Systems and methods for infusion of hydrogen

Abstract

This disclosure relates to hydrogen gas infusion (HI) systems and methods thereof. Systems described include ones comprising: a fluid reservoir containing a medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate the hydrogen gas with the medical solution to produce a hydrogen-infused solution; and a gas dilution module in fluid communication with and downstream of the fluid reservoir, the gas dilution module for continuously receiving and diluting excess hydrogen gas from the fluid reservoir without allowing the excess hydrogen gas to reach a combustible concentration. Such innovations described herein provide for an efficient system for infusing high concentrations of hydrogen gas into a medical solution for treatment of an organ or tissue.

Description

[0001] SYSTEMS AND METHODS FOR INFUSION OF HYDROGEN

[0002] CROSS REFERENCE TO RELATED APPLICATIONS

[0003] [1] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 556,631, filed February 22, 2024, entitled SYSTEMS AND METHODS FOR INFUSION OF HYDROGEN, the disclosure of which is incorporated herein by reference in its entirety.

[0004] BACKGROUND

[0005] [2] After a period of ischemia, anoxia, or hypoxia (e.g., lack of oxygen), tissue can be subject to damage by reperfusion injury when blood supply returns. The absence of oxygen and other nutrients from the tissue during the period of ischemia can create conditions where restoration of circulation can result in inflammation and oxidative damage due, in part, to reactive oxygen species that come into existence during reperfusion. This can occur, for example, through induction of oxidative stress instead of, or in addition to, restoration to normal tissue function.

[0006] [3] Hydrogen has been shown to be a viable molecule for reducing oxidative stress in blood, tissues, or organs, but hydrogen concentrations and methods of administering thereof have been limited so as to avoid risk of combustion. Further, such methods of administration often result in insufficient hydrogen delivery and uneven distribution throughout the affected regions.

[0007] BRIEF SUMMARY

[0008] [4] In view of the foregoing, there is a need for improved systems and methods of safe infusion of highly concentrated hydrogen gas into solutions for delivery to patients to reduce or prevent the occurrence of oxidative stress, while simultaneously reducing or preventing a risk of dangerous levels of hydrogen subject to combustion. The systems and methods of the present disclosure address this need and provide additional advantages as well. This section includes a summary of the claims in the commonly accepted definition of a comprehensive and usually brief recapitulation of the claims.

[0009] [5] In some aspects, the techniques described herein relate to a system for hydrogen gas infusion into a solution including: a hydrogen gas source for producing hydrogen gas; a fluid reservoir containing a medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate the hydrogen gas with the medical solution to produce a hydrogen-infused solution with a hydrogen gas concentration of about 0.25 - 2.5 mM; and a gas dilution module in fluid communication with and downstream of the fluid reservoir; wherein the gas dilution module is configured to continuously receive and dilute excess hydrogen gas from the fluid reservoir without allowing the excess hydrogen gas to reach a combustible concentration.

[0010] [6] In some aspects, the techniques described herein relate to a system, further including a gas exchanger in fluid communication with at least the hydrogen gas source and the fluid reservoir, wherein the gas exchanger is configured to infuse the hydrogen gas into the medical solution to equilibrium.

[0011] [7] In some aspects, the techniques described herein relate to a system, wherein the gas dilution module is configured to discharge a gas mixture having 4.5% or less hydrogen gas by volume.

[0012] [8] In some aspects, the techniques described herein relate to a system, wherein the hydrogen gas source includes an electrolysis cell configured to produce hydrogen gas and oxygen gas.

[0013] [9] In some aspects, the techniques described herein relate to a system, wherein the hydrogen gas is dissolved within the hydrogen-infused solution.

[0014]

[0010] In some aspects, the techniques described herein relate to a system, wherein the hydrogen gas in the hydrogen-infused solution is not in a gaseous phase.

[0015]

[0011] In some aspects, the techniques described herein relate to a system, wherein the medical solution includes saline, organ preservation solution, whole blood, partial blood, cardioplegia solution, drugs, anticoagulants, antimicrobials, Prolyl hydroxylase domain (PHD) inhibitors, valproic acid, impermeants, thrombolytics, or a combination thereof.

[0016]

[0012] In some aspects, the techniques described herein relate to a system, wherein the gas dilution module includes a static gas dilution module, the static gas dilution module including a dilution chamber that is preloaded with a volume of diluent gas.

[0017]

[0013] In some aspects, the techniques described herein relate to a system, wherein the volume of diluent gas is of sufficient quantity to render a final gas mixture nonflammable.

[0018]

[0014] In some aspects, the techniques described herein relate to a system, wherein the volume of diluent gas is at least 10-fold higher than a total amount of the excess hydrogen gas.

[0019]

[0015] In some aspects, the techniques described herein relate to a system, wherein the dilution chamber is surrounded by a gas-permeable material configured to allow for slow diffusion of the excess hydrogen gas to an external environment over a predetermined period of time.

[0020]

[0016] In some aspects, the techniques described herein relate to a system, further including a safety shutoff circuit coupled to at least the hydrogen gas source and the dilution chamber, wherein the safety shutoff circuit is configured to shut off or limit a rate of production of the hydrogen gas from the hydrogen gas source based at least on a volume of diluent gas in the dilution chamber.

[0017] In some aspects, the techniques described herein relate to a system, wherein the gas dilution module includes a dynamic gas dilution module including a dilution chamber in fluid communication with the fluid reservoir and a dynamic flow source, wherein the dilution chamber is configured to receive the excess hydrogen gas from the fluid reservoir.

[0021]

[0018] In some aspects, the techniques described herein relate to a system, wherein the dynamic flow source includes a mechanism for conveying a flowing diluent gas into the dilution chamber to mix with the excess hydrogen gas at a ratio of at least 10: 1, wherein the flowing diluent gas includes one or more of nitrogen, carbon dioxide, or ambient air.

[0022]

[0019] In some aspects, the techniques described herein relate to a system, further including a safety shutoff circuit coupled to at least the hydrogen gas source and the dilution chamber, wherein the safety shutoff circuit is configured to shut off or limit a rate of production of the hydrogen gas from the hydrogen gas source based at least on whether the dynamic flow source is diffusing flowing diluent gas at a flow rate at least 10-fold higher than the rate of production of the hydrogen gas.

[0023]

[0020] In some aspects, the techniques described herein relate to a system for hydrogen gas infusion into a medical solution including: a hydrogen gas source; a fluid reservoir containing a medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate hydrogen gas with the medical solution to produce a hydrogen-infused solution with a hydrogen gas concentration of about 0.25 - 2.5 mM; and a catalytic combustion module in fluid communication with and downstream of the fluid reservoir; wherein the catalytic combustion module is configured to continuously receive and convert excess hydrogen gas from the fluid reservoir into water without allowing the excess hydrogen gas to reach a combustible concentration outside of the catalytic combustion module or in a gas stream discharged to surrounding environment.

[0024]

[0021] In some aspects, the techniques described herein relate to a system, wherein the catalytic combustion module includes an inlet stream of oxygen gas or air, a catalyst for promoting recombination of the excess hydrogen gas with the oxygen gas or air to produce water vapor, and one or more outlet for discharging a waste mixture of one or more of the excess hydrogen gas, the oxygen gas or air, and the water vapor.

[0025]

[0022] In some aspects, the techniques described herein relate to a system, wherein the catalyst includes a platinum group metal.

[0026]

[0023] In some aspects, the techniques described herein relate to a system, wherein the inlet stream is in fluidic communication with the hydrogen gas source.

[0024] In some aspects, the techniques described herein relate to a system, further including a safety shutoff circuit coupled to at least the hydrogen gas source and the catalytic combustion module, wherein the safety shutoff circuit is configured to shut off or limit a rate of production of the hydrogen gas from the hydrogen gas source based at least on whether a concentration of the hydrogen gas in the waste mixture is less than 2% by volume.

[0027]

[0025] In some aspects, the techniques described herein relate to a system for hydrogen gas infusion into a medical solution including: a hydrogen gas source configured to produce or dispense hydrogen gas; and a degassed medical solution, the degassed medical solution being in fluid communication with the hydrogen gas source so as to receive and dissolve the hydrogen gas into the degassed medical solution to produce a hydrogen-infused solution at equilibrium hydrogen partial pressures ranging from about 230 mm Hg to about 2300 mm Hg.

[0028]

[0026] In some aspects, the techniques described herein relate to a system, wherein the degassed medical solution includes partial pressures of all dissolved gases adding to a total partial pressure ranging from 0 mm Hg to 380 mm Hg.

[0029]

[0027] In some aspects, the techniques described herein relate to a system, wherein the degassed medical solution is degassed of one or more of N2, or 02 gas.

[0030]

[0028] In some aspects, the techniques described herein relate to a system, further including a degassing module for degassing the medical solution to produce the degassed medical solution, wherein the degassing module includes at least a vacuum pump, a gas exchanger, and at least one valve connected to at least the hydrogen gas source, the vacuum pump, and the gas exchanger.

[0031]

[0029] In some aspects, the techniques described herein relate to a system, wherein the at least one valve is configured to switch between a first configuration and a second configuration, wherein the first configuration includes blocking the hydrogen gas source from the at least one valve such that the vacuum pump is in fluid communication with the gas exchanger to allow for degassing of the medical solution in the gas exchanger, and wherein the second configuration includes blocking the vacuum pump from the at least one valve such that the hydrogen gas source is in fluid communication with the gas exchanger to allow for infusing the hydrogen gas into the medical solution in the gas exchanger to produce the hydrogen-infused solution.

[0032]

[0030] In some aspects, the techniques described herein relate to a system, wherein the gas exchanger includes a gas permeable membrane, a hollow-fiber gas exchanger, or a bubble gas exchanger.

[0033]

[0031] In some aspects, the techniques described herein relate to a system for hydrogen gas infusion into a medical solution including: a hydrogen gas source configured to produce or dispense hydrogen gas; and a fluid reservoir containing a degassed medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate the hydrogen gas with the degassed medical solution to produce a hydrogen-infused solution with a hydrogen gas concentration of about 0.25 - 2.5 mM.

[0034]

[0032] In some aspects, the techniques described herein relate to a system, wherein the degassed medical solution is degassed of one or more of N2, or 02 gas.

[0035]

[0033] In some aspects, the techniques described herein relate to a system, further including a degassing module for degassing the medical solution to produce the degassed medical solution, wherein the degassing module includes at least a vacuum pump, a gas exchanger, and at least one valve connected to at least the hydrogen gas source, the vacuum pump, and the gas exchanger.

[0036]

[0034] In some aspects, the techniques described herein relate to a system, wherein the at least one valve is configured to switch between a first configuration and a second configuration, wherein the first configuration includes blocking the hydrogen gas source from the at least one valve such that the vacuum pump is in fluid communication with the gas exchanger to allow for degassing of the medical solution in the gas exchanger, and wherein the second configuration includes blocking the vacuum pump from the at least one valve such that the hydrogen gas source is in fluid communication with the gas exchanger to allow for infusing the hydrogen gas into the medical solution in the gas exchanger to produce the hydrogen-infused solution.

[0037]

[0035] In some aspects, the techniques described herein relate to a system, wherein the gas exchanger includes a gas permeable membrane, a hollow-fiber gas exchanger, or a bubble gas exchanger.

[0038]

[0036] In some aspects, the techniques described herein relate to a system, wherein the degassing module further includes a circulation pump in fluid communication with at least the fluid reservoir and the gas exchanger, the circulation pump configured to apply pressure for continuously circulating the medical solution from the fluid reservoir through the gas exchanger.

[0039]

[0037] In some aspects, the techniques described herein relate to a system, wherein the degassing module further includes a second fluid reservoir, wherein a fuller of the fluid reservoir and the second fluid reservoir is elevated relative to the gas exchanger and an emptier of the fluid reservoir and the second fluid reservoir is lowered relative to the gas exchanger to allow the medical solution to flow through the gas exchanger due to gravity.

[0040]

[0038] In some aspects, the techniques described herein relate to a system, further including an ultrasonic bath, wherein the fluid reservoir is immersed in the ultrasonic bath, wherein the ultrasonic bath is allowed to apply ultrasonic energy for a desired degassing period such that gases in the medical solution are forced to migrate to an upper portion of the fluid reservoir.

[0039] In some aspects, the techniques described herein relate to a system, wherein the gases are dissolved.

[0041]

[0040] In some aspects, the techniques described herein relate to a system, wherein the desired degassing period ranges from about 1 minutes to about 25 minutes.

[0042]

[0041] In some aspects, the techniques described herein relate to a system for hydrogen gas infusion into an organ or tissue including: a fluid circuit configured to circulate a medical solution through an organ or tissue; the fluid circuit including a perfusion solution, a gas exchanger, and a source of hydrogen.

[0043]

[0042] In some aspects, the techniques described herein relate to a system, wherein the source of hydrogen gas includes a 0.1 to 5 liter volume of aqueous solution having a dissolved hydrogen gas concentration of 0.25 - 2.5 mM.

[0044]

[0043] In some aspects, the techniques described herein relate to a system, further including a waste gas dilution module in fluid communication with and downstream of the gas exchanger; wherein the gas dilution module is configured to dilute waste hydrogen gas from the gas exchanger without allowing the waste hydrogen gas to reach a combustible concentration.

[0045]

[0044] In some aspects, the techniques described herein relate to a system, further including a waste gas recycling system for recovery of at least a portion of hydrogen gas from a gas outflow of the gas exchanger and for recycling of recovered hydrogen gas into a gas inlet of the gas exchanger.

[0046]

[0045] In some aspects, the techniques described herein relate to a system, wherein the waste gas recycling system includes a carbon dioxide sorbent.

[0047]

[0046] In some aspects, the techniques described herein relate to a system, wherein the waste gas recycling system includes a pressure swing adsorption system.

[0048]

[0047] In some aspects, the techniques described herein relate to a system, wherein the waste gas recycling system includes one or more selectively gas-permeable membranes.

[0049]

[0048] In some aspects, the techniques described herein relate to a method of treating or reducing a risk of an injury due to reactive oxygen species and / or free radicals in blood, an organ or a tissue, the method including: administering a hydrogen-infused solution to the blood, organ or tissue at a concentration and a rate sufficient to reduce the reactive oxygen species and / or free radicals in the blood, organ or tissue, wherein the hydrogen-infused solution includes hydrogen gas at a concentration of about 0.25 - 2.5 mM.

[0050]

[0049] In some aspects, the techniques described herein relate to a method, further including determining that the blood, organ or tissue has or is at risk of an injury due to the reactive oxygen species and / or free radicals in the blood, organ or tissue based on an amount of the reactive oxygen species and / or free radicals in the blood, organ or tissue as compared to a normal blood, organ or tissue.

[0051]

[0050] In some aspects, the techniques described herein relate to a method, wherein the hydrogen- infused solution is obtained by a method of infusing hydrogen gas, the method including: receiving hydrogen gas and a solution into a fluid reservoir; and infusing the solution with the hydrogen gas to produce a hydrogen-infused solution and a stream of excess hydrogen gas.

[0052]

[0051] In some aspects, the techniques described herein relate to a method, further including diluting the excess hydrogen gas to a desired safe disposal concentration after the infusing.

[0053]

[0052] In some aspects, the techniques described herein relate to a method, wherein the desired safe disposal concentration is less than 4.5% by volume.

[0054]

[0053] In some aspects, the techniques described herein relate to a method, wherein the desired safe disposal concentration is less than 2% by volume.

[0055]

[0054] In some aspects, the techniques described herein relate to a method, wherein the infusing including applying a positive pressure to force the hydrogen gas into the solution through a gas exchanger.

[0056]

[0055] In some aspects, the techniques described herein relate to a method, wherein the solution is driven to flow through the gas exchanger using gravity or a circulation pump.

[0057]

[0056] In some aspects, the techniques described herein relate to a method, further including a recovery of hydrogen gas from a gas outflow of the gas exchanger and recirculation of the recovered hydrogen gas to a gas inlet of the gas exchanger.

[0058]

[0057] In some aspects, the techniques described herein relate to a method, wherein the diluting includes conveying the stream of the excess hydrogen gas to a static gas dilution module, the static gas dilution module including a dilution chamber that is preloaded with a volume of diluent gas.

[0059]

[0058] In some aspects, the techniques described herein relate to a method, wherein the volume of diluent gas is of sufficient quantity to render a final gas mixture nonflammable.

[0060]

[0059] In some aspects, the techniques described herein relate to a method, wherein the volume of diluent gas is at least 10-fold higher than a total amount of the excess hydrogen gas.

[0061]

[0060] In some aspects, the techniques described herein relate to a method, wherein the diluting includes conveying the stream of the excess hydrogen gas to a dynamic gas dilution module, the dynamic gas dilution module including a dilution chamber in fluid communication with the fluid reservoir and a dynamic flow source, wherein the dilution chamber is configured to receive the excess hydrogen gas from the fluid reservoir.

[0062]

[0061] In some aspects, the techniques described herein relate to a method, further including degassing the solution prior to the infusing, wherein the degassing includes flowing the solution through a gas exchanger and applying a negative pressure to the gas exchanger to extract gases from the solution.

[0063]

[0062] In some aspects, the techniques described herein relate to a method, wherein the hydrogen gas is generated by an electrolysis reaction, wherein the electrolysis reaction includes separating water into hydrogen and oxygen gases.

[0064]

[0063] In some aspects, the techniques described herein relate to a method, wherein the hydrogen- infused solution is administered to the blood, organ, or tissue by injecting the hydrogen-infused solution into a perfusion circuit of an ex-vivo organ perfusion system.

[0065]

[0064] In some aspects, the techniques described herein relate to a method, in which a flow of gas through a gas exchanger of the ex-vivo organ perfusion system is arrested prior to and during injection of the hydrogen-infused solution into a perfusate flow path.

[0066]

[0065] In some aspects, the techniques described herein relate to a method of treating or reducing a risk of an injury due to reactive oxygen species and / or free radicals in blood, an organ or a tissue prior to administering oxygen gas to the blood, organ or tissue, the method including: administering a hydrogen-infused solution to the blood, organ or tissue at a concentration and a rate sufficient to reduce the reactive oxygen species and / or free radicals in the blood, organ or tissue; and subsequently contacting the blood, organ or tissue with the oxygen gas so as to reduce damage to cells in the blood, organ or tissue.

