WO2022167797A1

WO2022167797A1 - Vaping e-liquid composition and use thereof

Info

Publication number: WO2022167797A1
Application number: PCT/GB2022/050279
Authority: WO
Inventors: Christopher Barry Wood; Zuhal RAHMANI; Hugh Semple Munro
Original assignee: Thirty Respiratory Limited
Priority date: 2021-02-04
Filing date: 2022-02-03
Publication date: 2022-08-11
Also published as: GB202101543D0

Abstract

An aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder, the method including operating an e-cigarette or vaping device including an aqueous e-liquid composition and wherein the e-liquid composition comprises one or more nitrite salt.

Description

Vaping e-liquid composition and use thereof

The present invention relates to an aqueous vaping e-liquid composition for use in an vaping device, the e-liquid composition comprising water and a nitrite salt, and use thereof, in particular for the treatment or prevention of a respiratory disease or disorder.

Background

The popularity of vaping devices (commonly referred to as electronic cigarettes or e- cigarettes and) has risen in the last ten years as an alternative to smoking conventional tobacco cigarettes. Vaping devices typically include an atomizer, a power source such as a battery, and a container such as a cartridge or tank. The atomizer is a heating element that atomizes a liquid solution called e-liquid. The e-liquid typically contains nicotine, for example, to assist smoking cessation.

Nitric oxide (NO) and nitric oxide precursors have been extensively studied as potential pharmaceutical agents. Nitric oxide is a potent vasodilator which is synthesised and released by vascular endothelial cells and plays an important role in regulating, inter alia, vascular local resistance and blood flow. In mammalian cells, nitric oxide is principally produced along with L-citrulline by the enzymatic oxidation of L-arginine. Nitric oxide is also released from the skin by a mechanism which appears to be independent of NO synthase enzyme. Nitric oxide is also involved in the inhibition of both platelet and leucocyte aggregation and adhesion, the inhibition of cell proliferation, the scavenging of superoxide radicals and the modulation of endothelial layer permeability. The role of nitric oxide in cancer treatment was discussed in Biochemistry (Moscow), 63(7), 802-809 (1998), the disclosure of which is incorporated herein by reference. Nitric oxide has been shown to possess antimicrobial properties, as reviewed by F C Fang in J. Clin. Invest. 99(12), 2818-2825 (1997) and as described for example in WO 95/22335 and WO 02/20026 (Aberdeen University), the disclosures of which are incorporated herein by reference. Other known uses and applications of systems for generation of nitric oxide, other oxides of nitrogen and precursors thereof are given below in the description of the present invention.

There remain substantial problems in connection with the efficient generation and delivery of nitric oxide, other oxides of nitrogen and precursors thereof to organisms and cells for treatment. A widely adopted system for the generation of nitric oxide relies on the acidification of nitrite salts using a mineral acid to produce initially nitrous acid (HNO2) in equimolar amounts in comparison with the starting nitrite, which nitrous acid then readily decomposes to nitric oxide and nitrate with hydrogen ions and water. The decomposition can be represented by the following balanced equation (1):

3 HNO₂ 2 NO + NOT + H⁺ + H₂O (1)

It has been conventional to perform the acidification of the nitrite at a pH of less than about 4, at which the formation of nitrous acid is generally favoured, in order to seek to maximise the yield of NO. However, the use of pH < 4 is not suitable for in vivo use where the acid is in contact with animal tissue. A higher pH would be more benign to cells and living systems, but at pH greater than 4 the prior systems have not produced satisfactory yields of NO. To seek to increase the amount of NO generated above pH 4, large quantities of nitrite are required, which is impractical in therapeutic applications and uneconomic. In addition, the conversion represented by Equation (1) is not readily controllable in view of the short half-life of nitrous acid, so that controlled release of nitric oxide for therapeutic use is difficult.

Summary of the Invention

The present invention is based on our surprising finding that nitric oxide, optionally other oxides of nitrogen and/or optionally precursors thereof (collectively referred to as NOx) can be generated using a vaping device containing an e-liquid composition, wherein the e-liquid composition includes a nitrite salt.

At its most general, the present invention relates to the generation of nitric oxide gas from a vaping device using an aqueous e-liquid composition comprising a nitrite salt. This has particular application in the treatment or prevention of respiratory diseases and disorders, in particular to respiratory diseases and disorders that are responsive to nitric oxide.

In a first aspect, the present invention provides an aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder, the method including operating a vaping device including the e-liquid composition and wherein the e-liquid composition comprises a nitrite salt. In a second aspect, the present invention provides a method of treatment or prevention of a respiratory disease or disorder, the method including operating a vaping device including an aqueous e-liquid composition and wherein the e-liquid composition comprises a nitrite salt.

In a third aspect, the present invention provides a vaping device containing an aqueous e-liquid composition, the e-liquid composition comprising a nitrite salt.

In a fourth aspect, the present invention provides an aqueous e-liquid composition for use in a vaping device, wherein the e-liquid composition comprises a nitrite salt.

In particular embodiments of the above aspects, the e-liquid composition further comprises one or more organic alcohols.

Detailed description

The term “vaping device” is used to refer to any device adapted to generate vapour and/or gas for inhalation by a subject and include, but are not limited to, electronic cigarettes and e-cigarettes.

The word “NOx” is used to refer to the products of generated from nitrite salts compositions, particularly nitric oxide, other oxides of nitrogen and precursors thereof both individually and collectively in any combination. It will be understood that each component of the generated NOx can be evolved as a gas, or can pass into solution in the reaction mixture, or can initially pass into solution and subsequently be evolved as a gas, or any combination thereof.

The features described below may be included in any one of the first to fourth aspects of the present invention as appropriate.

Aqueous e-liquid composition

The present invention uses an aqueous e-liquid composition. The e-liquid composition includes water and a nitrite salt. The nitrite salt is typically dissolved in the water of the composition. As such, the e-liquid composition may be referred to as an aqueous nitrite salt solution.

