System for the Detection and Quantification of Ammonia and Ammonium in Fluids

Abstract

To provide systems and methods for the detection and quantification of "total ammonia" (the sum of ammonia (NH3) and ammonium (NH4+)).SOLUTION: A system with an analyzer device in fluid communication with a sample of a body fluid is configured to chemically or electrochemically convert at least a portion of ammonium (NH4+) contained in the body fluid into ammonia (NH3) and discharge the converted ammonia (NH3) into a gas sensing chamber. An ammonia (NH3) sensor located within the gas sensing chamber in conjunction with a processor can quantify the amount of ammonia (NH3) present in the gas sensing chamber in relation to the total ammonia of the body fluid.SELECTED DRAWING: None

Description

Technical Field

[0001] Cross - reference to Related Applications This application claims the benefit of U.S. Provisional Application No. 62 / 617,053, filed on Jan. 12, 2018. The disclosure of the prior application is considered a part of the disclosure of this application (and is incorporated herein by reference).

[0002] Field of the Invention The present invention relates to systems and methods for the detection and quantification of "total ammonia" (defined herein as the sum of ammonia (NH3) and ammonium (NH4 + +)).

Background Art

[0003] In - hospital acute kidney injury (AKI) is a significant medical problem that can affect up to 30% of children and 20% of adults in the hospital. AKI is associated with increased mortality and can lead to chronic kidney disease (CKD) that is strongly correlated with future hospitalizations, cardiovascular events, and shortened life expectancy.

[0004] In most cases, in-hospital AKI is very difficult to diagnose rapidly (i.e., within minutes or hours) because the symptoms and signs of AKI generally do not become apparent when AKI begins to develop. Therefore, medical care for AKI is delayed in its development because the recognition of AKI is delayed. Furthermore, the “diagnostic features” of AKI (i.e., an increase in serum creatinine concentration, a decrease in urine output rate) are not necessarily rapidly changing and may be sufficiently delayed compared to the first period (hours or days) of early AKI, and thus are actually very poor markers of early kidney tissue distress or early AKI. Also, changes in serum creatinine or urine volume are not specific to AKI. In fact, in many commonly encountered clinical scenarios where renal perfusion to the kidney tissue (“effective circulating blood volume”) is sufficiently decreased (e.g., decreased oral water intake, gastrointestinal water loss, transdermal water loss), serum creatinine values may increase and the rate of urine volume may decrease (without actual renal tissue damage occurring). Adaptive responses to decreased effective circulating blood volume include an increase in renal water reabsorption in the collecting ducts of the kidney as a response to a decrease in glomerular filtration rate (resulting in a decrease in creatinine clearance and an increase in serum creatinine) and an increase in the circulating levels of antidiuretic hormone (resulting in the production of more concentrated urine and a decrease in urine volume). Serum creatinine values can also be significantly affected by the administration of certain drugs without actual renal tissue damage. These drugs include cimetidine, trimethoprim, pyrimethamine, salicylates, furosemide, corticosteroids, and some vitamin D derivatives. The use of angiotensin-converting enzyme inhibitors, angiotensin II receptor blockers, and diuretics also affects creatinine clearance (and thus serum creatinine values) without causing AKI. Therefore, improving the diagnostic process for recognizing early AKI in hospitalized patients is an unmet clinical need.

[0005] There is a strong interest in the discovery and validation of novel methods for detecting AKI. Conventional methods for detecting AKI may include serial testing of lagging blood markers (i.e., creatinine) and monitoring of urine flow rate (i.e., urine output per hour). Subjecting patients to serial blood draws to monitor changes in serum creatinine is inherently problematic because each sample taken necessarily involves the removal of tissue from the body. In addition to the latency and lack of specificity for AKI, serum creatinine values that change significantly as a result of AKI indicate tissue damage and overall organ dysfunction rather than early renal tissue cellular insufficiency. Novel urinary biomarkers for AKI, including neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), insulin-like growth factor binding protein 7 (IGFBP7), and tissue metalloprotease inhibitor 2 (TIMP2), correlate with AKI (as defined by conventional characteristic markers) and can change more rapidly than serum creatinine, but share the same limitation as creatinine in that they change relatively late (i.e., hours rather than minutes after kidney injury occurs). Furthermore, novel urinary AKI biomarkers perform more poorly in predicting and / or diagnosing AKI in patients with certain health conditions, including chronic kidney disease (CKD) and sepsis. Considering that it is often clinically important to determine whether a particular episode of AKI can be optimally treated by the rapid application of a treatment to increase the effective circulating blood volume, the inherent inability of each and all standard and novel AKI laboratory tests to assist in that determination also limits their overall usefulness in the medical setting. Accordingly, there is a need for AKI detection systems and methods that can rapidly detect at-risk renal tissue, renal tissue insufficiency, and / or early AKI before the onset of significant renal tissue damage (AKI).

[0006] For that purpose, urinary "total ammonia" (ammonia (NH3) and ammonium (NH4 +)) can be used as a novel urinary biomarker for the detection of at-risk renal tissue, renal tissue insufficiency, early AKI, and / or rapid detection of AKI. In a fluid, ammonia (NH3) and ammonium (NH4 + ) exist in equilibrium, and the amount of each species depends on the ambient environmental conditions (e.g., pH, temperature, pressure). The production of total ammonia in the kidney (renal ammonia production) depends mainly on the metabolism of glutamine within renal proximal tubular cells in addition to other intrarenal states. Renal ammonia production normally varies or adapts in response to various systemic states, such as the systemic acid / base status, potassium status, and fluctuations in dietary protein intake. Changes in the total ammonia level in the body due to changes in liver function, etc., also lead to changes in the total ammonia level in urine. A decrease in effective circulating blood volume and / or AKI rapidly affects renal ammonia production, reducing the total amount of ammonia excreted in urine and the total ammonia excreted from renal tissue. Therefore, by using continuous, automatic, predictive monitoring of the dynamic changes in total ammonia concentration and / or content in urine, potential changes in renal tissue, renal cellular insufficiency, early AKI, and / or renal blood flow (effective circulating blood volume) at risk of AKI can be detected rapidly. Furthermore, continuous automatic monitoring of the dynamic changes (increase or decrease in level) of total ammonia in urine can be used to detect potential changes in any systemic state that affects renal ammonia production and / or total ammonia in urine, including the systemic acid / base status, potassium status, dietary protein intake, and liver function status.

[0007] Therefore, there is a need for a system and method that enables continuous, automatic monitoring of changes in the amount of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in a fluid, including that excreted from the body (i.e., urine).

[0008] Ammonia (NH3) and ammonium (NH4 +Conventional systems and methods for the measurement of ) are often inaccurate, difficult to use, cumbersome, and not suitable for bedside urine testing or continuous monitoring. Conventional systems for ammonia or ammonium measurement have (1) colorimetric sensors, (2) spectroscopy-based sensors, (3) nanomaterial-based sensors, (4) non-contact conductive sensors, and (5) reagent sticks.

[0009] Colorimetric sensors are used for the measurement of ammonia (NH3) and / or ammonium (NH4 + ) in wastewater and blood. This type of sensor consists of a thin film embedded with an ammonia (NH3)-sensitive pH dye attached to the end of a detection unit (e.g., an optical fiber). These devices have high sensitivity, but further instrument improvement is required for use with biological samples.

[0010] For example, the blood analysis technology in ROCHE COBAS INTEGRA® is based on an enzymatic method that requires the use of two reagents and analysis within 30 minutes of blood sample collection. This method used for blood analysis cannot be directly applied to urine measurement. When used for urine, urine (total urinary ammonia concentration in millimoles) has to be diluted at least 10 - 1000 times to achieve a concentration (total blood ammonia concentration in micromoles) within the limits of the blood analysis technology. Therefore, blood analysis technology is not suitable for bedside urine testing.

[0011] In another example, a handheld portable device can detect the level of ammonium (NH4 + ) in blood. The handheld portable device can use a color-based sensor and a partition membrane that converts ammonium (NH4 + ) to ammonia (NH3). However, the handheld device may be disposable and ammonium (NH4 +It may only provide a single-point measurement value at a given time and is not suitable for continuous, automated, or repeated measurements. Additionally, the described handheld portable device may be used with blood, which requires invasive blood sampling and makes evaluation more difficult and cumbersome when continuous patient monitoring is desired. Further, the described handheld portable device is not adaptable for use outside of a medical environment (i.e., within a patient's home) since it is used with blood. Moreover, this device does not provide the at least 24-hour continuous quantification required to monitor clinically important conditions. Further, conventional handheld portable devices have been demonstrated to have insufficient accuracy and tend to produce both false positive and false negative test results.

[0012] Ammonium (NH4 + ) or ammonia (NH3) in a sample can be detected by conventional methods that also involve laboratory-based methods with manual or pump-based handling of liquids and samples, and may include external detection instruments such as scanners or photomultiplier tubes and optical fibers. These conventional systems can use paper-based extraction membranes or solution mixtures for the analysis of ammonium (NH4 + ) or ammonia (NH3) in water or pure samples. Laboratory-based methods cannot accurately detect ammonium (NH4 + ) levels or ammonia (NH3) levels from complex body fluid samples where the amount of other dissolved components (i.e., urine) varies widely.

[0013] Furthermore, semi-continuous measurements of ammonium (NH4 + ) / ammonia (NH3) performed in water quality monitoring applications using commercial technologies based on amperometric sensors or colorimetric sensors are not adaptable to a medical environment. For example, the ANALYTICAL TECHNOLOGY Q45N device weighs 15 pounds and measures ammonium (NH4 +) / Ammonia (NH3) is converted to stable monochloramine measured by a current measurement sensor. A minimum flow rate of 200 mL / min (Note: The minimum or absolute urine volume in humans is 0.5 mL / Kg / hr. For adults, typical urine volume is 800 - 2000 mL / day (0.6 - 1.4 mL / min).) is required and it is reliable in the range of 0 - 5 ppm NH3 (0 - 270 micromolar concentration). The AZTEC 600 color analyzer (ABB) is designed to be used semi - continuously in wastewater. The AZTEC 600 color analyzer can only measure 4 samples per hour using indophenol blue chemistry and requires a continuous flow rate of 200 - 500 mL / min. The AZTEC 600 color analyzer measures ammonia (NH3) up to 3 ppm. AWA INSTRUMENTS CX4000 also operates based on the colorimetric principle. These large - scale commercially available semi - continuous measurement devices from the water treatment industry cannot be easily adapted for use in a medical environment, and the sensors utilized by these devices must be calibrated regularly for different concentration ranges. Ammonium (NH4 + ) / Batch measurements of ammonia (NH3) are generally performed in water treatment applications via either an ammonium ion probe or a colorimeter coupled with a spectrophotometer.

[0014] Ammonium (NH4 + ) ion - selective electrodes (either solid - state or membrane - based) operate based on the principle of a membrane with an ammonium (NH4 + ) - selective ion exchanger, which creates different potentials across the membrane by comparing an unknown solution to a reference solution. Most ammonium (NH4 + ) colorimetric methods involve adding a reagent to a water sample and evaluating the color of the liquid solution with a special device. A few ammonium (NH4 +)The colorimetric test involves a strip (similar to a pH measurement strip) where a reagent is added to a solution, the strip is immersed in the solution, and a color change is visually observed. Commercially available batch measurement products require the use of reagents, enzymes, and / or large analytical devices. The Roche enzymatic method requires a minimum sample volume of 20 μL and is not designed for urine. For aqueous solutions of ammonium (NH4 + )Selective electrodes require a minimum sample volume of several milliliters, and the electrodes must be calibrated every 1 - 2 hours (in continuous measurements). For accuracy, the electrodes must also be calibrated with various solutions depending on the expected concentration range of ammonium (NH4 + ) in the solution being measured.