[0067]

[0066] In some aspects, the techniques described herein relate to a method, wherein the hydrogen- infused solution includes hydrogen gas at a concentration of about 0.25 - 2.5 mM.

[0068]

[0067] In some aspects, the techniques described herein relate to a method, wherein the injury is related to ischemia or hypoxia.

[0069]

[0068] In some aspects, the techniques described herein relate to a method, further including determining that the blood, organ or tissue has or is at risk of an injury due to the reactive oxygen species and / or free radicals in the blood, organ or tissue based on an amount of the reactive oxygen species and / or free radicals in the blood, organ or tissue as compared to a normal blood, organ or tissue.

[0070]

[0069] In some aspects, the techniques described herein relate to a method, wherein the hydrogen- infused solution is obtained by a method of infusing hydrogen gas, the method including: receiving hydrogen gas and a solution into a fluid reservoir; and infusing the solution with the hydrogen gas to produce a hydrogen-infused solution and a stream of excess hydrogen gas.

[0071]

[0070] In some aspects, the techniques described herein relate to a method, further including diluting the excess hydrogen gas to a desired safe disposal concentration after the infusing.

[0071] In some aspects, the techniques described herein relate to a method, wherein the desired safe disposal concentration is less than 4.5% by volume.

[0072]

[0072] In some aspects, the techniques described herein relate to a method, wherein the desired safe disposal concentration is less than 2% by volume.

[0073]

[0073] In some aspects, the techniques described herein relate to a method, wherein the diluting includes conveying the stream of the excess hydrogen gas to a static gas dilution module, the static gas dilution module including a dilution chamber that is preloaded with a volume of diluent gas.

[0074]

[0074] In some aspects, the techniques described herein relate to a method, wherein the volume of diluent gas is of sufficient quantity to render a final gas mixture nonflammable.

[0075]

[0075] In some aspects, the techniques described herein relate to a method, wherein the volume of diluent gas is at least 10-fold higher than a total amount of the excess hydrogen gas.

[0076]

[0076] In some aspects, the techniques described herein relate to a method, wherein the diluting includes conveying the stream of the excess hydrogen gas to a dynamic gas dilution module, the dynamic gas dilution module including a dilution chamber in fluid communication with the fluid reservoir and a dynamic flow source, wherein the dilution chamber is configured to receive the excess hydrogen gas from the fluid reservoir.

[0077]

[0077] In some aspects, the techniques described herein relate to a method, wherein the infusing including applying a positive pressure to force the hydrogen gas into the solution through a gas exchanger.

[0078]

[0078] In some aspects, the techniques described herein relate to a method, wherein the solution is driven to flow through the gas exchanger using gravity or a circulation pump.

[0079]

[0079] In some aspects, the techniques described herein relate to a method, further including degassing the solution prior to the infusing, wherein the degassing includes flowing the solution through a gas exchanger and applying a negative pressure to the gas exchanger to extract gases from the solution.

[0080]

[0080] In some aspects, the techniques described herein relate to a method, wherein the hydrogen gas is generated by an electrolysis reaction, wherein the electrolysis reaction includes separating water into hydrogen and oxygen gas using electric current.

[0081] BRIEF DESCRIPTION OF THE DRAWINGS

[0082]

[0081] The novel features of the disclosure are set forth with particularity in the appended claims. The patent or application file contains at least one drawing executed in color. Copies of this patent or patent application publication with color drawing(s) will be provided by the Office upon request and payment of the necessary fee. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the disclosure are utilized, and the accompanying drawings of which:

[0083]

[0082] FIGURES 1A-1C illustrate exemplary embodiments of a hydrogen gas infusion (HI) system with a static dilution module, and FIGURE ID illustrates a process for infusing hydrogen gas into a medical solution using an HI system according to exemplary embodiments of this disclosure.

[0084]

[0083] FIGURES 2A-2B illustrate embodiments of an HI system with a dynamic dilution module according to exemplary embodiments of this disclosure.

[0085]

[0084] FIGURES 3A-3B illustrate exemplary embodiments of an HI system with a catalytic combustion module, according to exemplary embodiments of this disclosure.

[0086]

[0085] FIGURES 4A-4B illustrate exemplary embodiments of an HI system with a degassing module using a circulation pump, and FIGURE 4C illustrates a process for degassing and infusing hydrogen gas into a medical solution using an HI system according to exemplary embodiments of this disclosure.

[0087]

[0086] FIGURES 5A-5B illustrate exemplary embodiments of an HI system with a degassing module using a gravity feed. FIGURES 5C-5D illustrate exemplary embodiments of an HI system with a degassing module using external pressure on the solution reservoirs, and FIGURE 5E illustrates a process for degassing and infusing hydrogen gas into a medical solution using an HI system according to exemplary embodiments of this disclosure.

[0088]

[0087] FIGURES 6A-6B illustrate exemplary embodiments of an HI system with an ultrasound degassing module, FIGURE 6C illustrates a process for degassing and infusing hydrogen gas into a medical solution using an HI system according to exemplary embodiments of this disclosure, and FIGURE 6D illustrates a process for infusion of hydrogen into degassed medical solution so as to be packaged for later administration or to be directly administered to an organ or tissue.

[0089]

[0088] FIGURE 7 is a block diagram of a system with which some embodiments may operate.

[0090]

[0089] FIGURE 8 illustrates is a block diagram of a computing device with which some embodiments may operate.

[0091]

[0090] FIGURES 9A-9C illustrate an HI system with reusable modules, according to exemplary embodiments of this disclosure.

[0092]

[0091] FIGURE 10A illustrates a typical process for organ recovery, flushing, transportation, and transplantation using static cold storage. FIGURE 10B depicts an exemplary process in which the flushing solution is infused with hydrogen using the HI system prior to flushing the organ. FIGURE IOC depicts an exemplary process in which the hydrogen-infused flush is administered to the organ after the static cold storage interval.

[0093]

[0092] FIGURES 11A-11H illustrate exemplary embodiments of HI systems and methods adapted to ex-vivo organ perfusion systems and methods. FIGURE HA illustrates the essential elements of a non-oxygenating ex-vivo perfusion system. FIGURE 11B depicts an exemplary process 4000 of steps in which a hydrogen-infused solution can be flushed through the organ after ex-vivo perfusion, prior to reperfusion with the patient’s oxygenated blood. FIG. 11C depicts a typical ex-vivo oxygenated perfusion system, in which a gas exchanger and an oxygen source are included in the perfusion circuit. FIGURES 11D-11E depict an embodiment and process, respectively, in which a hydrogen-rich flush solution is administered to the organ after cessation of gas flow through the gas exchanger and prior to removing the organ from the perfusion system. FIGURES 11F-11G depict an embodiment and process, respectively, for safely administering dissolved hydrogen to the organ in the perfusion system by means of a gas exchanger. FIGURE 11H depicts an embodiment of an ex-vivo perfusion system in which the waste gases from a gas exchanger are separated by type and some constituents of the waste gas are recirculated through the gas exchanger to minimize production or emission of hydrogen waste gas.

[0094]

[0093] FIGURES 12A-12B describe an exemplary embodiment and process, respectively, for administration of hydrogen during recovery of a donated heart after a prolonged period of warm ischemia.

[0095]

[0094] While the above-identified drawings set forth presently disclosed embodiments, other embodiments are also contemplated, as noted in the discussion. This disclosure presents illustrative embodiments by way of representation and not limitation. Numerous other modifications and embodiments can be devised by those skilled in the art which fall within the scope and spirit of the principles of the presently disclosed embodiments.

[0096] DETAILED DESCRIPTION

[0097]

[0095] Described herein are systems and techniques for hydrogen gas infusion into a solution with increased concentrations and efficiency relative to those previously available. This disclosure relates generally to hydrogen gas infusion (HI) systems such that clinically-relevant concentrations of hydrogen are safely infused into solutions and delivered to patients to reduce the occurrence of oxidative stress and subsequent dysfunction or death of cells and / or tissues, as well as cell apoptosis and tissue inflammation. In particular, the HI systems address the need for efficient control over levels of dissolved oxygen, as well as the lack of efficient, low-cost methods for treatments of high-concentration hydrogen gas. Further, the HI systems provide for homogenous delivery of hydrogen with precision and within safe concentrations rather than existing methods that provide heterogeneous delivery to treated areas. A key distinction of the HI system is its ability to eliminate the hydrogen gas flammability hazard by both minimizing the volume of hydrogen gas needed for infusion, and neutralizing any hydrogen-containing waste gases produced by the infusion process.

[0098]

[0096] After a period of ischemia, anoxia, or hypoxia, live organs or tissue can be subject to damage by reperfusion injury when blood supply returns. The absence of oxygen and other nutrients from the tissue during the period of ischemia can create conditions where restoration of circulation can result in inflammation and oxidative damage due, in part, to reactive oxygen species that come into existence during reperfusion. This can occur through induction of oxidative stress instead of, or in addition to, restoration to normal tissue function.

[0099]

[0097] Analogous to reperfusion injury in organs and tissues, reoxygenation injury is reported for live cells harvested or isolated from tissues and organs, such as adipocytes from fat, hepatocytes or Kupffer cells from liver, or islet cells from the pancreas. The harvested or isolated cells are exposed to temporary period of hypoxia during the isolation or harvesting process. Reintroduction of oxygen by subsequent processing or grafting can trigger oxidative stress and subsequent dysfunction or death of the harvested or isolated cells.

[0100]

[0098] Molecular hydrogen gas, also called “diatomic hydrogen” or simply H2, has antioxidant properties for treatment or prevention of injury from reperfusion (in the case of organs or vascular tissue) or reoxygenation (in the case of harvested or isolated cells). Hydrogen gas may be administered through multiple methods, such as through inhalation or administration via dissolution of hydrogen gas into an aqueous, therapeutic or medical solution such as saline or organ preservation solution. Hydrogen-saturated solutions, for example, may be administered for treatment of a target tissue, cells, or organs by immersion, suspension, or perfusion.

[0101]

[0099] The inventors have recognized and appreciated various unmet needs in the pursuit of high- concentration infusion of hydrogen gas into medical solutions. For example, administration of high concentrations of molecular hydrogen via hydrogen-saturated aqueous solutions or blood in medical clinics for human treatment has been limited by the explosive danger posed by hydrogen gas. Hydrogen / air or hydrogen / oxygen gas mixtures are flammable and in some cases explosive for hydrogen concentrations exceeding 4.5% hydrogen. This has resulted primarily in methods that implement dilution of high concentrations of hydrogen upstream of infusion into aqueous solutions, thereby avoiding dangerous levels of hydrogen that are subject to combustion. Consequently, although aqueous solutions with low concentrations of dissolved hydrogen are easy to produce, aqueous solutions having very high concentrations hydrogen concentrations at or near the saturation level are difficult to produce in a healthcare setting without also producing a dangerous quantity of flammable or explosive gas. Further, administration of aqueous solutions with low concentrations of hydrogen have been known to result in heterogeneous microperfusion, in that not all targets, e.g., tissues or organs, receive the same amount of blood flow and thus hydrogen levels. In particular, to achieve a functional benefit in the impacted area, a minimum dose of hydrogen may be required, though remains unmet as a result of local underperfusion. This may lead to under delivery of hydrogen-saturated aqueous solutions. Timing of administering hydrogen gas to a patient has also been shown to be crucially important, as well as the duration of the hydrogen gas to be acting on the target, thus requiring high concentrations of hydrogen gas to be accessible on demand.

[0102]

[0100] The inventors have recognized and appreciated methods and systems thereof for administering hydrogen gas at such high concentrations via dissolution of hydrogen gas into aqueous, therapeutic or medical solutions, such as saline or organ preservation solution, for administration to target tissue, cells, or organs by immersion, suspension, or perfusion. The inventors have also recognized and appreciated, through experimentation, system parameters required for achieving optimal localized distribution of the high concentration hydrogen gas for effective durations that preserve target tissues or organs and result in desired outcomes.

[0103]

[0101] For clinical applications dominated by acute oxidative stress, it is clinically advantageous to maximize the hydrogen concentration in therapeutic or medical solutions beyond those levels achievable using the traditional methods that dilute the hydrogen gas prior to infusion into the medical solution. In addition, clinical preferences and logistical constraints favor the on-demand preparation of freshly hydrogen-saturated solutions as opposed to using medical solutions prepared and stored for future emergent need. The HI systems disclosed herein may provide a uniquely safe and convenient method to quickly infuse any aqueous therapeutic liquid with high hydrogen concentrations that are unachievable, unsafe, or inconvenient to produce using previous methods. Further, the HI systems of the present disclosure may provide a safe means to virtually eliminate dissolved oxygen from the hydrogen-saturated therapeutic or medical solution which can further increase effectiveness in preventing reperfusion injury.

[0104]

[0102] Provided herein are hydrogen gas infusion systems with (1) excess gas dilution modules; (2) catalytic combustion modules; (3) degassing modules; and (4) systems and methods of computer implementation.

[0105] I. Excess gas dilution Modules

[0106]

[0103] In various aspects, this disclosure provides hydrogen gas infusion (HI) systems that may be used for delivering high concentrations of dissolved hydrogen gas into medical solutions. The HI systems are configured to dilute waste hydrogen gas streams to prevent discharge of mixtures of hydrogen and air or hydrogen and oxygen to the clinical environment may cause risks of explosion hazards. FIGS. 1 A- 7 illustrate HI systems according to exemplary embodiments of this disclosure.

[0107]

[0104] As illustrated in FIG. 1A, an HI system 100 may comprise at least a hydrogen gas source 101, at least one fluid reservoir 102 downstream of the hydrogen gas source 101, and a gas dilution module 103 downstream of the at least one fluid reservoir 102. The hydrogen gas source 101 may provide hydrogen gas that is to be infused into a desired medical solution to produce a hydrogen- infused solution.

[0108]

[0105] In some embodiments, the hydrogen gas source 101 may comprise an electrolysis cell, where electric current splits water into oxygen as at the anode and hydrogen gas at the cathode. Production of hydrogen gas is initiated upon application of an electric current, wherein a tube carries the hydrogen gas from the cathode to the desired components of the HI system. In some embodiments, the hydrogen is channeled toward the gas infusion components of the device. In some embodiments, the oxygen gas is vented to the ambient environment. In some embodiments, the oxygen is directed to recombine with hydrogen waste gas in the catalytic combustion module, described in more detail below.

[0109]

[0106] In some embodiments, the HI system 100 further comprises a gas exchange module between the hydrogen gas source 101 and the at least one fluid reservoir 102 so as to infuse the medical solution with the hydrogen gas. In some embodiments, the gas exchange module comprises a gas permeable membrane, a hollow-fiber gas exchanger, a bubble gas exchanger, a thin film gas exchanger, or a combination thereof. In some embodiments, the gas permeable membrane may be, for example, silicone or polysulfone, as in a standard hollow-fiber membrane oxygenator design. In some embodiments, the gas exchange module comprises a liquid channel and a gas channel that are separated by the gas permeable membrane. The liquid channel may be coupled to the fluid reservoir 102 via an additional lumen, and an inlet of the gas channel may be coupled to the hydrogen gas source 101 via a first lumen 104. The configuration of the gas channel and liquid channel allow for unwanted gases like nitrogen and oxygen to (a) be extracted from the solution flowing through the liquid channel, resulting in degassed solution, and / or (b) hydrogen gas diffusion through the gas permeable membrane into the solution flowing through the liquid channel, resulting in a hydrogen-infused solution. In some embodiments, the hydrogen gas continues to diffuse through the gas permeable membrane until equilibrium or saturation within the solution has been reached. In some embodiments, the hydrogen gas is dissolved in an aqueous phase within the solution. In some embodiments, the dissolved hydrogen gas may reach concentrations as high as 1600 ppb at normal atmospheric pressure. Simultaneously, hydrogen gas that is not diffused through the gas permeable membrane may exit the gas exchange module as excess or waste hydrogen gas.

[0110]

[0107] In some embodiments, the volume of the solution is within a range of about 0.1 to about 3 liters. In some embodiments, the hydrogen gas is allowed to equilibrate until the hydrogen reaches a desired concentration, i.e., a clinically relevant concentration. In some embodiments, the desired concentration comprises about 0.25 - 2.5 mM. In some embodiments, the hydrogen gas is dissolved into the solution in the reservoir. In some embodiments, any oxygen gas that remains dissolved in the solution is controlled within a range of about 0 to about 250 mm Hg. In some embodiments, the temperature of the solution in the fluid reservoir 102 is maintained within a range of about 4°C to about 37°C. In some embodiments, the plurality of ports may further include a first outlet port that is proximally in fluid communication with the gas dilution module 103 via a second lumen 105. In some embodiments, the gas dilution module 103 may comprise an inlet port distally coupled to the second lumen 105. As the solution in the reservoir saturates with hydrogen gas, the first outlet port conveys hydrogen-containing waste gas from the reservoir safely to the inlet port of the gas dilution module 103 without exposure to any oxygen or air. In some embodiments, any number of lumens in the HI system may include at least one valve, such that the at least one valve may be controllable to be opened or closed to cut off fluid communication therein. In some embodiments, the at least one valve may be controllable manually or via automated methods or systems.

[0111]

[0108] Generally, the waste hydrogen gas may be a hydrogen-rich gas stream that may be at risk of mixing with oxygen or air to cause an explosion hazard. However, the dilution of the waste hydrogen gas immediately after infusion with the aqueous solution may allow for prevention of fire / explosion by the dilution of the effluent hydrogen gas. Further, this sequence eliminates an undesirable limitation on hydrogen concentrations within the solution, especially compared to previous methods that dilute the hydrogen gas prior to infusion into the solution. In some embodiments, the gas dilution module 103 comprises at least a dilution chamber for diluting the waste hydrogen with a diluent gas 106. In some embodiments, the diluent gas 106 may comprise one or more of nitrogen, carbon dioxide, or ambient air. In some embodiments, the gas dilution module 103 comprises a chamber with gas-permeable sides or vents to allow the diluted gas mixture to slowly diffuse out through the sides or vents into the atmosphere. In some embodiments, the gas dilution module 103 comprises a grounded and electrically conductive silicone reservoir configured to slowly release the diluted waste gas to the atmosphere by diffusion. In some embodiments, the gas dilution module 103 comprises a pump for conveying diluted waste hydrogen gas from the dilution chamber into the atmosphere.