Nitrite salt

The choice of nitrite salt is not particularly limited. Specific examples of nitrite salts that may be used in the compositions of the present invention include alkali metal nitrites or alkaline earth metal nitrites. In some embodiments, the one or more nitrite salt is selected from LiNO₂, NaNO₂, KNO₂, RbNO₂, CsNO₂, FrNO₂, AgNO₂, Be(NO₂)₂, Mg(NO₂)₂, Ca(NO₂)₂, Sr(NO₂)₂, Mn(NO₂)₂, Ba(NO₂)₂, Ra(NO₂)₂ and any mixture thereof.

In particular embodiments, the nitrite salt is NaNO₂ or KNO₂. In one embodiment, the nitrite salt is NaNO₂.

The molarity of nitrite ion in the e-liquid composition may be in the range of about 0.001 M to about 5 M. In some embodiments, the molarity of nitrite ion in the e-liquid composition is in the range of about 0.01 M to about 2 M. In some embodiments, the molarity of nitrite ion in the e-liquid composition is in the range of about 0.05 M to about 1.6 M. In more particular embodiments, the molarity of nitrite ion in the e-liquid composition is in the range of about 0.1 M to about 1.2 M. In embodiments, the molarity of nitrite ion in the e-liquid composition can be in the range of 0.8 to 1 .2 M. For example, the molarity of nitrite ion in the e-liquid composition may be about 0.8 M, about 0.9 M, about 1.0 M, about 1.1 M or about 1.2 M. In alternative embodiments, the molarity of nitrite ion in the e-liquid composition can be in the range of 0.2 to 0.8 M. For example, the molarity of nitrite ion in the e-liquid composition may be about 0.2 M, about 0.3 M, about 0.4 M, about 0.5 M, about 0.6 M or about 0.7 M.

Water

Water is included the e-liquid composition. The water may be any form of water that is suitable for use in an e-liquid. In certain embodiments, the water is deionised water. Organic alcohol

In particular embodiments, the e-liquid composition further includes one or more organic alcohol. The expression “organic alcohol” herein refers to an organic molecule with one or more hydroxyl groups that is not a proton source, particularly for a nitrite salt reaction, and is not a saccharide or polysaccharide (the terms “saccharide” and “polysaccharide” include oligosaccharide, glycan and glycosaminoglycan). The organic alcohol will thus have a pKai of about 7 or greater, for example 7.0 or greater.

The organic alcohol may be cyclic or acyclic or may be a mixture of one or more cyclic organic polyol and one or more acyclic organic alcohol. For example, the one or more organic alcohol may be selected from one or more alkane substituted by one or more OH groups, one or more cycloalkane substituted by one or more OH groups, one or more cycloalkylalkane substituted by one or more OH groups, and any combination thereof. Most preferably the alkane, cycloalkane or cycloalkylalkane do not carry any substituents other than OH.

In particular embodiments, the e-liquid composition includes one or more organic polyol. The expression “organic polyol” herein refers to an organic molecule with two or more hydroxyl groups that is not a proton source, particularly for a nitrite salt reaction, and is not a saccharide or polysaccharide (the terms “saccharide” and “polysaccharide” include oligosaccharide, glycan and glycosaminoglycan). The organic polyol will thus have a pKai of about 7 or greater, for example 7.0 or greater.

The expressions “organic alcohol” and “organic polyol” herein preferably exclude reductants. In one embodiment of the invention in all its aspects, therefore the organic alcohol (e.g. the organic polyol) excludes reductants. Examples of reductants which are organic molecules with one or more hydroxyl groups and not a saccharide or polysaccharide are formic acid, thioglycerol (for example, 1 -thioglycerol), hydroquinone, butylated hydroquinone, ascorbic acid, ascorbate, erythorbic acid and erythorbate. Thioglycerol (for example, 1-thioglycerol), hydroquinone, butylated hydroquinone, ascorbate and erythorbate are thus preferably excluded from the expression “organic polyol” because they are reductants. Ascorbic acid and erythorbic acid are excluded from the expression anyway because they are proton sources, particularly for the nitrite salt reaction. In some embodiments, the one or more organic alcohol is one or more acyclic organic alcohol. The acyclic organic alcohol may be an acyclic organic alcohol having 2 to 12 carbon atoms. In particular embodiments, the acyclic organic alcohol is an acyclic organic alcohol having 2 to 4 carbon atoms and 1 to 3 hydroxy groups that are not a proton source. In more particular embodiments, the acyclic organic alcohol is selected from ethanol (ethyl alcohol), glycerol, propylene glycol and combinations thereof.

Alternatively, the one or more acyclic organic polyol may be selected from the sugar alcohols having 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. A preferred one or more acyclic organic polyol is selected from the alditols, for example the alditols having 4, 5,

6, 7, 8, 9, 10, 11 or 12 carbon atoms. It is preferred that the one or more organic polyol does not include a saponin, sapogenin, steroid or steroidal glycoside.

In other embodiments, the one or more organic alcohol is one or more cyclic organic alcohol. The cyclic organic alcohol may be a cyclic organic alcohol having 5 to 18 carbon atoms. In particular embodiments, the cyclic organic alcohol is a cyclic organic alcohol having 8 to 12 carbon atoms and 1 to 3 hydroxy groups that are not a proton source. In more particular embodiments, the cyclic organic alcohol is menthol.

The one or more cyclic organic polyol may be a cyclic sugar alcohol or a cyclic alditol. For example the one or more cyclic polyol may be a cyclic sugar alcohol having 4, 5, 6,

7, 8, 9, 10, 11 or 12 carbon atoms or a cyclic alditol having 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. A specific example of a cyclic polyol is inositol.

In some embodiments the one or more organic polyol has 7 or more hydroxy groups. In particular embodiments the one or more organic polyol is a sugar alcohol or alditol having 7 or more hydroxy groups. In more particular embodiments the one or more organic polyol has 9 or more hydroxy groups. In further embodiments the one or more organic polyol is a sugar alcohol or alditol having 9 or more hydroxy groups. In some embodiments the one or more organic polyol has 20 or fewer hydroxyl groups. In particular embodiments the one or more organic polyol is a sugar alcohol or alditol having 20 or fewer hydroxy groups. In more particular embodiments the one or more organic polyol has 15 or fewer hydroxyl groups. In further embodiments the one or more organic polyol a sugar alcohol or alditol having 15 or fewer hydroxyl groups. The one or more organic polyol may have a number of hydroxyl groups in the range of 7 to 20, more particularly in the range of 9 to 15. In certain embodiments the one or more organic polyol includes 9, 12, 15 or 18 hydroxy groups.