[0015] Most measurement techniques for the detection of ammonium (NH4 + ) / ammonia (NH3) in human samples rely on exhaled breath or blood (plasma). However, conventional techniques are not suitable for continuous monitoring because the stability of the sensing membrane is limited and there is drift in the measurement output, so ion - selective electrodes require time - consuming calibration between the analysis of each sample. For example, the ORION (trademark) high - performance ammonium (NH4 + ) electrode needs to be calibrated before each new measurement to minimize measurement drift due to the limited stability of the sensing membrane.

[0016] Furthermore, considering the complexity of biological samples, there are few methodologies approved by the US Food and Drug Administration (FDA) for clinical research. For the detection of ammonium (NH4 + ) in blood and urine, the enzymatic assay is the only technique that meets the FDA standards. Generally, enzymatic assays have a limited shelf life, involve multiple incubation steps, require a processing time of more than one hour, and involve significant operator effort.

[0017] Spectroscopic methods for ammonia gas (NH3) measurement include pulsed quantum cascade laser spectroscopy and optical micro-ring resonators. There are large (tabletop) measurement systems, but these lack the small size, light weight, and low cost desirable for portable individual monitoring (such as at the hospital bedside). For example, ammonia (NH3) can be detected via absorption spectroscopy by devices such as NEPHROLUX (trademark) that use tunable lasers and acoustic detectors to perform sub-ppb zero-background measurements of ammonia (NH3) in the presence of interfering substances such as carbon dioxide and water vapor (such as in exhaled breath). Spectroscopic techniques are very sensitive, but they usually have bulky components and are inconvenient for personal use. Furthermore, the optical components in absorption spectroscopy are prone to misalignment and are unsuitable for personal use.

[0018] Gas chromatography-mass spectrometry (GC-MS) and selected ion flow tube mass spectrometry (SIFT-MS) may be accurate for ammonia (NH3) measurement, but they are expensive and difficult to maintain. GC-MS separates and identifies both ammonia (NH3) and ammonium (NH4 + ) from complex mixtures, but requires expensive instrumentation (~$300,000) and a pre-concentration step that makes high reproducibility and real-time implementation impossible. SIFT-MS was developed to detect low molecular weight volatile substances, including ammonia (NH3), in various biological samples (such as headspace of skin and urine, exhaled breath) in real time, but it is expensive (~$200,000) and quite difficult to maintain.

[0019] Nanomaterial-based chemiresistors and electrochemical sensors show detection limits that match clinically relevant ammonia (NH3) levels (exhaled ammonia (NH3) (ppb)) under well-defined near-ideal laboratory conditions. However, further improvement is needed to obtain the selectivity and lifetime required for continuous monitoring to detect ammonia (NH3) in complex samples using these sensors.

[0020] Furthermore, conventional ammonia (NH3) detectors may have a non-contact conductive sensor. However, the acidic liquids used in conventional non-contact conductive ammonia (NH3) sensors need to be replaced after each measurement, and continuous measurement is not practical.

[0021] Finally, urine reagent strips are widely used to determine ten different urine parameters (including pH, specific gravity, leukocyte esterase, nitrite, urobilinogen, protein, hemoglobin, glucose, ketone, bilirubin), but commercially available electronic readers for these urine dipsticks do not include the measurement of ammonia (NH3) or ammonium (NH4 + ). However, ammonium (NH4 + ) detection reagent strips are commercially available for use in water samples. The operation of these ammonium (NH4 + ) detection reagent strips is based on irreversible chemical reactions, and therefore they are disposable devices. These reagent strips are rapid and easy to use, but only provide a semi-quantitative assessment of the parameters and do not have the accuracy and continuous real-time monitoring capabilities desired in important applications.

[0022] Furthermore, ammonia (NH3) and ammonium (NH4 + ) in biological solutions are in equilibrium with each other and spontaneously convert to one another depending on changes in the local conditions (i.e., pH, temperature, pressure) within the biological sample being tested. Therefore, there is a need for systems and methods for detecting and quantifying total ammonia (ammonia (NH3) and ammonium (NH4 + )) in biological samples (e.g., body fluids, urine) in the medical field to account for the presence of both ammonia (NH3) and ammonium (NH4 + ) in the sample being tested.

[0023] Therefore, ammonium (NH4 +There is a need for a system and method that enables continuous and automatic monitoring of changes in the concentration and / or amount of ammonia (NH3) in body fluids, taking into account the presence of SUMMARY OF THE INVENTION

[0024] The present disclosure describes a system and method for detecting total ammonia (ammonia (NH3) and ammonium (NH4 + )) in a fluid.

[0025] In some embodiments, the systems and methods described herein can convert ammonium (NH4 + ) to ammonia (NH3). Body samples such as urine can contain varying amounts of ammonia (NH3) and ammonium (NH4 + ) depending on the pH. The systems and methods described herein can extract ammonia (NH3) from body fluids (e.g., urine, sweat, blood, etc.) through an extraction membrane, such that the total ammonia (NH3) and ammonium (NH4 + ) contained in the biological sample is measured substantially as ammonia (NH3). In some embodiments, this enables continuous samples of the fluid (e.g., when urine is produced) to be measured continuously and almost continuously for total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration. The extraction membrane can convert substantially all of the ammonium (NH4 + ) of the fluid sample to ammonia (NH3) chemically or electrochemically.

[0026] In some embodiments, the system has an analyzer. The analyzer may be in fluid communication with a sample of body fluid. The analyzer has an intelligent controlled sample conditioning and transport system, an extraction membrane (ammonium (NH4 +)substantially all of which is converted to ammonia (NH3), a sensing chamber, and an ammonia (NH3) sensor. An intelligently controlled sample conditioning and transport system can control the amount of body fluid contacting the extraction membrane to prevent sensor performance from varying over multiple consecutive uses and long periods of time. The sample conditioning and transport system can be operated by an intelligently programmable valve system based on an intelligent algorithm that includes sample volume, time, and sensor signal change information.

[0027] In some embodiments, the sample conditioning and transport system may have a signal saturation and drift avoidance mechanism having a microcontrolled actuated valve system. The microcontrolled actuated valve system may control the volume of body fluid in contact with the analyzer. The microcontrolled actuated valve system may have valves configured to control the transport of body fluid, headspace gas, and gas from a zero adjustment channel. The sample conditioning and transport system may be formed by at least two inlets, namely, a sampling channel in contact with the body fluid and a purge channel in contact with a zeroing material that allows the system to record a baseline. The baseline may be essential for correcting sensor signal drift. The extraction membrane can be disposed between the region in fluid communication with the body fluid and the sensing chamber, and 1) ammonium (NH4 +) at least partially convert it to ammonia (NH3), and 2) be configured to discharge the converted ammonia (NH3) into the sensing chamber. The ammonia (NH3) sensor disposed in the sensing chamber may be pre-treated by heat under specific conditions and may have a pre-calibration algorithm to ensure the performance of the sensor under a wide range of temperature, relative humidity, and pressure conditions. The ammonia (NH3) sensor processor, when executed, may have a non-transitory storage device that stores instructions for causing the processor to quantify the amount of ammonia (NH3) present in the sensing chamber. The analyzer may, based on the quantified amount of ammonia (NH3) present in the sensing chamber and how that amount may change over time, detect changes in organ or tissue function, the occurrence of organ or tissue damage, changes in the total body ammonia (ammonia (NH3) and ammonium (NH4 + )) physiological function, or other in-body processes in which the level of total ammonia (ammonia (NH3) and ammonium (NH4 + )) changes. Optionally, the system may also have a user interface device. In some embodiments, the ammonia (NH3) sensor may be further configured to transmit the quantified amount of ammonia (NH3) present in the sensing chamber to the user interface device. In some embodiments, the user interface device may be configured to receive at least the transmission from the analyzer and may have a display with a graphical user interface configured to display the transmission received from the analyzer.

[0028] In some embodiments, the method includes, in the analyzer, receiving a sample of body fluid and, via an extraction membrane located between a region in fluid communication with the sample of body fluid and the sensing chamber of the analyzer, ammonium (NH4 +) is at least partially converted to ammonia (NH3), and the converted ammonia (NH3) is discharged into the sensing chamber through an extraction membrane. The amount of ammonia (NH3) present in the sensing chamber is determined via an ammonia (NH3) sensor located within the sensing chamber. Changes in organ or tissue function, organ or tissue damage, changes in physiological functions that affect the total ammonia concentration in body fluids, or other body fluid processes in which the total ammonia level in body fluids changes can be detected if the determined amount of ammonia (NH3) present in the sensing chamber changes or is interpreted as being outside the normal or expected concentration or range for an individual at the time of measurement. Optionally, the method can further include the step of transmitting the amount of ammonia (NH3) present in the sensing chamber to a user interface device, and the user interface device further includes a display having a graphical user interface. Optionally, the method can also include, at the user interface device, receiving the amount of ammonia (NH3) present in the sensing chamber and displaying the amount of ammonia (NH3) present in the sensing chamber via the graphical user interface.

[0029] In some embodiments, a method for determining kidney function includes detecting, in a first analyzer, the concentration of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in a subject sample. Thereafter, the analyzer can transmit the detected total ammonia (ammonia (NH3) and ammonium (NH4 + )) level to a second user interface device. The second user interface device can then associate the detected level of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in the subject sample with a diagnosis of altered kidney function. This association is based on the total ammonia (ammonia (NH3) and ammonium (NH4 +The detection level of ((, or the total ammonia (ammonia (NH3) and ammonium (NH4 + )) in the subject sample can be considered in comparison with the detection level of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in the previous sample of the subject.