[0112]

[0109] In reference to FIG. 1A, in some embodiments, the gas dilution module 103 is a static dilution module in which the gas flows through the solution while the solution remains in a single container. In some embodiments, the static gas dilution module comprises a dilution chamber that is preloaded with a volume of diluent gas 106 that is at least 10- to 100-fold higher than a total amount of hydrogen gas produced from the hydrogen gas source 101, such that the diluent gas 106 and the waste hydrogen gas are mixed at a ratio of at least 10: 1, or optionally at least 25:1 (4% waste hydrogen gas). In some embodiments, the dilution chamber is surrounded by a gas- permeable material, such as, for example, silicone, that may allow for slow diffusion of the waste hydrogen gas to an external environment over an extended period of time. This slow diffusion allows for dilution of the waste hydrogen gas from inside the dilution chamber. In some embodiments, the preloaded volume of diluent gas 106 may comprise one or more of nitrogen, carbon dioxide, or ambient air. In some embodiments, the second lumen 105 is in a second tube separate from the first tube. In some embodiments, the first tube includes multiple lumens, such as at least the first lumen 104 and the second lumen 105. In some embodiments, the first lumen 104 is longer than the second lumen 105. In particular, the use of a longer lumen releases inflowing hydrogen gas at the bottom of the solution bag so that a resulting bubble path is longer as it travels towards the top of the bag. This facilitates further gas mixing between the bubbles from the hydrogen gas and the solution for improving gas exchange efficiency. Since the hydrogen gas collects at the top of the bag and has to be either recirculated as an intermediate step or disposed as a final step, the shorter lumen penetrates into the gas pocket at the top of the bag so hydrogen gas is sucked up instead of any liquid. The use of two separate lumens using two separate ports are conveniently simple. In some embodiments, the first lumen 104 and second lumen 105 may be combined into a single port penetrator, requiring the use of a single bag port.

[0113] [HO] In some embodiments, the plurality of ports in the solution container may further include a second outlet port for conveying the saturated solution to be administered to the patient, organ, tissue, or cells. In some embodiments, a medical infusion line may be connected to the second outlet port, such that resulting hydrogen-infused solution may flow from the fluid reservoir 102 into the medical infusion line for downstream administration to the patient, organ, tissue, or cells. In some embodiments, the solution can be a fluid used to treat or maintain a patient during surgery, to maintain tissue requiring perfusion, or some other solution. In some embodiments, the solution can be, for example, saline, organ preservation solution, whole blood, partial blood, cardioplegia solution or an aqueous solution with other chemical components, such as drugs or tissue stabilizers for the designated use. In some cases, whole blood, partial blood, or mixtures of blood components with other solutions can be used. In some embodiments, the medical solution comprises anticoagulants, antimicrobials, Prolyl hydroxylase domain (PHD) inhibitors, valproic acid, impermeants, thrombolytics, or a combination thereof

[0114] [Hl] In reference to FIG. IB, in some embodiments, the HI system further comprises a safety shutoff circuit 107. The safety shutoff circuit 107 may be coupled to at least the hydrogen gas source 10 and the gas dilution module 103. In particular, the safety shutoff circuit 107 may be configured to shut off or limit the rate of production of hydrogen gas from the hydrogen gas source 10 based at least on a volume of diluent gas 106 in the dilution chamber. In some embodiments, hydrogen production is shut off by cessation of the electrolysis current. In some embodiments, the hydrogen production rate is controlled by the amperage of electric current through the electrolysis cell, wherein the hydrogen production rate is related to the electrolysis current according to the following relation: H2 production (cubic centimeters per second) = 418 cc of hydrogen per amp- hour of electrolysis current. The volume of diluent gas 106 in the dilution chamber may be within a predetermined volume range so as to ensure adequate dilution of a predetermined quantity of hydrogen gas. The safety circuit may include a sensor to measure the quantity or pressure of diluent gas 106 and function to prevent the production of hydrogen gas beyond the volume that can be safely diluted into the measured volume of diluent gas 106.

[0115]

[0112] In reference to FIG. 1C, in some embodiments, the gas dilution module 103 may comprise a bag or container made at least partly from a gas permeable material. In some embodiments, the gas permeable material comprises silicone. In some embodiments the gas permeable material or other parts of the gas dilution module 103 may comprise electrically conductive materials to prevent electrostatic discharge. In some embodiments, the gas dilution module 103 may be electrically grounded. After filling the container with diluent gas 106 and waste hydrogen gas, the gases can be allowed to slowly diffuse through the gas-permeable membrane to dissipate into the surrounding air at hydrogen concentrations below 4.5%. In some embodiments the gas dilution module 103 may comprise a diluent gas source 108 to pre-fill the gas dilution module 103 with diluent gas 106. In some embodiments, the diluent gas source 108 may comprise a compressed gas container. In other embodiments, the diluent gas source 108 may comprise a nitrogen gas concentrator which concentrates nitrogen from the air by selective separation of oxygen from nitrogen by a gas permeable membrane. In some embodiments, the diluent gas source 108 may comprise a nitrogen concentrator which concentrates nitrogen from the air using the pressureswing adsorption method. In other embodiments, the diluent gas source 108 may comprise a chemical reaction to produce a fixed volume of carbon dioxide for dilution.

[0113] In some embodiments, as illustrated in FIG. ID, hydrogen gas infusion using the HI system may be performing according to process 130. In step 131, electric current is applied to the electrolysis cell to initiate an electrolysis reaction to generate hydrogen gas. In step 132, the generated hydrogen gas is allowed to be conveyed in a desired volume or flow rate to the fluid reservoir 102, such that the desired volume results in a desired concentration of hydrogen gas in the fluid reservoir 102. In step 133, hydrogen gas is allowed to infuse into the solution to produce hydrogen gas-infused solution and a stream of waste hydrogen gas. This may be done through agitation of the fluid, recirculation of the gas to make multiple passes through the fluid as in a bubbler, or using single or multiple passes through a gas exchange module to infuse hydrogen gas from a separate gas line into a fluid line containing the solution. In some embodiments, the gas exchange module may further comprise a vacuum to apply negative pressure for degassing the solution prior to infusing the solution with the hydrogen gas. In step 134, any waste hydrogen gas that does not infuse into the solution is allowed to flow out of the first outlet of the fluid reservoir 102 and into an inlet port of the gas dilution module 103. The waste hydrogen gas is then diluted further in the gas dilution module 103 to a desired safe disposal concentration. In some embodiments, the desired safe disposal concentration of waste hydrogen gas is less than 4.5% by volume, optionally wherein the desired safe disposal concentration of waste hydrogen gas is less than 2% by volume. In step 135, the hydrogen gas-infused solution is administered for treatment of a target patient, organ, tissue, or cells.

[0116]

[0114] In reference to FIG. 2A, in some embodiments, the gas dilution module comprises a dynamic dilution module 203. In some embodiments, the dynamic gas dilution module 203 comprises a dilution chamber 203 A in fluid communication with a second lumen 205 from a fluid reservoir 202 and a dynamic flow source 203B. The fluid reservoir 202 may be in fluid communication with a hydrogen gas source 201 via a first lumen 204. The dynamic flow source 203B may comprise a compressed gas container, a blower, a compressor, a pump, a venturi, a nitrogen concentrator, a gas-producing chemical reaction, or other mechanism for conveying a flowing diluent gas into the dilution chamber 203 A. In some embodiments, the flowing diluent gas comprises one or more of nitrogen, carbon dioxide, water vapor, or ambient air. Waste hydrogen gas from the fluid reservoir 202 flows into the dilution chamber 203A via the second lumen 205, and the dynamic flow source 203B forces the flowing diluent gas into the dilution chamber 203 A to mix with the waste hydrogen gas. In some embodiments, the dilution chamber 203A comprises shear-inducing or turbulence-inducing elements to facilitate mixing of the hydrogen gas with the dilution gas to produce a diluted mixture 206. In some embodiments, the flowing diluent gas and the waste hydrogen gas are mixed at a ratio of at least 10: 1, or optionally at least 25: 1. In some embodiments, the dilution chamber 203 A comprises an outlet for releasing the diluted mixture 206. In some embodiments, the diluted mixture 206 comprises a hydrogen concentration of less than 4.5% by volume, optionally wherein the hydrogen concentration is less than 2% by volume. Hydrogen concentrations ranging from 0% to 4.5% are below the combustion limits and are considered safe for discharge to ambient room air.

[0117]

[0115] In reference to FIG. 2B, in some embodiments, the HI system 200 comprises a safety shutoff circuit 207 coupled to at least the hydrogen gas source 201 and the dynamic dilution module 203. The safety shutoff circuit 207 may be coupled to at least the hydrogen gas source 201 and the dynamic flow source 203B. The safety shutoff circuit 207 may be configured to shut off or limit the rate of production of hydrogen gas from the hydrogen gas source 201 based at least on whether the dynamic flow source 203B is operating and providing diluent gas to the dilution chamber 203 A at a flow rate at least 10-fold higher than the rate of production of the hydrogen gas.

[0118] II. Catalytic Combustion Modules

[0119]

[0116] In some embodiments, the HI systems of the present disclosure may comprise a catalytic combustion module. FIGS. 3A-3B illustrate such an HI system 300 according to exemplary embodiments. Specifically, the HI system 300 may comprise at least a hydrogen gas source 301, a fluid reservoir downstream of the hydrogen gas source 301, and a catalytic combustion module 303 downstream of both of the hydrogen gas source 301 and the fluid reservoir 302. In particular, a waste hydrogen gas stream from the fluid reservoir 302 may be subject to risk of combustion, and may produce a hot hydrogen combustion flame if there is reactivity with any local oxygen. A catalytic combustion module 303 may be incorporated to convert the waste hydrogen gas into harmless water vapor at room temperature or alternatively up to 150 °C by combining with a stream of oxygen gas or air within the controlled environment of the catalytic combustion chamber.

[0120]

[0117] The fluid reservoir 302 may operate similar to those illustrated in FIGS. 1-2, in that the hydrogen gas source 301 may comprise a reaction chamber with an outlet port that is in fluid communication with a fluid reservoir via a first lumen 304, and that the reaction chamber hosts an electrolysis reaction that breaks down water into at least hydrogen gas and oxygen gas. Here, however, the hydrogen gas source 301 may further comprise an oxygen outlet port in fluid communication with the catalytic combustion module via a third lumen 306 such that oxygen is conveyed from the electrolytic cell to the catalytic combustion module 303. The catalytic combustion module 303 may be in fluid communication with the fluid reservoir 302 via a second lumen 305, thereby receiving waste hydrogen gas from the fluid reservoir 302. The catalytic combustion module 303 may include a catalytic reaction chamber, the catalytic reaction chamber comprising a catalyst and operate to mix the waste hydrogen gas incoming via the second lumen 305 with the oxygen gas incoming via the third lumen 306. The catalyst may serve to promote the recombination of the hydrogen gas with the oxygen gas to produce water vapor at room temperatures or other sufficiently safe combustion temperatures such as 100 - 150 °C. In some embodiments, the recombination is performed at room temperature up to 500 °C. For hotter temperatures, a heat sink, water bath, or equivalent is needed to keep the temperature of any exhaust or released residual gases or vapors below about 100 ° for their release into the environment. In some embodiments, the catalyst comprises a platinum group metal. In some embodiments, the platinum group metal comprises one or more of a Pd-based metal alloy or a metal oxide such as Pd, Pt, Ag, Ru, Co, Ni, Mn, Cu, or Fe. In some embodiments, the catalytic combustion module 303 further comprises a heat sink. In some embodiments, the catalytic converter further comprises a warming element to initiate the catalytic combustion process. Similar to the gas dilution modules 103 or 203, the catalytic combustion module 303 further comprises one or more outlets for diffusing a waste mixture 307 of any excess gas, i.e., any additional waste hydrogen and / or any waste oxygen, as well as any waste water vapor. The outlet may be configured to diffuse the waste mixture 307 having hydrogen residues less than 4.5% by volume, optionally wherein the hydrogen residues are less than 2% by volume. In some embodiments, the waste mixture 307 further comprises diluent gas that is preloaded into the catalytic combustion module 303 or flowed in from a dynamic flow source. In some embodiments, the diluent gas or the flowing diluent gas may comprise one or more of nitrogen, carbon dioxide, or ambient air. In some embodiments a diluent gas may be used to regulate the temperature of the catalytic reaction chamber.

[0121]

[0118] In some embodiments, the third lumen 306 may be connected to a separate oxygen source rather than the hydrogen gas source. In some embodiments, instead of a third lumen 306, the catalytic combustion module 303 may instead comprise a vent or a port for receiving a stream of air from the external environment. The air may be conveyed to the catalytic reaction chamber so that oxygen in the air reacts with the waste hydrogen gas in the presence of a catalyst to produce the water vapor.

[0122]

[0119] In reference to FIG. 3B, in some embodiments, the HI system 300 comprises a safety shutoff circuit 308 coupled to at least the hydrogen gas source 301 and the catalytic combustion module 303. The safety shutoff circuit 308 may be configured to shut off or limit the rate of production of hydrogen gas from the hydrogen gas source 301 based at least on whether the catalytic combustion module 303 is operating such that the hydrogen concentration in the final waste mixture 307 discharged from the catalytic reaction chamber to ambient air is less than 4.5% by volume, optionally wherein the hydrogen concentration is less than 2% by volume. In some embodiments the safety shutoff circuit 308 may comprise sensors to measure the temperature or infrared signature of the catalytic reaction chamber or exhaust gas, a gaseous hydrogen sensor to monitor the concentration of hydrogen in the gaseous exhaust.

[0123] III. Degassing Embodiments

[0124]

[0120] In some embodiments, the HI systems of the present disclosure may comprise a degassing module configured for degassing the solution prior to infusing the solution with hydrogen. FIGS. 4-5 illustrate such HI systems according to exemplary embodiments.

[0125]

[0121] In reference to FIGS. 4A-4B, the HI system 400 may comprise at least a hydrogen gas source 401 and a fluid reservoir 402 separated by a degassing module. Fluid that has equilibrated with air can contain dissolved oxygen and dissolved nitrogen gases, which can dilute the hydrogen gas used for infusing hydrogen into the fluid. The presence of dissolved oxygen and nitrogen gas in the fluid and their dilution of the hydrogen gas can prevent the production of fluid with high concentrations of dissolved hydrogen without also producing hazardous quantities of waste gas containing hydrogen and oxygen. In some embodiments, use of a degassing step to remove or substantially reduce the concentrations of dissolved oxygen and nitrogen prior to a hydrogen infusion step may allow for minimizing the volume of hydrogen gas required to infuse the solution with a high concentration of hydrogen, and may also allow for minimizing or eliminating the production of any hydrogen-containing waste gas during infusion of the solution with the hydrogen gas.

[0126]

[0122] In some embodiments, the degassing step uses at least a gas exchanger 403 and a vacuum pump 404. In some embodiments, the gas exchanger 403 and the vacuum pump 404 are separated by at least one valve 405. The at least one valve 405 may be configured such that a first port is in fluid communication with the hydrogen gas source 401 via a first lumen 406, a second port is in fluid communication with the gas exchanger 403 via a second lumen 407, and the third port is in fluid communication with the vacuum pump via a third lumen 408. In some embodiments, the gas exchanger 403 is in fluid communication with the fluid reservoir 402 via a fourth lumen 409 and in some embodiments a fifth lumen 410. In some embodiments, the at least one valve 405 may comprise, for example, a three-way valve or a at least one two-way valve.

[0127]

[0123] As discussed above, in some embodiments, the gas exchanger 403 comprises a gas permeable membrane, a hollow-fiber gas exchanger, or a bubble gas exchanger. In some embodiments, the gas permeable membrane may be, for example, silicone or polysulfone, as in a standard hollow-fiber membrane oxygenator design. In some embodiments, the gas exchanger 403 comprises at least one liquid channel and at least one gas channel that are separated by the gas permeable membrane. The liquid channel is coupled to the fluid reservoir via the fourth lumen 409 and in some embodiments, fifth lumen 410. In some embodiments, an inlet of the gas channel is coupled to the at least one valve 405 via the second lumen 407. The configuration of the gas channel and liquid channel (a) allow for oxygen and nitrogen gases to be extracted from the solution flowing through the liquid channel, resulting in degassed solution, or (b) allow for the hydrogen to diffuse through the gas permeable membrane into the solution flowing through the liquid channel, resulting in a hydrogen-infused solution. In some embodiments, the hydrogen gas continues to diffuse through the gas permeable membrane until equilibrium or saturation within the solution has been reached. In some embodiments, the dissolved hydrogen gas may reach equilibrium concentrations as high as 1600 ppb at normal atmospheric pressure of 760 mmHg. In some embodiments, the hydrogen may be pressurized above atmospheric pressure on the gas side of the gas exchange membrane to produce a supersaturated solution with dissolved hydrogen gas at concentrations up to 2400 ppb. Simultaneously, hydrogen gas that is not diffused through the gas permeable membrane may exit the gas exchanger 403 as waste hydrogen gas.

[0128]

[0124] In some embodiments, as illustrated in FIGS. 4A-4B, the degassing module further comprises a circulation pump 411 in fluid communication with the fluid reservoir 402 separate from the fourth lumen 409. In reference to FIG. 4A, in a first configuration of the at least one valve, the first lumen 406 is disconnected such that the vacuum pump 404 is connected to the gas exchanger 403 through consecutive connection of the third lumen 408, the at least one valve 405 and the second lumen 407. The first configuration is used to facilitate degassing of the medical solution prior to infusion with hydrogen gas from the hydrogen gas source 401. In reference to FIG. 4B, in a second configuration of the at least one valve 405, the third lumen 408 is disconnected such that the hydrogen gas source 401 is connected to the gas exchanger 403 through consecutive connection of the first lumen 406, the at least one valve 405 and the second lumen 407. The second configuration is used to facilitate the infusion of the degassed solution with the hydrogen gas from the hydrogen gas source 401 to produce the hydrogen-infused solution for downstream administration to the patient, organ, tissue, or cells.

