - Sensor membrane for the reversible detection of analytes

Info

Publication number: WO2015013731A1
Application number: PCT/AT2014/000144
Authority: WO
Inventors: Gerhard Mohr
Original assignee: Joanneum Research Forschungsgesellschaft Mbh
Priority date: 2013-07-31
Filing date: 2014-07-21
Publication date: 2015-02-05
Also published as: AT514611A1; AT514611B1

Abstract

In a sensor membrane for the reversible detection of analytes, comprising a polymeric carrier membrane, an indicator dye covalently bonded to the carrier membrane, and optionally a covering layer, a further dye having a different colour from the indicator dye is covalently fixed to the carrier membrane, and the further dye and the indicator dye are covalently bonded to in particular nano- or microparticles different from one another and consisting of cellulose, polyurethane hydrogel, poly(hydroxyethyl methacrylate), amino polymer, polyacrylates, silicones, epoxides, sol gel glasses or polyolefins, and then are embedded into a polymeric carrier membrane or are physically or chemically coupled.

Description

SENSOR MEMBRANE FOR THE REVERSIBLE DETECTION OF ANALYTES

The present invention relates to a sensor membrane for the reversible detection of analytes comprising a polymeric support membrane, an indicator dye covalently bound to the support membrane, and optionally a cover layer.

Sensor membranes which have a covalently fixed indicator dye for detecting analytes on a carrier membrane have been known for some time and are used for a very wide variety of purposes depending on the bound indicator tartar dyes.

Indicator dyes are substances that indicate changes in state in chemical systems through a color change. So well-known purposes in addition to the conventional absorption indicators (for example, pH indicators and redox indicators), for example, fluorescence indicators. Particularly in the case of redox indicators and pH indicators, the measuring principle is based on a color change of the indicator dye, for example at a specific pH or a specific redox potential. At present, a variety of indicator dyes are known which either have a single color change or, such as methylene blue, also show a plurality of color changes depending on the pH. A disadvantage of these known indicators, however, is that the color change often takes place either between two poorly distinguishable colors, or that a relatively slow transition takes place from one color to another, such as Bromthymolblau, in which an envelope from yellow to green to takes place blue, so that it is not or very difficult to see, especially with the naked eye, when exactly the change between the two colors has taken place.

Especially with sensors such as oxygen sensors, C0 ₂ sensors, NH ₃ sensors and the like, it is of eminent importance that an exact color change and in particular a precise detection of the time at which the color change takes place is possible to be able to monitor a chemical reaction accurately or precisely. In order to improve the distinctness of the two colors of the indicators or to increase the scope of the indicators, mixed indicators consisting of two or more indicator dyes have been produced a long time ago, which on the one hand can show a plurality of color changes and, on the other hand, the individual color changes within a narrower range be it pH range or rich redox potential, so that a wide range of applications has been created and a more accurate detection has been made possible. The disadvantage of all these indicators is that there is still a non-negligible range in which a color change happens and not an exact point, which can be defined as a turnover point.

It has also been known for some time to provide test strips or sensor elements on which indicator dyes are physically fixed and which sensors for monitoring chemical reactions are either immersed or introduced into the reaction medium. A disadvantage of such sensor elements on which the indicator dyes are physically bound, however, is the fact that when in contact with fluids to be examined media there is a risk that the indicator dye is washed out, so that especially in a prolonged use or multiple use the risk is to lose excessively large amounts of indicator dye, so that a color change can not or only with difficulty be determined.

The document M. Roses, Analytioca Chimica Acta, 204 (1988) 31 1 ff describes a computer program which calculates chromaticity parameters from spectrophotometric data, since the visual evaluation of color changes can lead to doubtful results and is furthermore subjective. In addition, it is possible with such a program to quickly and reliably provide a comparison of the indicator color changes available. US 5,853,669 already discloses a pH indicator element in which the indicator dye is covalently bonded to a matrix. Due to the covalent binding of the indicator dye to the matrix or sensor membrane, in such indicators washing out of the indicator is withheld for multiple uses, but the problem of an inaccurate or over a relatively large range extending color change with such a pH sensor not be solved.

WO 03/036293 A1 discloses an optical method by means of which it is possible to determine the pH of a solution and its content of dissolved oxygen. It is based on the use of two optical indicators in only one sensor matrix, wherein the two indicators provide two differentiable optical signals, which can be assigned to the respective measured variables. DE 10 2007 053 664 A1 discloses optical sensors for detecting ions, gases and biomolecules, comprising a matrix with one or more indicator substances contained therein, in which one or more indicator substances are present in at least one thin layer of ionic liquid Matrix are included.