In some embodiments the one or more organic polyol is sugar alcohol compound comprising, for example consisting of, one or more monosaccharide units and one or more acyclic sugar alcohol units. The one or more organic polyol may be a sugar alcohol compound comprising, for example consisting of, a straight chain of one or more monosaccharide units and one or more acyclic sugar alcohol units or a branched chain of one or more monosaccharide units and one or more acyclic sugar alcohol units.

A monosaccharide unit as used herein refers to a monosaccharide covalently linked to at least one other unit (whether another monosaccharide unit or an acyclic sugar alcohol unit) in the compound. An acyclic sugar alcohol unit as used herein refers to an acyclic sugar alcohol linked covalently to least one other unit (whether a monosaccharide unit or another acyclic sugar alcohol unit) in the compound. The units in the compound may be linked through ether linkages. In some embodiments, one or more of the monosaccharide units are covalently linked to other units of the compound through a glycosidic bond. In particular embodiments, each of the monosaccharide units are covalently linked to other units of the compound through a glycosidic bond. In certain embodiments, the sugar alcohol compound is a glycoside with a monosaccharide or oligosaccharide glycone and an acyclic sugar alcohol aglycone.

The acyclic sugar alcohol units may be sugar alcohol units having 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. In particular embodiments the acyclic sugar alcohol unit is selected from the group consisting of units of erythritol, threitol, arabitol, xylitol, ribitol, mannitol, sorbitol, galactitol, fucitol, iditol and volemitol.

The one or more of the monosaccharide units may be a Cs or Ce monosaccharide unit. In other words, one or more of the monosaccharide units are a pentose or hexose unit. Each monosaccharide unit may be a Cs or Ce monosaccharide unit. The one or more of the sugar alcohol units may be a Cs or Ce sugar alcohol unit. Each sugar alcohol unit may be a Cs or Ce sugar alcohol unit.

The sugar alcohol compound may comprise, for example consist of, n monosaccharide units and m acyclic sugar alcohol units, where n is a whole number and at least one, m is a whole number and at least one and (n + m) is no more than 10. In certain embodiments the sugar alcohol compound comprises, for example consists of, a chain of n monosaccharide units terminated with one acyclic sugar alcohol unit, where n is a whole number between one and nine. In these embodiments, the chain of monosaccharide units may be covalently linked by glycosidic bonds. In particular embodiments, each monosaccharide unit is covalently linked to another monosaccharide unit or the acyclic sugar alcohol unit by a glycosidic bond. In certain embodiments the sugar alcohol compound comprises, for example consists of, a chain of 1 , 2 or 3 monosaccharide units terminated with one acyclic alcohol unit. 1 , 2, 3 or each monosaccharide unit may be a Cs or Ce monosaccharide unit. The acyclic alcohol unit may be a Cs or Ce sugar alcohol unit. Examples of the sugar alcohol compound include but are not limited to: isomalt, maltitol and lactitol (n = 1); maltotriitol (n = 2); and maltotetraitol (n = 3).

Such sugar alcohol compounds may be described as sugar alcohols derived from a disaccharide or an oligosaccharide. Oligosaccharide, as used herein, refers to a saccharide consisting of three to ten monosaccharide units. Sugar alcohols derived from disaccharides or oligosaccharides may be synthesised (e.g. by hydrogenation) from disaccharides, oligosaccharides or polysaccharides (e.g. from hydrolysis and hydrogenation), but are not limited to compounds synthesised from disaccharides, oligosaccharides or polysaccharides. For example, sugar alcohols derived from a disaccharide may be formed from the dehydration reaction of a monosaccharide and a sugar alcohol. The one or more organic polyol may be a sugar alcohol derived from a disaccharide, trisaccharide or tetrasaccharide. Examples of sugar alcohols derived from disaccharides include but are not limited to isomalt, maltitol and lactitol. An example of a sugar alcohol derived from a trisaccharide includes but is not limited to maltotriitol. An example of a sugar alcohol derived from a tetrasaccharide includes but is not limited to maltotetraitol.

As suitable organic alcohols there may be mentioned any selected from ethanol, glycerol, propylene glycol, menthol, linalool, maltol, erythritol, threitol, arabitol, xylitol, ribitol, mannitol, sorbitol, galactitol, fucitol, iditol, inositol, volemitol, isomalt, maltitol, lactitol, maltotriitol, maltotetraitol, polyglycitol, and any combination thereof. In particular, suitable organic alcohols may be any selected from ethanol, glycerol, propylene glycol, menthol, and combinations thereof. Many organic polyols contain one or more chiral centre and thus exist in stereoisomeric forms. All stereoisomeric forms and optical isomers and isomer mixtures of the organic polyols are intended to be included within the scope of this invention. In particular, the D and/or L forms of all chiral organic polyols and all mixtures thereof may be used.

The total amount of organic alcohol in the e-liquid composition may be in the range of 1% to 75% by weight based on the total weight of the e-liquid composition. In some embodiments, the total amount of polyol in the e-liquid composition is in the range of 5% to 15% by weight based on the total weight of the e-liquid composition. In certain embodiments, the total amount of polyol in the e-liquid composition is about 10 % by weight based on the total weight of the e-liquid composition. Alternatively, the total amount of polyol organic alcohol in the e-liquid composition may be in the range of 1% to 75% by volume based on the total volume of the e liquid composition. In some embodiments, the total amount of polyol in the e-liquid composition is in the range of 5% to 15% by volume based on the total volume of the e liquid composition. In certain embodiments, the total amount of polyol in the e-liquid composition is about 10 % by volume based on the total volume of the e liquid composition. pH of the composition

The pH of the e-liquid composition may be greater than about 6.0. A pH of lower than 6.0 is likely to generate NOx at times when it is not desired. In some embodiments, the pH is about 6.4 or greater, about 7.0 or greater or about 7.4 or greater. The pH may be in the range of about 6.0 to about 9.0, or about 6.4 to about 8.0 or about 7.0 to about 8.0.