[0129]

[0125] In the first configuration, the circulation pump 411 applies a pressure so as to convey solution from the fluid reservoir 402 into the gas exchanger 403. Solution entering the gas exchanger 403 via the liquid channel is subject to negative pressure applied by the vacuum pump 404, causing gases to be extracted into the gas channel from the solution and thereby form degassed solution. In some embodiments, the extracted gases are in the form of dissolved gases in aqueous phase or bubbles in gaseous phase. In some embodiments, the extracted gases comprise one or more of nitrogen, carbon dioxide, oxygen, hydrogen, water vapor, or ambient air. After extraction, the degassed solution may be conveyed back into the fluid reservoir 402 via the fourth lumen 409 or in some embodiments, the fifth lumen 410. As a result, the circulation pump 411 allows the solution to continuously circulate from the fluid reservoir 402, through the gas exchanger 403 for further extraction, and back into the fluid reservoir 402 until the degassed solution has reached a desired low oxygen and nitrogen gas concentration. In some embodiments, the desired low oxygen and nitrogen gas concentrations are each monitored via a sensor, and may be monitored until the concentrations reach a threshold amount, at which point the circulation pump 411 may refrain from further continuing circulation. In some embodiments, the circulation pump 411 continues to circulate the solution and / or the degassed solution for a pre-determined number of cycles. In some embodiments, the desired gas partial pressure of the residual oxygen and nitrogen combine to about 0 - 100 mm Hg.

[0130]

[0126] In the second configuration, the circulation pump 411 again applies a negative pressure to recirculate the previously degassed solution through the gas exchanger 403. However, positive pressure from the hydrogen gas source 401 conveys hydrogen gas through the first lumen, the at least one valve 405 and the second lumen 407 to the gas exchanger 403 and infuses into the degassed solution to produce the hydrogen-infused solution. As hydrogen gas is infused into the previously degassed solution, the hydrogen-infused solution is conveyed back into the fluid reservoir 402 via the fourth lumen 409 or in some embodiments the fifth lumen 410. As a result, the circulation pump 411 allows the hydrogen-infused solution to continuously circulate from the fluid reservoir 402, through the gas exchanger 403 for further infusion, and back into the fluid reservoir 402 until the hydrogen-infused solution has reached a desired hydrogen gas concentration. In some embodiments, the desired hydrogen gas concentration is monitored via a sensor, and may be monitored until the hydrogen gas concentration reaches a threshold amount, at which point the circulation pump 411 may refrain from further continuing circulation. In some embodiments, the circulation pump 411 continues to circulate the degassed solution and / or the hydrogen-infused solution for a pre-determined number of cycles. In some embodiments, the desired hydrogen gas concentration comprises about 0.25 - 2.5 mM.

[0131]

[0127] In some embodiments, as illustrated in FIG. 4C, degassing with the vacuum pump may be performed according to process 430. In step 431, the at least one valve is adjusted to be in the first configuration such that the vacuum pump is in fluid communication with the gas exchanger. The circulation pump is then initiated to circulate the solution from the fluid reservoir through the gas exchanger. In step 432, the vacuum pump is allowed to apply continuous negative pressure to the gas exchanger, allowing for continuous degassing of the recirculating solution. In step 433, the at least one valve is adjusted to be in the second configuration such that the hydrogen gas source is in fluid communication with the gas exchanger. The hydrogen gas source is then activated, such as through the activation of electric current to an electrolysis cell, to produce the pure hydrogen gas that is conveyed to the gas exchanger using hydrogen gas pressure. The hydrogen gas is then infused into the recirculating, previously degassed solution without producing any waste hydrogen gas. In step 434, the hydrogen gas-infused solution is administered to a target patient, organ, tissue, or cells.

[0132]

[0128] In some embodiments, as illustrated in FIGS. 5A-5B, the degassing module comprises a gravity feed for conveying solution through the gas exchanger rather than or in addition to the circulation pump in the embodiments in FIGS. 4A-4B. In particular, the gravity feed comprises a first fluid reservoir 502A and a second fluid reservoir 502B separated by the gas exchanger 503, wherein the first fluid reservoir 502A is filled to a desired volume with the solution, while the second fluid reservoir 502B is empty or relatively empty of solution compared to the first fluid reservoir. The first fluid reservoir 502A may be in fluid communication with the gas exchanger 503 via a fourth lumen 509, and the second fluid reservoir 502B may be in fluid communication with the gas exchanger 503 via a fifth lumen 510. In some embodiments, each of the first fluid reservoir 502A and the second fluid reservoir 502B is in fluid communication with the gas exchanger 503 via separate lumens.

[0133]

[0129] In reference to FIG. 5 A, in a first configuration of the gravity feed, the first fluid reservoir 502A may begin at an elevated height relative to the gas exchanger 503 and the second fluid reservoir 502B, and the second fluid reservoir 502B is at a lower height relative to the gas exchanger 503 and the first fluid reservoir 502A. At least one valve 505 is also in the same configuration as illustrated in FIG. 4A, in that a first lumen 506 is disconnected such that a vacuum pump 504 is connected to the gas exchanger 503 through consecutive connection of a third lumen 508, the at least one valve 505 and a second lumen 507. Again, the first configuration is used to facilitate degassing of the medical solution prior to infusion with hydrogen gas from a hydrogen gas source 501 so as to fill either of the first fluid reservoir 502A or second fluid reservoir 502B with a degassed solution. In reference to FIG. 5B, in a second configuration of the gravity feed, a fuller fluid reservoir of the first and second fluid reservoirs 502A,B may begin at an elevated height relative to the gas exchanger 503 and an emptier fluid reservoir of the first and second fluid reservoirs 502A,B. In some embodiments, the fuller fluid reservoir is the second fluid reservoir 502B and the emptier fluid reservoir is the first fluid reservoir 502A. In some embodiments, the fuller fluid reservoir is the first fluid reservoir 502A and the emptier fluid reservoir is the second fluid reservoir 502B. The at least one valve 505 is also in the same configuration as illustrated in FIG. 4B, the third lumen 508 is disconnected such that the hydrogen gas source 501 is connected to the gas exchanger 503 through consecutive connection of the first lumen 506, the at least one valve 505 and the second lumen 507. Similarly, the second configuration is used to facilitate the infusion of the degassed solution with the hydrogen gas from the hydrogen gas source 501 to produce the hydrogen-infused solution for downstream administration to the patient, organ, tissue, or cells.

[0134]

[0130] In the first configuration, the positioning of one fluid reservoir relative to the other allows gravity to act upon the solution so as to be conveyed from the filled first fluid reservoir 502A into the gas exchanger 503. Solution entering the gas exchanger 503 is subject to negative pressure applied by the vacuum pump 504, causing gases to be extracted from the solution and thereby form degassed solution. In some embodiments, the extracted gases are in the form of dissolved gases in aqueous phase or bubbles in gaseous phase. In some embodiments, the extracted gases comprise one or more of nitrogen, carbon dioxide, oxygen, hydrogen, water vapor, or ambient air. After extraction, the degassed solution may be conveyed into the empty second fluid reservoir 502B. As a result, the empty second fluid reservoir 502B fills with degassed solution and the previously full first fluid reservoir 502A is emptied of solution. In some embodiments, while the at least one valve 505 remains in the first configuration, the second fluid reservoir 502B may be adjusted to an elevated height relative to the gas exchanger 503 and the first fluid reservoir 502A, and the first fluid reservoir 502A may be adjusted to a lower height relative to the gas exchanger 503 and the second fluid reservoir 502B, thereby allowing the degassed solution to flow through the gas exchanger 503. This adjusting may be performed any number of times until the degassed solution has reached a desired gas concentration. In some embodiments, the desired gas partial pressure of the residual oxygen and nitrogen combine to about 0 - 100 mm Hg.

[0135]

[0131] In the second configuration, the positioning of one fluid reservoir relative to the other allows gravity to act upon the degassed solution so as to be conveyed from the fuller fluid reservoir into the gas exchanger 503. However, positive pressure from the hydrogen gas source 501 conveys hydrogen gas through the first lumen 506, the at least one valve 505 and the second lumen 507 to the gas exchanger 503 and infuses into the degassed solution to produce the hydrogen-infused solution. As hydrogen gas is infused into the degassed solution, the hydrogen-infused solution is conveyed into the emptier fluid reservoir. As a result, the empty first fluid reservoir 502A fills with hydrogen-infused solution and the filled second fluid reservoir 502B is emptied of degassed solution. In some embodiments, while the at least one valve 505 remains in the second configuration, the first fluid reservoir 502A may be adjusted to an elevated height relative to the gas exchanger 503 and the second fluid reservoir 502B, and the second fluid reservoir 502B may be adjusted to a lower height relative to the gas exchanger 503 and the first fluid reservoir 502A, thereby allowing the hydrogen-infused solution to flow through the gas exchanger 503. This adjusting may be performed any number of times until the hydrogen-infused solution has reached a desired gas concentration. In some embodiments, the desired concentration comprises about 0.25 - 2.5 mM.

[0136]

[0132] In some embodiments, as illustrated in FIG. 5C and 5D, the solution may be pumped from the first fluid reservoir 512A to the second fluid reservoir 512B without gravity assistance by applying external pressure to one solution reservoirs to move solution through the gas exchanger 513 and into another unpressurized solution reservoir. In some embodiments the solution reservoirs are flexible reservoirs. In other embodiments pressurized cuffs may apply squeezing pressure to a solution reservoir to force the solution through the gas exchanger and into the other solution reservoir.

[0137]

[0133] In reference to FIG. 5C, the first fluid reservoir 512A is pressurized by the squeezing force of a first pressurized cuff, while the second fluid reservoir 512B remains unpressurized within a second, unpressurized cuff. The solution is forced from the first fluid reservoir 512A through the gas exchanger 513 and into the second fluid reservoir 512B.

[0138]

[0134] In reference to FIG. 5D, the second fluid reservoir 512B is pressured by the squeezing force of the second, pressurized cuff while the first fluid reservoir 512A remains unpressurized within the first, unpressurized cuff. The solution is forced from the second fluid reservoir 512B through the gas exchanger 513 and into the first fluid reservoir 512A.

[0139]

[0135] The same sequence of vacuum degassing and hydrogen infusion from FIGS. 5 A and 5B may coincide with the sequences of squeezing the fluid from one reservoir to the other as depicted in FIGS. 5C and 5D, respectively, without requiring elevation of either fluid reservoir above the other or any use gravity to convey fluid through the gas exchanger 503,513.

[0140]

[0136] In some embodiments, as illustrated in FIG. 5E, degassing with the vacuum pump may be performing according to process 530. In step 531, the at least one valve is adjusted to be in the first configuration such that the vacuum pump is in fluid communication with the gas exchanger. The vacuum pump may be activated to apply negative pressure to the gas exchanger, allowing for iterative degassing of the solution flowing through the gas exchanger. In step 532, the fuller, first fluid reservoir is elevated above the gas exchanger to allow solution to flow subsequently from the first fluid reservoir, through the gas exchanger, and into the emptier, second fluid reservoir. The fluid flowing through the gas exchanger while vacuum is applied to the gas exchanger will be degassed to enhance effectiveness of the following hydrogen infusion step. In step 533, the at least one valve is adjusted to be in the second configuration such that the hydrogen gas source is in fluid communication with the gas exchanger. The fuller of the first fluid reservoir and second fluid reservoir is also elevated to be above the gas exchanger while the emptier of the fluid reservoirs is lowered to below the gas exchanger. The hydrogen gas source is then initiated, such as through the addition of water, to produce the hydrogen gas that is conveyed to the gas exchanger using pure hydrogen gas pressure. The hydrogen gas is then iteratively forced to infuse into the degassed solution without producing waste hydrogen gas. In step 534, the hydrogen gas-infused solution is administered to a target patient, organ, tissue, or cells.

[0141]

[0137] In some embodiments, the degassing module may comprise an ultrasound degassing module. FIGS. 6A-6B illustrate such an HI system 600 according to exemplary embodiments. Specifically, the HI system 600 may comprise at least a hydrogen gas source 601 and a fluid reservoir 602, such that at least the fluid reservoir 602 is immersed in the ultrasound degassing module. The ultrasound degassing module may allow for similar benefits as the previously mentioned vacuum degassing steps, while further minimizing the number of elements required to obtain high concentrations of hydrogen in the solution while minimizing waste hydrogen gas.

[0142]

[0138] In reference to FIG. 6A, in some embodiments, the ultrasound degassing module comprises an ultrasonic bath 603. Once the fluid reservoir 602 is immersed into the ultrasonic bath 603, the ultrasonic bath 603 is allowed to apply ultrasonic energy for a desired degassing period such that gases in the solution, e.g., oxygen, nitrogen, etc., are forced to migrate to an upper portion 602A of the fluid reservoir 602. In some embodiments, the gases are dissolved gases. In some embodiments, the gases are bubbles of gas phase in the liquid phase. In some embodiments, the desired degassing period ranges from about 5 minutes to about 10 minutes. After the degassing period, an outlet port 604 of the fluid reservoir 602 is opened to allow for the migrated gases to evacuate into the ambient atmosphere.

[0143]

[0139] In reference to FIG. 6B, the fluid reservoir 602 may comprise an inlet port 605 for connecting to a hydrogen gas source 601 via a connector tube comprising a lumen 606. The hydrogen gas source 601 is configured to release hydrogen gas through the lumen and into the inlet port 605 for infusion into the solution. In some embodiments, the hydrogen gas forms a volume of pure hydrogen gas in at least the upper portion 602A of the fluid reservoir 602 upon injection into the fluid reservoir 602. In some embodiments, the hydrogen gas may infuse into the solution when the fluid reservoir 602 is agitated, such as, for example, through shaking.

[0144]

[0140] In some embodiments, as illustrated in FIG. 6C, degassing with the ultrasound degassing module may be performing according to process 630. In step 631, the fluid reservoir containing solution is placed into the ultrasonic bath, which is triggered to apply ultrasonic energy for a desired length of time to force migration of the gases toward an outlet port of the fluid reservoir. In step 632, the migrated gases are released from the fluid reservoir, such as through opening of the outlet port. In step 633, the fluid reservoir is connected to the hydrogen gas source using a connector tube such that the fluid reservoir is in fluid communication with the hydrogen gas source. The hydrogen gas source is triggered to generate hydrogen gas that is conveyed to the fluid reservoir via the connector tube until a desired volume and / or concentration of hydrogen gas is reached within the fluid reservoir. In step 634, the fluid reservoir is then agitated to allow the hydrogen gas to infuse into the solution.

[0145]

[0141] With reference to FIG. 6D, in some embodiments, infusion of hydrogen into degassed medical solution may be performed via process 640 so as to be packaged for later administration or to be directly administered to an organ or tissue. Step 641 comprises applying a degassing process to the medical solution so as to remove gas or gasses from the medical solution. In some embodiments, the degassing process comprises exposing the medical solution to a vacuum, heat, ultrasonic energy, or a combination thereof. In some embodiments, the degassing process is performed according to processes 430, 530, 630, or a combination thereof. In some embodiments, the degassed medical solution comprises partial pressures of all dissolved gases adding to a total partial pressure ranging from 0 mm Hg to 380 mm Hg. Step 642 comprises infusing the degassed medical solution with hydrogen gas from a hydrogen gas source to produce a hydrogen-infused solution. In some embodiments, the infusing utilizes a gas exchanger or any of the gas dissolution embodiments previously mentioned. In some embodiments, the infusing may comprise simply injecting hydrogen gas into a closed container of degassed medical solution and waiting for the hydrogen gas to passively dissolve into the degassed medical solution. In some embodiments, the hydrogen gas dissolves into the degassed medical solution to produce a hydrogen-infused solution at equilibrium hydrogen partial pressures ranging from about 230 mm Hg to about 2300 mm Hg or concentrations ranging from 0.25 mM - 2.5 mM. The user may then proceed to either step 643 A or 643B, depending on when the hydrogen-infused solution is needed. Step 643A comprises packaging the hydrogen-infused solution. In some embodiments, the packaging comprises receiving the degassed medical solution and the hydrogen gas from the gas source such that the hydrogen gas passively dissolves into the degassed medical solution. Contrarily, step 643B comprises administering the hydrogen-infused solution directly to the target organ or tissue.

[0146]

[0142] Each of the embodiments illustrated in FIGS. 1-6 can stand on its own or can be combined in various permutations or combinations with one or more of the other embodiments. For example, each of the hydrogen gas source and the fluid reservoir may be combined with one or more of the gas dilution module - i.e., the static gas dilution module, the dynamic dilution module, or a combination thereof - the safety shutoff circuit, the catalytic combustion module, the degassing module - i.e., with the circulating pump, the gravity feed, or a combination thereof - and the ultrasonic degassing module, along with any associated tubing and / or valves.

[0147]

[0143] In addition to the aforementioned features, the HI systems of this disclosure provide several advantages over traditional systems for infusing hydrogen gas into solutions. For example, hydrogen may be infused to achieve up to 100% saturation (i.e., 0.8 mM of hydrogen) while reducing dissolved oxygen in existing solution to 1-20% atmospheric levels. The sterility and biochemical integrity of the solutions are further preserved during use of the HI systems, and combustion hazards associated with the risk of hydrogen and oxygen mixing are negligible to nonexistent. Additionally, the infusion of hydrogen gas may be performed rapidly as compared, i.e., reasonably within 10 minutes of assembling the HI system, without the need for any pre- saturation of the solution that may be shipped from a factory. The HI systems require minimal handling, and particular operations may be automated using control instrumentation, discussed in further detail below. Each of the respective modules of the HI systems disclosed herein may also be reusable.

[0148] IV. Reusable and Disposable Modules

[0149]

[0144] In some embodiments, any number of the aforementioned modules or components of the HI systems may be reusable or disposable. The division of the HI system into reusable and disposable components allows for re-use of the more expensive, durable, or nonsterilized hardware and electronic components, while simultaneously allowing for the less costly, less durable, or terminally sterilized components to be designated as single-use and / or disposable. The single-use components can be provided in a sterile condition, thereby ensuring sterility of the hydrogen- infused solution produced by the HI system.