[0030] In some embodiments, the non-invasive device can semi-continuously detect the concentration of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in the fluid. The non-invasive device can be miniaturized, conveniently placed, and can automatically transmit data to provide near real-time and / or semi-continuous analysis. In some embodiments, the device may be used for the automatic monitoring and rapid detection of acute kidney injury (AKI) in inpatients with indwelling urinary catheters. Alternatively, the device can be used to detect changes in renal ammonia production and / or total urinary ammonia (ammonia (NH3) and ammonium (NH4 + )) due to the following: 1) changes in renal function, 2) acute kidney injury or failure, 3) chronic kidney disease, 4) changes in liver function, 5) acute liver injury or failure, 6) chronic liver disease (e.g., cirrhosis), 7) acute gastrointestinal bleeding, 8) chronic gastrointestinal bleeding, 9) genetic or inborn metabolic diseases (e.g., urea cycle disorders, organic acidurias, carnitine deficiency due to defects in fatty acid oxidation, dibasic aminoaciduria, and defects in pyruvate metabolism) involved in or affecting the generation, handling, and / or excretion of ammonia physiology, 10) changes in normal metabolic processes (e.g., increased ammonia production and excretion after a protein meal), 11) acute or chronic systemic acid / base changes or imbalances due to metabolic processes or disease states, 12) acute or chronic systemic acid / base changes or imbalances due to respiratory processes or disease states, 13) changes in effective circulating blood volume, 14) changes in renal blood flow, or 15) renal plasma flow. BRIEF DESCRIPTION OF THE DRAWINGS

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Mode for Carrying Out the Invention

[0056] Some embodiments of the systems and methods described herein are wireless, solid state, and portable, and have a continuous total ammonia (ammonia (NH3) and ammonium (NH4 + )) sensing and quantification device. In addition to other potential uses, medical providers can use the systems, methods, and devices described herein to more quickly and accurately measure total ammonia (ammonia (NH3) and ammonium (NH4 + )) in a biological sample than was possible with conventional systems. In some embodiments, the systems, methods, and devices described herein can reliably determine the exact concentration of total ammonia (ammonia (NH3) and ammonium (NH4 + )) contained in a biological sample within 5 seconds and wirelessly transmit the data to other devices. In some embodiments, the wireless transmission may be performed using Bluetooth®. In some embodiments, the systems, methods, and devices described herein include an extraction membrane, an ammonia (NH3) sensor composed of a hydrophobic material such as a polytetrafluoroethylene (PTFE) substrate impregnated with a pH indicator such as bromophenol blue, a light emitting diode (LED) at the maximum absorption wavelength of the indicator, and a photodiode configured to measure the change in absorbance after ammonia (NH3) exposure. Further, LEDs of different wavelengths at which the indicator does not absorb light may be composed of corresponding photodiodes to generate a second reading that allows for further correction of sensor signal drift. The photodiode can convert the color change of the sensor into an electronic signal and transmit this (either wired or wirelessly) to a smart device for reading out. The described systems, methods, and devices can exhibit high sensitivity, high specificity, high-speed reversibility, and fast response times compared to conventional systems.

[0057] As described above, total ammonia in urine (ammonia (NH3) and ammonium (NH4 +)) can be used as a biomarker for the early detection of acute kidney injury (AKI) and other physiological states and diseases. The systems and methods described herein are for total ammonia (ammonia (NH3) and ammonium (NH4 + )) in urine or other body fluids and / or for the detection of ammonia (NH3) gas in the headspace of urine or the headspace of other body fluids. The body fluid can include one or more of whole blood, plasma, serum, intracellular fluid, interstitial fluid, lymph (lymph), sweat, urine, pleural fluid, pericardial fluid, peritoneal fluid, bile (bile), feces, cerebrospinal fluid, synovial fluid, saliva, sputum, nasal fluid, or ocular fluid.

[0058] As further discussed below, the systems and methods described herein can have an analyzer. The analyzer is optionally referred to herein as a colorimetric optoelectronic mechanical analyzer (or simply "CODA") and uses very small amounts of urine or body fluid to provide real-time and continuous detection and quantification of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in urine. The analyzer can use a sensor with a pH dye-based ammonia (NH3)-sensitive sensing probe. Different from conventional detection methods for human body fluids that directly measure dissolved ammonium (NH4 + ) in blood or urine, the sensing chamber of the analyzer can detect and measure ammonia (NH3) gas in the urine headspace by converting fluid ammonium (NH4 + ) to gaseous ammonia (NH3) by exposing the body fluid (or body fluid sample) to an alkali before measurement.

[0059] Referring now to FIG. 1, total ammonia (ammonia (NH3) and ammonium (NH4 +A schematic diagram of a system for detecting )) is shown. As shown in FIG. 1, the wearable analyzer can have an analyzer device 100 that communicates wirelessly with a user interface device 102. In some embodiments, the analyzer device 100 can include an extraction membrane 104, an ammonia (NH3) sensor 106, a photodiode 108, a light emitting diode 110, a microcontroller 112, a Bluetooth (registered trademark) transmitter / receiver 114, a flexible printed circuit board 116, and / or a flexible battery 118. The extraction membrane 104 may be configured to be in fluid communication with a body fluid sample such as urine or the headspace of the body fluid sample 120.

[0060] In some embodiments, the extraction membrane 104 may be disposed between a region in fluid communication with the body fluid and a sensing chamber (including the ammonia (NH3) sensor 106). The extraction membrane 104 is configured to convert at least a portion of the ammonium (NH4 + ) in the body fluid to ammonia (NH3) and discharge the converted ammonia (NH3) into the sensing chamber. As will be further described below, in some embodiments, the extraction membrane can have a distribution layer, an alkali layer, a hydrophobic layer, and an indicator layer. The distribution layer can be configured to distribute the body fluid sample along the extraction membrane. The alkali layer can be configured to convert at least a portion of the ammonium (NH4 + ) in the body fluid sample to ammonia (NH3). In some embodiments, the alkali layer can have an organic hydroxide and / or sodium hydroxide. The hydrophobic layer can be configured to filter the ammonia (NH3) converted from the body fluid sample and discharge the converted ammonia (NH3) into the sensing chamber. In some embodiments, the hydrophobic layer can include polytetrafluoroethylene or the like. The indicator layer can include bromophenol blue, a plant-derived pH indicator (e.g., anthocyanin), or any other suitable material. The indicator layer is exposed to the ammonia (NH3) gas and / or the alkali layer of the body fluid and interacts with the alkali layer to form fluid ammonium (NH4 +) It may be configured to change color in response to the amount and / or concentration of ammonia (NH3) gas extracted from

[0061] In some embodiments, the ammonia (NH3) sensor 106 may have a colorimetric nanocomposite sensor that uses a composite sensing nanomaterial for the detection of ammonia (NH3) on the sensing region 106A and the reference region 106B (without a sensing probe) to evaluate the absorption color change. In some embodiments, the absorbance is calculated as the negative logarithm of the signal from the sensing region divided by the signal from the reference region. The light-emitting diode 110 and the photodiode can be combined to form a detection unit (or a hybrid sensor) as further described below. The ammonia (NH3) sensor 106 may also have a processor that, when executed, stores instructions for the processor to quantify the amount of ammonia (NH3) present in the sensing chamber in a non-transitory storage device.

[0062] As further described in detail below, the ammonia (NH3) sensor can have four photodiodes, namely, two sensing photodiodes arranged in the sensing region 106A and two reference photodiodes arranged in the reference region 106B. The two light-emitting diodes may be configured to illuminate the indicator layer. In some embodiments, the light-emitting diode may emit red light. In some embodiments, the light source and the light detector may be configured to use a CMOS chip (camera).

[0063] The ammonia (NH3) sensor 106 calculates an absorbance metric of the indicator layer based on the signal from the first photodiode and the signal from the second photodiode, and converts the calculated absorbance metric into an amount of ammonia (NH3) that can be quantified by comparing the absorbance metric with one or more reference values indicating the relationship between absorbance and ammonia (NH3) concentration, thereby quantifying the amount of ammonia (NH3) present in the sensing chamber. Further, the absorbance signal can be further corrected from an LED and a corresponding photodiode designed to record the sensor signal at a wavelength at which the indicator has no light absorption (minimum absorption wavelength), for example, a wavelength higher than 675 nm.

[0064] In some embodiments, the user interface 102 is presented on a computing device. The computing device may be built into the detection system or may be built into an external device. The built-in computing device can be associated with a display. In the case of an external device, the user interface 102 can include one or more software applications that can obtain data from the analyzer 100 and generate one or more reports for display on the graphical user interface of the user interface 102. The generated reports may require the execution of one or more analytical calculations on the data obtained from the analyzer 100. The computing device can be a mobile device such as a tablet computer (e.g., Apple iPad®, Samsung Galaxy Tab, etc.), a smartphone (e.g., Apple iPhone®, Blackberry Phone, Android Phone, etc.), a smartwatch (e.g., Apple Watch, etc.), a personal digital assistant (PDA), a personal computer device (PC; via a web browser and installable software), and / or other similar devices. The computing device may be wired or communicably connected to the analyzer 100 via a network such as a local area network (LAN), a wide area network (WAN), a digital subscriber line (DSL), a wireless network (e.g., 3G or 4G network), or other equivalent connection means. A Bluetooth® communication configuration is shown in FIG. 1.

[0065] A computing device can have a processing device, a memory, a data storage device, and a communication interface. The components can communicate with each other via data and control buses. The processing device can have, but is not limited to, a microprocessor, a central processing unit, an application specific integrated circuit (ASIC), a field programmable gate array (FPGA), a digital signal processor (DSP), and / or a network processor. The processing device may be configured to execute processing logic for performing the operations described herein. Generally, the processing device may have any suitable dedicated processing device that is specially programmed with processing logic to perform the operations described herein.

[0066] The memory stores computer-readable instructions executable by the processing device and can have, but is not limited to, at least one of, for example, read-only memory, random access memory, flash memory, dynamic RAM, and static RAM. Generally, the memory may have any suitable non-transitory computer-readable storage medium that stores computer-readable instructions executable by the processing device to perform the operations described herein. In some examples, the computing device may have two or more memory devices (e.g., dynamic memory and static memory).

[0067] The computing device can have a communication interface device for direct communication with other computers (including wired and / or wireless communication) and / or communication with a network. In some examples, the computing device may have a display device (e.g., a liquid crystal display, a touch-sensitive display, etc.). In some examples, the computing device may have a user interface (e.g., an alphanumeric input device, a cursor control device, etc.).

[0068] In some examples, a computer device can have a data storage device that stores instructions (e.g., software) for performing any one or more of the functions described herein. The data storage device can have any suitable non-transitory computer-readable storage medium including, but not limited to, solid-state memory, optical media, and magnetic media.

[0069] As shown in FIG. 1, the analyzer 100 can have various configurations. For example, in panel C of FIG. 1, a tube version of the analyzer 100 is shown. In the tube version of the analyzer 100, a wireless flexible printed circuit board 116 is configured to have a flexible battery 118 and a flexible display located under the wireless flexible printed circuit board 116. The tube version of the analyzer 100 is configured such that the extraction membrane is in fluid contact with urine within the tube. In panel C of FIG. 1, the analyzer 100 is arranged in series with the urine of a patient within a catheter tube or collection bag.

[0070] In some embodiments, to reduce fouling of the analyzer, the sensor surface can be arranged in a direction parallel to the urine flow to avoid deposition of urinary solids (see panel C of FIG. 1).

[0071] In some embodiments, hydrophilic modifications to the connector walls are included to enhance easy wetting of the connector and the membrane and reduce clogging of the sample into the membrane by inserting the analyzer into the catheter. Further, one embodiment of the system described herein can use a leak-free standard tube fitting of Jaco™ to prevent urine leakage.

[0072] Alternatively, as shown in panel D of FIG. 1, using an adhesive version of the analyzer 100, the device 100 can be adhered to a diaper or skin such that the extraction membrane 104 is in contact with urine or sweat, respectively.