Optional additional components

The e-liquid may further comprise one or more additional components. The one or more additional components may be one or more components typically used in e-liquids. Examples of additional components include, but are not limited to, one or more food grade flavourings and nicotine. In particular embodiments, the e-liquid composition is substantially free of nicotine.

Optional additional components may, for example, be selected from sweetening agents, taste-masking agents, thickening agents, viscosifying agents, wetting agents, lubricants, binders, film-forming agents, emulsifiers, solubilising agents, stabilising agents, colourants, odourants, salts, coating agents, antioxidants, pharmaceutically active agents and preservatives. Such components are well known in the art and a detailed discussion of them is not necessary for the skilled reader. Examples of auxiliary substances such as wetting agents, emulsifying agents, lubricants, binders, and solubilising agents include, for example, sodium phosphate, potassium phosphate, gum acacia, polyvinylpyrrolidone, cyclodextrrin derivatives, sorbitan monolaurate, triethanolamine acetate, triethanolamine oleate, and the like. A sweetening agent or a taste-masking agent may, for example, include a sugar, saccharin, aspartame, sucralose, neotame or other compound that beneficially affects taste, after-taste, perceived unpleasant saltiness, sourness or bitterness, that reduces the tendency of an oral or inhaled formulation to irritate a recipient (e.g. by causing coughing or sore throat or other undesired side effect, such as may reduce the delivered dose or adversely affect patient compliance with a prescribed therapeutic regimen). Certain tastemasking agents may form complexes with one or more of the nitrite salts. Examples of thickening agents, viscosifying agents and film-forming agents have been given above.

Vaping device

The vaping device used in the method disclosed herein is not particularly limited. The vaping device typically has an e-liquid chamber (or tank) for receiving the e-liquid composition. Vaping devices typically also include an atomizer and a power source such as a battery. The atomizer may be a heating element for atomizing the e-liquid composition.

Method of operating the vaping device

Methods of operating a vaping device in order to produce a vapour from an e-liquid are generally known. The method of operating the vaping device typically includes vaporising at least part of the aqueous e-liquid composition containing the nitrite salt. Such operation typically produces a vapour including at least nitric oxide (NO). Vaporising of the aqueous e-liquid composition containing the nitrite salt may occur by heating the aqueous e-liquid composition containing the nitrite salt. The vaping device may be operated by an operation means for generating vapour from the e-liquid composition. For example, the operation of the vaping device may be instigated and/or controlled by actuation of a power button. The operation of the vaping device may be stopped by release of the power button. Alternatively, operation of the vaping device may be instigated by a subject inhaling on the vaping device mouthpiece. The operation of the vaping device may be stopped by cessation of the inhalation.

The operation of the vaping device may occur for an operation time period. The operation time period is typically the time during which the user is operating the device to vaporize the e-liquid composition. The operation time period may be in the range of about one to about ten seconds. In some embodiments, the operation time period is in the range of about three to about seven seconds. In particular embodiments, the operation time period is in the range of about four to about six seconds, such as about five seconds.

The method of treatment or prevention of the present invention may involve a single treatment session. Alternatively, the method of treatment or prevention of the present invention may involve more than one treatment session. Each treatment session includes at least one operation of the vaping device. In some embodiments, each treatment session includes a single operation of the vaping device. Alternatively, each treatment session includes a plurality of operations of the vaping device in succession. In some embodiments, each treatment session includes two to ten operations, such as four to eight operations or five, six or seven operations of the vaping device in succession.

Typically, each operation is separated by a separation time period of at least about one second. In some embodiments, each operation is separated by a separation time period of at least about ten seconds. In particular embodiments, each operation is separated by a separation time period of at least about thirty seconds. In some embodiments, each operation is separated by a separation time period of less than about five minutes. In particular embodiments, each operation is separated by a separation time period of less than about two minutes. In certain embodiments, each operation is separated by a separation time period of less than about 90 seconds. The separation time period may be in the range of about one second to about five minutes, about ten seconds to about two minutes, or about thirty seconds to about 90 seconds. The method of treatment or prevention may include two or more treatment sessions as described herein. In some embodiments, the method of treatment or prevention includes at least two treatment sessions separated by a treatment session separation period. In other words, each treatment session may be separated by a treatment session separation period. The treatment session separation period may be at least about 12 hours. The treatment session separation period may be at least about 24 hours. The treatment session separation period may be at least about one week.

NOx generated by operation of the vaping device

Each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO of at least about 50 ppm. In some embodiments, each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO of at least about 200 ppm. In particular embodiments, each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO of at least about 500 ppm. In other embodiments, each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO of at least about 2000 ppm. In some embodiments, each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO in the range of about 50 ppm to about 10,000 ppm. In particular embodiments, each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO in the range of about 200 ppm to about 5,000 ppm. In certain embodiments, each operation of the vaping device containing the e-liquid composition described herein may generate a peak concentration of NO in the range of about 500 ppm to about 2,000 ppm.

Each treatment session may generate at least about 5,000 nmolnmol of NO in total. In some embodiments, each treatment session generates at least about 20,000 nmol of NO in total. In particular embodiments, each treatment session generates at least about 40,000 nmol of NO in total. In some embodiments, each treatment session generates in the range of about 5,000 to about 300,000 nmol of NO in total. In particular embodiments, each treatment session generates in the range of about 20,000 to about 100,000 nmol of NO in total. In certain embodiments, each treatment session generates in the range of about 40,000 to about 60,000 nmol of NO in total. Each treatment session may generate no more than about 50,000 nmol of NO2 in total. In some embodiments, each treatment session generates no more than about 5,000 nmol of NO2 in total. In particular embodiments, each treatment session generates no more than about 2,000 nmol of NO2 in total. In some embodiments, each treatment session generates in the range of about 0 to about 50,000 nmol of NO2 in total. In particular embodiments, each treatment session generates in the range of about 50 to about 5,000 nmol of NO2 in total. In certain embodiments, each treatment session generates in the range of about 100 to about 2,000 nmol of NO2 in total.

