WO2016109329A2 - Compositions, apparatus and methods for determining ph of an analyte solution - Google Patents
Compositions, apparatus and methods for determining ph of an analyte solution Download PDFInfo
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- WO2016109329A2 WO2016109329A2 PCT/US2015/067339 US2015067339W WO2016109329A2 WO 2016109329 A2 WO2016109329 A2 WO 2016109329A2 US 2015067339 W US2015067339 W US 2015067339W WO 2016109329 A2 WO2016109329 A2 WO 2016109329A2
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N31/00—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods
- G01N31/22—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods using chemical indicators
- G01N31/221—Investigating or analysing non-biological materials by the use of the chemical methods specified in the subgroup; Apparatus specially adapted for such methods using chemical indicators for investigating pH value
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
- G01N21/79—Photometric titration
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
- G01N21/80—Indicating pH value
Definitions
- kits and methods of the present invention are used to determine the pH of an analyte composition by adding the analyte composition to two different lyophilized titrant compositions in two different microwells and determining the pH of the analyte based on the response to the lyophilized titrant compositions.
- pH affects many chemical and biological processes in water.
- Conventional methods for determining pH include analyzing water samples in the field and in a laboratory setting. Analysis of water in a laboratory setting requires that the sample be titrated within 2 hours of receipt due to carbon dioxide from the air dissolving in the water, which may bring the pH toward 7.
- Field testing can be done using electronic hand-held devices that are dipped in the water and provide a digital readout of the pH. These devices can be calibrated to one pH buffer. Other methods involve adding a reagent to the sample that colors the sample water. The intensity of the color is proportional to the pH of the sample. The color is then matched against a standard color chart.
- Still other methods can include titrating solutions of interest to a pH where the entire titratable base is consumed.
- the end point of the titration is determined by adding a colorimetric pH indicator and using naked-eye detection of a change in color to determine the endpoint.
- These techniques suffer from many disadvantages.
- the visual technique requires the analyst to manually titrate a solution of acid, drop wise, into the analyte solution of interest.
- the visual technique requires the subjective determination of a color change.
- These two disadvantages often result in errors resulting from the analyst overshooting the endpoint due to adding too much strong acid or misjudging the color change at the endpoint.
- the analyst has to in many cases redo the titration method.
- manual titration is time-consuming.
- the pH devices usually include glass electrodes which are breakable and require calibration with buffer solutions.
- the present invention eliminates the drawbacks of traditional manual titrations by eliminating the subjective naked-eye determination and provides an accurate value as a user simply has to add the analyte solution or analyte solutions to each microwell in the plate instead of manually titrating each analyte solutions and/or having to calibrate the electrode in a laboratory setting prior to use. Furthermore, the present invention removes the subjective naked-eye determination of an endpoint by using a spectrophotometer to determine the endpoint.
- a pH assay kit can include a microwell plate, a first lyophilized titrant composition that includes a first pH sensitive dye, where the first pH sensitive dye is maximally sensitive to acidic solutions having a pH of 6 or less, and a second lyophilized titrant composition that includes a second pH sensitive dye, where the second pH sensitive dye is maximally sensitive to solutions having a pH of 4 or more.
- a plurality of the microwells contain the first lyophilized titrant composition that include the first pH sensitive dye and a plurality of different microwells contain the second lyophilized titrant composition that can include the second pH sensitive dye such that when an analyte composition is added to the lyophilized titrant compositions in each well of the plurality of wells, a solution forms and the pH of the solution is determined based on the response of the pH sensitive dyes to the solution.
- Each pH sensitive dye has an equivalence point and the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye.
- the microwell plate can include 6, 24, 96, 384, or 1536 microwells.
- the microwell plate includes one or more removable strips. Each of the strips that include 8 microwells and 3 microwells of the 8 microwells contain the first lyophilized titrant composition comprising the first pH sensitive dye and 4 microwells contain the second lyophilized titrant composition that include the second pH sensitive dye.
- the amounts of lyophilized titrant in the microwells can have different absorbance values in response to the pH of the analyte and the pH of the solution is determined based on the absorbance and a calibration chart.
- the plurality of microwells are sealed with a plastic film or a foil.
- the pH assay kit can also include a spectrophotometer capable of measuring wavelengths between 400 and 700 nanometers (nm).
- the titrant composition can include a pH sensitive dye capable of having a colorimetric response in response to a change in pH of the solution, an acid compound, and an excipient compound.
- the pH sensitive dyes can be any pH sensitive dye.
- the pH sensitive dyes can have a colorimetric response in a particular pH range.
- the pH sensitive dyes can have an acid form that has a different absorbance value than an absorbance value of a base form of the pH sensitive dyes.
- Non-limiting examples of the first pH sensitive dye and the second pH sensitive dyes are bromocresol green and neutral red, respectively.
- the first pH sensitive dye can have an absorbance from 440 to 460 nm and the second pH sensitive dye can have an absorbance from 510 to 530 nm.
- the composition can be a powder.
- the powder can be made by providing an aqueous solution of the titrant composition to one or more containers and subjecting at least one of the containers to lyophilizing conditions sufficient to remove the water from the aqueous solution to form the powder.
- the one or more containers are microwells of a microwell plate.
- the powder can be packaged (for example, a bag, vial, or encapsulated). The powder can be sold separately from the kit.
- other pH sensitive indicators can be used in place of or in combination with the first and second indicators.
- Non-limiting examples of other pH sensitive dyes include gentian violet, malachite green, thymol blue, Methyl yellow, Methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein.
- one or both of the titrant compositions can include an excipient.
- a non-limiting example of an excipient is poly glycol.
- polyethylene glycol is used as an excipient.
- An amount of excipient added to a titrant composition ranges from 1 to 10 wt.%, from 1.5 to 5 wt.%, or from 2 to 3 wt.% of the titrant composition.
- a system for determining the pH of an analyte composition can include the pH assay kit of the present invention, at least one detector configured to detect absorbances from the first and second pH sensitive dyes, a controller configured to monitor signals from the at least one detector, and a display device configured to display the pH of the analyte composition.
- the controller can include an algorithm for: comparing the maximum absorbance values of the first and second pH sensitive values with pH calibration curves for the first and second pH sensitive dyes to determine the pH of the solution.
- the detector is capable of measuring wavelengths between 400 and 700 nanometers. In some instances, the detector is capable of simultaneously measuring two wavelengths between 400 and 700 nanometers.
- the pH assay kit of the present invention and/or the system of the present invention can be used to determine the pH of an analyte composition or a plurality of analyte compositions.
- the method can include obtaining any one of the pH assay kits or systems described throughout this specification; obtaining an analyte composition; adding substantially the same volume of the analyte composition to each of the plurality of microwells of the microwell plate to form solutions from the analyte composition and the lyophilized titrate compositions in each of the plurality of microwells; and measuring the absorbance value for each solution in each of the plurality of microwells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values.
- the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye.
- the display value is compared to the value to a calibration curve of the first pH sensitive dye to quantify the pH. If the pH value for the samples is between 6 and 8, the display value is compared to the value to a calibration curve of the second pH sensitive dye to quantify the pH.
- the analyte can be obtained from a variety of sources such as a subsurface well, a hydrocarbon subsurface, a water well in a subsurface hydrocarbon formation, a pool, a pond, a drilling or tracking process, water treatment, a chemical processing site, a swimming pool, a lake, a paper processing site, or any combination thereof.
- the analyte solution is obtained from a hydrocarbon drilling or tracking process.
- pluralities of solutions having the same analyte are obtained, and each analyte solution is obtained from a different well of a plurality of subsurface wells.
- the pH assay kits described throughout the specification can be made by obtaining a microwell plate; obtaining a first lyophilized titrant composition that includes a first pH sensitive dye; obtaining a first lyophilized titrant composition that includes a second pH sensitive dye; and adding amounts of the lyophilized titrant composition to a plurality of wells of the microwell plate such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different.
- the kit is made by adding the first and second pH sensitive dyes to different microwells of a strip of microwells and b) lyophilizing the titrant solutions.
- the plurality of microwells can be sealed with a cap, foil, or a plastic film to inhibit or prevent the titrant composition from exiting the plurality of microwells.