[0073] In some embodiments, the analyzer 100 can provide specific, rapid response and accurate measurement for ammonia (NH3) gas concentrations in the range of 2 ppm to 1000 ppm (corresponding to 0.1 mmol / L to 50 mmol / L of ammonium (NH4 + ) in the liquid fluid). The ammonia (NH3) sensor 106 can be highly selective for ammonia (NH3), especially considering the large number of interfering substances in the urine headspace. As described below, the sensor 106 constructed according to the methods and systems described herein can exhibit good reusability over long sampling periods, enabling routine use for medical applications. Thus, the analyzer 100 can accurately monitor the total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration in urine and / or ammonia (NH3) gas extracted from urine, as demonstrated by comparison with measurements from commercially available reference methods (ISE electrodes) further described in the experimental section below. In some embodiments, the ammonia (NH3) sensor 106 is durable and can last for at least 10 weeks. As described below, the synthesis process of the sensor 106 is simple and easily reproducible. Further, the analyzer 100 can be wirelessly connected to a smart device, thereby providing measurement flexibility for inpatients, outpatients, or personal health monitoring.

[0074] In some embodiments, the analyzer 100 may be particularly well - suited for hospital or outpatient settings. As described above in connection with FIG. 1, the analyzer 100 can have a replaceable cartridge that includes a combination of an extraction membrane / sensor and optoelectronic components for the detection of ammonia (NH3), signal conditioning, and wireless communication with a user interface device. Software for data acquisition, signal processing algorithms, display, transmission, and the user interface may be included in the analyzer 100 and / or the user interface 102.

[0075] In some embodiments, a sample of body fluid (such as urine or sweat) may be routed onto an extraction membrane / sensor cartridge (replaceable cartridge) from which ammonia (NH3) is extracted. The extracted ammonia (NH3) can then interact with a colorimetric sensor in the cartridge, thereby changing its color according to the ammonia (NH3) concentration. Software including one or more signal processing algorithms can then determine the ammonia (NH3) concentration. In some embodiments, the total ammonia in urine or sweat (ammonia (NH3) and ammonium (NH4 + )) excretion rate can be determined by knowing the concentration of the extracted ammonia (NH3), the pH of the fluid, and / or the flow rate of the fluid. In some embodiments, by knowing the concentration of the extracted ammonia (NH3), and the density, specific gravity, osmolality, or osmolarity of the fluid, the total ammonia in urine, the headspace of the skin, or sweat (ammonia (NH3) and ammonium (NH4 + )) excretion rate can also be estimated. In some embodiments, the test can be performed in an automated, continuous manner, and the test is performed every few minutes. Next, the data may be automatically transmitted from the analyzer 100 to the user interface 102, where the data is processed and graphically displayed.

[0076] For example, the user interface 102 can show the change in total ammonia (ammonia (NH3) and ammonium (NH4 + )) or ammonia (NH3) concentration, or total ammonia (ammonia (NH3) and ammonium (NH4 + )) or ammonia (NH3) excretion rate over a long period of time. The user interface 102 may be configured to be regularly reviewed by a healthcare provider, a patient, etc. In some embodiments, the analyzer 100 or the user interface 102 is the total ammonia in urine (ammonia (NH3) and ammonium (NH4 +))Identify sudden or unexpected changes in concentration and / or excretion rate and trigger an automated warning, thereby alerting healthcare providers to the total ammonia in urine (ammonia (NH3) and ammonium (NH4 + )) changes in related and associated health or metabolic states that have led to changes in parameters (e.g., either a possible acute kidney injury or a possible acute kidney impairment (AKI) event). The total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration and / or past measured excretion rates can be stored in a database and compared.

[0077] As described above, the composite extraction membrane / sensor is based on a colorimetric sensor and an extraction membrane that measures ammonia (NH3), both of which are assembled on the same substrate / unit, whereby the detection principle is scalable and miniaturized (as shown in FIGS. 1 and 5). The extraction membrane with alkaline buffer extraction ability can reduce its dimensions from centimeters to millimeters. In some embodiments, the extraction membrane may be configured to extract ammonia (NH3) from ammonium (NH4 + ) into the gas phase at a pH of 10 or higher according to Henry's law partitioning behavior. The colorimetric component may be configured to detect light absorption per unit area, whereby the sensitivity is determined by the binding sites per unit area, independent of the total sensing area. In other words, the systems and methods described herein can scale the combination of extraction membrane / sensor size without sacrificing extraction quality, sensitivity, and detection limit. The sensor detection method uses high-performance flexible optoelectronic components such as LEDs and photodiodes (PDs) and is low-cost (see Panel D of FIG. 5).

[0078] Next, refer to FIG. 2 showing a schematic diagram of a sensor for ammonia (NH3) according to one aspect of the present invention. As shown, the sensing chamber may have a red light-emitting diode 110 arranged to illuminate a sensing region 106A and a reference region 106B positioned parallel to the flow direction. The photodiode 108 can be arranged below the sensing region 106A and the reference region 106B. The target gas can be directed into the sensing chamber where it is exposed to the sensor, and then the sensor exhibits a color change proportional to the concentration of ammonia (NH3) in the target gas. The photodiode 108 may be mounted on a printed circuit board having a resistor to obtain a photodiode signal sensitivity. By using Beer's law, the concentration of ammonia (NH3) gas can be determined through absorbance using the change in the negative logarithm of the signal ratio between the sensing region and the reference region.

[0079] FIG. 3 shows an analysis cartridge for total ammonia (ammonia (NH3) and ammonium (NH4 + )) according to one aspect of the present invention. As shown in FIG. 1, the analyzer 100 can communicate wirelessly with the user interface 102. Also, wired communication may be used. Further, as shown in FIG. 3, the sensing region is highly sensitive to the ammonia (NH3) concentration. In the sections before and after FIG. 3, the color change of the sensing region 106A indicates after exposure to ammonia (NH3).

[0080] Next, referring to FIG. 4, a graph of absorbance change according to one aspect of the present disclosure is shown. It shows the change in the absorption spectrum of the sensor obtained in the sensor chamber of a JAZ spectrophotometer before and after exposure to ammonia (NH3). As shown, the sensor has a maximum absorption range wavelength of 600 nm to 630 nm. The maximum absorbance of the ammonia sensor (NH3) occurs between 600 and 630 nm, and the minimum absorbance occurs at wavelengths higher than 675 nm.

[0081] FIG. 5 shows total ammonia (ammonia (NH3) and ammonium (NH4+ )) shows a schematic diagram and an assembly diagram of the sensor for [purpose]. Panel A of FIG. 5 shows a nanocomposite sensor of bromophenol blue (BPB) contained in the indicator layer of the extraction membrane. As shown in Panel A of FIG. 5, the sensing element can change color after exposure to ammonia (NH3). The sensing element may be made of chemically selective nanocrystals of BPB deposited on a porous hydrophobic substrate that provides a rapid and reversible reaction to ammonia (NH3). The sensor fabrication process is shown in Panel B of FIG. 5. As shown, the sensor manufacturing process may include a lamination and laser cutting process. Panel C of FIG. 5 shows the absorption spectrum of the sensor during exposure to ammonia (NH3). A maximum absorption wavelength of 630 nm is shown. Panel D of FIG. 5 shows a sensor assembly (referred to as a hybrid sensor) by the lamination process shown in Panel B of FIG. 5. A schematic diagram of this sensor assembly shows optoelectronic components for simultaneous detection at the maximum and minimum absorption wavelengths.

[0082] As shown in Panels A and C of FIG. 5, the nanocomposite for the ammonia (NH3) sensor may be manufactured using a pH indicator (e.g., bromophenol blue, BPB) as a molecular probe. Any suitable alternative pH indicator can be used. In some embodiments, the colorimetric sensor substrate may be composed of a custom or commercially available (e.g., polytetrafluoroethylene (PTFE)) membrane that is immersed in a BPB (colorimetric detector) solution that generates nanocrystals when deposited on the substrate. Next, the substrate of the modified PTFE can be dried at 25 °C. This process can generate a nanocrystalline structure formed on PTFE in the molecular probe (BPB). In some embodiments, this can enable an important or rapid reaction with the analyte ammonia (NH3). In some embodiments, the molecular probe can be used to rapidly and selectively detect ammonia (NH3) from the headspace of urine or skin. Different combinations of chemicals and substrate preparation methods were screened and studied as discussed below in relation to the experiments. In some embodiments, the porous hydrophobic substrate for the immobilization of BPB (PTFE) can promote not only fast ammonia (NH3) reactions but also reversible reactions because the substrate does not retain ambient water, thereby avoiding the permanent solubilization of ammonia (NH3). As shown, the resulting nanocomposite showed fast and highly responsive nanocrystals (<200 ms) with high specificity in the presence of urine / sweat interfering molecules (see Panel C of FIGS. 5 and 6) (see Panel A of FIG. 6).

[0083] In panel D of FIG. 5, the structured sensor is depicted with its main components. The main components have a laminated film on a spacer, and the laminated film and the spacer are disposed on the sensor. Alternatively, the extraction film may be physically separated from the sensor or the sensor chamber. Further, the assembly has a feed dispenser that is in direct contact with body fluids (i.e., urine, sweat). The illustrated components may be integrated into the sensor cartridge using a mask layer via the lamination process illustrated in panel B of FIG. 5. The laminated components may form a single assembly that fits within the sensor chamber together with optoelectronic components as shown in panel D of FIG. 5 and panel A of FIG. 1. Signals generated by the optoelectronic components may be electronically captured and processed with calibration data. The data may then be transmitted to the user interface 102 either wired or wirelessly. Then, to show trends in the total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration and / or excretion rate in body fluids, the data can be graphically displayed over time. In some embodiments, an automatic warning signal for rapid changes in the total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration and / or excretion may be sent to the attending clinician to alert the attending clinician of changes in the metabolic state and / or potentially harmful conditions.

[0084] The cartridge is designed with flexible electronics integrated to fit a tubular system or an adhesive strip, thereby allowing the user to easily "plug and play" as shown in two configurations of the analyzer illustrated in panels C and D of FIG. 1. In some embodiments, to eliminate the need for focusing optics, the LED and PD may be placed on a flexible printed circuit board (PCB) as close as possible to the sensor cartridge, thereby reducing size and cost (see panels A and D of FIG. 5).

[0085] The LED and PD may be used in reflection mode. Two LEDs may be used to reduce sensor drift signals. The analyzer 100 can form an integrated unit adapted to fit into a tube or an adhesive strip. In both versions of the wearable analyzer (see Panels C and D of FIG. 1), electronics including electro-optical components, a microcontroller, power from a small thin-film flexible battery (e.g., Blue Spark Technologies Inc.), a small display, a power switch, and low-energy Bluetooth® are mounted on a flexible PCB. The flexible PCB has one microcontroller for controlling and reading colorimetric sensor signals, general functions for data collection, minimal data processing for the embedded display (when used as an option for the total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration), and may have transmission via Bluetooth®. In some embodiments, the entire described assembly can be placed within a sleek housing designed to be portable, functional, and ergonomic.

[0086] In some embodiments, to reduce baseline drift of an ammonia (NH3) sensor during use due to temperature changes, changes in mechanical operation, changes in the stability of electronic components, etc., the sensor may be composed of two identical sensing regions and two identical reference regions. Each pair of the sensing region and the reference region may be illuminated by an LED. The LED may have a specific wavelength. One LED can have a wavelength of 630 nm and can be used to capture the maximum absorbance change (Abs max) of the sensing probe. The second LED can have a non-absorbing wavelength (e.g., 700 nm) and can be used to capture the baseline minimum absorbance (Abs min) of the sensing probe. The difference in absorbance: Δ absorbance = maximum absorbance - minimum absorbance can be used as the sensor signal. The use of two wavelengths corrects for additional baseline drift in the sensor system (see Panel D of Figure 5).