[0150]

[0145] In some embodiments, the reusable components may include the hydrogen gas source, the vacuum pump, the dilution chamber, the dynamic flow source, the diluent gas source, manual or automatic valves, a pump for moving or circulating the fluid, pressure cuffs for circulating fluid, an ultrasonic bath for degassing the solution, a temperature-controlled water bath to maintain solution temperatures, a catalytic reaction chamber, safety circuits and sensors, a user interface, or any combination of the above, including the valves, tubing, wires and lumens connecting said elements to each other.

[0151]

[0146] In some embodiments, the single-use or disposable components of the HI system may include a reservoir full of medical solution that is to be infused with hydrogen, one or more extra solution reservoirs, a gas exchanger, tubing and connectors to convey fluids and gases from different elements of the system, sampling ports for measuring dissolved gas concentrations, sterile filters to ensure sterility of a final product (i.e., the hydrogen-infused solution), spikes or needles to penetrate the ports of the fluid reservoirs, or any combination of the above.

[0152]

[0147] In some embodiments, each of the reusable modules and disposable modules may comprise one or more cartridges for mounting into an HI system.

[0153]

[0148] In some embodiments, any number of the following components for each respective reusable module may be disposable for incorporation into the cartridges. In some embodiments, the disposable components may include one or more of the at least one fluid reservoir and the solution. In some embodiments, the solution comprises one or more of saline, organ preservation solution, whole blood, partial blood, cardioplegia solution or an aqueous solution with other chemical components, such as drugs or tissue stabilizers for the designated use.

[0154]

[0149] In some embodiments, the cartridges or their components may vary in size so as to produce a required volume of hydrogen-infused solution. For example, disposable cartridges may be sized to infuse hydrogen into discrete volumes ranging from 50-5000 mL (e.g., 500 ml, 1000 ml, 2000 ml, or more) of solution. The surface area of the gas exchanger membrane, the diameter of tubing, and the capacity of any extra solution reservoirs may be sized specifically to the desired volume of hydrogen-infused solution.

[0155]

[0150] In some embodiments, the disposable cartridges may include an identification component to communicate important information to any computer-implemented controls of the reusable HI system, discussed in more detail below. The cartridge identifier may communicate information regarding volume of solution, desired gas concentrations, ultrasonic degassing time, desired temperature, amount of recirculation time required in the gas exchanger, serial and or model number of the cartridge, number of previous uses, and any combination of the above or other control parameters. The identification component may be a barcode, QR code, RFID chip, or other optical, textual, electronic, electromagnetic, or mechanical means of communication. In some embodiments the HI system will not operate if it detects that the cartridge is damaged, expired, obsolete, incompatible, or has already been used, in order to prevent use of components that are dysfunctional or no longer sterile, or to prevent use of unauthorized or incompatible cartridges.

[0156] V. Clinical Implementation

[0157]

[0151] In many instances of organ transplantation, an organ is transported from an organ recovery site to an organ transplant site in a hypothermic temperature range (0 - 10 °C), with no recirculation of blood or fluids through the vasculature of the organ aside from short flushes either before or after static cold storage. This organ transport method, referred to as “static cold storage” or “cold static storage”, imposes a period of ischemia or hypoxia on the organ during transport, after which the organ is surgically grafted into the transplant patient and reperfused with the patient’s oxygenated blood. The reperfusion of the previously ischemic organ with oxygenated blood initiates the production of damaging free radicals within the cells of the organ. The extent of damage (reperfusion injury) and severity of physiologic effects caused by the free radical production is related to the temperature and duration of the ischemic interval, the metabolic demands of the organ or tissue type, the presence of antioxidants or free radical scavengers, and the age and condition of the donated organ prior to static cold storage, as well as other factors. The use of a hydrogen-infused flushing solution either before or after static cold storage of an organ can reduce the likelihood of severe reperfusion injury and downstream medical complications as further described in Example 2 below.

[0158]

[0152] In other instances such as those for organ, tissue, or cell transplant, the organ, tissue, or cells must be transported for several hours between the place of organ recovery and the place of organ transplantation into the recipient patient. Also, in many organ recovery scenarios, organs may endure a prolonged period of warm ischemia or hypoxia in the donor body prior to the removal of the organ from the donor body. In either case, and especially when both cases are true, the transplant team may use an ex-vivo organ perfusion system to flush accumulated metabolites from the organ and in some cases to also perfuse the organ with a perfusate solution containing dissolved oxygen. In some cases, the organ may be perfused in an ex-vivo perfusion system either before transport at the organ recover site or after transport at the organ transplant site.

[0159]

[0153] Although ex-vivo organ perfusion systems can shorten the duration and degree of hypoxia by supplying oxygen to the organ, they do not neutralize the free radicals that are inevitably produced within the cells when oxygenated blood or oxygenated perfusate enter the organ after a period of ischemia or hypoxia. Although beneficial overall, the use of ex-vivo organ perfusion systems can introduce additional reperfusion steps into the organ transplant sequence. Each additional episode of reperfusion with oxygenated solution or blood after a period of ischemia or hypoxia can introduce a risk for causing oxidative stress or reperfusion injury. Therefore, it can be advantageous to incorporate the HI systems and methods into the systems and methods for ex-vivo organ perfusion, as further described in Example 3 below, in order to achieve neutralization of harmful free radicals resulting from reintroduction of oxygen to hypoxic or ischemic organs, tissues, and cells.

[0160]

[0154] In other instances, hydrogen-infused solutions can be administered to organs and tissues at different steps and in different settings in the organ transplant process, as further described in Example 4 below. In some settings, the hydrogen-infused solution may be administered to the donated organs while they are still in the donor body, by introducing the hydrogen-infused solution into an artery upstream of the organ. In other settings the hydrogen-infused solution may be flushed through the aorta of the donor body prior to removal of some or all organs, thereby flushing multiple organs with hydrogen-infused solution simultaneously. In other settings the donor circulatory system may be reperfused after the donor’s circulatory death by an external circulatory assistance device such as extracorporeal membrane oxygenation (ECMO) or cardiopulmonary bypass (CPB) to shorten the exposure of donated organs to warm ischemia prior to organ recovery.

[0161] VI. System and Computer Implementation

[0162]

[0155] An exemplary embodiment of a block diagram of a system 700 with which some embodiments may operate for receiving the plurality of datasets is illustrated in FIG. 7. The system 700 can analyze user inputs, system operational status, or cartridge identifier information for obtaining desired solution parameters based on any number of targets (i.e., blood, tissue, organ), gas concentrations, solution types, solution volumes, and solution usage rate. The system 700 can include a user computing device 710, which may be a desktop or laptop personal computer, smart mobile phone, server, or other suitable device. The user computing device 710 may include a user interface 711 by which the user 750 may interact with the user computing device 710. For example, the user 750 can use the user interface 711 to interface with a test target database 730 or test target analysis facility 721 of the server computing device 720. For example, the user 750 may operate the user interface 711 to initiate analysis of a dataset from the test target database 730 and display analysis results such as calculating gas distribution curves based on concentrations of gases over time for gas administration studies in the interface 711. The user 750 may additionally or alternatively operate the user interface 711 to input datasets obtained from the test target database 730, such as output to the user 750 in another interface. Those values may be provided to the test target analysis facility 721. As a further example, the user 750 may operate the user interface 711 to initiate analysis of the test target by the test target database 730 and provision of analysis results (e.g., extraction, infusion, a minimum effective concentration (MEC), etc.) from the test target database 730 to the test target analysis facility 721. Results of analysis of the results (received from the test target database 730 or from the interface 711) by the test target analysis facility 721 may be output to the user interface 711, such as by being received at the user interface 711 and displayed on the device 710. In some embodiments, as mentioned above, the user interface 711 may include a web interface, such as one or more web pages into which values may be output and which may display results of the analysis by the test target analysis facility 721, but embodiments are not so limited. The user interface 711 may accept input in a variety of different formats, such as through speech recognition, text input, or other means, as embodiments are not limited in this respect.

[0156] The system 700 can include a server computing device 720, which may include a test target analysis facility 721 configured to analyze factors (e.g., derived from the datasets, such as by the test target database 730) for the user 750 to characterize gas infusion or solution parameters of the test targets. In some embodiments, the test target analysis facility 721 may receive information on the factors from the test target database 730 and / or from the user interface 711. In some embodiments, the test target analysis facility 721 may output selected test targets that satisfy predetermined criteria.

[0163]

[0157] The system 700 can include a network 740 to facilitate communications among the test target database 730, the user computing device 710, and the server computing device 720. The network 740 can be or include any one or more wired and / or wireless, local- and / or wide-area network, including one or more enterprise networks and / or the Internet.

[0164]

[0158] While the example of FIG. 7 includes the client interface on a device 710 separate from the sample analyzer 112, it should be appreciated that embodiments are not so limited. In other embodiments, the user interface 711 may be an interface of the test target database 730 and may be operated by the user 750. Additionally or alternatively, while the test target analysis facility 721 is illustrated on a different computing device from the user computing device 710 and the test target database 730, embodiments are not so limited. In other embodiments, the analysis facility may be implemented on the client computing device or the test target database 730. In some embodiments, the user interface 711 may not be separate from the test target analysis facility 721, but instead may be implemented as a single program or software application. In some embodiments, a test target database 730 may include the user interface 711 and the test target analysis facility 721, and the interface 711 and facility 116 may be implemented within the same program or application executed on the test target database 730.

[0165]

[0159] In some embodiments, the HI systems described herein may further comprise an electronics module. The electronics module may be any machine or computer-implemented device, which may be used as a control system for controlling any number of processes associated with the HI systems, such as, for examples, processes 130, 430, 530, and 630. In some embodiments, the electronics module includes, but is not limited to, a printed circuit board assembly (PCBA) that houses and electrically interfaces many of the components of the HI system including, but not limited to, integrated circuits (ICs), at least one microcontroller (MCU) (with a processor, memory and input / output peripherals), a network adapter and a plurality of sensors. Some of the functions of the electronics module in conjunction with a firmware each of the components of the HI system include, but are not limited to, turning the components on / off; recognizing user input; processing data; storing data; managing power supply; sending and receiving data; and pairing / unpairing to an external device / application via a wired or wireless communication.

[0166]

[0160] In some embodiments, the electronics module comprises a power supply. In some embodiments, the power supply comprises a battery. In some embodiments, the plurality of sensors comprises a plurality of sensors designated for a diagnostic function, such as a concentration, volume, and / or partial pressure of a gas within or between each module the HI system. For example, the levels of H2, O2, CO2, N2, or combinations thereof are monitored and reported to a user and / or a monitoring system. In some embodiments, the plurality of sensors includes temperature sensors.

[0167]

[0161] FIG. 8 illustrates one exemplary implementation of an electronics module in the form of a computing device 800 that may be used in a system implementing techniques described herein, although others are possible. It should be appreciated that FIG. 8 is intended neither to be a depiction of necessary components for a computing device to execute an analysis facility in accordance with the principles described herein, nor a comprehensive depiction. Computing device 800 may comprise at least one processor 801, a network adapter 802, and computer- readable storage media 803. Computing device 800 may be, for example, a desktop or laptop personal computer, a personal digital assistant (PDA), a smart mobile phone, a server, a wireless access point or other networking element, or any other suitable computing device. Network adapter

[0168] 802 may be any suitable hardware and / or software to enable the computing device 800 to communicate wired and / or wirelessly with any other suitable computing device over any suitable computing network. The computing network may include wireless access points, switches, routers, gateways, and / or other networking equipment as well as any suitable wired and / or wireless communication medium or media for exchanging data between two or more computers, including the Internet. Computer-readable media 803 may be adapted to store data to be processed and / or instructions to be executed by processor 801. Processor 801 enables processing of data and execution of instructions. The data and instructions may be stored on the computer-readable storage media 803.

[0169]

[0162] Computer-executable instructions implementing the techniques described herein (when implemented as one or more functional facilities or in any other manner) may, in some embodiments, be encoded on the one or more computer-readable media 803 to provide functionality to the media. The data and instructions stored on computer-readable storage media

[0170] 803 may comprise computer-executable instructions implementing techniques which operate according to the principles described herein. In the example of FIG. 8, computer-readable storage media 803 stores computer-executable instructions implementing various facilities and storing various information as described above. Computer-readable storage media 803 may store test target analysis facility 804 for analyzing data obtained from each of the plurality of sensors.

[0171]

[0163] Computer-readable media include magnetic media such as a hard disk drive, optical media such as a Compact Disk (CD) or a Digital Versatile Disk (DVD), a persistent or non-persistent solid-state memory (e.g., Flash memory, Magnetic RAM, etc.), or any other suitable storage media. Such a computer-readable medium may be implemented in any suitable manner, including as computer-readable storage media 803 of FIG. 8 (i.e., as a portion of a computing device 800) or as a stand-alone, separate storage medium. As used herein, “computer-readable media” (also called “computer-readable storage media”) refers to tangible storage media. Tangible storage media are non-transitory and have at least one physical, structural component. In a “computer- readable medium,” as used herein, at least one physical, structural component has at least one physical property that may be altered in some way during a process of creating the medium with embedded information, a process of recording information thereon, or any other process of encoding the medium with information. For example, a magnetization state of a portion of a physical structure of a computer-readable medium may be altered during a recording process.

[0172]

[0164] In some, but not all, implementations in which the techniques may be embodied as computer-executable instructions, these instructions may be executed on one or more suitable computing device(s) operating in any suitable computer system, one or more computing devices (or one or more processors of one or more computing devices) may be programmed to execute the computer-executable instructions. A computing device or processor may be programmed to execute instructions when the instructions are stored in a manner accessible to the computing device or processor, such as in a data store (e.g., an on-chip cache or instruction register, a computer-readable storage medium accessible via a bus, a computer-readable storage medium accessible via one or more networks and accessible by the device / processor, etc.). Functional facilities comprising these computer-executable instructions may be integrated with and direct the operation of a single multi-purpose programmable digital computing device, a coordinated system of two or more multi-purpose computing device sharing processing power and jointly carrying out the techniques described herein, a single computing device or coordinated system of computing devices (co-located or geographically distributed) dedicated to executing the techniques described herein, one or more Field-Programmable Gate Arrays (FPGAs) for carrying out the techniques described herein, or any other suitable system.

[0173]

[0165] While not illustrated in FIG. 8, a computing device may additionally have one or more components and peripherals, including input and output devices. These devices can be used, among other things, to present a user interface. Examples of output devices that can be used to provide a user interface include printers or display screens for visual presentation of output and speakers or other sound generating devices for audible presentation of output. Examples of input devices that can be used for a user interface include keyboards, and pointing devices, such as mice, touch pads, and digitizing tablets. As another example, a computing device may receive input information through speech recognition or in other audible format.

[0174] EXAMPLES

[0175] Example 1. Exemplary HI system with reusable modules.

[0176]

[0166] The purpose of this example is to provide an embodiment of an HI system with reusable modules as described above. FIG. 9A illustrates an embodiment of an HI system 900 comprising at least a reusable module and a disposable cartridge. The components of each of the reusable module and disposable cartridge are divided into reusable components and disposable components, respectively, designated according to a provided color key. The disposable cartridge components include a first solution reservoir 902 A full of medical solution (i.e., a first fluid reservoir), a second solution reservoir 902B that is empty to receive the solution (i.e., a second fluid reservoir), a gas exchanger 903, one or more sterile filters 905 to ensure sterility of the final solutions, and tubing to connect the full and empty solution reservoirs 902A,B to the gas exchanger 903 and to fit into a roller pump 904. In some embodiments, any number of fluid-contacting surfaces of the disposable components are sterilized prior to use. In some embodiments, the tubing further comprises a number of tubing connections 908 between disposable components and reusable components.

[0177]

[0167] The reusable module components include the hydrogen gas source 901, a solution cooling well 909, the roller pump 904, an air blower 907 (i.e., a dynamic flow source), a waste gas dilution chamber 906 with an outlet 906A, any number of lumens or tubing conveying gases to and from the gas exchanger 903 and any other reusable components, and associated valves or control systems for electrical or mechanical operation. In some embodiments the reusable module includes pressure cuffs to push solution from one fluid reservoir to another instead of using the roller pump 904. In some embodiments the hydrogen gas source 901 comprises a disposable module.

[0178]

[0168] In some embodiments, the disposable cartridge components interact with the reusable module as follows: (a) each of the original and empty reservoirs 902 A, B rest in the solution cooling well 909 with a bath of cooling water; (b) the tubing connecting one or more of the reservoirs to the gas exchanger 903 is installed between rollers of the roller pump 904 for conveyance of solution through the gas exchanger 903; (c) the hydrogen gas source 901 is connected to a gas inlet of the gas exchanger 903; and (d) the gas outlet of the gas exchanger 903 is connected to the waste gas dilution chamber.

[0179]

[0169] The gas exchanger 903 comprises at least one liquid channel and at least one gas channel that are separated by at least one gas permeable membrane. In this example, the hydrogen gas flow rate can be 30 to 150 ml per minute. Here, the liquid channel is in fluid communication with each of the first and second fluid reservoirs 902A,B via a first lumen and a second lumen, respectively, and the gas channel is in fluid communication with the hydrogen gas source 901 and the dilution chamber 906 via a third lumen and a fourth lumen, respectively.

[0180]

[0170] As shown, the roller pump 904 is configured between at least one of the reservoirs and the gas exchanger 903 to convey the solution from the respective reservoir through the gas exchanger 903 for hydrogen infusion. In this example the solution flow rate through the gas exchanger can be 100 to 500 ml / min. The roller pump 904 can change direction to transfer the solution from the first solution reservoir 902A to the second solution reservoir 902B, or from the second solution reservoir 902B to the first solution reservoir 902A. The dilution chamber 906 further comprises an additional inlet for air to enter. In some embodiments, the air blower 907 can be replaced by a nitrogen gas source or carbon dioxide gas source as the source of diluent gas. In this example, the diluent gas flow rate can be 1 to 4 liters per minute. Further, in some embodiments, the nitrogen gas source comprises a compressed gas reservoir, a nitrogen enrichment membrane module, or a pressure swing adsorption module to deliver nitrogen gas for dilution. Further, the ideal nitrogen diluent gas should contain less than 6% oxygen by volume, which can be economically achieved by either of the nitrogen enrichment membrane or pressure swing adsorption technologies.