- the first titrant composition consists essentially of bromocresol green and the second titrant composition consists essentially of neutral red.
- a pH assay kit in the context of the present invention 37 embodiments are described.
- the pH assay kit can include (a) a microwell plate; (b) a first lyophilized titrant composition that includes a first pH sensitive dye, wherein the first pH sensitive dye is sensitive to acidic solutions having a pH of 6 or less; and (c) a second lyophilized titrant composition that includes a second pH sensitive dye, wherein the second pH sensitive dye is sensitive to solutions having a pH of 4 or more; wherein a plurality of the microwells contain the first lyophilized titrant composition that includes the first pH sensitive dye and a plurality of different microwells contain the second lyophilized titrant composition comprising the second pH sensitive dye such that when an analyte composition is added to the lyophilized titrant compositions in each well of the plurality of wells, a solution forms and the pH of the solution in is determined based on the response of the pH sensitive dye
- Embodiment 2 is the pH assay kit of embodiment 1, wherein the microwell plate can include 6, 24, 96, 384, or 1536 wells.
- Embodiment 3 is the pH assay kit of embodiment 2, wherein the microwell plate includes one or more removable strips, wherein each of the strips comprise 8 microwells, wherein 3 microwells of the 8 microwells contain the first lyophilized titrant composition that includes the first pH sensitive dye and 4 microwells contain the second lyophilized titrant composition that include the second pH sensitive dye.
- Embodiment 4 is the pH assay kit of embodiment 3, wherein the microwell plate includes 3 strips or 12 strips or more.
- Embodiment 5 is the pH assay kit of embodiment 4, wherein a portion of the strips are sealed to prevent the titrant composition from exiting the plurality of wells.
- Embodiment 6 is the pH assay kit of embodiment 5, wherein the plurality of wells is sealed with a cap, a plastic film or a foil.
- Embodiment 7 is the pH assay kit of any one of embodiments 4 to 6, wherein at least one of the strips is used as to calibrate the assay.
- Embodiment 8 is the pH assay kit of embodiment 7, wherein the strip used as to calibrate the assay is not sealed.
- Embodiment 9 is the pH assay kit of any one of embodiments 1 to 8, wherein each pH sensitive dye has an equivalence point, wherein the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye.
- Embodiment 10 is the pH assay kit of any one of embodiments 1 to 9, wherein the absorbance of the first pH sensitive dye is 440 to 460 nm.
- Embodiment 11 is the pH assay kit of any one of embodiments 1 to 10, wherein the absorbance of the second pH sensitive dye is 510 to 530 nm.
- Embodiment 12 is the pH assay kit of embodiments 1 to 11, wherein the first pH sensitive dye is bromocresol green.
- Embodiment 13 is the pH assay kit of any one of embodiments 1 to 12, wherein the second pH sensitive dye is neutral red. gentian violet, malachite green, thymol blue, methyl yellow, methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein.
- Embodiment 14 is the pH assay kit of embodiment 14, wherein the second pH sensitive dye is neutral red.
- Embodiment 15 is the pH assay kit of any one of embodiments 1 to 14, further includes an excipient.
- Embodiment 16 is the pH assay kit of embodiment 15, wherein the excipient includes one or more poly glycol compounds, preferably polyethylene glycol.
- Embodiment 17 is the pH assay kit of any one of embodiments 1 to 16, further including calibration solutions or calibration curves for each of the pH sensitive dyes.
- Embodiment 18 is a system for determining the pH of an analyte composition.
- the system can include (a) a pH assay kit as claimed in any one of claims 1 to 15; (b) at least one detector configured to detect absorbances from the first and second pH sensitive dyes; (c) at controller configured to monitor signals from the at least one detector, wherein the controller includes an algorithm for: comparing the maximum absorbance values of the first and second pH sensitive values with pH calibration curves for the first and second pH sensitive dyes to determine the pH of the solution; and (d) a display unit configured to display the pH of the analyte composition.
- Embodiment 19 is the system of embodiment 18, wherein the detector is capable of measuring wavelengths between 400 and 700 nanometers.
- Embodiment 20 is the system of embodiment 18, wherein the detector is capable of simultaneously measuring two wavelengths between 400 and 700 nanometers.
- Embodiment 21 is a method of determining the pH of an analyte composition. The method can include (a) obtaining any one of the pH assay kits of embodiments 1 to 17 or the system of any one of embodiments 18 to 20; (b) obtaining an analyte composition; (c) adding substantially the same volume of the analyte composition to each of the plurality of wells of the microwell plate to form solutions from the analyte composition and the lyophilized titrate compositions in each of the plurality of wells, wherein the pH the solution in at least two of the microwells is different; and (d) measuring the absorbance value for each solution in each of the plurality of wells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values.
- Embodiment 22 is the method of embodiment 21, wherein each pH sensitive dye has an equivalence point, wherein the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye.
- Embodiment 23 is the method of any of embodiments 21 to 22, wherein the pH sensitive dye is bromocresol green.
- Embodiment 24 is the method of any one of embodiments 21 to 23, wherein the second pH sensitive dye is neutral red, gentian violet, malachite green, thymol blue, methyl yellow, methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein, or any combination thereof.
- Embodiment 25 is the method of embodiment 24, wherein the second pH sensitive dye is neutral red.
- Embodiment 26 is the method of any one of embodiments 21 to 25, wherein determining the pH can include (i) determining if the pH value for the samples is between 3 and 6, and comparing the value to a calibration curve of the first pH sensitive dye to quantify the pH; (ii) determining if the pH value for the samples is between 6 and 8, and (iii) comparing the value to a calibration curve of the second pH sensitive dye to quantify the pH.
- Embodiment 27 is the method of any one of embodiments 21 to 26, wherein the analyte in is an aqueous composition obtained from a subsurface well.
- Embodiment 28 is the method of any one of embodiments 21 to 27, wherein the analyte solution includes a plurality of solutions having the same analyte, and each analyte solution is obtained from a different well of a plurality of subsurface wells.
- Embodiment 29 is the method of embodiments 21 to 28, wherein the well is a hydrocarbon well or a water well in a subsurface hydrocarbon formation.
- Embodiment 30 is the method of any one of embodiments 21 to 29, wherein the analyte solution is obtained from a drilling or fracking process, water treatment, a chemical processing site, a swimming pool, a lake, a paper processing site, or any combination thereof.
- Embodiment 31 is a method of making any one of the pH assay kits of embodiments 1 to 17.
- the method can include (a) obtaining a microwell plate; (b) obtaining a first lyophilized titrant composition that includes a first pH sensitive dye; (c) obtaining a second lyophilized titrant composition that includes a second pH sensitive dye; and (d) adding amounts of the lyophilized titrant composition to a plurality of wells of the microwell plate such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different.
- Embodiment 32 is the method of embodiment 31, wherein at least a majority of the plurality of wells are sealed to prevent the titrant composition from exiting the plurality of wells.
- Embodiment 33 is the method of embodiment 32, wherein the plurality of wells are sealed with a cap, a plastic film or a foil.
- Embodiment 34 is a method of making any one of the pH assay kits of embodiments 1 to 17.
- the method can include (a) obtaining a microwell plate; (b) obtaining a first titrant composition that includes a first pH sensitive dye; (c) obtaining a second titrant composition that includes a second pH sensitive dye; and (d) subjecting the microwell plate to lyophilizing conditions such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different.
- Embodiment 35 is the method of embodiment 34, wherein at least a majority of the plurality of wells are sealed to prevent the titrant composition from exiting the plurality of wells.
- Embodiment 36 is the method of embodiment 35, wherein the plurality of wells are sealed with a cap, a plastic film or a foil.
- Embodiment 37 is the method of any one of embodiments 34 to 36, further including providing an excipient to at least one of the titrant compositions. [0016] The following includes definitions of various terms and phrases used throughout this specification.
- acidic solution refers to a solution that has a concentration of hydrogen ions greater than the concentration of hydroxide ion ([H+] > [OH ]).