[0087] Furthermore, as also shown in Panel D of Figure 5, in some embodiments, a single LED is used to illuminate the sensing region and the reference region to reduce fluctuations in LED light intensity. The absorbance reading of the sensing probe can be calculated as follows: absorbance = -log (sensing region reading / reference region reading).

[0088] Figure 6 shows specificity analysis (Panel A), ammonia (NH3) detection accuracy (Panel B), reversibility and response time (Panel C), and sensor life (Panel D).

[0089] As shown in Panel A of Figure 6, the sensor can be configured to be specific to ammonia (NH3) and not react to other materials.

[0090] Furthermore, as shown in Panel B of FIG. 6, the assembly of the extraction membrane and the ammonia (NH3) sensor can exhibit high sensitivity and a high level of accuracy for the detection of ammonia (NH3) in the range from parts per billion (ppb) to parts per million (ppm) of ammonia (NH3) detection when compared to commercial methods such as enzyme reference methods or ion-selective electrodes. For example, as shown in Panel B of FIG. 6, the correlation coefficient (r2) = 0.998, and the accuracy is 88% with a 2% error (95% confidence interval).

[0091] Panel C of FIG. 6 shows the reversibility and time response of the sensor. It shows the sensing region potential response over time to periodic exposure to high and low parts per million of ammonia (NH3).

[0092] Panel D of FIG. 6 shows the sensor sensitivity (absorbance vs. concentration) after 24-hour exposure to ammonia (NH3) levels. As shown, the sensor demonstrates reusability after pretreatment. Pretreatment includes placing the analyzer at 45 °C for two weeks. This enables strong immobilization of components to the support substrate and allows for stringent shipping and operating conditions. In connection with this test, ammonia (NH3) extraction was performed on a high-concentration glycine buffer membrane of polystyrene / PTFE with pH = 10 cured with an organic hydroxide.

[0093] In some embodiments, as shown in FIG. 7, the analyzer can further include a temperature sensor 120. The illustrated analyzer is configured to contact the skin for measuring total ammonia (ammonia (NH3) and ammonium (NH4 + )) in sweat. In some embodiments, the temperature sensor 120 may be mounted adjacent to the assembly sensor surface to determine the in-situ temperature of the assembly sensor and provide correction of the ammonia (NH3) level readings due to temperature changes. Furthermore, the embodiment of the analyzer shown in FIGS. 7 and D of FIG. 1 may have an adhesive layer that provides a hermetic seal of the skin headspace compartment.

[0094] In some embodiments, the analyzer may have one or more sensors for at least one of fluid pH, fluid density, fluid specific gravity, fluid osmotic pressure, fluid temperature, oxygen (O2) partial pressure, carbon dioxide (CO2) partial pressure, nitrogen (Na + ) partial pressure, sodium (Na + ), potassium (K + ), chloride (Cl - ), bicarbonate (HCO3 - ), calcium (Ca 2+ ), magnesium (Mg 2+ ), phosphate ions (H2PO4 - , HPO4 2- , PO4 3- (including)), creatinine, urea, uric acid, cystatin C, amino acids, renal tubular brush border enzymes, albumin, Tamm-Horsfall protein, insulin, cortisol, cortisone, creatinine, lactate, cyclic adenosine monophosphate, neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), insulin-like growth factor binding protein 7 (IGFBP7), and tissue inhibitor of metalloproteinase 2 (TIMP2). The analyzer may have a flow sensor configured to determine the total volume of the fluid and / or the fluid production rate. In some embodiments, the rate of fluid production may be expressed in units of urine volume per unit time.

[0095] In some embodiments, the analyzer 100 may also have one or more signal processing algorithms configured to process raw data from the sensors and calibrate for any memory effects within the sensors. In some embodiments, the signal processing algorithms can account for any memory effects within the sensors when the concentration of the supply solution changes rapidly.

[0096] The measurement of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in biological samples has conventionally presented technical challenges. Total urinary ammonia (ammonia (NH3) and ammonium (NH4 + )) and / or ammonium (NH4 +)Concentrations are generally not measured, and physicians are trained to calculate and utilize a flawed indirect indicator (i.e., the "urine anion gap") to estimate the concentration of total ammonia (ammonia (NH3) and ammonium (NH4 + )) and / or ammonium (NH4 + ). However, more reliable methods for determining total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels in blood, urine, and other body fluids (e.g., breath, sweat) are very beneficial in certain treatment scenarios. For example, blood total ammonia (ammonia (NH3) and ammonium (NH4 + )) is a critical marker used to inform treatment decisions in patients with urea cycle disorders, organic acidurias, carnitine deficiency due to fatty acid oxidation defects, dibasic aminoacidurias, pyruvate metabolism defects, and liver diseases (e.g., cirrhosis) (although sampling is more inconvenient than urine). Urine total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels are known to vary with blood levels of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in UCD patients, and thus serial measurements of urine total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels are very useful for individualizing treatment of UCD patients without the need for very frequent blood sampling. Dynamic changes in renal total ammonia (ammonia (NH3) and ammonium (NH4 + )) production (i.e., renal ammonia production) are stimulated by acid-base balance, potassium balance, and other systemic states. Thus, an immediate understanding of urine total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels in patients prone to acid-base or potassium disorders (i.e., hospitalized patients with serious illnesses) can be utilized to increase immediate clinical knowledge and function as an early warning signal for rapidly changing (and otherwise unrecognized or unacknowledged) systemic states. Total ammonia (ammonia (NH3) and ammonium (NH4+ Considering the complex interplay of the adaptations of the kidneys and the liver to homeostasis, a disorder in either of these organs can result in a rapid change in the total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels in the body fluids. For example, acute liver dysfunction or decompensation is associated with an increase in the total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels in the plasma, and acute kidney dysfunction is associated with a rapid decrease in the total ammonia (ammonia (NH3) and ammonium (NH4 + )) in the urine. In ambulatory patients, continuous monitoring of the total ammonia (ammonia (NH3) and ammonium (NH4 + )) levels in biological samples (including breath, sweat, blood, and urine) can provide a baseline level of total ammonia (ammonia (NH3) and ammonium (NH4 + )), and a deviation therefrom is a strong predictive signal of unresolved liver decompensation or a predictive signal of unresolved renal insufficiency in patients with advanced liver disease. In hospitalized patients, patients with an indwelling urinary catheter at high risk of acute kidney injury can be monitored for a rapid change in the total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration or excretion rate in the urine as the first sign of renal tissue damage or acute kidney injury. Utilizing this technique can lead to the rapid identification of acute renal failure (compared to the lagging of conventional markers including serum creatinine, which takes minutes to hours or days). In any of these scenarios, specific treatments to improve the underlying organ failure or dysfunction can be employed in a much more rapid and individualized manner than currently practiced in modern medical settings. Therefore, the systems and methods described herein for detecting total ammonia (ammonia (NH3) and ammonium (NH4 + )) can be adapted for use in the clinical setting.

[0097] In some embodiments, the systems and methods described herein can improve the health outcomes of hospitalized patients experiencing acute kidney injury (AKI) events and reduce associated healthcare costs. For hospitalized patients with indwelling urinary catheters, the systems and methods described herein can continuously monitor for AKI and, when a suspected AKI event begins and when it begins, automatically send a signal to healthcare team members. Previous studies have shown that patient outcomes improve when AKI events are recognized more quickly.

[0098] In some embodiments, the systems and methods described herein can assist clinical researchers in testing new therapeutic agents for AKI in humans. The lack of the ability to rapidly diagnose AKI (outside of a controlled laboratory environment in animal models) has significantly hindered most past attempts in AKI treatment research in human subjects and continues to severely impede AKI care in the clinical setting. This is at least partially because new therapies being tested in human study populations are almost universally administered outside of the ideal treatment window, and there are several reports even in studies where administration occurred several days after an AKI event was known to have started. Interestingly, many new therapeutic agents have shown great promise in animal experiments where the timing of AKI was precisely known and the drug was rapidly administered within 90 minutes after the AKI event occurred. In the clinical setting, the timing of AKI is unknown because 1) symptoms and signs are almost always absent, 2) markers are present with a significant delay (i.e., hours or days), and 3) detection systems for identifying the earliest moments of acute renal failure and / or early AKI have not been developed. With appropriate testing, it is possible to find a place in future clinical practice, such as the systems and methods described herein, that can rapidly detect human AKI for one of the new therapeutic agents that has shown great promise in animal models of AKI.

[0099] The systems and methods described herein for the detection of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in body fluids may also be used in physiological studies where renal ammonia production and / or excretion of total ammonia in the kidney (ammonia (NH3) and ammonium (NH4 + )) changes rapidly. Further, the systems and methods described herein can be used for the detection of medical conditions where current diagnostic tools are limited and changes in total ammonia in urine (ammonia (NH3) and ammonium (NH4 + )) may correlate with the onset or activity of a disease. These conditions include: 1) changes in renal function, 2) acute kidney injury or failure, 3) chronic kidney disease, 4) changes in liver function, 5) acute liver injury or failure, 6) chronic liver disease (e.g., cirrhosis), 7) acute gastrointestinal bleeding, 8) chronic gastrointestinal bleeding, 9) genetic or inborn metabolic disorders (e.g., urea cycle disorders, organic acidurias, carnitine deficiency due to defects in fatty acid oxidation, dibasic aminoacidurias, and defects in pyruvate metabolism) that involve or affect the production, handling, and / or excretion of ammonia (NH3) and / or ammonium (NH4 + ), 10) changes in normal metabolic processes (e.g., increased production and excretion of ammonia (NH3) and / or ammonium (NH4 + ) after a protein meal), 11) acute or chronic systemic acid / base changes or imbalances due to metabolic processes or disease states, 12) acute or chronic systemic acid / base changes or imbalances due to respiratory processes or disease states, but are not limited thereto.

[0100] In some embodiments, the systems and methods described herein may have a "plug and play", reversible, continuous use, and rapid response assembly sensor cartridge with a specific configuration of an extraction membrane and a colorimetric sensor.

[0101] In some embodiments, the systems and methods described herein may also have signal processing algorithms based on a particular optoelectronic system design for two wavelengths, and built-in mechanisms for removing drifts (built-in sensing regions and reference regions, as well as temperature sensors).

[0102] Furthermore, the systems and methods described herein can be applied to industrial uses such as the measurement of total ammonia (ammonia (NH3) and ammonium (NH4 + )) in wastewater such as groundwater discharge, recycled water, industrial wastewater, sanitary wastewater, and produced water from oil and gas wells.

Examples

[0103] The following examples are provided to illustrate exemplary embodiments of the present disclosure. However, it should be understood that the present disclosure should not be limited to the specific conditions or details described in these examples.

[0104] Example 1: Field Performance of the Analyzer The response of an analyzer constructed according to the systems and methods described herein was tested using actual human urine samples in comparison with the ion-selective electrode method. Urine samples were evaluated from subjects who consumed 1 g of protein / Kg body weight in a single meal (shake). After the meal, the samples were analyzed every hour for several hours. The ion-selective method required a two-point calibration before analyzing each sample. For the analyzer, a single assembly sensor was used for the analysis of a complete experiment. As shown in FIG. 8, both the reference method and the assembly sensor of the analyzer gave results with a correlation close to 1. This example shows that an analyzer using a single sensor can perform similarly to a reference method using actual samples of human urine, and this example reconfirmed the successful performance of the sensor by reusing it several times a day.