DE 1603454 A discloses a fiber material which is dyed on the one hand with an indicator dye which is covalently bonded to the fiber material and which indicator should be moved to a color change with materials commonly used in the household such as lemon juice. According to DE 1603454 A, such materials are to be used, for example, in the toy industry in order to make the game interesting for a long time by achieving a color change. In such an indicator provided with color material is therefore completely irrelevant whether and how far the color change takes place within a range or exactly at one point, as long as the color change is reversible.

Finally, US Pat. No. 6,531,322 B1 discloses an optically evaluable blood glucose test strip in which an indicator dye or a mixture of indicator dyes is integrated on a test strip consisting of two membranes and an inert dye is incorporated in another layer in order to obtain a mixed color from which, in any case, a color shows a pH change, to allow a more accurate determination of the pH. A disadvantage of this patent, however, is the fact that both dyes are only physically integrated and thus can easily be washed out of the membrane, so that the reversibility or multiple use of such a sensor membrane appears only to a very limited extent possible.

The present invention now aims to provide a sensor membrane which, even after a plurality of uses, exhibits consistent color changes and color intensities during the color change and, in particular, provides a sensor membrane which exhibits a precise color change, which only occurs over one extends very narrow range of values, in order to make an exact measurement possible in this way.

To solve this problem, the sensor membrane according to the invention is essentially characterized in that on the support membrane, another one of the indicator dye having different color dye is covalently fixed and that the further dye and the indicator dye are covalently bound to, in particular, different nanoparticles or microparticles consisting of cellulose, polyurethane hydrogel, poly (hydroxyethyl methacrylate), amino-polymer, polyacrylates, silicones, epoxides, sol-gel glasses or polyolefins, and then into a polymer support membrane embedded or physically or chemically coupled.

By further covalently fixing a dye having a different color from the indicator dye to the support membrane, it is possible to provide a sensor membrane which is available for a plurality of measurements without either the indicator dye or the second sensor membrane Dye are washed out. Furthermore, by covalent fixing of both the indicator dye and the second dye, it is possible to provide a sensor element which has a reversible optical property in order to be able to continuously monitor, for example, a reaction proceeding over a long period of time or a chemical compound , Of course, the optical property of the sensor element, to which the indicator dye and the second dye are covalently fixed, can also be formed irreversibly, with which, for example, exceeding a limit value of a component to be monitored can be reliably detected.

The fact that the indicator dyes and other dyes are covalently bonded or copolymerized to particular different nano and microparticles consisting of cellulose, polyurethane hydrogel, poly (hydroxyethyl methacrylate), amino, polyacrylates, silicones, epoxies, sol-gel glasses or polyolefins can Any sensor membranes or sensor elements are produced, wherein the micro- and nanoparticles are physically or chemically coupled to the polymeric support membrane. Indicator dyes and further dyes can be covalently bound to the same or to different nano- and microparticles. For a particularly simple production of a sensor membrane according to the invention, the further colorant and the indicator dye are fixed to mutually different layers of the polymeric support membrane. With such a construction of a sensor membrane, it is possible to fix both dyes, both the indicator dye and the further dye separately on a layer of the polymeric support membrane and only after fixing the dye molecules to connect these membrane layers or only one of the dyes on a layer of the support membrane and deposit a second layer thereon and only then the second indicator dye covalently on this second layer. In this way, the differences in the chemical reactivity of the various dyes can be taken into account and it is safe and reliable to provide a homogeneous and uniformly doped sensor membrane available

According to one embodiment of the invention, the sensor element is substantially designed so that the further dye with the indicator dye results in a clear color contrast in a color change of the indicator dye having mixed color. The fact that the further dye with the indicator dye results in a mixed color which allows a clear color contrast in a color change of the indicator dye, it is possible to determine the envelope and thus the measured optical property of the monitored component or the substance to be monitored significantly more accurate than with an indicator dye alone. With an indicator dye alone the turning point with the naked eye can often not be clearly recognized, since until the actual color change a slow or continuous color change takes place, which is often very poorly recognized or distinguished by the naked eye. For example, when applying a yellow-to-red indicator dye, before a definite transfer, the yellow color slowly becomes more intense and the red component in the yellow dye slowly becomes higher. For a viewer, it is often difficult to detect in such a dye whether the color change has already taken place or not, but if, for example, a dye with a dark color, such as blue, is added to such an indicator dye, the color change from green to dark red or Violet, which envelope can be perceived significantly better with the naked eye than, for example, a color change from yellow to red.