- basic solution or "alkaline solution” refers to a solution that has a concentration of hydrogen ions less than the concentration of hydroxide ion ([H+] ⁇ [OH ]).
- substantially and its variations are defined as being largely but not necessarily wholly what is specified as understood by one of ordinary skill in the art, and in one non-limiting embodiment substantially refers to ranges within 10%, within 5%, within 1%, or within 0.5%.
- the pH assay kits and the methods of using and making the pH assay kits of the present invention can "comprise,” “consist essentially of,” or “consist of particular ingredients, components, compositions, etc. disclosed throughout the specification.
- a basic and novel characteristic of the kits of the present invention is the ability to determine the pH of an aqueous solution using spectrometric analysis.
- FIGS. 1A and IB are schematics of pH assay kits of the present invention.
- FIG. 2 is a schematic of a system for determining pH of an analyte solution.
- FIG. 3 is a graph of pH versus absorbance at 450 nm for bromocresol green.
- FIG. 4 is a graph of pH versus absorbance a 520 nm for neutral red.
- Each microwell of the microwell of the microwell plates has at least two microwells having different pH sensitive dyes.
- the analyte solution is added to the titrant to form a solution and the pH of the solution is determined by measuring the absorbance value for each solution in each of the plurality of microwells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values.
- FIGS. 1A and IB depict schematics of embodiments of pH assay system 100.
- the pH assay system or kit includes microwell plate 102 having a plurality of microwells 104 and 106.
- the plurality of microwells 104 can be assembled in the removable holders 106.
- Holders 106 may include members 108 that position on top of the side wall 110. Holders 106 may rest on, or be suspended above, bottom wall 112 of the microwell plate 102.
- holder 106 includes eight (8) microwells 104, however, it should be understood that the number of microwells can be adjusted to the size of the microwell plate 102.
- the number of the microwells 104 can be 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, etc.
- the microwell plate 102 does not include any titrant composition.
- FIG. IB depicts a schematic of the microwells having titrant composition 116 and 118 in the microwells.
- the microwells 104 can hold a volume of 20, 50, 300, 500 microliters, preferably 300 microliters or 400 microliters.
- the microwell plate 102, microwells 104, holders 106 can be made of any material having chemical resistance to acid.
- Non-limiting examples of materials include polymers, copolymers of polymers, polystyrene, polypropylene, cyclo-olefins and the like.
- the holders 106 may be polymeric or plastic tape with the microwells 104 embossed on the tape. Microwell plates are commercially available from Thermo Fisher Scientific (Waltham, MA, USA).
- the microwells 104 and 106 can be filled with two different pH sensitive dyes.
- four microwells 104 can be filled with the an aqueous solution of a desired amount of the first pH sensitive dye and three microwells 106 can be filled with an aqueous solution of a desired amount the second pH sensitive dye.
- the microwells 104 are each filled with 100 microliter ( ⁇ ) of a 40 micromolar ( ⁇ ) solution of bromocresol green and microwells 106 are each filled with a 100 ⁇ ⁇ of a 100 ⁇ solution of neutral red.
- the microwells 104 in each holder 106 can include the first pH sensitive dye
- microwells 104' can include the second pH sensitive dye
- microwell 104" is empty.
- dyes and excipients can be lyophilized in the microwells 104, 104' in the microwell plate 102. Lyophilizing conditions include -60 degree Celsius at 100 mtorr.
- the microwells 104, 104', microwell holders 106, and/or the microwell plate can be sealed with a known sealing agent (for example, caps, plastic film or foil) to allow the microwell plate 102 or the microwell holders 106 to stored or transported.
- a known sealing agent for example, caps, plastic film or foil
- the pH assay system includes a spectrophotometer that is capable of measuring the absorbance of the chosen colorimetric dye.
- the titrant composition can include a pH sensitive dye capable of having a colorimetric response in response to a change in pH of the solution, an acid compound, and an excipient compound.
- the pH sensitive dyes can be any pH sensitive dye.
- the pH sensitive dyes can have a colorimetric response in a particular pH range.
- the pH sensitive dyes can have an acid form that has a different absorbance value than an absorbance value of a base form of the pH sensitive dyes.
- Non-limiting examples of the first pH sensitive dye and the second pH sensitive dyes are bromocresol green and neutral red, respectively.
- Non- limiting examples of other pH sensitive dyes include gentian violet, malachite green, thymol blue, Methyl yellow, Methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein. pH sensitive dyes are commercially available from several sources. A non- limiting example of a commercial source is Sigma- Aldrich® (USA).
- one or both of the titrant compositions can include an excipient.
- an excipient is poly glycol.
- polyethylene glycol is used as an excipient.
- the polyethylene glycol can have an average molecular weight from 3600 to 4400.
- Polyethylene glycol is commercially available from many sources.
- a non-limiting example of a commercially available polyethylene glycol is CARBOWAXTM manufactured by Dow Chemical Company (Midland, MI, USA).
- the excipient can aid in the lyophilization process.
- An amount of excipient added to a titrant composition ranges from 1 to 10 wt.%, from 1.5 to 5 wt.%, or from 2 to 3 wt.% of the titrant composition.
- the composition can be a powder.
- the powder can be made by providing an aqueous solution of the titrant composition to one or more containers and subjecting at least one of the containers to lyophilizing conditions sufficient to remove the water from the aqueous solution to form the powder.
- the one or more containers are microwells of a microwell plate.
- the powder can be packaged (for example, a bag, vial, or encapsulated).
- the powder can be sold separately from the kit.
- other pH sensitive indicators can be used in place of or in combination with the first and second indicators.
- the pH assay system and kit described throughout the specification can be used to determine the pH of an analyte solution.
- the solution can be a sample from a water body such as a subsurface water well in a hydrocarbon formation, a lake, a river, a canal, a pool, a pond, or the like.
- a schematic for determining pH is depicted.
- system 200 the microwell plate 102 containing the lyophilized titrant compositions in microwells 104, 104 is obtained.
- a known amount of analyte solution 202 (for example 300 microliters) is added to the lyophilized titrant composition in the microwells 104, 104', using a delivery apparatus (for example, multichannel pipette) to produce microwell plate 204.
- a delivery apparatus for example, multichannel pipette
- the microwell plate 204 is placed in a detector 206 (for example, a plate reader) and the absorbances of each microwell at the wavelengths of the colorimetric dyes are measured.
- the colorimetric dyes are bromocresol green and neutral red, the absorbances at 450 nm and 520 nm are measured.
- At least one of the solutions in microwells 104, 104' has an absorbance value at the measured wavelengths that is not statistically differentiable from the absorbance values of a solution with a pH value below that at which the dye shows a colorimetric response.
- a controller system 208 coupled to the detector 206 includes a software algorithm that is used to determine whether to use the neutral red or bromocresol green data. The data collected at both wavelengths is collected and plotted against the calibration curves. If the pH returned is within 3 to 6, then the bromocresol green data is used. If the pH returned is 6 to 8, the neutral red data is used.
- the pH of the analyte taken from a single well can be determined using one sample rather than using multiple samples and many indicators and/or buffer systems.
- the controller system 208 can include components such as CPUs or applications with an associated machine readable medium or article which may store an instruction or a set of instructions that, if executed by a machine, may cause the machine to perform a method and/or operations in accordance with the methods of the present invention.
- the measured absorbance from microwell plate 204 can be stored in a computer system in the spectrophotometer and/or transmitted to another computer system.
- Either computer may be capable of processing the absorbance and displaying or printing an alkalinity value for a series of analytes.
- Such a machine may include, for example, any suitable processing platform, computing platform, computing device, processing device, computing system, processing system, computer, processor, or the like, and may be implemented using any suitable combination of hardware and/or software.
- the machine-readable medium or article may include, for example, any suitable type of memory unit, memory device, memory article, memory medium, storage device, storage article, storage medium and/or storage unit, for example, memory, removable or non-removable media, erasable or non-erasable media, writeable or re-writeable media, digital or analog media, hard disk, floppy disk, Compact Disk Read Only Memory (CD-ROM), Compact Disk Recordable (CD-R), Compact Disk Rewriteable (CD-RW), optical disk, magnetic media, magneto-optical media, removable memory cards or disks, various types of Digital Versatile Disk (DVD), a tape, a cassette, or the like.