[0105] Example 2: Preparation of the Sensor In one example, an ammonia (NH3) sensor according to the systems and methods described herein was constructed based on Sigma-Aldrich's bromophenol blue (BpB). The sensor was synthesized by immersing the sensor substrate in a BpB solution. The sensor substrate in the solution was then vortexed for 10 minutes using a Scientific Industries Vortex Genie 2 and dried at room temperature for 5 minutes. To test the effect of the substrate on the detection sensitivity, the sensor was constructed on five different sensor substrates including Omnipore™ polyvinylidene fluoride (PVDF) [pore size: 0.1 μm and porosity: 80%], Sterlitech's polytetrafluoroethylene (PTFE) / polyethylene (PE) [pore size: 0.2 or 0.45 μm], Interstate Specialty Products' hydrophobic PTFE [pore size: 10 μm], Omnipore™ hydrophilic PTFE [pore size: 0.1 and porosity: 70%], and Whatman no. 1 filter paper [pore size: 11 μm]. The sensor substrates were cut into a rectangular shape (2.7 cm × 1.2 cm), stacked so that they fit into the sensing chamber of the analyzer, and arbitrarily named a colorimetric optoelectronic mechanical analyzer (CODA). A portion of the constructed sensors was enclosed in a black Mylar™ bag and placed in an oven at 45 °C for 2 days to test their performance stability.

[0106] Example 3: Preparation of the Analyzer An analytical device, namely, a colorimetric optoelectronic mechanical analyzer (CODA), was constructed according to the systems and methods described herein. The analytical device has a horizontal flow path passing through the sensing chamber, and this flow path has a red LED above the sensor and four photodiodes (sensing / reference pair and sensing / reference backup pair) below the sensor. The target gas is introduced into the sensing chamber where it is exposed to the sensor, and the sensor showed a color change proportional to the concentration of ammonia (NH3) in the target gas. The photodiodes (manufactured by Vishay Semiconductor Opto Division) were mounted on a PCB integrated with a Bluetooth unit together with a 5MΩ resistor to obtain the photodiode (PD) signal sensitivity, enabling signal transmission to an Android phone. The application was created to provide a user interface for displaying the signal read by the PD within the range of 0 to 3V. The sensor had a reference region and a sensing region. The background responses from the reference region and the sensing region when the sensor was in the chamber were measured to be approximately 1.2V. A pair of PDs continuously read the responses from the reference region and the sensing region simultaneously every 0.2 seconds.

[0107] Figure 9 shows the results of Example 3, particularly the change in the sensor signal before and after exposure to ammonia (NH3). The reproducibility of the sensor was determined using (a) four different PVDF sensors for detecting 2 ppm of ammonia (NH3), and (b) four different PTFE sensors for detecting 40 ppm of ammonia (NH3) using the analytical device. The four different substrates showed similar signal responses. The PTFE response signal has higher noise compared to the PVDF substrate.

[0108] The absorbance was calculated based on Beer's law by taking the negative logarithm of the ratio of the signal response from the sensing region (S sens. ) to the signal response from the reference region (S ref. ) as shown in Equation 1 below.

Equation

[0109] PTFE and PVDF sensors were cut into rectangles using a laser cutter (Universal Laser System) and then laminated with a Fellowes Jupiter 125 Laminator. A calibration curve for PTFE was created by plotting the measured absorbance changes against the known concentrations of the samples in the concentration range of 2 - 1000 ppm of ammonia (NH3).

[0110] A crosstalk test was conducted to ensure no interference between the PD readings from the sensing and reference regions of the sensor. In this test, either the reference or sensing region was individually masked with thick black ink to block the light. The measurement was carried out for 30 seconds to confirm that the response to the blocked region was zero and that the unblocked sensor region was not affected. The crosstalk test results are shown in Table 1 and Table 2.

Table 1

Table 2

[0111] For both masked substrates, the crosstalk test showed small signal changes (<0.1% for the sensing region and <15% for the reference region), which were not significantly important under the sensing conditions and could be further improved by creating a thicker barrier between the PDs or reducing the distance from the sensor to the detector.

[0112] Example 4: Optoelectronic Devices To conduct sensitivity tests and spectral measurements of various sensor materials before and after exposure to ammonia (NH₃), a JAZ spectrophotometer (JS) manufactured by Ocean Optics was used. Figure 10 shows a schematic diagram of the JS measuring device. The optical fiber is at the top of the chamber, while the tungsten light source is at the bottom of the chamber. The gas enters from the left tube and is released into the ambient environment from the right tube. When the response of sensors synthesized from various materials after being exposed to 10 ppm ammonia (NH₃) for 180 seconds was measured by JS, PVDF showed an excellent absorbance response compared to other materials.

[0113] The filter paper was cut into a round shape to fit the JS sensing chamber. An ammonia (NH₃) sensor integrated with an ammonia (NH₃) extraction membrane was used for spectral measurement. A schematic diagram of sensor and sample transport from the measured liquid fluid to gas is shown in Figure 11. The extraction membrane / sensor assembly consisted of five parts, namely: 1) a distribution layer (e.g., filter paper) to evenly disperse the liquid fluid, 2) an alkalization membrane layer (e.g., a PE membrane impregnated with 40 μL of 2M NaOH solution) to extract ammonia (NH₃) from the sample, 3) a polytetrafluoroethylene (PTFE) membrane to prevent the liquid fluid from reaching the indicator layer, 4) an indicator layer (filter paper impregnated with bromothymol blue) that reacts with the extracted ammonia (NH₃), and 5) a tape mask to protect the sensing probe. Synthetic urine feed (a solution containing ammonium (NH₄ + ) and other ions simulating urine: NaCl, KH₂PO₄, CaCl₂, MgSO₄) was injected from the top of the integrated ammonia (NH₃) sensor membrane. JS quantified the ammonia (NH₃) level in the sample. The ammonia (NH₃) sensing mechanism will be further explained below.

[0114] The dispenser evenly disperses the sample feed. The alkali layer converts the ammonium (NH₄ +) is converted to ammonia (NH3), its conjugate base. The PTFE membrane selectively filters ammonia (NH3) gas based on the hydrophobicity of the membrane. The ammonia (NH3) sensor has an indicator that changes color from yellow to blue based on how much ammonia (NH3) gas it is exposed to.

[0115] Example 5: Optoelectronic Sensor Signals As previously mentioned, bromophenol blue (BpB) was used as a colorimetric detection probe for ammonia (NH3) detection. The BpB solution has a yellow / orange color when exposed to pH levels below 3 and a blue color when exposed to pH above 4.6. Ammonium (NH4 + )(acid) and ammonia (NH3)(conjugate base) and the acid / base balance between them is determined by the pH of the solution in the overall reaction OH - +NH4 + ←→H2O + NH3. Ammonia (NH3) has a vapor pressure of 1062 kPa and a pKa of 9.25 at room temperature. Biologically relevant pH conditions are below the pKa of the NH4 + / NH3 equilibrium. For example, in human urine at a relatively high pH of 8, only 6.6% of the total NH4 + / NH3 exists as NH3 (gas). Due to the dynamic nature of body fluid pH (e.g., urine) and the variable ratio of urinary NH4 + to urinary NH3, an alkaline solution is required to raise the fluid sample pH above ~10 to ensure 100% conversion of NH4 + (liquid) to NH3 (gas). Ammonia (NH3) makes the sensing surface more alkaline, shifts the pH value higher, and causes a change from yellow to blue. By quantifying the color change using an analyzer (CODA), the corresponding ammonia (NH3) concentration obtained from the sample can be determined.

[0116] Example 6: Gas Sample Preparation - Ammonia Bag The ammonia (NH3) gas samples used in this study were diluted with 100 ppm and 1000 ppm calibration ammonia (NH3) gases purchased from Calibration Technologies, Inc. Dilutions of the gas samples in laboratory compressed air were prepared from 100 ppm and 1000 ppm ammonia (NH3) calibration gases. These calibration gases were introduced into a 40 L bag for a predetermined time using a TOPSFLO (flow rate: 1.6 LPM) microdiaphragm gas pump. Additional clean air was also introduced into the bag for a controlled period until the concentration of ammonia (NH3) in the bag reached the desired level. The target ammonia (NH3) gas concentration was prepared by manipulating the ratio of ammonia (NH3) gas injection to air injection (0.02 - 0.8). Another ammonia (NH3) bag was prepared by injecting 5 μL of ammonium hydroxide (NH4OH) into a 1 L Tedlar (trademark) bag and leaving it at ambient room temperature for 30 minutes to confirm the calibration curve of the sensor.

[0117] Example 7: Gas Sample Preparation - Urine Headspace Bag The urine test samples were pretreated by adding 0.3 mL of 10 M NaOH to 2.7 mL of urine samples so that the pH of the samples exceeded 12. Then, the pretreated urine samples were added to a 4 L Tedlar (trademark) bag and purged with dry air until the bag was full. The Tedlar bag was left at room temperature for 30 minutes to confirm that all ammonium (NH4 + ) reacted with the base and became its conjugate phase ammonia (NH3) in the urine headspace. The subjects in this part of the study received approval from the Institutional Review Board of Arizona State University (IRB protocol #1012005855). The subjects participated voluntarily and gave written consent to participate in this trial. All tests in this study were conducted from February 2016 to July 2017. The subjects drank "ON High protein Gainer protein shake" at 1 g of protein per kg of body weight and urinated regularly after drinking. Urine samples were collected and immediately stored in a -80 °C freezer for later analysis.

[0118] Example 8: Detection Procedure of the Sensor The sensitivity, reversibility, and reusability of an ammonia (NH₃) sensor were tested using an ammonia (NH₃) flow system having a microdiaphragm gas pump (flow rate: 1.6 LPM), a three-way valve, one 40 L airbag, one 40 L sample bag, and a sensing chamber. The test was conducted each time with one sensor placed in the sensing chamber. The three-way valve was first switched to connect to the airbag for several seconds, whereby the sensor was purged in air before being exposed to the sample for several seconds. To study the sensor's sensitivity for different sample exposure times, the sampling time was varied to include 1, 5, 20, and 180 seconds. After exposure to ammonia (NH₃), to test the reversibility of the sensor, the valve was switched to allow dry air to pass through the system for several seconds.

[0119] Example 9: Results and Discussion - Selection of the Wavelength of the Colorimetric Optoelectrochemical Analyzer (CODA) The color of the light source for an analytical device, which can be called a colorimetric optical dynamics analyzer (CODA), was selected based on the spectral changes induced on the sensing probe (BpB) upon ammonia (NH₃) exposure. Circular sensors formed of filter paper impregnated with BpB were placed in the sensing chamber of the JS device, and the spectrum of each sensor was recorded before and after exposure to ammonia (NH₃). Figure 4 shows the visible spectrophotometric changes of the BpB-based sensor, and a significant increase in absorbance in the range of 575 - 625 nm is clearly observed. Based on these results, the LED color was selected to be red, wavelength: 610 nm. Once the detection wavelength and the first screening of the sensor substrate were selected, the analytical device, CODA, was constructed and used to proceed with the remaining research.