According to one embodiment of the invention, in a manner known per se, the indicator dye is a pH indicator dye or a redox indicator dye or other selective indicator dye. Both pH indicator dyes and redox indicator dyes can be used to track a wide variety of chemical reactions and thus for a wide variety of uses. With selective indicator dyes, for example, oxygen, reactive oxygen species, such as hydrogen peroxide, CO2, amines, carboxylic acids, ketones, Na +, K +, Ca ₂ +, Mg ₂ +, ions, saccharides, alcohols, diols, thiols, nitrogen oxides, proteins, metabolites, organophosphorus Compounds and formic acid are investigated or these compounds and ions are traced. A particularly clearly visible and smooth color change can be achieved according to the invention. be characterized in that the indicator dye is, for example, an indicator dye turning from yellow to red, in particular 2 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) -4-methoxyphenol, 4-fluoro-2 - ((4 - (2-hydroxyethylsulfonyl) phenyl) dia- zenyl) phenol or 4-bromo-2 - ((4- (2-hydroxyethylsulfonyl) cyclohexa-1,4-dienyl) diazenyl) phenol. By using an indicator dye turning from, for example, yellow to red and adding an inert dye which has a darker base color, it is possible to create a clear color change, so that the optical property in the form of a color change of the indicator element can be reliably perceived and, in particular, the smallest changes are perceived can be. Particularly clearly visible and in particular only a very narrow range extending color envelopes can be achieved according to the invention characterized in that the indicator dye, for example, 1-hydroxy-4- [4- (2-hydroxyethylsulfonyl) phenylazo] naphthale ne-2-sulfonic acid potassium salt (Chromoionophore XVI I), dilithium (1 +) ion 10-amino-3- (vinylsulfonyl) -2,4-dioxo-3-azatricyclo [7.3.1.0 ^A {5, 13}] trideca-1 ( 13), 5,7,9, 11-pentaene-7,11-disulfonate (Lucifer Yellow VS dilithium salt), 4-hydroxy-3 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) benzonitrile, 4- (trifluoroacetyl) -4 '- [N- (1-methacryloxyundecyl) -N-ethylamino] azobenzene, N-allyl-4- (N-methylpiperazinyl) -1, 8-naphthalimide or 4- [N, N-bis (1-methacryloxyundecyl) amino] -4 '- (trifluoroacetyl) -stilbene. By, as corresponds to a development of the invention, the further dye, for example, 2- (3- (4-amino-9, 10-dihydro-3-sulfo-9, 10-dioxoanthracen-4-yl) aminobenzenesulfonyl ) vinyl) disodium sulfate, trade name Reactive Blue 19, trisodium 5-acetamido-4-hydroxy-3- (2- {2-hydroxy-5- [2- (sulfonatooxy) ethanesulfonyl] phenyl} diazene-1 - yl) - naphthalenes-2-disulfonate, trade name Remazol Brilliant Violet 5R or tetrasodium (3Z) -5-amino-4-oxo-6- [4- (2-sulfonato-oxyethylsulfonyl) -phenyl] diazenyl-3 - [[4 (2-sulfonato-oxyethylsulfonyl) phenyl] -hydrazinyldienes] -naphthalenes-2,7-disulfonate, trade name Reactiv Black 5, it is possible to achieve color changes with a very clear contrast and thus an exact detection of, for example, the pH value. Value, a certain redox potential or the like. To enable. Furthermore, it is possible with a combination of a specific indicator dye and another dye. Test strips and sensors for a variety of measuring ranges produce and it can also be set arbitrarily contrast at the transition point, as corresponds to a development of the invention. For this purpose, the sensor membrane is essentially characterized in that the further dye and the indicator dye in a ratio of 1: 20 to 1: 1 are used. By choosing the ratio of the further dye and the indicator dye, it is possible to adjust the color depth and thus to maximize the color depth. adjust possible contrast of the two colors of the indicator dye before and after the turnover by adding another dye.

In order to prevent further unintentional change in the color of the sensor membrane or of a sensor, the invention is preferably developed in such a way that the polymeric carrier membrane is formed from a colorless, transparent polymer. By forming the polymeric membrane from a colorless, transparent polymer, no additional color effect is introduced so that the desired colors before and after the turnover of the indicator dye are achieved only by the mixture of the indicator dye with the further dye.