- memory removable or non-removable media, erasable or non-erasable media, writeable or re-writeable media, digital or analog media, hard disk, floppy disk, Compact Disk Read Only Memory (CD-ROM), Compact Disk Recordable (CD-R), Compact Disk Rewriteable (CD-RW), optical disk, magnetic
- the instructions may include any suitable type of code, such as source code, compiled code, interpreted code, executable code, static code, dynamic code, and the like.
- the instructions may be implemented using any suitable high-level, low-level, object- oriented, visual, compiled and/or interpreted programming language, such as C, C++, Java, BASIC, Perl, Matlab, Pascal, Visual BASIC, assembly language, machine code, and so forth.
- the computer system may further include a display device such as monitor, an alphanumeric input device such as keyboard, and a directional input device such as mouse.
- Calibration curves were established for the two pH sensitive dyes by adding water of a known pH and measuring the absorbance. Calibration curves for the wells containing bromocresol green were developed using the absorbance at 450 nm (FIG. 3), and the curves for wells containing neutral red were developed using the absorbance at 520 nm (FIG 4).
- Table 1 lists pH data of 14 oil and gas fluids determined using the assay of the present invention and a pH glass electrode. As shown in Table 1, the assay of the present invention provides similar results to that of the pH determined using a conventional glass electrode.
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Abstract
Compositions, kits and methods of using the kits and compositions to determine the pH of an analyte solution are described. The kit can include a lyophilized titrant.
Description
DESCRIPTION
COMPOSITIONS, APPARATUS AND METHODS FOR DETERMINING PH OF AN
ANALYTE SOLUTION
CROSS REFERENCE TO RELATED APPLICATIONS [0001] This application claims the benefit of U.S. Provisional Application No. 62/097,922, filed December 30, 2014, which is incorporated by reference herein.
BACKGROUND OF THE INVENTION
A. Field of the Invention
[0002] The invention generally concerns the determination of pH of a solution. In particular, the kits and methods of the present invention are used to determine the pH of an analyte composition by adding the analyte composition to two different lyophilized titrant compositions in two different microwells and determining the pH of the analyte based on the response to the lyophilized titrant compositions.
B. Description of Related Art [0003] pH affects many chemical and biological processes in water. Conventional methods for determining pH include analyzing water samples in the field and in a laboratory setting. Analysis of water in a laboratory setting requires that the sample be titrated within 2 hours of receipt due to carbon dioxide from the air dissolving in the water, which may bring the pH toward 7. Field testing can be done using electronic hand-held devices that are dipped in the water and provide a digital readout of the pH. These devices can be calibrated to one pH buffer. Other methods involve adding a reagent to the sample that colors the sample water. The intensity of the color is proportional to the pH of the sample. The color is then matched against a standard color chart. Still other methods can include titrating solutions of interest to a pH where the entire titratable base is consumed. The end point of the titration is determined by adding a colorimetric pH indicator and using naked-eye detection of a change in color to determine the endpoint. These techniques suffer from many disadvantages. First, the visual technique requires the analyst to manually titrate a solution of acid, drop wise, into the analyte solution of interest. Secondly, the visual technique requires the subjective determination of a color change. These two disadvantages often result in errors resulting
from the analyst overshooting the endpoint due to adding too much strong acid or misjudging the color change at the endpoint. Thus, the analyst has to in many cases redo the titration method. Also, manual titration is time-consuming. The pH devices usually include glass electrodes which are breakable and require calibration with buffer solutions. SUMMARY OF THE INVENTION
[0004] A solution to the disadvantages of a visual titration and glass electrode methods has been discovered. In particular, the solution resides in the use of two different lyophilized titrant samples in different microwells of a microwell plate. Analyte samples are added to the lyophilized samples and the absorbance of the resulting samples at two different wavelengths is measured and the pH of the analyte composition is determined based on the measured absorbance values. Notably, the present invention eliminates the drawbacks of traditional manual titrations by eliminating the subjective naked-eye determination and provides an accurate value as a user simply has to add the analyte solution or analyte solutions to each microwell in the plate instead of manually titrating each analyte solutions and/or having to calibrate the electrode in a laboratory setting prior to use. Furthermore, the present invention removes the subjective naked-eye determination of an endpoint by using a spectrophotometer to determine the endpoint.
[0005] In one aspect of the invention, there is disclosed a pH assay kit. The kit can include a microwell plate, a first lyophilized titrant composition that includes a first pH sensitive dye, where the first pH sensitive dye is maximally sensitive to acidic solutions having a pH of 6 or less, and a second lyophilized titrant composition that includes a second pH sensitive dye, where the second pH sensitive dye is maximally sensitive to solutions having a pH of 4 or more. A plurality of the microwells contain the first lyophilized titrant composition that include the first pH sensitive dye and a plurality of different microwells contain the second lyophilized titrant composition that can include the second pH sensitive dye such that when an analyte composition is added to the lyophilized titrant compositions in each well of the plurality of wells, a solution forms and the pH of the solution is determined based on the response of the pH sensitive dyes to the solution. Each pH sensitive dye has an equivalence point and the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye. The microwell plate can include 6, 24, 96, 384, or 1536 microwells. In some aspects of the invention, the microwell plate includes one or more removable strips. Each of the strips that
include 8 microwells and 3 microwells of the 8 microwells contain the first lyophilized titrant composition comprising the first pH sensitive dye and 4 microwells contain the second lyophilized titrant composition that include the second pH sensitive dye. The amounts of lyophilized titrant in the microwells can have different absorbance values in response to the pH of the analyte and the pH of the solution is determined based on the absorbance and a calibration chart. In some instances, the plurality of microwells are sealed with a plastic film or a foil. The pH assay kit can also include a spectrophotometer capable of measuring wavelengths between 400 and 700 nanometers (nm).
[0006] In some instances, the titrant composition can include a pH sensitive dye capable of having a colorimetric response in response to a change in pH of the solution, an acid compound, and an excipient compound. The pH sensitive dyes can be any pH sensitive dye. The pH sensitive dyes can have a colorimetric response in a particular pH range. In some instances, the pH sensitive dyes can have an acid form that has a different absorbance value than an absorbance value of a base form of the pH sensitive dyes. Non-limiting examples of the first pH sensitive dye and the second pH sensitive dyes are bromocresol green and neutral red, respectively. The first pH sensitive dye can have an absorbance from 440 to 460 nm and the second pH sensitive dye can have an absorbance from 510 to 530 nm. The composition can be a powder. The powder can be made by providing an aqueous solution of the titrant composition to one or more containers and subjecting at least one of the containers to lyophilizing conditions sufficient to remove the water from the aqueous solution to form the powder. In some instances, the one or more containers are microwells of a microwell plate. The powder can be packaged (for example, a bag, vial, or encapsulated). The powder can be sold separately from the kit. In some embodiments, other pH sensitive indicators can be used in place of or in combination with the first and second indicators. Non-limiting examples of other pH sensitive dyes include gentian violet, malachite green, thymol blue, Methyl yellow, Methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein. In some instances, one or both of the titrant compositions can include an excipient. A non-limiting example of an excipient is poly glycol. In a preferred aspect, polyethylene glycol is used as an excipient. An amount of excipient added to a titrant composition ranges from 1 to 10 wt.%, from 1.5 to 5 wt.%, or from 2 to 3 wt.% of the titrant composition.
[0007] In one aspect of the present invention, a system for determining the pH of an analyte composition is described. The system can include the pH assay kit of the present
invention, at least one detector configured to detect absorbances from the first and second pH sensitive dyes, a controller configured to monitor signals from the at least one detector, and a display device configured to display the pH of the analyte composition. The controller can include an algorithm for: comparing the maximum absorbance values of the first and second pH sensitive values with pH calibration curves for the first and second pH sensitive dyes to determine the pH of the solution. The detector is capable of measuring wavelengths between 400 and 700 nanometers. In some instances, the detector is capable of simultaneously measuring two wavelengths between 400 and 700 nanometers.