The percentage ratio of NO generated in each treatment session to NO2 generated in each treatment session (referred to herein as “the percentage ratio of NO to NO2”) may be about 15 % or less. In some embodiments, the percentage ratio of NO to NO2 is about 10 % or less. In particular embodiments, the percentage ratio of NO to NO2 is about 6 % or less. In more particular embodiments, the percentage ratio of NO to NO2 is about 5 % or less. In some embodiments, the percentage ratio of NO to NO2 is in the range of about 0 to about 15 %. In some embodiments, the percentage ratio of NO to NO2 is in the range of about 0.1 to about 10 %. In particular embodiments, the percentage ratio of NO to NO2 is in the range of about 0.5 % to about 6 %. In certain embodiments, the percentage ratio of NO to NO2 is in the range of about 1 % to 5 %. The percentage ratio of NO to NO2 is calculated by dividing the total molar amount of NO2 generated in the treatment session by the sum of the total molar amount of NO generated in the treatment session and the total molar amount of NO2 generated in the treatment session (and the figure is multiplied by 100 to be expressed as a percentage).

The peak concentration of NO and the cumulative amount of NOx (NO and NO2) generation may be measure by SIFT-MS.

Treatment or prevention of a respiratory disease or disorder

Many physiological effects of nitric oxide and nitric oxide generating compositions and medical treatments based thereon have been reported in the literature, and as a result many therapeutic treatments have been developed. A non-exhaustive list is provided on pages 57 to 61 of WO 2020/245574 as illustration and are incorporated herein by reference. The gas evolved from the operation of the vaping device containing the e-liquid composition may be used to treat or prevent diseases or disorders associated with the mucosae and tissues of the nose, mouth, respiratory tract and lungs.

The conditions treatable using the present invention include lung diseases such as viral infections for example influenza, SARS-CoV or SARS-CoV-2, pulmonary arterial hypertension, ischemic reperfusion injury of the heart, brain and organs involved in transplantation, chronic obstructive pulmonary disease (COPD) (particularly, emphysema, chronic bronchitis), asthma including severe asthma and viral and bacterial induced exacerbations of asthma and refractory (non-reversible) asthma, intra-nasal or pulmonary bacterial infections such as pneumonia, tuberculosis, nontuberculosis mycobacterial infections and other bacterial and viral lung infections, for example secondary bacterial infections following virus infections of the respiratory tract.

The property of nitric oxide to induce vasodilation characterises some of the treatments using e-liquid composition of the present disclosure and the gas evolved therefrom.

A particular example of diseases, disorders and conditions responsive to vasodilation includes, but is not limited to conditions associated with ischaemia.

Conditions associated with tissue ischaemia include Raynauld syndrome, severe primary vasospasm, and tissue ischaemia, for example tissue ischaemia caused by surgery, septic shock, irradiation or a peripheral vascular disease (for example diabetes and other chronic systemic disease).

When used in the treatment or prevention of conditions associated with tissue ischaemia as a result of surgery, the vaping device containing the e-liquid composition of the present disclosure may be operated to administer the evolved gas to a subject before, during or after the surgery. The evolved gas is typically administered to a subject by inhalation by the subject.

The operation of the vaping device containing the may be used in the treatment or prevention of ischemic reperfusion injury of an organ by administering the evolved gas from the vaping device according to the present disclosure to a lung (e.g. to treat or prevent ischemic reperfusion injury of the lung). The surgery may be the transplantation of an organ. Administration of the evolved gas may follow an ischemic episode or may be prophylactic.

In some embodiments, the respiratory disease or disorder may be associated with the presence of one or more microbes in the subject to be treated. In other words, the respiratory disease or disorder may be associated with one or more microbial infections in the subject. The gas evolved from operation of the vaping device containing the e- liquid composition may have a biocidal or biostatic effect on a potentially wide range of microorganisms, leading to many anti-microbial treatments. The microbes may, for example, be any one or more selected from bacterial cells, viral particles and/or fungal cells, or microparasites, and may be individual cells, organisms or colonies.

When the microbe is present in a bacterial infection, a fungal infection, viral or microparasitic infection of a human or other animal, the infection may, for example, be in the context of a disease such as the common cold, influenza, tuberculosis, SARS, COVID-19, pneumonia or measles.

Bacterial Cells

The bacterium may be a pathogenic bacterial species. The microbial infection may be an infection caused by a pathogenic bacterial species, including Gram positive and Gram negative, aerobic and anaerobic, antibiotic-sensitive and antibiotic-resistant bacteria.

Examples of bacterial species which may be targeted using the present invention include species of the Actinomyces, Bacillus, Bartonella, Bordetalla, Borrelia, Brucella, Campylobacter, Chlamydia, Chlamydophila, Clostridium, Corynebacterium, Enterococcus, Escherichia, Francisella, Haemophilus, Heliobacter, Legionella, Leptospira, Listeria, Mycobacterium, Mycoplasma, Neisseria, Pseudomonas, Rickettsia, Salmonella, Shigella, Staphylococcus, Streptococcus, Treponema, Ureaplasma, Vibrio, or Yersinia genera. Any combination thereof can also be targeted by the present invention.