[0120] Example 10: Results and Discussion - Sensitivity of the Sensing Probe Table 3 shows the characteristics of various sensing substrates embedded with BpB, and Figure 10 summarizes the sensitivities of the sensing substrates tested with the JS device after exposure to 10 ppm ammonia (NH3) gas for 180 seconds. The sensitivity of the sensor to ammonia (NH3) strongly depends on the nature of the substrate. The graph in Figure 10 shows that the PVDF substrate has the largest measured sensitivity, which is about 10 times greater than that of all other substrates tested. The normal total ammonia (ammonia (NH3) and ammonium (NH4 + )) concentration in urine is typically higher than 6 mmol / L, which (according to the complete conversion from ammonium (NH4 + ) to ammonia (NH3) and ammonia (NH3) gas extraction) results in an ammonia (NH3) gas concentration in the urine headspace higher than 100 ppm at a temperature of 25°C based on the ideal gas state equation. Since the sensor rapidly saturates with a single use, the high sensitivity of PVDF prevents the sensor from being used multiple times. Therefore, the sensitivity, specificity, and reversibility characteristics of the hydrophobic PTFE, another substrate, were investigated compared to PVDF for ammonia (NH3) monitoring in urine.

Table 3

[0121] Example 11: Results and Discussion - Reproducibility of the Sensor Response Figure 12 compares the absorbance responses of sensors based on PTFE and PVDF support materials as used in CODA. Four replicate sensors were fabricated using each material and placed in CODA. Next, the sensors were exposed to ammonia (NH3) for 180 seconds and then further exposed to dry air for 60 seconds to measure the recovery rate. The sensors show similar response characteristics that the absorbance increases when ammonia (NH3) is injected and decreases when purged with dry air. The absorbance noise of the PTFE substrate was higher compared to the PVDF substrate.

[0122] Table 4 summarizes the percentage of sensor recovery after purge, which is the ratio of the change in absorbance during the recovery period to the change in absorbance during the sensor response and exposure period. The sensor response included 0.64 a.u. with a standard deviation of 0.02 for PVDF and 0.58 a.u. with a standard deviation of 0.03 for PTFE, and the response dispersion across the entire sensor substrate was 5% or less. Even though PVDF had similar reproducibility to PTFE, it is important to note that the lower the ammonia (NH3) concentration required for comparison (20 times lower ammonia (NH3) concentration), the more similar the recovery rate was to that of PTFE. The recovery characteristics of the sensor response using PTFE within the concentration range of realistic urine-derived ammonia (NH3) concentrations made PTFE a more attractive candidate for further research on the analytical performance of this sensor substrate. As a result, in the remaining studies, PTFE sensors were investigated.

Table 4

[0123] Example 12: Results and Discussion - Ammonia (NH 3 ) Sensor Calibration As shown in Figure 13, two calibration curves were created for PTFE sensors using CODA with a sampling time of 5 seconds at ammonia (NH3) gas levels in the range of 2 ppm to 1000 ppm. For the first upper calibration curve, the Langmuir model was applied, showing an R 2 value greater than 0.99. For a sampling time of 5 seconds, the calibration equation is as follows, where A L represents the absorbance derived from the Langmuir model and C represents the corresponding concentration.

Equation

[0124] For the other lower calibration curve, the calibration curve was divided into two ranges for fitting a linear regression, namely 2 - 150 ppm and 150 - 1000 ppm. Both measurement ranges showed an R 2 value greater than 0.98. The calibration equation is as follows, where A1 represents the absorbance derived from a linear model of 0 to 150 ppm, and A 2 represents the absorbance derived from a linear model of 150 to 1000 ppm, and C represents the corresponding concentration.

Number

Number

[0125] In another set of fittings, a linear regression of the change in absorbance evaluated with 1 - second ammonia (NH3) exposure was also obtained and compared with that obtained with 5 - second ammonia (NH3) exposure. Using these regression analyses, the unknown sample concentrations obtained from a mixture of ammonium hydroxide (NH4OH) and the air in the bag were tested. Table 5 shows the results evaluated by the sensor for unknown concentration samples using 1 - second and 5 - second sample exposures, and the corresponding calibration curves. Both calibration curves (obtained from 1 - second and 5 - second exposure data) yielded the same concentration of the prepared ammonia (NH3) bags of unknown concentration, showing self - consistency in calibration. Furthermore, these results show the consistency between each pair of photodiodes in the system (PD1(sensing) / PD3(reference) shown as PD1 and PD2(sensing) / PD4(reference) shown as PD2) since both pairs of photodiodes produced the same response.

Table 5

[0126] Example 13: Results and Discussion - Ammonia (NH 3 ) Sensor Selectivity To confirm that the sensor is selective only to ammonia (NH3), the sensor was exposed to several interfering substances (e.g., acetone, 2-butanone, and methylene chloride) reported to be present in the urine headspace. Figure 14 shows the selectivity of the sensor to ammonia (NH3). Even with a relatively high concentration of interfering substance (e.g., 100 ppm acetone), the sensor only showed a significant response to ammonia (NH3). This test confirmed the selectivity of the sensor in the harsh environment of urine headspace samples.

[0127] Example 14: Results and Discussion - Reversibility and Reusability of the Sensor Healthy adults may urinate every 2 - 3 hours (8 - 9 times a day). Current methods for quantifying ammonium (NH4+) in urine for clinical medicine involve requiring patients to collect all the urine excreted over 24 hours. There is no clinically used method for the instantaneous measurement of urinary ammonia (NH3) or total urinary ammonia (ammonia (NH3) and ammonium (NH4+)). The upper panel of Figure 15 shows the absorbance response of a PTFE-based sensor to repeated alternate exposure to 100 ppm ammonia (NH3) and dry air over 1.2 hours. The sensor was continuously subjected to a repeating cycle of 5-second exposure to ammonia (NH3) followed by 120-second exposure to dry air. This sensor was reused for over 60 detection cycles without performance degradation. In actual clinical applications, to cover 24-hour monitoring, ammonia measurements can be taken for 5 seconds once every 24 minutes, but much more frequent testing is also possible.

[0128] The lower panel of Figure 15 shows the measured concentrations derived from consecutive tests after signal analysis and the use of a calibration equation. It is important to note that the sensor required an adjustment period of 5 - 7 exposures. After this adjustment time, the concentration output remained fairly constant through multiple exposures to ammonia (NH3) and detection events at the same concentration. The detection concentration error was less than 20% and could be further improved with a better enclosure of CODA, which reduces ambient light interference.

[0129] Example 15: Results and Discussion - Sensor Stability To test the stability of the PTFE sensors, a new set of sensors was prepared and used in an ammonia (NH3) test immediately after synthesis. The same set of sensors was prepared, sealed in a black Mylar (trademark) bag, and aged in a convection oven at 45 °C for two weeks. According to the aging protocol (ASTM F1980), two weeks of aging at 45 °C is equivalent to two months of aging at room temperature (25 °C). Both sets of sensors were exposed to ammonia (NH3) at concentrations of 2, 10, 15, and 20 ppm. Figure 16 is a diagram showing a sensitivity comparison between fresh sensors and aged sensors. Figure 16 shows the linear regression of all data and the averaged data. A slope of 0.001 a.u. / ppm and an R 2 greater than 0.99 were obtained for the average data. Figure 16 also shows another set of fresh and aged sensors obtained from different synthesis batches. In a t-test between these responses from the membranes of each batch for fresh and aged sensors, a p-value equal to 0.87 was obtained, which showed no significant difference. These tests confirmed the stability of the sensing probe (BpB) on the PTFE substrate during long-term thermal exposure, which is relevant to sensor application and storage. In the case of commercial products, the sensor sensitivity needs to be guaranteed after the thermal exposure period that can occur under actual shipping or storage conditions.

[0130] Example 16: Results and Discussion - Use of the Sensor for Urine Samples To confirm the feasibility of CODA and the use of the sensor under actual conditions, urine sample analysis was performed and measurements from a calibrated batch of the sensor were recorded. As a reference method for ammonia (NH3) detection, an ion-selective electrode (ISE) [Thermo Fisher Scientific's ammonia high-performance ion-selective electrode (no. 9512HPBNWP)] was used. Subjects were first asked to urinate and then given a protein shake to drink. Urine samples from the subjects were collected before and after drinking the shake at 0, 0.5, 2.5, and 3.5 hours. These samples were stored at -80 °C before measurement. Next, the samples were measured with the ISE electrode and then with CODA. The upper panel of Figure 17 shows an example of the measurements from one subject. Similar results can also be seen in the literature using SIFT-MS. The lower panel of Figure 17 shows a correlation plot of the results evaluated from the CODA method and the ISE method. A good agreement was seen with an accuracy close to 100% between the measurements by CODA and the ISE electrode.

[0131] Example 17: Results and Discussion - Use of the Sensor for Urine Samples as an Insert in Diapers or Adhesive Patches or Cards As shown in Figure 18, in some embodiments, the sensors described herein may be used in the form of inserts for diapers or wearable fabrics. The use of the sensor with the extraction membrane can be implemented in the form of an insert for a diaper or wearable fabric or device (e.g., a bracelet), or an adhesive patch for the skin, or a card (e.g., a badge) for in vitro testing. Under these conditions, the quantification of the sensor response (i.e., color change) due to the detection of total ammonia (ammonia (NH3) and ammonium (NH4 + )) and / or ammonium (NH4 + ) as ammonia (NH3) can be performed by any method capable of detecting slight color changes, such as RGB deconvolution software.

[0132] Example 18: Results and Discussion - Use of the Sensor for Continuous Samples FIG. 19 shows the components of an example CODA device. In some embodiments, the sensors described herein can be used for continuous ammonia monitoring from a biological sample and can be inserted into a device that can manage the liquid from the sample and the gas from the extraction process within the sensor, and the sensor surface (sensing probe) can be regenerated with a clean source of air, such as from a scrubber.

[0133] Example 19: Results and Discussion - Continuous Use of the Sensor for Extraction Membrane and Quantification of Samples As shown in FIGS. 20a - 20o, a model of the extraction membrane from the sensor can be constructed to optimize the geometry, chemical / physical design, and lifetime. Along this line, models with extraction membranes that chemically convert NH4+ to NH3 via an alkaline substance, or electrochemically generate an alkaline substance to convert NH4+ to NH3, or electrochemically convert NH4+ directly to NH3 can be utilized. Based on this model, in the range of 3.6 - 100.0 mM, which is the concentration typically seen in urine samples for the incoming NH4 + concentration, it is concluded that the incoming NH4 + concentration does not affect the concentration profile of the alkaline substance source (e.g., fixed hydroxide (OH - sites)). In other words, the concentration of the alkaline substance in the chemical extraction membrane is mainly affected by the inlet velocity, which is a parameter modeled as a change in the limiting state (e.g., inlet velocity u0). Also, this model can determine the number of times the chemical extraction membrane can be continuously used until the chemical alkaline substance (OH - ) is depleted. Embodiments such as the CODA device shown in FIG. 19 can be modeled assuming the sensor has a source of alkaline substance that depletes after each use. FIGS. 20a - 20o also show how different variables in the extraction membrane can be tested. These variables include porosity, boundary conditions, the initial NH4 + concentration in the sample, and the geometry. All variables can be analyzed to determine how they affect the alkaline substance concentration profile of the extraction membrane.