By, as corresponds to a development of the invention, as a polymeric support membrane, for example cellulose membranes, polyurethane hydrogels, poly (hydroxyethyl methacrylates), amino polymers, polyacrylates, silicones, epoxies, sol-gel glasses or polyolefins are used, the parameter of the membrane and by selecting the porosity, the diffusion properties and the layer thickness of the membrane, in particular the response speed of the sensor membrane are controlled and thus a fine adjustment of the measuring range can be achieved. By, as corresponds to a development of the invention, the polymeric carrier membrane is formed multi-layered, any sensor membranes or sensor elements can be produced. Thus, a support membrane layer may constitute a rigid layer, which is provided to protect the sensitive sensor membrane and the actual sensor membrane then applied only on this sensor, or it may be arranged one above the other several layers of the support membrane, of which further dyes and the Indicator dye are arranged only in one layer.

In order to optimize or increase the recyclability and the lifetime of a sensor membrane according to the invention as much as possible, the invention is developed so that the polymeric support membrane is covered with a porous, transparent cover layer at least on one side having the covalently bound indicator dye , By applying a porous, transparent cover layer on the sensor membrane or support membrane containing the indicator dye, it is possible, on the one hand, to prevent damage to the sensor membrane and, on the other hand, to ensure that For example, in the case of a redox indicator, the redox potentials can be reliably and reliably detected and detected despite the presence of a cover layer.

The invention will be explained in more detail below with reference to embodiments and figures. These show in a covalent coupling to polymer membranes:

Example 1 2- (3- (4-Amino-9,10-dihydro-3-sulfo-9,10-dioxoanthracen-4-yl) aminobenzenesulfonyl) vinyl) disodium sulfate in combination with 1-hydroxy 4- [4- (2-hydroxyethylsulfonyl) phenylazo] naphthalene-2-sulfonic acid potassium salt at a dye ratio of 1: 5 a color change from olive green to violet:

Example 2 and FIG. 1: Tetrasodium (3Z) -5-amino-4-oxo-6- [4- (2-sulfonato-oxyethylsulfonyl) -phenyl] diazenyl-3 - [[4- (2-sulfonatooxyethylsulfonyl) -phenyl] -hydrazinylidenes] -naphthalenes-2,7-disulfonates in combination with 2 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) -4-methoxyphenol at a dye ratio of 1:10 a color change from yellow-green to deep red wherein Figure 1 shows both the indicator set alone and the covalently bound indicator and further dye fixedly shown;

Example 3 2- (3- (4-Amino-9,10-dihydro-3-sulfo-9,10-dioxoanthracen-4-yl) aminobenzenesulfonyl) vinyl disodium sulphate in combination with 4-fluoro 2 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) phenol at a dye ratio of 1:10 a change in color from green to deep red.

The same procedure was followed in all examples, the covalent coupling taking place as follows:

100 mg of the indicator dye are thoroughly mixed with 1 g of concentrated sulfuric acid in a mortar and allowed to stand for 30 minutes in a desiccator. Thereby, the 2-hydroxyethylsulfonyl group of the indicator dye could be converted into a reactive sulfonate. This mixture is then emptied into 900 ml of water and neutralized with 1 .6 ml of 32% sodium hydroxide solution. Thereafter, 25.0 g of sodium carbonate in 100 ml of water and subsequently 5.3 ml of a 32% sodium hydroxide solution are added. At the same time, a solution of 20 mg of the further dye in 100 ml of water is added. The polymer membranes are placed in this staining solution. Under the basic conditions of the dyeing solution, the dye sulfonate is converted into a chemically reactive vinyl sulfonyl derivative which covalently binds to the reactive groups of the poly- meres (for example, hydroxyl groups of cellulose or amino groups of the polyurethane) binds. The additional dye thus binds to the reactive groups of the polymer. After 60 minutes, the dyed polymer membranes are removed from the dyebath and washed several times with water. Thereafter, the dyed polymer membranes are dried in air.