In particular embodiments, the microbe may be a pathogenic species of Corynebacterium, Mycobacterium, Streptococcus, Staphylococcus, Pseudomonas or any combination thereof. In more particular embodiments, the microbe to be targeted can be selected from Actinomyces israelii, Bacillus anthracis, Bacteroides fragilis, Bordetella pertussis, Borrelia burgdorferi, Borrelia garinii; Borrelia afzelii; Borrelia recurrentis; Brucella abortus; Brucella canis; Brucella melitensis; Brucella suis; Campylobacter jejuni; Chlamydia pneumoniae; Chlamydia trachomatis; Chlamydophila psittaci; Clostridium botulinum; Clostridium difficile; Clostridium perfringens; Clostridium tetani; Corynebacterium diphtheria; Ehrlichia canis; Ehrlichia chaffeensis; Enterococcus faecalis; Enterococcus faecium; Escherichia coli, such as Enterotoxigenic E. coli (ETEC), Enteropathogenic E. coli, Enteroinvasive E.coli (El EC), and Enterohemorrhagic (EHEC), including E. coli O157:H7; Francisella tularensis; Haemophilus influenza; Helicobacter pylori; Klebsiella pneumoniae; Legionella pneumophila; Leptospira species; Listeria monocytogenes; Mycobacterium leprae; Mycobacterium tuberculosis; Mycobacterium abscessus;Mycobacterium ulcerans; Mycoplasma pneumoniae; Neisseria gonorrhoeae; Neisseria meningitides;

Pseudomonas aeruginosa; Nocardia asteroids; Rickettsia rickettsia; Salmonella typhi; Salmonella typhimurium; Shigella sonnei; Shigella dysenteriae; Staphylococcus aureus; Staphylococcus epidermidis; Staphylococcus saprophyticus; Streptococcus agalactiae; Streptococcus pneumoniae; Streptococcus pyogenes; Streptococcus viridans; Treponema pallidum subspecies pallidum; Vibrio cholera; Yersinia pestis; and any combination thereof.

In particular, the microbe may be selected from Chlamydia pneumoniae, Bacillus anthracis, Corynebacterium diphtheria, Haemophilus influenza, Mycobacterium leprae, Mycobacterium tuberculosis, Mycobacterium abscessus, Mycobacterium ulcerans, Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus pneumoniae, or any combination thereof.

The microbe may be an antibiotic-resistant or antibiotic-sensitive pathogenic bacterial species or an antibiotic-resistant or antibiotic-sensitive strain of a bacterial species. The use of nitric oxide to treat methicillin resistant Staphylococcus aureus (MRSA) and methicillin sensitive Staphylococcus aureus (MSSA) is described, for example, in WO 02/20026, the disclosure of which is incorporated herein by reference. An example of an antibiotic-resistant or antibiotic-sensitive pathogenic bacterial species which may be killed or treated using the present invention is thus methicillin resistant Staphylococcus aureus (MRSA) or methicillin sensitive Staphylococcus aureus (MSSA). Fungal Cells

The microbe may be a pathogenic fungal species. The microbial infection may be an infection caused by a pathogenic fungal species, including pathogenic yeasts.

Examples of fungal species which may be targeted using the present invention include species of Aspergillus, Blastomyces, Candida (for example Candida auris), Coccidioides, Cryptococcus (in particular, Cryptococcus neofromans or Cryptococcus gattii), Hisoplamsa, Murcomycetes, Pneumocystis (for example Pneumocystis jirovecii), Sporothrix, Talaromyces, or any combination thereof.

Examples of fungal infections include aspergillosis (such as allergic bronchia pulmonary aspergillosis), tinea pedis (athlete’s foot), infections caused by a pathogenic species of Candida, such as vaginal yeast infections, fungal toenail infections and diaper rash, tinea cruris (jock itch), and tinea corporis (ringworm).

Virus Particles

The microbe may be a virus particle. The infection may be cause by a pathogenic virus.

Examples of viruses which may be targeted using the present invention include influenza viruses, parainfluenza viruses, adenoviruses, noroviruses, rotaviruses, rhinoviruses, coronaviruses, respiratory syncytial virus (RSV), astroviruses, and hepatic viruses. In particular, the compositions of the present invention may be used in the treatment or prevention of an infection caused by one of the group selected from H1 N1 influenza virus, Infectious Bovine Rhinotracheitis virus, Bovine Respiratory Syncytial virus, Bovine Parainfluenza-3 virus, SARS-CoV, SARS-CoV-2, and any combination thereof.

In particular, the invention may be applied to treat of a disease or disorder caused by a viral infection. Examples of such diseases which may be targeted by the present invention include respiratory viral diseases, gastrointestinal viral diseases, exanthematous viral diseases, hepatic viral disease, cutaneous viral diseases, hemorrhagic viral diseases, and neurological viral diseases. Respiratory viral infections include influenza, rhinovirus (i.e. common cold virus), respiratory syncytial virus, adenovirus, coronavirus infections, for example, COVID-19, and severe acute respiratory syndrome (SARS). Gastrointestinal viral diseases include norovirus infections, rotavirus infections, adenovirus infections and astrovirus infections. Exanthematous viral diseases include measles, rubella, chickenpox, shingles, roseola, smallpox, fifth disease and chikungunya virus disease. Hepatic viral diseases include hepatitis A, hepatitis B, hepatitis C, hepatitis D and hepatitis E. Cutaneous viral diseases include warts, such as genital warts, oral herpes, genital herpes and molluscum contagiosum. Hemorraghic viral diseases include Ebola, Lassa fever, denghue fever, yellow fever, Marbug hemorrhagic fever and Crimean-Congo hemorrhagic fever. Neurological viral diseases which may be targeted using the present invention include polio, viral meningitis, viral encephalitis and rabies.

Parasitic Microorganisms

The microbe may be a parasitic microorganism (microparasite). The infection may be cause by a pathogenic parasitic microorganism.

Examples of parasitic microorganisms which may be targeted using the present invention include protozoa.

In particular, the invention may target the protozoa groups of Sarcodina (e.g. amoeba, for example Entamoeba such as Entamoeba histolytica or Entamoeba dispar), Mastigophora (e.g. flagellates, for example Giardia and Leishmania), Ciliophora (e.g. ciliates, for example Balantidium), Sporozoa (e.g. Plasmodium and Cryptosporidium), and any combination thereof.

Parasitic infections that may be treated using the present invention include malaria, amoebic dysentery and leishmaniasis (e.g. cutaneous leishmaniasis, mucocutaneous leishmaniasis or visceral leishmaniasis).

Human/Animal Hosts or Subjects

The subject may be an animal or human subject. The term “animal” herein generally can include human; however, where the term “animal” appears in the phrase “an animal or human subject” or the like, it will be understood from the context to refer particularly to non-human animals or that the reference to “human” merely particularises the option that the animal may be a human to avoid doubt.

In particular embodiments, the subject is a human subject. The human subject may be an infant or adult subject.