[0134] As an example, FIGS. 21a and 21b show the modeled results of the cross-sectional concentration profiles of alkaline substances in the chemical extraction membrane after the first cycle (shown in FIG. 21a) and after the 20th cycle (shown in FIG. 21b).

[0135] Furthermore, other design aspects of the extraction membrane are important. One of these is the elimination of potential ammonia leakage. Based on practice and simulation, multiple but narrow liquid paths, and a narrower exposure area to the liquid / air interface minimize ammonia gas leakage from the membrane. Additionally, further modification of the extraction membrane with potential agents for chelating amine groups eliminates interference from the non-enzymatic decomposition of primary amine group molecules that would otherwise make ammonia present in the sample and thus physiologically irrelevant. This modification eliminates the problem of overestimation of ammonia (FIG. 22a) that is known to be a problem under current test conditions (FIG. 23). In fact, the combination of all of the above factors gives an extraction membrane with high specificity, as shown in FIGS. 22a and 22b.

[0136] FIGS. 20a - 20o show examples of how different variables, namely porosity, boundary conditions, initial NH4 + concentration in the sample, and geometric shape, affect the concentration profile of alkaline substances in the extraction membrane. The NH4 + concentration, inlet velocity, and porosity were set to values of 37.8 mM, 0.05 m / s -1 and 0.34 respectively. The results shown here are the concentration profiles for the first (FIG. 20a), tenth (FIG. 20b), and twentieth (FIG. 20c) measurements. The NH4 + concentration and inlet velocity were set to 37.8 mM and 0.05 m / s -1 respectively. The results shown here are the twentieth measurement at porosities of (FIG. 20d) 0.34, (FIG. 20e) 0.66, and (FIG. 20f) 0.90. The NH4 + concentration and porosity were set to 37.8 mM and 0.34. The results shown here are (FIG. 20g) 0.0035 m / s -1 (FIG. 20h) 0.05 m / s-1 and (Figure 20i) 0.5 ms -1 This is the 20th measurement at the inlet velocity. The inlet velocity and porosity of the sample were set to 0.05 m / s -1 and 0.34, respectively. The results shown here are the 20th measurements at NH4 concentrations of (Figure 20j) 37.8 mM, (Figure 20k) 100.0 mM, and (Figure 20l) 3780.0 mM + . The concentration and porosity of NH4 were set to 37.8 mM and 0.34, respectively. This result shows the concentration profiles of how the hydroxide (OH + ) depletes in configurations of (Figure 20m) length 3 cm and diameter 2.5 cm, (Figure 20n) length 1.5 cm and diameter 2.5 cm, and (Figure 20o) length 1.5 cm and diameter 2.0 cm - .

[0137] Figures 22a and 22b show the analytical performance of the extraction membrane and the overall sensor for complex body fluids. In Figure 22a, the fluid was treated with a copper ion chelating material. These copper ions chelated (bonded) with the amine groups of amino acids and primary amine residues in other molecules, thus preventing enzymatic and non-enzymatic degradation (the degradation of these molecules causes an incorrect increase in ammonia levels). Figure 22b shows the overall selectivity of the extraction membrane in the sensor for whole blood using 170 μM ammonia (as ammonium). Figure 22b shows that the extraction membrane and the sensor have a negligible response to other blood components at known maximum concentrations: 5,000 mg / dL albumin, 3 mg / dL ascorbic acid, 5 mg / dL creatinine, 2656 mg / dL glucose, 370 mg / dL (5 mM) potassium ions, 228 mg / dL (3.9 mM) sodium ions, 1,000 mg / dL phosphate, 107 mg / dL urea, and 6.8 mg / dL uric acid

[0138] Figure 23 shows the reaction of the enzymatic reference method (Roche Cobas (registered trademark)) with plasma collected from the body, plasma spike 1 (spiked with glutamine), and plasma spike 2 (added with amino acids glutamine, L-arginine, L-asparagine, and creatinine, urea-ammonia metabolites). The concentration of the spiking agent ranges from 10 to 100 μM levels, simulating physiologically expected levels. This demonstrates that the enzymatic method is disrupted by the spontaneous non-enzymatic deamination of amino acids, resulting in an overly high reading value for ammonia.

[0139] Example 20: Results and Discussion - Use of the Sensor for Free Calibration for Ammonia Quantification The intelligent algorithm is constructed based on the quantified general sensor sensitivity and can be used as a means to avoid sensor calibration (either every time before using the sensor or for either the device or the sensor being used). The intelligent algorithm is supplied with physical / chemical behaviors such as the sensor sensitivity for the initial operating conditions of different sensors, like the initial signal (V) before sample detection. Figure 24 shows the demonstration of a quantifiable general sensor sensitivity that enables a calibration-free strategy for ammonia quantification.

[0140] Figure 24 shows the relationship between the measured initial signals from the sensing regions of different sensors having corresponding calibration curve slopes in the range of 3.6 mM to 18.6 mM for the sensor. The relationship between these two variables is linear with a regression coefficient of R 2 = 0.88. This linear relationship helps to construct a calibration-free intelligent algorithm for ammonia quantification.

[0141] Example 21: Results and Discussion - Use of the Sensor for High-Precision Continuous Quantification of Ammonia Figure 25 shows the continuous measurement of NH4 from 37.6 mM to 3.6 mM + and is consistent with the actual ammonia concentration having an error smaller than <15%. As shown in Figure 25, the above application can successfully achieve the continuous quantification of ammonia with a slight difference between the ammonia value measured by the sensor in this application and the true ammonia concentration. "Speed" is NH4 +It is a parameter defined for the CODA used to quantify the concentration.

[0142] Although the present disclosure has been discussed with respect to specific embodiments, it should be understood that the present disclosure is not so limited. There are numerous modifications, variations, and other embodiments that can be adopted which are described herein by way of example and remain within the scope of the present disclosure.

Claims

1. A system comprising an analyzer configured to communicate with a sample of body fluid, wherein the analyzer comprises a sensing chamber, An extractor disposed between the source of the body fluid and the sensing chamber, which extracts ammonia (NH ４ ＋ ) from ammonium (NH ３ ) present in the sample of the body fluid and discharges the ammonia (NH ３ ) into the sensing chamber, Separate from the sensing chamber, a first sensor is disposed within the sensing chamber and configured to quantify the amount of ammonia (NH ３ ) present in the sensing chamber, and is provided with The analysis device is configured to detect at least one of a change in organ function, a change in tissue function, and a change in metabolic state, based on the amount of the quantified ammonia (NH ３ ). A system.

2. wherein the extractor At least a part of the ammonium (NH ４ ＋ ) present in the sample of the body fluid is converted into ammonia (NH ３ ), and an alkali layer configured to perform the conversion The ammonia (NH ３ ) converted by the alkali layer is filtered, and a hydrophobic layer configured to discharge the ammonia (NH ３ ) to the sensing chamber, the system according to claim 1.

3. The system according to claim 2, wherein the alkaline layer contains at least one of an organic hydroxide, sodium hydroxide, and a buffer solution having a pH of 10 or higher.

4. The system according to claim 2, wherein the hydrophobic layer contains polytetrafluoroethylene, a polytetrafluoroethylene derivative, or a cellulose derivative.

5. The system according to claim 1, further comprising a distribution layer disposed between the source of the body fluid and the extractor and configured to distribute a sample of the body fluid along the extractor.

6. The system according to claim 1, wherein the first sensor has an indicator layer formed on a hydrophobic or hydrophilic substrate so as to reversibly or irreversibly change color according to the amount of ammonia (NH ３ ).

7. wherein the first sensor comprises a CMOS camera configured to capture an image of the indicator layer over time, a processor configured to detect discoloration of the image according to the amount of the ammonia (NH ３ ), and the system according to claim 6, further comprising.

8. wherein the first sensor further comprises at least one photodiode configured to measure a change in absorbance of the indicator layer, and at least one light emitting diode configured to illuminate the indicator layer. The system according to claim 6, further comprising at least one light emitting diode configured to emit light at a maximum absorption wavelength of the indicator layer and another one of the at least one light emitting diode configured to emit light at a minimum absorption wavelength of the indicator layer.

9. The system according to claim 8, wherein at least one of the at least one light emitting diode emits light at a maximum absorption wavelength of the indicator layer, and another one of the at least one light emitting diode emits light at a minimum absorption wavelength of the indicator layer.

10. wherein the first sensor is further configured to quantify at least one of carbon dioxide or bicarbonate present in the sample of body fluid, The system according to claim 1, wherein the analyzer is configured to detect at least one of a change in organ function, a change in tissue function, and a change in metabolic state based on the quantified amount of ammonia and the quantified amount of at least one of carbon dioxide or bicarbonate.

11. The system according to claim 1, wherein the analyzer further comprises a second sensor configured to quantify at least one of carbon dioxide, bicarbonate, or urea present in the sample of body fluid, The system according to claim 1, wherein the analyzer is configured to detect at least one of a change in organ function, a change in tissue function, and a change in metabolic state based on the quantified amount of ammonia and the quantified amount of at least one of carbon dioxide, bicarbonate, or urea.

12. The system according to claim 1, wherein the analysis device further comprises an inlet configured to receive the body fluid.

13. The system further comprises a fluid sensor associated with the inlet and configured to determine at least one of the total volume of fluid passing through the inlet and the flow rate of fluid passing through the inlet. The system according to claim 12, wherein the analysis device is configured to detect at least one of a change in organ function, a change in tissue function, and a change in metabolic state based on the quantified amount of ammonia and the flow rate of the fluid.

14. The system according to claim 13, wherein the fluid sensor is one of a fluid flow rate sensor, a fluid viscosity sensor, a fluid density sensor, a fluid osmotic pressure sensor, a fluid osmolality sensor, and a fluid specific gravity sensor.

15. The analysis device is configured to be disposed within a catheter that receives a sample of the body fluid. The system according to claim 12, wherein the inlet has an opening through which the sample of the body fluid passes.

16. The analysis device is configured to be disposed in fluid communication with a body surface corresponding to the source of the body fluid. The system according to claim 12, wherein the inlet has a surface of the extractor.

17. The system according to claim 1, further comprising an inlet configured to receive the body fluid and a first valve configured to control the volume of the body fluid passing through the inlet.

18. Connected to a source of zeroing substance, controlling the transport of the zeroing substance through the analyzer, and discharging the sample of the body fluid and the ammonia (NH ３ ) from the analyzer, the system according to claim 1, further comprising a second valve configured to do so.

19. The extractor extracts ammonia (NH ４ ＋ ) from ammonium (NH ３ ) present in a next, second sample of the body fluid discharged from the analyzer, and is configured to discharge the ammonia (NH ３ ) into the sensing chamber. The system according to claim 18, wherein the first sensor is configured to quantify the amount of ammonia (NH ３ 3) in the sample of the second body fluid present in the sensing chamber.

20. The system according to claim 1, further comprising a user interface device configured to receive at least one transmission from the analysis device, the user interface device further having a display including a graphical user interface configured to display an output from the analysis device.

21. The analysis device is further configured to Transmitting the amount of the quantified ammonia (NH ３ ) to the user interface device, and Identifying the change over time in the amount of the quantified ammonia (NH ３ ) perform at least one of

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