[0008] The pH assay kit of the present invention and/or the system of the present invention can be used to determine the pH of an analyte composition or a plurality of analyte compositions. The method can include obtaining any one of the pH assay kits or systems described throughout this specification; obtaining an analyte composition; adding substantially the same volume of the analyte composition to each of the plurality of microwells of the microwell plate to form solutions from the analyte composition and the lyophilized titrate compositions in each of the plurality of microwells; and measuring the absorbance value for each solution in each of the plurality of microwells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values. The absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye. In one instance, if the pH value for the samples is between 3 and 6, the display value is compared to the value to a calibration curve of the first pH sensitive dye to quantify the pH. If the pH value for the samples is between 6 and 8, the display value is compared to the value to a calibration curve of the second pH sensitive dye to quantify the pH.
[0009] In one instance, the analyte can be obtained from a variety of sources such as a subsurface well, a hydrocarbon subsurface, a water well in a subsurface hydrocarbon formation, a pool, a pond, a drilling or tracking process, water treatment, a chemical processing site, a swimming pool, a lake, a paper processing site, or any combination thereof. In some instances, the analyte solution is obtained from a hydrocarbon drilling or tracking process. In some instances, pluralities of solutions having the same analyte are obtained, and each analyte solution is obtained from a different well of a plurality of subsurface wells.
[0010] The pH assay kits described throughout the specification can be made by obtaining a microwell plate; obtaining a first lyophilized titrant composition that includes a
first pH sensitive dye; obtaining a first lyophilized titrant composition that includes a second pH sensitive dye; and adding amounts of the lyophilized titrant composition to a plurality of wells of the microwell plate such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different. In some instances, the kit is made by adding the first and second pH sensitive dyes to different microwells of a strip of microwells and b) lyophilizing the titrant solutions. The plurality of microwells can be sealed with a cap, foil, or a plastic film to inhibit or prevent the titrant composition from exiting the plurality of microwells. In some embodiments, the first titrant composition consists essentially of bromocresol green and the second titrant composition consists essentially of neutral red.
[0011] In the context of the present invention 37 embodiments are described. In a first embodiment a pH assay kit is described. The pH assay kit can include (a) a microwell plate; (b) a first lyophilized titrant composition that includes a first pH sensitive dye, wherein the first pH sensitive dye is sensitive to acidic solutions having a pH of 6 or less; and (c) a second lyophilized titrant composition that includes a second pH sensitive dye, wherein the second pH sensitive dye is sensitive to solutions having a pH of 4 or more; wherein a plurality of the microwells contain the first lyophilized titrant composition that includes the first pH sensitive dye and a plurality of different microwells contain the second lyophilized titrant composition comprising the second pH sensitive dye such that when an analyte composition is added to the lyophilized titrant compositions in each well of the plurality of wells, a solution forms and the pH of the solution in is determined based on the response of the pH sensitive dyes to the solution. Embodiment 2 is the pH assay kit of embodiment 1, wherein the microwell plate can include 6, 24, 96, 384, or 1536 wells. Embodiment 3 is the pH assay kit of embodiment 2, wherein the microwell plate includes one or more removable strips, wherein each of the strips comprise 8 microwells, wherein 3 microwells of the 8 microwells contain the first lyophilized titrant composition that includes the first pH sensitive dye and 4 microwells contain the second lyophilized titrant composition that include the second pH sensitive dye. Embodiment 4 is the pH assay kit of embodiment 3, wherein the microwell plate includes 3 strips or 12 strips or more. Embodiment 5 is the pH assay kit of embodiment 4, wherein a portion of the strips are sealed to prevent the titrant composition from exiting the plurality of wells. Embodiment 6 is the pH assay kit of embodiment 5, wherein the plurality of wells is sealed with a cap, a plastic film or a foil. Embodiment 7 is the pH assay kit of any one of embodiments 4 to 6, wherein at least one of the strips is used as to calibrate the assay.
Embodiment 8 is the pH assay kit of embodiment 7, wherein the strip used as to calibrate the assay is not sealed. Embodiment 9 is the pH assay kit of any one of embodiments 1 to 8, wherein each pH sensitive dye has an equivalence point, wherein the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye. Embodiment 10 is the pH assay kit of any one of embodiments 1 to 9, wherein the absorbance of the first pH sensitive dye is 440 to 460 nm. Embodiment 11 is the pH assay kit of any one of embodiments 1 to 10, wherein the absorbance of the second pH sensitive dye is 510 to 530 nm. Embodiment 12 is the pH assay kit of embodiments 1 to 11, wherein the first pH sensitive dye is bromocresol green. Embodiment 13 is the pH assay kit of any one of embodiments 1 to 12, wherein the second pH sensitive dye is neutral red. gentian violet, malachite green, thymol blue, methyl yellow, methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein. Embodiment 14 is the pH assay kit of embodiment 14, wherein the second pH sensitive dye is neutral red. Embodiment 15 is the pH assay kit of any one of embodiments 1 to 14, further includes an excipient. Embodiment 16 is the pH assay kit of embodiment 15, wherein the excipient includes one or more poly glycol compounds, preferably polyethylene glycol. Embodiment 17 is the pH assay kit of any one of embodiments 1 to 16, further including calibration solutions or calibration curves for each of the pH sensitive dyes.
[0012] Embodiment 18 is a system for determining the pH of an analyte composition. The system can include (a) a pH assay kit as claimed in any one of claims 1 to 15; (b) at least one detector configured to detect absorbances from the first and second pH sensitive dyes; (c) at controller configured to monitor signals from the at least one detector, wherein the controller includes an algorithm for: comparing the maximum absorbance values of the first and second pH sensitive values with pH calibration curves for the first and second pH sensitive dyes to determine the pH of the solution; and (d) a display unit configured to display the pH of the analyte composition. Embodiment 19 is the system of embodiment 18, wherein the detector is capable of measuring wavelengths between 400 and 700 nanometers. Embodiment 20 is the system of embodiment 18, wherein the detector is capable of simultaneously measuring two wavelengths between 400 and 700 nanometers. [0013] Embodiment 21 is a method of determining the pH of an analyte composition. The method can include (a) obtaining any one of the pH assay kits of embodiments 1 to 17 or the system of any one of embodiments 18 to 20; (b) obtaining an analyte composition; (c)
adding substantially the same volume of the analyte composition to each of the plurality of wells of the microwell plate to form solutions from the analyte composition and the lyophilized titrate compositions in each of the plurality of wells, wherein the pH the solution in at least two of the microwells is different; and (d) measuring the absorbance value for each solution in each of the plurality of wells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values. Embodiment 22 is the method of embodiment 21, wherein each pH sensitive dye has an equivalence point, wherein the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye. Embodiment 23 is the method of any of embodiments 21 to 22, wherein the pH sensitive dye is bromocresol green. Embodiment 24 is the method of any one of embodiments 21 to 23, wherein the second pH sensitive dye is neutral red, gentian violet, malachite green, thymol blue, methyl yellow, methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein, or any combination thereof. Embodiment 25 is the method of embodiment 24, wherein the second pH sensitive dye is neutral red. Embodiment 26 is the method of any one of embodiments 21 to 25, wherein determining the pH can include (i) determining if the pH value for the samples is between 3 and 6, and comparing the value to a calibration curve of the first pH sensitive dye to quantify the pH; (ii) determining if the pH value for the samples is between 6 and 8, and (iii) comparing the value to a calibration curve of the second pH sensitive dye to quantify the pH. Embodiment 27 is the method of any one of embodiments 21 to 26, wherein the analyte in is an aqueous composition obtained from a subsurface well. Embodiment 28 is the method of any one of embodiments 21 to 27, wherein the analyte solution includes a plurality of solutions having the same analyte, and each analyte solution is obtained from a different well of a plurality of subsurface wells. Embodiment 29 is the method of embodiments 21 to 28, wherein the well is a hydrocarbon well or a water well in a subsurface hydrocarbon formation. Embodiment 30 is the method of any one of embodiments 21 to 29, wherein the analyte solution is obtained from a drilling or fracking process, water treatment, a chemical processing site, a swimming pool, a lake, a paper processing site, or any combination thereof. [0014] Embodiment 31 is a method of making any one of the pH assay kits of embodiments 1 to 17. The method can include (a) obtaining a microwell plate; (b) obtaining a first lyophilized titrant composition that includes a first pH sensitive dye; (c) obtaining a second lyophilized titrant composition that includes a second pH sensitive dye; and (d)
adding amounts of the lyophilized titrant composition to a plurality of wells of the microwell plate such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different. Embodiment 32 is the method of embodiment 31, wherein at least a majority of the plurality of wells are sealed to prevent the titrant composition from exiting the plurality of wells. Embodiment 33 is the method of embodiment 32, wherein the plurality of wells are sealed with a cap, a plastic film or a foil.