In particular embodiments, the subject is a vertebrate animal subject. The vertebrate animal may be in the Class Agnatha (jawless fish), Class Chondrichthyes (cartilaginous fish), Class Osteichthyes (bony fish), Class Amphibia (amphibians), Class Reptilia (reptiles), Class Aves (birds), or Class Mammalia (mammals). In particular embodiments, the subject is an animal subject in the Class Mammalia or Aves.

In particular embodiments, the subject is a domestic species of animal. The domestic species of animal may be one of: commensals, adapted to a human niche (e.g., dogs, cats, guinea pigs) prey or farm animals sought or farmed for food (e.g., cows, sheep, pig, goats); and animals for primarily draft purposes (e.g., horse, camel, donkey)

Examples of domestic animals include, but are not limited to: alpaca, addax, bison, camel, canary, capybara, cat, cattle (including Bali cattle), chicken, collared peccary, deer (including fallow deer, sika deer, thorold's deer, and white-tailed deer), dog, donkey, dove, duck, eland, elk, emu, ferret, gayal, goat, goose, guinea fowl, guinea pig, greater kudu, horse, llama, mink, moose, mouse, mule, muskox, ostrich, parrot, pig, pigeon, quail, rabbit, rat (including the greater cane rat), reindeer, scimitar oryx, sheep, turkey, water buffalo, yak and zebu.

Preparation of the e-liquid composition

The e-liquid composition may be prepared by dissolving a nitrite salt in water.

The solid nitrite salt used to prepare the e-liquid composition may be a pharmaceutically acceptable grade of solid nitrite salt. In some embodiments, the solid nitrite salt is pharmacopoeia grade. In other words, the solid nitrite salt may adhere to one or more active pharmacopoeia monographs for the nitrite salt. For example, the nitrite salt may adhere to the monograph of the nitrite salt of one or more of the United States Pharmacopoeia (USP), European Pharmacopoeia or Japanese Pharmacopoeia.

In particular embodiments, the nitrite salt used has one or more of the following limitations on its characteristics: (i) the nitrite salt contains no more than about 0.02 %, about 0.01 % or about 0.001 % by weight of sodium carbonate;

(ii) the nitrite salt contains no more than about 10 ppm (0.001 % by weight) of an anti-caking agent, such as sodium alkyl-naphthalene sulfonate;

(iii) the nitrite salt is a white to off-white solid;

(iv) the nitrite salt has a positive identification for the cation determined according to the relevant method in the relevant USP;

(v) the nitrite salt has a positive identification test for nitrite determined according to the relevant method in the relevant USP;

(vi) the nitrite salt contains no less than about 97 % or no less than 98 % by weight of the nitrite salt and/or no more than 102 % or no more than 101 % by weight of the nitrite salt, optionally as determined by the relevant USP calorimetric assay, for example, as determined by ion chromatography, such as ion chromatography coupled with suppressed conductivity detection;

(vii) the nitrite salt has a pH between about 7 and about 9 or between about 8 and about 9 when measured in a 10 % solution at 25 °C, optionally measured according to the relevant USP and/or using a pH meter;

(viii) the nitrite salt has a loss on drying of no more than about 0.25 % or about 0.01 % by weight;

(ix) the nitrite salt has a water content of no more than about 0.5 % by weight, optionally as determined by the Karl Fischer method;

(x) the heavy metal content in the nitrite salt is no more than about 10 ppm of a heavy metal, optionally the heavy metal content in the nitrite salt is no more than about 10 ppm;

(xi) the nitrite salt contains no more than about 0.4 % by weight of a nitrate salt, optionally no more than about 0.4 % by weight sodium nitrate when the nitrite salt is sodium nitrite and no more than about 0.4 % by weight potassium nitrate when the nitrite salt is potassium nitrite;

(xii) the nitrite salt contains no more than about 0.005 % or about 0.001 % by weight of insoluble matter;

(xiii) the nitrite salt contains no more than about 0.005 % by weight of chloride;

(xiv) the nitrite salt contains no more than about 0.01 % by weight of sulphate; (xv) the nitrite salt contains no more than about 0.001 % by weight of iron;

(xvi) the nitrite salt contains no more than about 0.01 % by weight of calcium;

(xvii) the nitrite salt contains no more than about 0.005 % or about 0.001 % by weight of potassium when the nitrite salt is not potassium nitrite or no more than about 0.005 % or about 0.001 % by weight of sodium when the nitrite salt is not sodium nitrite;

(xviii) the nitrite salt contains no more than about 0.1 % by weight, no more than about 5000 ppm, no more than about 1000ppm, no more than about 500 ppm, no more than about 100 ppm or no more than about 10 ppm of organic volatile compounds;

(xix) the nitrite salt contains no more than about 0.1 % by weight, no more than about 5000 ppm, no more than about 1000ppm, no more than about 500 ppm, no more than about 100 ppm or no more than about 10 ppm of ethanol;

(xx) the nitrite salt contains no more than about 3000 ppm, no more than about 1000 ppm, no more than about 500 ppm, no more than about 100 ppm or no more than about 10 ppm of methanol;

(xxi) the nitrite salt contains no more than about 50 ppm, no more than about 25 ppm, no more than about 20 ppm, no more than about 10 ppm, no more than about 7.9 ppm, no more than about 8 ppm, no more than about 6 ppm, no more than about 5.6 ppm, or no more than about 2.5 ppm of non-volatile organic carbon;

(xxii) the nitrite salt contains no more than about 0.05 ppm of mercury;

(xxiii) the nitrite salt contains no more than about 2 ppm or 0.2 ppm of aluminium;

(xxiv) the nitrite salt contains no more than about 3 ppm or 1 ppm of arsenic;

(xxv) the nitrite salt contains no more than about 0.003 % or 0.001 % by weight of selenium;

(xxvi) the total aerobic count of microbial load in the nitrite salt is no more than about 100 CFU/g;

(xxvii) the total yeast and mold count in the nitrate salt is no more than about 20 CFU/g; (xxviii) the nitrite salt contains no more than about 0.25 Ell/rng or 0.018 Ell/rng of bacterial endotoxins; and

(xxix) the nitrite salt contains less than about 0.1 ppm of a phosphate salt, such as sodium phosphate, disodium hydrogen phosphate or trisodium phosphate, and preferably the nitrite salt contains no detectable amount of phosphate salt.