As Fig. 1 is an example of an indistinct color change of a pure covalent bound to a polymer membrane indicator dye compared to a significant color change in a mixture of indicator dye and further dye covalently bonded to a polymer membrane. Herein, in the upper series, the color change of the pure indicator dye is shown with a color change from orange to red and in the lower series the color change of an indicator covalently attached to the support and a further dye covalently bonded to the support, with a color change from green to red ,

Claims

Patent claim:

1 . Sensor membrane for the reversible detection of analytes, comprising a polymeric support membrane, an indicator dye covalently bound to the support membrane and optionally a cover layer, characterized in that on the support membrane another dye having a different color of the indicator dye is covalently fixed and that the further Dye and the indicator dye in particular different from each other nano- or icroparticles consisting of cellulose, polyurethane hydrogel, poly (hydroxyethyl methacrylate), amino, polyacrylates, silicones, epoxies, sol-gel glasses or polyolefins are covalently attached, and then embedded in a polymeric support membrane or be physically or chemically coupled.

2. Sensor membrane according to claim 1, characterized in that the further dye with the indicator dye gives a clear color contrast in a color change in the indicator dye having mixed color.

3. Sensor membrane according to claim 1 or 2, characterized in that the indicator dye in a conventional manner is a pH indicator or redox indicator dye or other selective indicator dyes.

4. Sensor membrane according to one of claims 1, 2 or 3, characterized in that the indicator dye is, for example, a yellow to red umschlagender indicator dye, in particular 2 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) -4-methoxyphenol, 4-fluoro-2 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) phenol or 4-bromo-2 - ((4- (2-hydroxyethylsulfonyl) cyclohexa-1,4-dienyl) diazenyl) phenol.

5. Sensor membrane according to one of claims 1, 2 or 3, characterized in that the indicator dye, for example, from 1-hydroxy-4- [4- (2-hydroxyethylsulfonyl) phenyl azo] naphthalene-2-sulfonic acid potassium salt (Chromoionophore XVII Dilithium (1 +) ion 10-amino-3- (vinylsulfonyl) -2,4-dioxo-3-azatricyclo [7.3.1.0 {5, 13}] trideca-1 (13), 5,7, 9,1 1 -pentaene-7,1,1-disulfonate (Lucifer Yellow VS dilithium salt), 4-hydroxy-3 - ((4- (2-hydroxyethylsulfonyl) phenyl) diazenyl) benzonitrile, 4- (trifluoroacetyl) -4 '- [N- (1-methacryloxy-and-ecyl) -N-ethylamino] azobenzene, N-allyl-4- (N-methylpiperazinyl) -1, 8-naphthalimide or 4- [N, N-bis (1 1 - methacryloxyundecyl) amino] -4 '- (trifluoroacetyl) -stilbene.

6. Sensor membranes according to one of claims 1 to 5, characterized in that the further dye with a plurality of indicator dyes (mixing indicators) is combined.

7. Sensor membrane according to one of claims 1 to 5, characterized in that the further dye, for example, 2- (3- (4-amino-9, 10-dihydro-3-sulfo-9, 10-dioxoanthracene-4- yl) aminobenzenesulfonyl) vinyl) disodium sulfate (Reactive Blue 19), trisodium 5-acetamido-4-hydroxy-3- (2- {2-hydroxy-5- [2- (sulfonatooxy) ethanesulfonyl] phenyl } diazen-1-yl) naphthalene-2,7-disulfonates (Remazol Brilliant Violet 5R) or tetrasodium (3Z) -5-amino-4-oxo-6- [4- (2-sulfonato-oxyethylsulfonyl) -phenyl ] diazenyl-3 - [[4- (2-sulfonatooxyethylsulfonyl) phenyl] -hydrazinyldiene] -naphthalenes-2,7-disulfonate (Reactiv Black 5).

8. Sensor membrane according to one of claims 1 to 7, characterized in that the further dye and the indicator dye in a ratio of 1: 20 to 1: 1 are used.

9. sensor membrane according to one of claims 1 to 8, characterized in that the polymeric support membrane is formed from a colorless, transparent polymer.

10. Sensor membrane according to one of claims 1 to 9, characterized in that as polymeric carrier membrane, for example, cellulose membranes, polyurethane hydrogels, poly (hydroxyethyl methacrylates) or amino polymers, polymer acrylates, silicones, epoxies, sol-gel glasses or polyolefins are used.

1 1. Sensor membrane according to one of the claims 1 to 10, characterized in that the polymeric carrier membrane is formed in multiple layers.

12. Sensor membrane according to claim 11, characterized in that the further dye and the indicator dye are fixed to mutually different layers of the polymeric support membrane.

13. Sensor membrane according to one of claims 1 to 12, characterized in that the polymeric support membrane is covered at least on one, the covalently bound indicator dye having, side with a porous, transparent cover layer.