[0015] Embodiment 34 is a method of making any one of the pH assay kits of embodiments 1 to 17. The method can include (a) obtaining a microwell plate; (b) obtaining a first titrant composition that includes a first pH sensitive dye; (c) obtaining a second titrant composition that includes a second pH sensitive dye; and (d) subjecting the microwell plate to lyophilizing conditions such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different. Embodiment 35 is the method of embodiment 34, wherein at least a majority of the plurality of wells are sealed to prevent the titrant composition from exiting the plurality of wells. Embodiment 36 is the method of embodiment 35, wherein the plurality of wells are sealed with a cap, a plastic film or a foil. Embodiment 37 is the method of any one of embodiments 34 to 36, further including providing an excipient to at least one of the titrant compositions. [0016] The following includes definitions of various terms and phrases used throughout this specification.
[0017] The term "acidic solution" refers to a solution that has a concentration of hydrogen ions greater than the concentration of hydroxide ion ([H+] > [OH ]).
[0018] The terms "basic solution" or "alkaline solution" refers to a solution that has a concentration of hydrogen ions less than the concentration of hydroxide ion ([H+] < [OH ]).
[0019] The term "pH" refers to the measurement of the concentration of hydrogen ions in water or other media. pH is generally expressed as a log scale based on 10 where pH = - log[H+].
[0020] The term "about" or "approximately" are defined as being close to as understood by one of ordinary skill in the art, and in one non-limiting embodiment the terms are defined
to be within 10%, preferably within 5%, more preferably within 1%, and most preferably within 0.5%.
[0021] The term "substantially" and its variations are defined as being largely but not necessarily wholly what is specified as understood by one of ordinary skill in the art, and in one non-limiting embodiment substantially refers to ranges within 10%, within 5%, within 1%, or within 0.5%.
[0022] The terms "inhibiting" or "reducing" or "preventing" or "avoiding" or any variation of these terms, when used in the claims and/or the specification includes any measurable decrease or complete inhibition to achieve a desired result. [0023] The term "effective," as that term is used in the specification and/or claims, means adequate to accomplish a desired, expected, or intended result.
[0024] The use of the word "a" or "an" when used in conjunction with the term "comprising" in the claims or the specification may mean "one," but it is also consistent with the meaning of "one or more," "at least one," and "one or more than one." [0025] The words "comprising" (and any form of comprising, such as "comprise" and "comprises"), "having" (and any form of having, such as "have" and "has"), "including" (and any form of including, such as "includes" and "include") or "containing" (and any form of containing, such as "contains" and "contain") are inclusive or open-ended and do not exclude additional, unrecited elements or method steps. [0026] The pH assay kits and the methods of using and making the pH assay kits of the present invention can "comprise," "consist essentially of," or "consist of particular ingredients, components, compositions, etc. disclosed throughout the specification. With respect to the transitional phase "consisting essentially of," in one non-limiting aspect, a basic and novel characteristic of the kits of the present invention is the ability to determine the pH of an aqueous solution using spectrometric analysis.
[0027] Other objects, features and advantages of the present invention will become apparent from the following figures, detailed description, and examples. It should be understood, however, that the figures, detailed description, and examples, while indicating specific embodiments of the invention, are given by way of illustration only and are not
meant to be limiting. Additionally, it is contemplated that changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. In further embodiments, features from specific embodiments may be combined with features from other embodiments. For example, features from one embodiment may be combined with features from any of the other embodiments. In further embodiments, additional features may be added to the specific embodiments described herein.
BRIEF DESCRIPTION OF THE DRAWINGS
[0028] Advantages of the present invention may become apparent to those skilled in the art with the benefit of the following detailed description and upon reference to the accompanying drawings.
[0029] FIGS. 1A and IB are schematics of pH assay kits of the present invention.
[0030] FIG. 2 is a schematic of a system for determining pH of an analyte solution.
[0031] FIG. 3 is a graph of pH versus absorbance at 450 nm for bromocresol green.
[0032] FIG. 4 is a graph of pH versus absorbance a 520 nm for neutral red.
[0033] While the invention is susceptible to various modifications and alternative forms, specific embodiments thereof are shown by way of example in the drawings and may herein be described in detail. The drawings may not be to scale.
DETAILED DESCRIPTION OF THE INVENTION
[0034] Conventional technologies used to determine the pH of a solution involve visual titration methods and/or glass electrodes that are time-consuming, often inaccurate, and expensive. Many time, manual visual titrations result in error resulting from the analyst overshooting the endpoint due to adding too much strong acid or misjudging the color change at the endpoint. Glass electrodes require that the electrodes be kept in a buffered solution and calibrated prior to use. A discovery has been made that avoids overshooting the endpoint and eliminating the need for a visual titration and/or the use of glass electrodes The discovery lies in the use of a lyophilized titrant sample used in a microwell plate. The titrant sample
can include a lyophilized pH sensitive dye. Each microwell of the microwell of the microwell plates has at least two microwells having different pH sensitive dyes. The analyte solution is added to the titrant to form a solution and the pH of the solution is determined by measuring the absorbance value for each solution in each of the plurality of microwells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values.
[0035] These and other non-limiting aspects of the present invention are discussed in further detail in the following sections.
A. pH Assay Kit [0036] FIGS. 1A and IB depict schematics of embodiments of pH assay system 100. The pH assay system or kit includes microwell plate 102 having a plurality of microwells 104 and 106. The plurality of microwells 104 can be assembled in the removable holders 106. Holders 106 may include members 108 that position on top of the side wall 110. Holders 106 may rest on, or be suspended above, bottom wall 112 of the microwell plate 102. As shown, holder 106 includes eight (8) microwells 104, however, it should be understood that the number of microwells can be adjusted to the size of the microwell plate 102. For example, the number of the microwells 104 can be 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, etc. As shown in FIG. 1A, the microwell plate 102 does not include any titrant composition. FIG. IB depicts a schematic of the microwells having titrant composition 116 and 118 in the microwells. The microwells 104 can hold a volume of 20, 50, 300, 500 microliters, preferably 300 microliters or 400 microliters. The microwell plate 102, microwells 104, holders 106, can be made of any material having chemical resistance to acid. Non-limiting examples of materials include polymers, copolymers of polymers, polystyrene, polypropylene, cyclo-olefins and the like. The holders 106 may be polymeric or plastic tape with the microwells 104 embossed on the tape. Microwell plates are commercially available from Thermo Fisher Scientific (Waltham, MA, USA).
[0037] As shown in FIG. IB, the microwells 104 and 106 can be filled with two different pH sensitive dyes. For example, four microwells 104 can be filled with the an aqueous solution of a desired amount of the first pH sensitive dye and three microwells 106 can be filled with an aqueous solution of a desired amount the second pH sensitive dye. In a non- limiting example, the microwells 104 are each filled with 100 microliter (μί) of a 40
micromolar (μΜ) solution of bromocresol green and microwells 106 are each filled with a 100 μΐ^ of a 100 μΜ solution of neutral red. The solution in the eight well is used as a blank to allow any absorbance of the sample to be subtracted from the absorbance of the dyes. In other embodiments, the microwells 104 in each holder 106 can include the first pH sensitive dye, microwells 104' can include the second pH sensitive dye and microwell 104" is empty. In some instances, dyes and excipients can be lyophilized in the microwells 104, 104' in the microwell plate 102. Lyophilizing conditions include -60 degree Celsius at 100 mtorr. The microwells 104, 104', microwell holders 106, and/or the microwell plate can be sealed with a known sealing agent (for example, caps, plastic film or foil) to allow the microwell plate 102 or the microwell holders 106 to stored or transported. In some embodiments, the pH assay system includes a spectrophotometer that is capable of measuring the absorbance of the chosen colorimetric dye.