[0181]

[0171] In the embodiment illustrated in FIG. 9 A, at least the hydrogen gas source 901, dilution chamber 906, roller pump 904, and air blower 907 are reusable, and each of the gas exchanger 903, the first fluid reservoir 902A, the second fluid reservoir 902B, and tubes containing the lumens are disposable. Any number of the valves, pumps, or other components of each module is controlled manually or via automated systems.

[0182]

[0172] FIG. 9B illustrates a first pass of the solution through the gas exchanger 903. The roller pump 904 moves solution out of the first solution reservoir 902 A, through the gas exchanger 903, and into the second solution reservoir 902B. Hydrogen gas 910 flows from the hydrogen gas source 901 to the gas exchanger 903 where a substantial portion of the hydrogen gas 910 diffuses across the gas exchange membrane into the solution. Simultaneously, in some embodiments, dissolved nitrogen and oxygen gases diffuse out of the solution, through the gas exchange membrane and mix with the gas stream to create a stream of mixed waste gases 911. The waste gas 911 flows into the dilution chamber 906, where the waste gas 911 mixes with diluent gas or air resulting in a discharge mixture 912 having less than 4.5% hydrogen prior to release to the atmosphere. The first pass of solution through the gas exchanger can result in a hydrogen-infused solution 913 with dissolved hydrogen concentrations ranging from 200 ppb to 1600 ppb, depending upon parameters such as fluid flow rates, gas flow rates, gas exchanger design, and solution type.

[0183]

[0173] FIG. 9C illustrates an optional second pass of the solution through the gas exchanger 903 to further increase the hydrogen concentration in the hydrogen-infused solution 913, or to move the hydrogen-infused solution 913 back to its original reservoir. The HI system 900 can pass the hydrogen-infused solution 913 through the gas exchanger 903 more than once to increase the final hydrogen concentration, or to finish the process with the hydrogen-infused solution 913 in the preferred solution reservoir.

[0184]

[0174] By reversing the direction of the roller pump 904 the hydrogen-infused solution 913 is drawn from the second reservoir 902B through the gas exchanger 903 and back into the original first solution reservoir 902A. In some embodiments a sterile filter 905 is used to ensure the sterility of the solution in the final reservoir. The hydrogen gas source 901 can continue to supply hydrogen gas 910 to the gas exchanger 903 to increase the hydrogen concentration in the hydrogen-infused solution 913. The air blower 907 or other source of diluent gas can continue to deliver sufficient volume of air or other diluent gas to dilute the hydrogen waste gas 911 before its release to atmosphere.

[0185] Example 2. Incorporation of HI systems and methods into static cold storage organ transport methods.

[0186]

[0175] The use of a hydrogen-infused flushing solution either before or after static cold storage of an organ can reduce the likelihood of severe reperfusion injury and downstream medical complications as described above. In reference to FIGs 10A - 10C, the steps for preparation and administration of the hydrogen infused solution are depicted in relation to the nominal sequence of steps for static cold storage and transport of organs.

[0187]

[0176] FIG. 10A illustrates a typical process 1000 for organ recovery, flushing, transportation, and transplantation using static cold storage. In step 1001, surgery is performed to recover the organ from the donor. In step 1002, the organ is flushed with one or more solutions, such as preservation or other static cold storage solutions. In step 1003, the organ is stored at hypothermic temperature (0 - 10 °C) without any further perfusion. In step 1004, the organ is removed from storage and optionally flushed with a rinse solution to remove any constituents of the flushed solution. In step 1005, vasculature of the organ is grafted to the recipient patient’s vasculature. In step 1006, the organ is reperfused with the patient’s oxygenated blood. In step 1007, as a result of the reperfusion with the oxygenated blood, the cells of the donated organ are at risk of producing damaging free radicals, potentially leading to reperfusion injury that manifests in the form of one or more of inflammation, cell death, delayed graft function, medical complications with varying degrees of severity. Since flushing solutions are commonly used in the existing processes, both before and after static cold storage, the hydrogen-infused solution can substitute for existing solutions at various flush steps without disrupting a typical sequence of operational steps. FIG. 10A further illustrates which steps of process 1000 are ischemic or hypoxic for the organ being transplanted, and steps that have the potential for a reperfusion injury to be initiated if hydrogen has not yet dissolved into the tissue.

[0188]

[0177] FIG. 10B depicts an exemplary process 2000 of steps in which the flushing solution is infused with hydrogen using the HI system prior to flushing the organ. In this example process, the hydrogen-infused flush is administered either during organ recovery surgery while the organ is in the donor body, or after removal of the organ from the donor body but before static cold storage.

[0189]

[0178] Exemplary process 2000 comprises the following steps. Step 2001 is analogous to step 1001. In step 2002, the organ is flushed with one or more hydrogen-infused solutions produced via the HI system. Specifically, the hydrogen-infused solution in step 2002 is prepared by procuring a sterile container of solution for flushing or perfusing the organ, then installing the sterile container along with a disposable cartridge of the HI system into a reusable module of the HI system. The HI system is initiated to infuse hydrogen into the solution to produce the hydrogen- infused solution, which is then maintained at a preferred temperature until the organ is ready for flushing. Notably, as the organ is flushed, hydrogen diffuses into the organ tissue. In cases of long ischemic times prior to organ recovery, such as donation after cardiac death (DCD), any hydrogen- infused solution should be confirmed to have oxygen partial pressure lower than 40 mm Hg to minimize free radical production and hydrogen consumption during post-recovery flushing. Step 2003 is analogous to step 1003. In step 2004, vasculature of the hydrogen-infused organ is grafted to the recipient patient’s vasculature. In step 2005, the hydrogen-infused organ is reperfused with the patient’s oxygenated blood. In step 2006, hydrogen that previously was diffused into the organ neutralizes free radicals that are produced as a result of reperfusion, thereby reducing the chance of reperfusion injury. Any remaining unreacted hydrogen in the organ tissue diffuses into and is diluted by patient’s blood to be absorbed by other tissues or exhaled by the lungs.

[0190]

[0179] The solution selected for the hydrogen-infused flush solution can be any of the solutions already in common use by transplant teams, including University of Wisconsin Solution, Belzer’s Solution, Celsior Solution, Histidine-Tryptophan-Ketoglutarate (HTK) solution, Steen solution, blood, normal saline, cardioplegic solution, and others. In some cases the flush solution can be a customized mix of ingredients made by the transplant team or by the hospital pharmacy. In some embodiments the solution also contains heparin or other anticoagulants, steroids, antimicrobial agents, antioxidant molecules, impermeant molecules, sugars, insulin, buffering compounds and electrolytes. Aside from reduction of levels of dissolved oxygen and / or nitrogen in the flush solution, the infusion of hydrogen gas into the flush solution may not affect the intended chemical effects of any of the other solution ingredients. Dissolved hydrogen may not affect pH buffering, clotting, steroid action, or the molecular actions and effects of the other solution ingredients.

[0191]

[0180] In some embodiments, a bicarbonate buffered solution may be selected as the medical solution for hydrogen infusion. Since bicarbonate-buffered solutions are typically designed to maintain physiologic pH in equilibrium with dissolved carbon-di oxide gas having a partial pressure of about 40 mm Hg, the HI system may use a hydrogen / carbon dioxide gas mixture having 5% carbon dioxide by volume with the remaining 95% comprising hydrogen or a mixture of hydrogen and other gases.

[0192]

[0181] The hydrogen in the flush solution is imparted from the solution to the organ by capillary diffusion and by bulk diffusion. The hydrogen-enriched flush solution may also be used to bathe the external surfaces of the organ and provide buoyant support during cold storage. Under the static cold storage conditions, the dissolved hydrogen remains largely static in the organ tissues until the organ is reperfused with the organ recipient’s blood.

[0193]

[0182] Under some organ recovery conditions, the duration of the organ recovery surgery or the sequence of cardiac events during organ recovery surgery may impose an extended period of warm ischemia on the organs. A period of warm ischemia can be defined as either (a) the time period between cessation of oxygenated blood flow to the organ and the cooling of the organ to hypothermic temperatures by administration of a cold flushing and / or bathing solution; or (b) the time period between cessation of oxygenated blood flow to the organ and reestablishment of oxygenated perfusion by some other means such as ex-vivo organ perfusion or mechanical circulator support of the donor body. Under conditions of warm ischemia exceeding about 5 minutes, the recovered organ can become especially susceptible to reperfusion injury. Under the profoundly hypoxic conditions caused by warm ischemia, free radical production can be stimulated in the cells, especially in the vascular endothelium, by flushing the organ with solutions containing dissolved oxygen at partial pressures in excess of about 70 mmHg. Thus, the flushing of a profoundly hypoxic organ with a flush solution that has equilibrated with air, having an oxygen partial pressure of approximately 160 mm Hg, can cause oxidative stress in the organ.

[0183] In cases of profound organ hypoxia as described above, a secondary benefit of the HI system and methods can be realized with the use of flushing solutions having simultaneously high concentrations of hydrogen and low concentrations of oxygen. In some embodiments, an optimal oxygen partial pressure would be lower than 40 mm Hg. The system and methods for infusing high concentrations of hydrogen described in previous embodiments have the effect of significantly reducing dissolved oxygen concentrations while increasing hydrogen concentrations in the flush solution. The previously described embodiments of the HI system are suited to safely produce flush solutions on demand or at the point of use with a high ratio of dissolved hydrogen to dissolved oxygen such as 8: 1, 10: 1, 15: 1, 20: 1 or even 100: 1. In other words, the oxygen partial pressure may be reduced from the default 160 mm Hg (as equilibrated with air over long storage periods) down to 40 mm Hg or even as low as 1 mm Hg or less. The high ratio of dissolved hydrogen to dissolved oxygen allows for the hydrogen to saturate the tissue prior to exposing the tissue to concentrations of oxygen that would stimulate production of otherwise damaging free radicals.

[0194]

[0184] FIG. 10C depicts an exemplary process 3000 of steps in which the hydrogen-infused flush is administered to the organ after the static cold storage interval. Here, steps 3001-3003 and 3005 are analogous to steps 1001-1003. In step 3004, the organ is flushed with one or more hydrogen- infused solutions produced via the HI system. Steps 3005-3007 are analogous to steps 2004-2006.

[0195]

[0185] The hydrogen is imparted from the hydrogen-infused solution to the organ in like manner as described in Figure 10B, namely capillary diffusion and bulk diffusion. With high concentrations of dissolved hydrogen in the organ, the organ will retain a protective level of dissolved hydrogen through the first 5 to 10 minutes of reperfusion with the patient’s blood. Previous experiments have demonstrated that the first 5 to 10 minutes of reperfusion present the greatest opportunity for dissolved hydrogen to neutralize free radicals and mitigate reperfusion injury in the organ.

[0196]

[0186] The methods for administering hydrogen-infused flushing solutions depicted in FIGS. 10B and 10C can be combined, such that the organ is flushed more than once with hydrogen-infused solutions.

[0197] Example 3. Adaptation of HI systems and methods to ex-vivo organ perfusion systems and methods.

[0198]

[0187] FIGS. 11A-11H illustrate exemplary embodiments of HI systems and methods for adaptation to ex-vivo organ perfusion systems and methods as described above. Hydrogen can be infused into the organ at different times within the sequence of steps between organ recovery from the donor body, and organ grafting into the recipient patient. The hydrogen administration systems and methods can vary depending upon circumstances or preferences of the transplant teams, as explained in FIGS. 11 A-FIG 11H.

[0199]

[0188] FIG. 11 A illustrates the essential elements of a non-oxygenating ex-vivo perfusion system 1100; namely, a perfusate circuit comprising an appropriate perfusate solution 1101, a pump 1102, and the organ 1103. Without a gas exchanger, the system does not continuously supply dissolved oxygen to the organ 1103. The system comprises other elements including chemical sensors, pressure and flow sensors, temperature sensors, heating or cooling elements, and other devices. In an embodiment, the heating or cooling elements can include an ice bath, contact with a phase change material, or other like thermal mechanisms. In some embodiments, the heating or cooling elements can maintain a target temperature of the perfusate solution within approximately 2 degrees C - approximately 12 degrees C. In some embodiments, the heating or cooling elements can maintain a target temperature of the perfusate solution within approximately 4 degrees C - approximately 8 degrees C. At a minimum, the perfusion circuit recirculates a perfusate solution 1101 through the organ 1103 to supply the organ 1103 with fresh perfusate to maintain a desired pH and electrolyte balance, while also rinsing away or diluting cellular metabolites such as lactic acid, carbon dioxide, and others. A hydrogen-infused perfusate can be incorporated into this minimalist arrangement by infusing hydrogen into the perfusate solution 1101 using one of the previously described systems or methods described in FIGS. 1-9 prior to installing the organ 1103 and perfusate into the ex-vivo perfusion system 1100. In some cases, the organ 1103 can also be flushed with a hydrogen-infused solution prior to installing it into an ex-vivo perfusion system 1100. In a closed recirculation system lacking a gas exchanger as depicted in FIG 11 A, the amount of dissolved hydrogen can remain largely static within the system throughout the transport or storage interval. If the perfusate solution 1101 or the organ 1103 starts with a sufficiently high concentration of hydrogen, the organ 1103 can emerge from the perfusion system containing enough dissolved hydrogen to mitigate the effects of reperfusion injury when the organ 1103 is reperfused with oxygenated perfusate or blood. In some embodiments, the organ 1103 and the perfusate solution 1101 are both saturated with a high concentration of hydrogen, such as within a range of about 1200 ppb-1600 ppb, to maintain a high hydrogen concentration in the tissue up through the first 5-10 minutes of oxygenated reperfusion when the organ 1103 is grafted into the patient’s vasculature.

[0200]

[0189] FIG. 1 IB depicts an exemplary process 4000 of steps in which a hydrogen-infused solution can be flushed through the organ after ex-vivo perfusion, prior to reperfusion with the patient’s oxygenated blood. Process 4000 is similar to process 3000, the exception being that step 4003 comprises perfusing the organ with the ex-vivo perfusion system 1100 and removing the organ from the ex-vivo perfusion system 1100 for flushing with hydrogen-infused solution at step 4004, rather than storing the organ in a static state without perfusion. Using a hydrogen-infused solution after ex-vivo perfusion is advantageous if the perfusate from the ex-vivo perfusion system has to be flushed out of the organ 1103 prior to grafting the organ 1103 into the patient. In some embodiments, a perfusate solution for ex-vivo perfusion can contain high concentrations of potassium which would raise the organ recipient’s blood potassium to undesirable levels if the perfusate is not rinsed out from the organ prior to grafting. In some cases, the hydrogen-infused solution can be administered before and after the ex-vivo perfusion step.

[0201]

[0190] FIG. 11C depicts a typical ex-vivo oxygenated perfusion system 1110, in which a gas exchanger 1114 and a gas source 1115 are added to the perfusion circuit. The gas exchanger 1114 infuses dissolved oxygen into the perfusate solution 1111 while simultaneously extracting carbon dioxide from the perfusate solution 1111. In some embodiments the gas source 1115 comprises a mix of oxygen and other gases 1116 comprising one or more of carbon dioxide, nitrogen, nitric oxide, or other gases. The pump 1112 recirculates the perfusate solution 1111 through the organ 1113. The perfusate solution 1111 can flow in either pumping direction depending on system design. In some embodiments, the perfusate solution 1111 flows from the organ 1113 to the pump 1112 to the gas exchanger 1114. In some embodiments, the perfusate solution 1111 flows from the organ 1113 to the gas exchanger 1114 to the pump 1112. For ex-vivo perfusion systems using one or more gas exchangers, the potential for the system to produce a flammable or explosive hydrogen-rich waste gas 1117 must be considered and the risks mitigated.

[0202]

[0191] FIGS. 1 ID and 1 IE depict an embodiment of a perfusion system 1120 and process 5000, respectively, in which a hydrogen-infused solution is administered to the organ 1103 after cessation of gas flow through the gas exchanger 1124 and prior to removing the organ 1103 from the perfusion system 1120. Process 5000 is similar to process 4000, the exception being that step 5003 comprises introducing the hydrogen-infused solution into the perfusate pathway upstream of the organ 1103, to flow through the organ 1103 while the organ 1103 is still housed in the ex-vivo perfusion system 1120, rather than flushing the organ 1103 with hydrogen-infused solution outside of the perfusion system 1120. By stopping the flow of oxygen, air, and other gases through the gas exchanger 1124, the volume of hydrogen gas that can diffuse out of the solution to mix with the air outside the system is limited to the static gas volume, also called the gas priming volume, of the gas exchanger itself. Static gas volumes of gas exchangers can be minimized by design to a small volume ranging from about 1 ml - 25 ml, limiting the volume, and associated hazards, of any hydrogen gas that diffuses to the gas side of the gas exchanger membrane. In some embodiments, isolation valves 1152 can be employed at the inlet, outlet, or both sides of the gas flow path through the gas exchanger 1124, to prevent the flow of air or other gases through the gas exchanger 1124 after hydrogen-infused solution has been injected into the organ 1103 or perfusion system. In some embodiments, isolation valves 1153 can be deployed to prevent flow of hydrogen- infused solution through the gas exchanger.

[0203]

[0192] In some embodiments, a reservoir of hydrogen-infused flush solution 1128 can be introduced into the perfusion pathway upstream of the organ 1103. In some embodiments the temperature of the reservoir of hydrogen-infused solution 1128 can be maintained by a temperature controlled environment 1150, to bring the organ 1103 to a final desired temperature range after the hydrogen-infusion flush and prior to removal from the perfusion system 1120. The hydrogen- infused flush solution 1128 may be moved through the organ 1103 by the force of gravity. In some embodiments, the hydrogen infused solution 1128 may be pumped through the organ 1103 by the perfusion pump 1122. In some embodiments, after exiting the organ 1103, the hydrogen- infused solution 1128 may exit the perfusion fluid pathway by a drain line 1151 to be collected in an effluent solution container 1154. In some embodiments, the hydrogen-infused solution may be recirculated from the drain line 1151 or effluent container 1154 back through the organ 1103.

[0204]

[0193] In some embodiments, the pump 1122 continues to circulate perfusate solution 1121 through the organ 1103 while the hydrogen-rich flush solution 1128 is introduced into the recirculating perfusate circuit at any point between the pump 1122, organ 1103, and gas exchanger 1124. In some embodiments the hydrogen-infused solution can be pumped into the perfusate flow path by the pump 1122. In some embodiments the perfusion system 1120 incorporates the extra volume of solution into the recirculating perfusate solution 1121. In some embodiments the perfusion system 1120 reduces the recirculating perfusate volume to accommodate the volume of the added hydrogen-infused solution. In another embodiment, the hydrogen-rich flushing solution 1128 is injected into the perfusate pathway upstream of the organ 1103, while the solution downstream of the organ 1103 is discarded to a waste container 1154 or otherwise prevented from recirculating back into the organ 1103. In some embodiments, the perfusion system 1120 houses the hydrogen-infused flushing solution in a container or reservoir 1128 separately from the perfusate solution, to be introduced into the perfusate pathway or organ 1103 at a predetermined step in the process or at a time of the user’s choosing. In some embodiments, the perfusion system 1120 regulates the temperature of the hydrogen-infused flushing solution by either heating or cooling the hydrogen-infused solution by contact with a heating or cooling element 1150. In some embodiments, the perfusion system contains safety circuits, firmware, software, or any combination of these elements to prevent the flow of gases through the gas exchanger 1124 either during or after the introduction of hydrogen-rich solution into the organ 1103 or perfusate pathway.

[0205]

[0194] In an embodiment, a dissolved gas sensor 1165 may be positioned to monitor the concentration of dissolved oxygen, dissolved hydrogen, or both gases in the effluent solution stream 1151 coming from the organ 1103 during step 5003 of the process.

[0206]

[0195] In some embodiments the sensor element 1165 may include sensors for measurement of temperature and solution flow rate.

[0207]

[0196] In an embodiment, the measurement by sensor 1165 of any or all the following parameters (temperature, flow rate, dissolved oxygen concentration, or dissolved hydrogen concentration) may be compared to preferred setpoints for the same parameters by an electronic control system 1170. When predetermined parameter setpoints are achieved, the electronic control system may shut off, slow, or stop the flow of the hydrogen-enriched solution 1128 through the organ 1103. In the interest of time, this function may be advantageous for identifying the point of complete infusion by detecting either a sufficient rise in effluent hydrogen or a drop in effluent oxygen to indicate the desired concentration of dissolved hydrogen in the organ.

[0208]

[0197] In an embodiment, electronic control system 1170 prevents introduction of hydrogen- infused solution into the perfusion pathway until it is confirmed, either by the user or by onboard sensors and logic, that the oxygen gas flow from 1125 has ceased and that gas flow or liquid flow (or both) through the gas exchanger 1124 have been stopped.

[0209]

[0198] FIGS. 1 IF and 11G depict an embodiment of an ex-vivo perfusion system 1130 and process 6000, respectively, for safely administering dissolved hydrogen to the organ in the perfusion system by means of a gas exchanger. Process 6000 is similar to process 5000, the exceptions being that instead of introducing hydrogen-infused solution into the perfusate path, hydrogen and oxygen gas sources 1135 A, B are directly introduced, either internal or externally to the ex-vivo perfusion system 1130, to be in fluid communication with the gas exchanger 1134 for infusion into the perfusate. Once connected to the gas exchanger 1134, the hydrogen gas source 1135A is activated. The resulting hydrogen-infused perfusate allows for hydrogen gas 1136A to diffuse into the organ 1133 to achieve solution concentrations of about 40-2000 ppb. Any waste gas 1137 from the gas exchanger 1134 is treated by (a) dilution, or (b) catalytic combustion, using any of the aforementioned dilution or catalytic combustion modules, respectively, to achieve hydrogen concentration below 4.5% in the final waste gas stream. In some embodiments, the waste gas 1137 exits the gas exchanger through a flame arrester before treatment to reduce hydrogen concentration below 4.5%.

[0199] As illustrated in FIG. 1 IF, the gas exchanger 1134 may receive any mixture of hydrogen, oxygen, and other gases from the one or more gas sources 1135 A, B to impart dissolved gases into the perfusate solution. The gas feed to the gas exchanger 1134 can be carefully controlled to prevent mixing of hydrogen gas 1136A with oxygen gas 1136B in the feed gas flowing into the gas exchanger 1134, but the mixture of waste gases 1137 flowing out from the gas exchanger 1134 may still contain flammable mixtures of hydrogen and oxygen gas. In some embodiments, the ex- vivo perfusion system 1130 stops the flow of oxygen gas 1136B to the gas exchanger 1134, purges the gas exchanger 1134 with nitrogen gas, and then starts the flow of hydrogen gas 1136A through the gas exchanger 1134. The waste gas 1137 may still contain a potentially hazardous mixture of hydrogen and oxygen gas due to the diffusion of oxygen from the previously oxygenated perfusate solution through the gas exchange membrane and into the stream of waste gas 1137. To prevent the release of hazardous waste gas mixtures into the local environment, the waste gas 1137 would require neutralizing the flammability hazard using methods similar to those already described above, such as dilution or catalytic combustion of the waste hydrogen prior to releasing the waste gas into the surround environment, so as to render a final gas mixture nonflammable. Similar to the embodiment in FIG. 11C, in the embodiment in FIG. 1 IF, the perfusate solution 1131 can flow in either pumping direction depending on system design.

[0210]

[0200] FIG. 11H depicts and embodiment of an ex-vivo perfusion system 1140 for safely administering dissolved hydrogen to the organ in the perfusion system by means of a gas exchanger and a gas separator which combine to minimize or eliminate discharge of hydrogen gas to the ambient environment. Ex-vivo perfusion system 1140 is similar to ex-vivo perfusion system 1130 of FIG. 1 IF, with the only exceptions being the connection of the gas exchanger outlet gas stream comprising waste gas 1137 to a waste gas recycling system comprising a gas separator 1144, a final waste gas stream 1148 and a gas recycling stream 1149. The functions and interactions of the organ 1103, perfusate solution 1131, pump 1132, gas exchanger 1134, and gas sources 1135A and 1135B are as described in FIG. 1 IF.

[0211]

[0201] As illustrated in FIG. 11H, the waste gas 1137 from the gas exchanger 1134 flows through a gas separator 1144. The waste gas 1137 from the gas exchanger 1134 may comprise a mixture or subset of carbon dioxide, water vapor, hydrogen, oxygen, or nitrogen gases. The gas separator 1144 is configured to recover all or a substantial portion of the hydrogen gas from the gas exchanger waste gas 1137 and return the recovered hydrogen gas via the gas recycling stream 1149 to the gas inlet of the gas exchanger 1134. The amount or concentration of hydrogen in the final waste gas stream 1148 is thereby reduced to sub-flammable concentrations of less than 4% hydrogen for immediate discharge to atmosphere, or it may be reduced in hydrogen concentration sufficient to facilitate subsequent and final disposition by any of the dilution or catalytic combustion processes previously disclosed. In addition, the recirculation of the recovered hydrogen gas via the gas recycling stream 1149 lowers the total amount of hydrogen needed by the system to maintain a desired concentration of hydrogen in the perfusate solution 1131 even as the metabolic activity of the organ 1103 consumes oxygen from the supply stream of oxygen gas 1136B and adds carbon dioxide gas to the waste gas 1137 of the gas exchanger 1134.

[0212]

[0202] In some embodiments the gas separator 1144 recovers both hydrogen and oxygen from the gas exchanger waste gas 1137 and recycles them to the gas exchanger 1134 by the gas recycling stream 1149. A simultaneous recycling of hydrogen and oxygen gases may benefit the ex-vivo perfusion system 1140 by minimizing the total usage of both oxygen and hydrogen gases.

[0213]

[0203] It is anticipated that embodiments containing mixtures of hydrogen and oxygen gases in any of the illustrated gas handling elements (e.g., gas exchanger 1134, gas inlets 1136A,B, gas exchanger waste gas 1137, gas separator 1144, gas recycling stream 1149) would seek to minimize the total volume of mixed gases contained within their enclosed spaces.

[0214]

[0204] The gas separator 1144 of FIG. 11H may utilize various gas separation technologies to accomplish recovery of hydrogen gas. In an embodiment, the gas separator 1144 is configured primarily to remove carbon dioxide waste from the oxygenator waste gas 1137 by adsorption of carbon dioxide with a carbon dioxide sorbent or adsorbent such as activated carbon, zeolite, metal organic framework, or an alkaline chemical scrubber comprising some combination of sodium hydroxide, calcium hydroxide, and lithium hydroxide. After carbon dioxide adsorption, the remaining gases comprising a mixture or subset of hydrogen, oxygen, nitrogen, and water vapor are directed back to the gas exchanger as the gas recycling stream 1149.

[0215]

[0205] In another embodiment of FIG. 11H, the gas separator 1144 may comprise one or more pressure swing adsorption (PSA) systems to selectively divert carbon dioxide or nitrogen or a mixture of both to the final waste gas stream 1148 while the hydrogen or oxygen or a mixture of both are recycled via the gas recycling stream 1149.

[0216]

[0206] In another embodiment of FIG. 11H, the gas separator 1144 may comprise chemically selective diffusion membranes to recover hydrogen gas or a combination of gases including hydrogen for recirculation via the gas recycling stream 1149.

[0217] Example 4. Protocol for on-demand hydrogen infusion into saline and administration for treatment.

[0218]

[0207] The purpose of this example is to provide an exemplary protocol for hydrogen gas infusion into a solution and the downstream administration to an organ or tissue.

[0208] Use of ex-vivo perfusion machines or whole body circulatory support for organ recovery and transport, as described above, may present multiple opportunities to administer hydrogen- infused solutions into the organ(s), with the ideal hydrogen administration steps happening prior to steps that reintroduce oxygen to the organ(s) either by oxygenated perfusate solution or by oxygenated blood.

[0219]

[0209] FIGs 12A and 12B describe an exemplary embodiment 1200 and process 7000, respectively, for administration of hydrogen during recovery of a donated heart after a prolonged period of warm ischemia. Hearts recovered after warm ischemic periods exceeding 3-5 minutes can typify the need for hydrogen administration at more than one step. The first occurrence of hydrogen administration may be as a cold, hydrogen-infused flushing solution that can be used to flush and cool the heart during or immediately after removal from the donor body. This first hydrogen administration serves the purpose to infuse the antioxidant (hydrogen) into the cardiac tissue before reintroducing any oxygen back into the tissue, regardless of the oxygen source. The sources of oxygen that can initiate free radical production at this early point in the organ recovery process may come from donor blood pumped and oxygenated by extracorporeal circulatory support, the typical organ flushing solutions themselves (having equilibrated with 160 mm Hg atmospheric oxygen) or the oxygen-enriched perfusate solutions used in ex-vivo oxygenated perfusion systems.

[0220]

[0210] After one to six hours of oxygenated perfusion, the concentration of hydrogen in the heart and in the perfusate can decrease to 40 ppb or less due to diffusion of hydrogen out from the solution through the gas exchanger membrane and into the waste gas stream. A second administration of hydrogen to the heart can be beneficial prior to the second occurrence of reperfusion, which occurs with the flow of the patient’s oxygenated blood into the organ after the several minutes of warm ischemia imposed by surgical engraftment of the organ.

[0221] [2H] The second occurrence of hydrogen administration to the heart can utilize the gas exchanger and the recirculating perfusate solution to deliver dissolved hydrogen to the heart prior to the ischemic interval of the engraftment surgery. Hydrogen concentrations in the cardiac tissues would remain steady enough through the ischemic engraftment period to provide protection against reperfusion injury in the first 5 to 10 minutes of reperfusion with oxygenated blood.

[0222]

[0212] The preparation step for the first hydrogen administration is illustrated in FIG 12A. A full solution bag 1201 of flushing solution is suspended above a gas exchanger 1202, with a sterile, empty solution bag 1203 positioned below the gas exchanger. A lumen connects the port of the full solution bag 1201 to the fluid inlet of the gas exchanger 1202. A lumen connects the fluid outlet of the gas exchanger 1202 to the port of the empty solution bag 1203. An electrolytic hydrogen gas source 1204 is connected to the gas inlet of the gas exchanger 1202. A flexible reservoir 1205 containing 15 liters of nitrogen gas for dilution is connected to the gas outlet of the gas exchanger 1202. The flushing solution flows via force of gravity from the full solution bag 1201 through the gas exchanger to the empty solution bag 1203. Simultaneous with the downward flow of solution through the gas exchanger, the hydrogen gas flows in the opposite direction (upward) from a hydrogen gas source 1204 through the gas exchanger 1202 to the nitrogen-filled gas dilution reservoir 1205. The waste gas flowing to 1205 is diluted harmlessly into the flexible volume of nitrogen dilution gas. The solution flowing from the gas exchanger into the solution bag 1203 contains the full 1 liter of flushing solution, now having a hydrogen concentration of 1550 ppb and an oxygen partial pressure less than 40 mm Hg. The above procedure can be repeated for each liter of hydrogen-rich flushing solution that is needed. After infusing hydrogen into the flushing solutions they were stored at 4 deg. C for five hours until used for administration to the donated heart in the experiment described in FIG 12B.

[0223]

[0213] In FIG. 12B, process 7000 describes an experiment carried out by the inventor to observe the effects of dissolved hydrogen on the performance of ischemic pig hearts. Steps 7001 -7003 comprised steps for intentionally stressing an organ for simulation purposes. In step 7001, the porcine heart donor was anesthetized generally and prepared for surgical recovery of the heart. After administering systemic heparin to prevent blood clotting, in step 7002, the test subject was euthanized by injection of Euthasol, followed by step 7003 comprising waiting 30 minutes after cardiac arrest for ensuring a stressed organ.

[0224]

[0214] Steps 7004-7010 comprises specific steps for applying hydrogen-rich solution to the organ. In step 7004, the heart was surgically recovered from the subject and its aorta was cannulated to receive a cold, hydrogen-rich flushing solution. Del Nido cardioplegic solution was used for delivery of hydrogen to the heart due to the widespread use of Del Nido solution in clinical cardiac donor organ recovery.

[0225]

[0215] In step 7005, 2 liters of hydrogen-rich Del Nido cardioplegia solution were flushed through the heart over a period of four minutes. The cardioplegia solution simultaneously cooled the heart and infused hydrogen into the cardiac tissues. The cardioplegia solution comprised a hydrogen concentration of about 1550 ppb, and oxygen partial pressure < 20 mm Hg, and a temperature of about 4 °C.

[0226]

[0216] In step 7006, the cooled and hydrogen-infused heart was installed into a system for ex-vivo hypothermic oxygenated perfusion of hearts. The coronary arteries of the heart were fluidly connected to the perfusion solution flow path by the aortic cannula. Cold Belzer’s Machine Perfusion Solution (MPS, 4 °C) was used as the perfusate solution.

[0217] In step 7007, the heart was perfused in the ex-vivo hypothermic oxygenated perfusion system at 6 - 8 °C for six hours to simulate a longer-than-average timespan for transport of donated hearts. During perfusion, pure oxygen gas flowed into the gas exchanger to infuse the perfusate solution with oxygen to support the aerobic oxygen demand of the cardiac tissue. Simultaneous to the oxygen gas dissolving into the perfusate, the dissolved hydrogen gas in the cardiac tissue was diffusing from the tissue into the recirculating perfusate, and from the perfusate into the waste gas stream emitting from the gas outlet of the gas exchanger. The waste gas stream was continuously diluted with a flow of nitrogen gas to reduce hydrogen gas concentration to less than 1% by volume before discharging the waste gas to room air. Three hours after starting oxygenated perfusion, the hydrogen concentration in the waste gas from the gas exchanger was below the sensitivity of the gas measurement instrument.

[0227]

[0218] In step 7008, after six hours of ex-vivo hypothermic oxygenated perfusion, the oxygen gas source to the gas exchanger was shut off and a hydrogen gas source was connected to the gas exchanger inlet for five minutes. The perfusate pump continued to recirculate perfusate solution through the heart as before, and the pure hydrogen gas flowed into the gas inlet of the gas exchanger at a rate of 10 ml / min. The waste gas mixture from the outlet of the gas exchanger contained a mixture of hydrogen and oxygen and was diluted with a continuous stream of nitrogen gas as before, i.e., through a dynamic dilution chamber prior to discharge to ambient air. After five minutes of hydrogen infusion, the hydrogen concentration in the perfusate coming out of the heart was measured at 400 ppb.

[0228]

[0219] The perfusion pump was stopped and the heart was disconnected from the ex-vivo perfusion system. In step 7009, the heart was connected to a benchtop ex-vivo normothermic reperfusion apparatus, to reperfuse and reanimate the beating heart with warm, oxygenated blood. Finally, in step 7010, cardiac output and the myocardial contractility relaxation rates of the myocardium were monitored continuously for two hours after reperfusion to assess the effects of the profoundly ischemic organ recovery conditions on the viability of the heart. Myocardial contractility at resuscitation after using standard cardioplegia and perfusion solutions without hydrogen resulted in about 50% of contractility measured at baseline, while myocardial contractility at resuscitation after using hydrogen-infused cardioplegia and hydrogen-infused perfusion solutions resulted in about 100% of baseline measurements.

[0229] DEFINITIONS

[0230]

[0220] As used herein, “about” and its grammatical equivalents in relation to a reference numerical value and its grammatical equivalents as used herein can include a range of values plus or minus 10% from that value. For example, the amount “about 10” encompasses amounts from 9 to 11. The term “about” in relation to a reference numerical value can also include a range of values plus or minus 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% from that value.

[0231]

[0221] As used herein, a “cell” refers to a biological cell. Some non-limiting examples include: a prokaryotic cell, eukaryotic cell, a bacterial cell, an archaea cell, a cell of a single-cell eukaryotic organism, a protozoa cell, a cell from a plant, an algal cell, a fungal cell, a fungal protoplast cell, an animal cell, and the like. Sometimes a cell is not originating from a natural organism, e.g., a cell can be a synthetically made, sometimes termed an artificial cell.