B. Materials
[0038] In some instances, the titrant composition can include a pH sensitive dye capable of having a colorimetric response in response to a change in pH of the solution, an acid compound, and an excipient compound.
1. pH sensitive dyes
[0039] The pH sensitive dyes can be any pH sensitive dye. The pH sensitive dyes can have a colorimetric response in a particular pH range. In some instances, the pH sensitive dyes can have an acid form that has a different absorbance value than an absorbance value of a base form of the pH sensitive dyes. Non-limiting examples of the first pH sensitive dye and the second pH sensitive dyes are bromocresol green and neutral red, respectively. Non- limiting examples of other pH sensitive dyes include gentian violet, malachite green, thymol blue, Methyl yellow, Methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein. pH sensitive dyes are commercially available from several sources. A non- limiting example of a commercial source is Sigma- Aldrich® (USA).
2. Excipients
[0040] In some instances, one or both of the titrant compositions can include an excipient. A non-limiting example of an excipient is poly glycol. In a preferred aspect, polyethylene glycol is used as an excipient. The polyethylene glycol can have an average
molecular weight from 3600 to 4400. Polyethylene glycol is commercially available from many sources. A non-limiting example of a commercially available polyethylene glycol is CARBOWAX™ manufactured by Dow Chemical Company (Midland, MI, USA). The excipient can aid in the lyophilization process. An amount of excipient added to a titrant composition ranges from 1 to 10 wt.%, from 1.5 to 5 wt.%, or from 2 to 3 wt.% of the titrant composition.
[0041] The composition can be a powder. The powder can be made by providing an aqueous solution of the titrant composition to one or more containers and subjecting at least one of the containers to lyophilizing conditions sufficient to remove the water from the aqueous solution to form the powder. In some instances, the one or more containers are microwells of a microwell plate. The powder can be packaged (for example, a bag, vial, or encapsulated). The powder can be sold separately from the kit. In some embodiments, other pH sensitive indicators can be used in place of or in combination with the first and second indicators. C. Method of Determining pH of a Solution
[0042] The pH assay system and kit described throughout the specification can be used to determine the pH of an analyte solution. The solution can be a sample from a water body such as a subsurface water well in a hydrocarbon formation, a lake, a river, a canal, a pool, a pond, or the like. Referring to FIG. 2, a schematic for determining pH is depicted. In system 200, the microwell plate 102 containing the lyophilized titrant compositions in microwells 104, 104 is obtained. A known amount of analyte solution 202 (for example 300 microliters) is added to the lyophilized titrant composition in the microwells 104, 104', using a delivery apparatus (for example, multichannel pipette) to produce microwell plate 204. After the solids in the plate have fully dissolved, the microwell plate 204 is placed in a detector 206 (for example, a plate reader) and the absorbances of each microwell at the wavelengths of the colorimetric dyes are measured. In embodiments when the colorimetric dyes are bromocresol green and neutral red, the absorbances at 450 nm and 520 nm are measured. At least one of the solutions in microwells 104, 104', has an absorbance value at the measured wavelengths that is not statistically differentiable from the absorbance values of a solution with a pH value below that at which the dye shows a colorimetric response. A controller system 208 coupled to the detector 206 includes a software algorithm that is used to determine whether to use the neutral red or bromocresol green data. The data collected at both wavelengths is collected
and plotted against the calibration curves. If the pH returned is within 3 to 6, then the bromocresol green data is used. If the pH returned is 6 to 8, the neutral red data is used. If the pH is less than 3 or greater than 8 then 'less than 3' or 'greater than 8' is returned, respectively on a device display of the controller 208. Using the method and pH kit of the present invention, the pH of the analyte taken from a single well can be determined using one sample rather than using multiple samples and many indicators and/or buffer systems. The controller system 208 can include components such as CPUs or applications with an associated machine readable medium or article which may store an instruction or a set of instructions that, if executed by a machine, may cause the machine to perform a method and/or operations in accordance with the methods of the present invention. The measured absorbance from microwell plate 204 can be stored in a computer system in the spectrophotometer and/or transmitted to another computer system. Either computer may be capable of processing the absorbance and displaying or printing an alkalinity value for a series of analytes. Such a machine may include, for example, any suitable processing platform, computing platform, computing device, processing device, computing system, processing system, computer, processor, or the like, and may be implemented using any suitable combination of hardware and/or software. The machine-readable medium or article may include, for example, any suitable type of memory unit, memory device, memory article, memory medium, storage device, storage article, storage medium and/or storage unit, for example, memory, removable or non-removable media, erasable or non-erasable media, writeable or re-writeable media, digital or analog media, hard disk, floppy disk, Compact Disk Read Only Memory (CD-ROM), Compact Disk Recordable (CD-R), Compact Disk Rewriteable (CD-RW), optical disk, magnetic media, magneto-optical media, removable memory cards or disks, various types of Digital Versatile Disk (DVD), a tape, a cassette, or the like. The instructions may include any suitable type of code, such as source code, compiled code, interpreted code, executable code, static code, dynamic code, and the like. The instructions may be implemented using any suitable high-level, low-level, object- oriented, visual, compiled and/or interpreted programming language, such as C, C++, Java, BASIC, Perl, Matlab, Pascal, Visual BASIC, assembly language, machine code, and so forth. The computer system may further include a display device such as monitor, an alphanumeric input device such as keyboard, and a directional input device such as mouse.
EXAMPLES
[0043] The present invention will be described in greater detail by way of specific examples. The following examples are offered for illustrative purposes only, and are not intended to limit the invention in any manner. Those of skill in the art will readily recognize a variety of noncritical parameters which can be changed or modified to yield essentially the same results.
Example 1
(pH Assay Kit)
[0044] To four microwells of an 8 microwell strip of a 96-microwell plate was added a sample (100 microliters) of bromocresol green (a 40 micromolar, (Sigma-Aldrich®, USA)). To three microwells of the 8 microwell strip, was added a sample of (100 microliters) of neutral red (100 micromolar). The resulting aqueous titrant solutions were lyophilized at - 45 °C for 22 hours followed by secondary drying at 20 °C for 9 hours. Several hundred 8 well strips were produced in the way described above. A portion of the strips were sealed using strip caps and heat sealed in bags. The remaining strips were used to calibrate the assay by adding water of a known pH and measuring the absorbance.
Example 2
(Calibration Curves)
[0045] Calibration curves were established for the two pH sensitive dyes by adding water of a known pH and measuring the absorbance. Calibration curves for the wells containing bromocresol green were developed using the absorbance at 450 nm (FIG. 3), and the curves for wells containing neutral red were developed using the absorbance at 520 nm (FIG 4).
Example 3
(Determination of pH of a Water Body) [0046] Seven analyte solutions (300 microliters each) were filtered through a 0.2 μιη syringe and then pipetted using a multichannel pipette into the 8 well strip containing freeze- dried titrant compositions of Example 1. The 8 microwell strip was placed in a detector and the absorbance of the wells containing bromocresol green were read at 450 nm and the absorbances of the wells containing neutral red were read at 520 nm. The detector was
connected to computer that includes an algorithm, which was used to determine whether to use the neutral red or bromocresol green data. The data collected at both wavelengths is collected and plotted against the calibration curves. If the pH returned is within 3 to 6, then the bromocresol green data was used. If the pH returned is 6 to 8, the neutral red data was used. Seven analyte solutions of the same material filtered and the pH measured using a conventional glass electrode method. Table 1 lists pH data of 14 oil and gas fluids determined using the assay of the present invention and a pH glass electrode. As shown in Table 1, the assay of the present invention provides similar results to that of the pH determined using a conventional glass electrode.