[0232]

[0222] Although various features of the disclosure may be described in the context of a single embodiment, the features can also be provided separately or in any suitable combination. Conversely, although the disclosure may be described herein in the context of separate embodiments for clarity, various aspects and embodiments can be implemented in a single embodiment.

[0233]

[0223] While exemplary embodiments of the present disclosure have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the present disclosure. It should be understood that various alternatives to the embodiments described herein, or combinations of one or more of these embodiments or aspects described therein may be employed in practicing the present disclosure. It is intended that the following claims define the scope of the present disclosure and that methods and structures within the scope of these claims and their equivalents be covered thereby.

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A system for hydrogen gas infusion into an organ or tissue comprising: a fluid circuit configured to circulate a medical solution through an organ or tissue; wherein the fluid circuit includes a perfusion solution, the perfusion solution being infused with a dissolved hydrogen gas concentration of 0.25 - 2.5 mM, and a gas exchanger.

2. The system of claim 1, further comprising a source of hydrogen gas including a 0.1 to 5 liter volume of aqueous solution having dissolved hydrogen gas.

3. The system of claim 1, further comprising a waste gas dilution module in fluid communication with and downstream of the gas exchanger; wherein the gas dilution module is configured to dilute waste hydrogen gas from the gas exchanger without allowing the waste hydrogen gas to reach a combustible concentration.

4. The system of claim 1, further comprising a waste gas recycling system for recovery of at least a portion of hydrogen gas from a gas outflow of the gas exchanger and for recycling of recovered hydrogen gas into a gas inlet of the gas exchanger.

5. The system of claim 4, wherein the waste gas recycling system comprises a carbon dioxide sorbent.

6. The system of claim 4, wherein the waste gas recycling system comprises a pressure swing adsorption system.

7. The system of claim 4, wherein the waste gas recycling system comprises one or more selectively gas-permeable membranes.

8. The system of claim 1, the system further comprising, an oxygen source; a gas exchanger arranged within the fluid circuit; and an isolation valve including a controller, the controller configured to actuate the isolation valve to prevent oxygen from flowing through the gas exchanger simultaneously with the perfusion solution infused with hydrogen.

9. The system of claim 1, further comprising a reservoir, in fluid communication with the fluid circuit, for containing a hydrogen-infused flushing solution configured to be introduced into the fluid circuit.

10. The system of claim 9, further comprising a heating or cooling element configured to regulate a temperature of the hydrogen-infused flushing solution.

11. A method of treating or reducing a risk of an injury due to reactive oxygen species and / or free radicals in blood, an organ or a tissue, the method comprising: administering a hydrogen-infused solution to the blood, organ or tissue at a concentration and a rate sufficient to reduce the reactive oxygen species and / or free radicals in the blood, organ or tissue, wherein the hydrogen-infused solution comprises hydrogen gas at a concentration of about 0.25 - 2.5 mM.

12. The method of claim 11, further comprising determining that the blood, organ or tissue has or is at risk of an injury due to the reactive oxygen species and / or free radicals in the blood, organ or tissue based on an amount of the reactive oxygen species and / or free radicals in the blood, organ or tissue as compared to a normal blood, organ or tissue.

13. The method of claim 11, wherein the hydrogen-infused solution is obtained by a method of infusing hydrogen gas, the method comprising: receiving hydrogen gas and a solution into a fluid reservoir; and infusing the solution with the hydrogen gas to produce a hydrogen-infused solution and a stream of excess hydrogen gas.

14. The method of claim 13, further comprising diluting the excess hydrogen gas to a desired safe disposal concentration after the infusing step.

15. The method of claim 14, wherein the desired safe disposal concentration is less than 4.5% by volume.

16. The method of claim 14, wherein the desired safe disposal concentration is less than 2% by volume.

17. The method of claim 13, wherein the infusing step comprising applying a positive pressure to force the hydrogen gas into the solution through a gas exchanger.

18. The method of claim 17, wherein the solution is driven to flow through the gas exchanger using gravity or a circulation pump.

19. The method of claim 18, further comprising a recovery of hydrogen gas from a gas outflow of the gas exchanger and recirculation of the recovered hydrogen gas to a gas inlet of the gas exchanger.

20. The method of claim 14, wherein the diluting comprises conveying the stream of the excess hydrogen gas to a static gas dilution module, the static gas dilution module comprising a dilution chamber that is preloaded with a volume of diluent gas.

21. The method of claim 20, wherein the volume of diluent gas is of sufficient quantity to render a final gas mixture nonflammable.

22. The method of claim 20, wherein the gas dilution chamber includes an electrically conductive material configured to prevent electrostatic discharge within the chamber.

23. The method of claim 20, wherein the volume of diluent gas is at least 10-fold higher than a total amount of the excess hydrogen gas.

24. The method of claim 20, wherein the gas dilution chamber includes a gas permeable elastomer configured to allow for diffusion of the diluent gas and excess hydrogen gas into ambient atmosphere.

25. The method of claim 14, wherein the diluting comprises conveying the stream of the excess hydrogen gas to a dynamic gas dilution module, the dynamic gas dilution module comprising a dilution chamber in fluid communication with the fluid reservoir and a dynamic flow source, wherein the dilution chamber is configured to receive the excess hydrogen gas from the fluid reservoir.

26. The method of claim 13, further comprising degassing the solution prior to the infusing, wherein the degassing comprises flowing the solution through a gas exchanger and applying a negative pressure to the gas exchanger to extract gases from the solution.

27. The method of claim 13, wherein the hydrogen gas is generated by an electrolysis reaction, wherein the electrolysis reaction comprises separating water into hydrogen and oxygen gases.

28. The method of claim 11, wherein the hydrogen-infused solution is administered to the blood, organ, or tissue by injecting the hydrogen-infused solution into a perfusion circuit of an ex-vivo organ perfusion system.

29. The method of claim 28, in which a flow of gas through a gas exchanger of the ex-vivo organ perfusion system is arrested prior to and during injection of the hydrogen-infused solution into a perfusate flow path.

30. A method of treating or reducing a risk of an injury due to reactive oxygen species and / or free radicals in blood, an organ or a tissue, the method comprising: administering a hydrogen-infused solution to the blood, organ or tissue at a concentration and a rate sufficient to reduce the reactive oxygen species and / or free radicals in the blood, organ or tissue; and contacting the blood, organ or tissue with oxygen gas so as to reduce damage to cells in the blood, organ or tissue.

31. The method of claim 30, wherein the hydrogen-infused solution comprises hydrogen gas at a concentration of about 0.25 - 2.5 mM.

32. The method of claim 30, wherein the injury is related to ischemia or hypoxia.

33. The method of claim 30, further comprising determining that the blood, organ or tissue has or is at risk of an injury due to the reactive oxygen species and / or free radicals in the blood, organ or tissue based on an amount of the reactive oxygen species and / or free radicals in the blood, organ or tissue as compared to a normal blood, organ or tissue.

34. The method of claim 30, wherein the hydrogen-infused solution is obtained by a method of infusing hydrogen gas, the method comprising: receiving hydrogen gas and a solution into a fluid reservoir; and infusing the solution with the hydrogen gas to produce a hydrogen-infused solution and a stream of excess hydrogen gas.

35. The method of claim 34, further comprising diluting the excess hydrogen gas to a desired safe disposal concentration after the infusing.

36. The method of claim 35, wherein the desired safe disposal concentration is less than 4.5% by volume.

37. The method of claim 35, wherein the desired safe disposal concentration is less than 2% by volume.

38. The method of claim 35, wherein the diluting comprises conveying the stream of the excess hydrogen gas to a static gas dilution module, the static gas dilution module comprising a dilution chamber that is preloaded with a volume of diluent gas.

39. The method of claim 38, wherein the volume of diluent gas is of sufficient quantity to render a final gas mixture nonflammable.

40. The method of claim 38, wherein the volume of diluent gas is at least 10-fold higher than a total amount of the excess hydrogen gas.

41. The method of claim 35, wherein the diluting comprises conveying the stream of the excess hydrogen gas to a dynamic gas dilution module, the dynamic gas dilution module comprising a dilution chamber in fluid communication with the fluid reservoir and a dynamic flow source, wherein the dilution chamber is configured to receive the excess hydrogen gas from the fluid reservoir.

42. The method of claim 34, wherein the infusing comprising applying a positive pressure to force the hydrogen gas into the solution through a gas exchanger.

43. The method of claim 42, wherein the solution is driven to flow through the gas exchanger using gravity or a circulation pump.

44. The method of claim 34, further comprising degassing the solution prior to the infusing, wherein the degassing comprises flowing the solution through a gas exchanger and applying a negative pressure to the gas exchanger to extract gases from the solution.

45. The method of claim 34, wherein the hydrogen gas is generated by an electrolysis reaction, wherein the electrolysis reaction comprises separating water into hydrogen and oxygen gas using electric current.

46. The method of claim 30, further comprising, sensing a parameter of fluid exiting the blood, organ or tissue, the parameter being at least one of temperature, flow rate, dissolved oxygen concentration, oxidation reduction potential(ORP), and dissolved hydrogen concentration; comparing the sensed parameter with predetermined set point values; and adjusting the flow rate or infusion duration of the hydrogen-infused solution to the blood, organ or tissue as a function of the comparing step.

47. The method of claim 46, wherein the sensed parameter is dissolved hydrogen concentration and the set point value is 250 mm Hg.

48. The method of claim 46, wherein the sensed parameter is dissolved hydrogen concentration and the set point value is 160 mm Hg.

49. The method of claim 46, wherein the sensed parameter is dissolved hydrogen concentration and the set point value is 40 mm Hg.

50. A system for hydrogen gas infusion into a solution comprising: a hydrogen gas source for producing hydrogen gas; a fluid reservoir containing a medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate the hydrogen gas with the medical solution to produce a hydrogen-infused solution with a hydrogen gas concentration of about 0.25 - 2.5 mM; and a gas dilution module in fluid communication with and downstream of the fluid reservoir; wherein the gas dilution module is configured to continuously receive and dilute excess hydrogen gas from the fluid reservoir without allowing the excess hydrogen gas to reach a combustible concentration.

51. The system of claim 50, further comprising a gas exchanger in fluid communication with at least the hydrogen gas source and the fluid reservoir, wherein the gas exchanger is configured to infuse the hydrogen gas into the medical solution to equilibrium.

52. The system of claim 50, wherein the gas dilution module is configured to discharge a gas mixture having 4.5% or less hydrogen gas by volume.

53. The system of claim 50, wherein the hydrogen gas source comprises an electrolysis cell configured to produce hydrogen gas and oxygen gas.

54. The system of claim 50, wherein the hydrogen gas is dissolved within the hydrogen- infused solution.

55. The system of claim 50, wherein the hydrogen gas in the hydrogen-infused solution is not in a gaseous phase.

56. The system of claim 50, wherein the medical solution comprises saline, organ preservation solution, whole blood, partial blood, cardioplegia solution, drugs, anticoagulants, antimicrobials, Prolyl hydroxylase domain (PHD) inhibitors, valproic acid, impermeants, thrombolytics, or a combination thereof.

57. The system of claim 50, wherein the gas dilution module comprises a static gas dilution module, the static gas dilution module comprising a dilution chamber that is preloaded with a volume of diluent gas.

58. The system of claim 57, wherein the volume of diluent gas is of sufficient quantity to render a final gas mixture nonflammable.

59. The system of claim 57, wherein the volume of diluent gas is at least 10-fold higher than a total amount of the excess hydrogen gas.

60. The system of claim 57, wherein the dilution chamber is surrounded by a gas-permeable material configured to allow for slow diffusion of the excess hydrogen gas to an external environment over a predetermined period of time.

61. The system of claim 57, further comprising a safety shutoff circuit coupled to at least the hydrogen gas source and the dilution chamber, wherein the safety shutoff circuit is configured to shut off or limit a rate of production of the hydrogen gas from the hydrogen gas source based at least on a volume of diluent gas in the dilution chamber.

62. The system of claim 50, wherein the gas dilution module comprises a dynamic gas dilution module comprising a dilution chamber in fluid communication with the fluid reservoir and a dynamic flow source, wherein the dilution chamber is configured to receive the excess hydrogen gas from the fluid reservoir.

63. The system of claim 62, wherein the dynamic flow source comprises a mechanism for conveying a flowing diluent gas into the dilution chamber to mix with the excess hydrogen gas at a ratio of at least 10: 1, wherein the flowing diluent gas comprises one or more of nitrogen, carbon dioxide, or ambient air.

64. The system of claim 63, further comprising a safety shutoff circuit coupled to at least the hydrogen gas source and the dilution chamber, wherein the safety shutoff circuit is configured to shut off or limit a rate of production of the hydrogen gas from the hydrogen gas source based atleast on whether the dynamic flow source is diffusing flowing diluent gas at a flow rate at least 10-fold higher than the rate of production of the hydrogen gas.

65. A system for hydrogen gas infusion into a medical solution comprising: a hydrogen gas source; a fluid reservoir containing a medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate hydrogen gas with the medical solution to produce a hydrogen-infused solution with a hydrogen gas concentration of about 0.25 - 2.5 mM; and a catalytic combustion module in fluid communication with and downstream of the fluid reservoir; wherein the catalytic combustion module is configured to continuously receive and convert excess hydrogen gas from the fluid reservoir into water without allowing the excess hydrogen gas to reach a combustible concentration outside of the catalytic combustion module or in a gas stream discharged to surrounding environment.

66. The system of claim 65, wherein the catalytic combustion module comprises an inlet stream of oxygen gas or air, a catalyst for promoting recombination of the excess hydrogen gas with the oxygen gas or air to produce water vapor, and one or more outlet for discharging a waste mixture of one or more of the excess hydrogen gas, the oxygen gas or air, and the water vapor.

67. The system of claim 66, wherein the catalyst comprises a platinum group metal.

68. The system of claim 66, wherein the inlet stream is in fluidic communication with the hydrogen gas source.

69. The system of claim 66, further comprising a safety shutoff circuit coupled to at least the hydrogen gas source and the catalytic combustion module, wherein the safety shutoff circuit is configured to shut off or limit a rate of production of the hydrogen gas from the hydrogen gas source based at least on whether a concentration of the hydrogen gas in the waste mixture is less than 2% by volume.

70. A system for hydrogen gas infusion into a medical solution comprising: a hydrogen gas source configured to produce or dispense hydrogen gas; and a degassed medical solution, the degassed medical solution being in fluid communication with the hydrogen gas source so as to receive and dissolve the hydrogen gas into the degassed medical solution to produce a hydrogen-infused solution at equilibrium hydrogen partial pressures ranging from about 230 mm Hg to about 2300 mm Hg.

71. The system of claim 70, wherein the degassed medical solution comprises partial pressures of all dissolved gases adding to a total partial pressure ranging from 0 mm Hg to 380 mm Hg.

72. The system of claim 70, wherein the degassed medical solution is degassed of one or more of N2, or O2 gas.

73. The system of claim 70, further comprising a degassing module for degassing the medical solution to produce the degassed medical solution, wherein the degassing module comprises at least a vacuum pump, a gas exchanger, and at least one valve connected to at least the hydrogen gas source, the vacuum pump, and the gas exchanger.

74. The system of claim 73, wherein the at least one valve is configured to switch between a first configuration and a second configuration, wherein the first configuration comprises blocking the hydrogen gas source from the at least one valve such that the vacuum pump is in fluid communication with the gas exchanger to allow for degassing of the medical solution in the gas exchanger, and wherein the second configuration comprises blocking the vacuum pump from the at least one valve such that the hydrogen gas source is in fluid communication with the gas exchanger to allow for infusing the hydrogen gas into the medical solution in the gas exchanger to produce the hydrogen-infused solution.

75. The system of claim 73, wherein the gas exchanger comprises a gas permeable membrane, a hollow-fiber gas exchanger, or a bubble gas exchanger.

76. A system for hydrogen gas infusion into a medical solution comprising: a hydrogen gas source configured to produce or dispense hydrogen gas; and a fluid reservoir containing a degassed medical solution, the fluid reservoir being in fluid communication with and downstream of the hydrogen gas source so as to receive and equilibrate the hydrogen gas with the degassed medical solution to produce a hydrogen-infused solution with a hydrogen gas concentration of about 0.25 - 2.5 mM.

77. The system of claim 76, wherein the degassed medical solution is degassed of one or more of N2, or O2 gas.

78. The system of claim 76, further comprising a degassing module for degassing the medical solution to produce the degassed medical solution, wherein the degassing module comprises at least a vacuum pump, a gas exchanger, and at least one valve connected to at least the hydrogen gas source, the vacuum pump, and the gas exchanger.

79. The system of claim 78, wherein the at least one valve is configured to switch between a first configuration and a second configuration, wherein the first configuration comprises blocking the hydrogen gas source from the at least one valve such that the vacuum pump is in fluidcommunication with the gas exchanger to allow for degassing of the medical solution in the gas exchanger, and wherein the second configuration comprises blocking the vacuum pump from the at least one valve such that the hydrogen gas source is in fluid communication with the gas exchanger to allow for infusing the hydrogen gas into the medical solution in the gas exchanger to produce the hydrogen-infused solution.

80. The system of claim 78, wherein the gas exchanger comprises a gas permeable membrane, a hollow-fiber gas exchanger, or a bubble gas exchanger.

81. The system of claim 78, wherein the degassing module further comprises a circulation pump in fluid communication with at least the fluid reservoir and the gas exchanger, the circulation pump configured to apply pressure for continuously circulating the medical solution from the fluid reservoir through the gas exchanger.

82. The system of claim 78, wherein the degassing module further comprises a second fluid reservoir, wherein a fuller of the fluid reservoir and the second fluid reservoir is elevated relative to the gas exchanger and an emptier of the fluid reservoir and the second fluid reservoir is lowered relative to the gas exchanger to allow the medical solution to flow through the gas exchanger due to gravity.

83. The system of claim 76, further comprising an ultrasonic bath, wherein the fluid reservoir is immersed in the ultrasonic bath, wherein the ultrasonic bath is allowed to apply ultrasonic energy for a desired degassing period such that gases in the medical solution are forced to migrate to an upper portion of the fluid reservoir.

84. The system of claim 83, wherein the gases are dissolved.

85. The system of claim 83, wherein the desired degassing period ranges from about 1 minutes to about 120 minutes.

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