Table 1
Claims
1. A pH assay kit comprising:
(a) a microwell plate;
(b) a first lyophilized titrant composition comprising a first pH sensitive dye, wherein the first pH sensitive dye is sensitive to acidic solutions having a pH of 6 or less; and
(c) a second lyophilized titrant composition comprising a second pH sensitive dye, wherein the second pH sensitive dye is sensitive to solutions having a pH of 4 or more; wherein a plurality of the microwells contain the first lyophilized titrant composition comprising the first pH sensitive dye and a plurality of different microwells contain the second lyophilized titrant composition comprising the second pH sensitive dye such that when an analyte composition is added to the lyophilized titrant compositions in each well of the plurality of wells, a solution forms and the pH of the solution in is determined based on the response of the pH sensitive dyes to the solution.
2. The pH assay kit of claim 1, wherein the microwell plate comprises 6, 24, 96, 384, or 1536 wells.
3. The pH assay kit of claim 2, wherein the microwell plate comprises one or more removable strips, wherein each of the strips comprise 8 microwells, wherein 3 microwells of the 8 microwells contain the first lyophilized titrant composition comprising the first pH sensitive dye and 4 microwells contain the second lyophilized titrant composition comprising the second pH sensitive dye.
4. The pH assay kit of claim 3, wherein the microwell plate comprises 3 strips or 12 strips or more.
5. The pH assay kit of claim 4, wherein a portion of the strips are sealed to prevent the titrant composition from exiting the plurality of wells.
6. The pH assay kit of claim 5, wherein the plurality of wells is sealed with a cap, a plastic film or a foil.
7. The pH assay kit of claim 4, wherein at least one of the strips is used as to calibrate the assay.
8. The pH assay kit of claim 7, wherein the strip used as to calibrate the assay is not sealed.
9. The pH assay kit of claim 1, wherein each pH sensitive dye has an equivalence point, wherein the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye.
10. The pH assay kit of claim 1, wherein the absorbance of the first pH sensitive dye is 440 to 460 nm.
11. The pH assay kit of claim 1, wherein the absorbance of the second pH sensitive dye is 510 to 530 nm.
12. The pH assay kit of claim 1, wherein the first pH sensitive dye is bromocresol green.
13. The pH assay kit of claim 1, wherein the second pH sensitive dye is neutral red. gentian violet, malachite green, thymol blue, methyl yellow, methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein.
14. The pH assay kit of claim 14, wherein the second pH sensitive dye is neutral red.
15. The pH assay kit of claim 1, further comprising an excipient.
16. The pH assay kit of claim 15, wherein the excipient comprises one or more poly
glycol compounds, preferably polyethylene glycol.
17. The pH assay kit of claim 1, further comprising calibration solutions or calibration curves for each of the pH sensitive dyes.
18. A system for determining the pH of an analyte composition, the system comprising
(a) a pH assay kit as claimed in any one of claims 1 to 15.
(b) at least one detector configured to detect absorbances from the first and second pH sensitive dyes;
(c) at controller configured to monitor signals from the at least one detector, wherein the controller comprises an algorithm for: comparing the maximum
absorbance values of the first and second pH sensitive values with pH calibration curves for the first and second pH sensitive dyes to determine the pH of the solution; and
(d) a display unit configured to display the pH of the analyte composition.
19. The system of claim 18, wherein the detector is capable of measuring wavelengths between 400 and 700 nanometers.
20. The system of claim 18, wherein the detector is capable of simultaneously measuring two wavelengths between 400 and 700 nanometers.
21. A method of determining the pH of an analyte composition, the method comprising:
(a) obtaining any one of the pH assay kits of claim 1 or the system of claim 18;
(b) obtaining an analyte composition;
(c) adding substantially the same volume of the analyte composition to each of the plurality of wells of the microwell plate to form solutions from the analyte composition and the lyophilized titrate compositions in each of the plurality of wells, wherein the pH the solution in at least two of the microwells is different; and
(d) measuring the absorbance value for each solution in each of the plurality of wells at a first wavelength and a second wavelength and determining the pH of the analyte composition based on the measured absorbance values.
22. The method of claim 21, wherein each pH sensitive dye has an equivalence point, wherein the absorbance value of the equivalence point of the first pH sensitive dye is different from the absorbance value of the equivalence point of the second pH sensitive dye.
23. The method of claim 21, wherein the pH sensitive dye is bromocresol green.
24. The method of claim 21, wherein the second pH sensitive dye is neutral red, gentian violet, malachite green, thymol blue, methyl yellow, methyl orange, azolitmin, naphtholphthalein, phenolpthalein and thymolphthalein, or any combination thereof.
25. The method of claim 24, wherein the second pH sensitive dye is neutral red.
26. The method of claim 21, wherein determining the pH comprises:
(i) determining if the pH value for the samples is between 3 and 6, and comparing the value to a calibration curve of the first pH sensitive dye to quantify the pH;
(ii) determining if the pH value for the samples is between 6 and 8, and comparing the value to a calibration curve of the second pH sensitive dye to quantify the pH.
27. The method of claim 21, wherein the analyte in is an aqueous composition obtained from a subsurface well.
28. The method of claim 21, wherein the analyte solution comprises a plurality of
solutions having the same analyte, and each analyte solution is obtained from a different well of a plurality of subsurface wells.
29. The method of claim 21, wherein the well is a hydrocarbon well or a water well in a subsurface hydrocarbon formation.
30. The method of claim 21, wherein the analyte solution is obtained from a drilling or fracking process, water treatment, a chemical processing site, a swimming pool, a lake, a paper processing site, or any combination thereof.
31. A method of making any one of the pH assay kits of claim 1, the method comprising: (b) obtaining a micro well plate;
(c) obtaining a first lyophilized titrant composition comprising a first pH sensitive dye;
(d) obtaining a second lyophilized titrant composition comprising a second pH sensitive dye; and
(e) adding amounts of the lyophilized titrant composition to a plurality of wells of the microwell plate such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different.
32. The method of claim 31, wherein at least a majority of the plurality of wells are sealed to prevent the titrant composition from exiting the plurality of wells.
33. The method of claim 32, wherein the plurality of wells are sealed with a cap, a plastic film or a foil.
34. A method of making any one of the pH assay kits of claim 1, the method comprising:
(f) obtaining a microwell plate;
(g) obtaining a first titrant composition comprising a first pH sensitive dye;
(h) obtaining a second titrant composition comprising a second pH sensitive dye; and
(i) subjecting the microwell plate to lyophilizing conditions such that when an analyte composition is added to the lyophilized titrant composition in each well of the plurality of wells a solution forms and the pH of the solution in at least two wells is different.
35. The method of claim 34, wherein at least a majority of the plurality of wells are sealed to prevent the titrant composition from exiting the plurality of wells.
36. The method of claim 35, wherein the plurality of wells are sealed with a cap, a plastic film or a foil.
37. The method of claim 34, further comprising providing an excipient to at least one of the titrant compositions.
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US201462097922P | 2014-12-30 | 2014-12-30 | |
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CN112321485A (en) * | 2019-08-05 | 2021-02-05 | 尚科纺织企业工业及贸易公司 | PH-sensitive dye for textile material, preparation method and application thereof |
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US6387651B1 (en) * | 1995-04-12 | 2002-05-14 | Biolog, Inc. | Comparative phenotype analysis of two or more microorganisms using a plurality of substrates within a microwell device |
US7531319B2 (en) * | 2006-08-31 | 2009-05-12 | Kimberly-Clark Worldwide, Inc. | Array for rapid detection of a microorganism |
FI20096013A (en) * | 2009-10-02 | 2011-04-03 | Finnzymes Oy | Process for preparation of reaction mixture and related products |
US9228986B2 (en) * | 2010-08-03 | 2016-01-05 | General Electric Company | Simultaneous determination of multiple analytes in industrial water system |
CN105122047A (en) * | 2012-12-27 | 2015-12-02 | 赛诺瓦系统股份有限公司 | Ph meter |
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CN112321485A (en) * | 2019-08-05 | 2021-02-05 | 尚科纺织企业工业及贸易公司 | PH-sensitive dye for textile material, preparation method and application thereof |
CN112321485B (en) * | 2019-08-05 | 2024-03-08 | 尚科纺织企业工业及贸易公司 | PH-sensitive dye for textile material, preparation method and application thereof |
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