In certain embodiments, the nitrite salt has two or more of the characteristics of (i) to (xxix). In further embodiments, the nitrite salt has five or more of the characteristics of (i) to (xxix). In yet further embodiments, the nitrite salt has ten or more of the characteristics of (i) to (xxix). In even further embodiments, the nitrite salt has fifteen or more of the characteristics of (i) to (xxix). In some embodiments, the nitrite salt has twenty or more of the characteristics of (i) to (xxix). In a particular embodiment, the nitrite salt has all of the characteristics of (i) to (xxix). In a more particular embodiment, the nitrite salt is sodium nitrite having all of the characteristics of (i) to (xxix).

In some embodiments the nitrite salt contains in the range of about 97 % to about 101 % by weight of the nitrite salt, optionally as determined by the relevant USP calorimetric assay, for example, as determined by ion chromatography, such as ion chromatography coupled with suppressed conductivity detection. In alternative embodiments nitrite salt contains in the range of about 98 % to about 102 % by weight of the nitrite salt, optionally as determined by the relevant USP calorimetric assay, for example, as determined by ion chromatography, such as ion chromatography coupled with suppressed conductivity detection

In particular embodiments the nitrite salt has the following characteristics:

(i) the nitrite salt contains no more than about 0.02 % by weight of sodium carbonate;

(ii) the nitrite salt contains no more than about 10 ppm of an anti-caking agent;

(vi) the nitrite salt contains no less than 97 % by weight of the nitrite salt and no more than 101 % by weight of the nitrite salt as determined by USP calorimetric assay;

(viii) the nitrite salt has a loss on drying of no more than about 0.25 % by weight;

(ix) the nitrite salt has a water content of no more than about 0.5 % by weight;

(x) the heavy metal content in the nitrite salt is no more than about 10 ppm;

(xi) the nitrite salt contains no more than about 0.4 % by weight of a nitrate salt;

(xii) the nitrite salt contains no more than about 0.005 % by weight of insoluble matter;

(xiv) the nitrite salt contains no more than about 0.01 % by weight of sulphate;

(xv) the nitrite salt contains no more than about 0.001 % by weight of iron;

(xvi) the nitrite salt contains no more than about 0.01 % by weight of calcium;

(xviii) the nitrite salt contains no more than about no more than about 5000 ppm, no more than about 1000ppm, no more than about 500 ppm, no more than about 100 ppm or no more than about 10 ppm of organic volatile compounds;

(xxi) the nitrite salt contains no more than about 10 ppm or no more than about 2.5 ppm of non-volatile organic carbon;

(xxii) the nitrite salt contains no more than about 0.05 ppm of mercury; (xxiii) the nitrite salt contains no more than about 2 ppm of aluminium;

(xxiv) the nitrite salt contains no more than about 3 ppm of arsenic; (xxv) the nitrite salt contains no more than about 0.003 % by weight of selenium; (xxvi) the total aerobic count of microbial load in the nitrite salt is no more than about 100 CFU/g;

(xxvii) the total yeast and mold count in the nitrate salt is no more than about 20 CFU/g; and

(xxviii) the nitrite salt contains no more than about 0.25 Ell/rng of bacterial endotoxins.

In these embodiments, the nitrite salt may be sodium nitrite and contain no more than about 0.005 % by weight of potassium. Preferably the sodium nitrite also has one or more of the following limitations:

(iii) the sodium nitrite is a white to off-white solid;

(iv) the sodium nitrite has a positive identification for sodium determined according to the relevant method in the relevant USP;

(v) the sodium nitrite has a positive identification test for nitrite determined according to the relevant method in the relevant USP;

(vii) the sodium nitrite has a pH between about 7 and about 9 or between about 8 and about 9 when measured in a 10 % solution at 25 °C, optionally measured according to the relevant USP and/or using a pH meter;

(xix) the sodium nitrite contains no more than about 0.1 % by weight, no more than about 5000 ppm, no more than about 1000ppm, no more than about 500 ppm, no more than about 100 ppm or no more than about 10 ppm of ethanol;

(xx) the nitrite salt contains no more than about 3000 ppm, no more than about 1000 ppm, no more than about 500 ppm, no more than about 100 ppm or no more than about 10 ppm of methanol; and

The characteristics of (i) to (xxix) may be determined according to the relevant method in USP XXXII (2009). Methods for determining the characteristics of (i) to (xxix) are provided in WO 2010/093746, the disclosure of which is incorporated herein by reference in its entirety. Methods of preparing sodium nitrite having one or more of the characteristics of (i) to (xxix) are also described in WO 2010/093746. Preferred Embodiments

Preferred embodiments of the aspects of the present invention are those wherein on or more of the following is present:

- the one or more nitrite salt comprises (for example, includes or consists essentially of or consists only of) one or more alkali metal or alkaline earth metal nitrite salt, for example: sodium nitrite; potassium nitrite; or any combination thereof;

- The nitrite salt has a concentration in the e-liquid composition in the range of about 0.1 to about 1.2 M;

- The nitrite salt has a concentration in the e-liquid composition in the range of about 0.4 to about 0.6 M, such as about 0.5 M;

- the e-liquid composition includes one or more organic alcohol;

- the one or more organic alcohol comprises (for example, includes or consists essentially of or consists only of) an acyclic organic alcohol having 2 to 4 carbon atoms and 1 to 3 hydroxy groups that are not a proton source, such as ethanol (ethyl alcohol), glycerol, propylene glycol and combinations thereof;

- the one or more organic alcohol comprises (for example, includes or consists essentially of or consists only of) a cyclic organic alcohol having 8 to 12 carbon atoms and 1 to 3 hydroxy groups that are not a proton source, such as menthol;

- the one or more organic alcohol comprises (for example, includes or consists essentially of or consists only of) a straight-chain sugar alcohol or alditol having from 4 to 12 carbon atoms and from 4 to 12 OH groups per molecule; for example sorbitol; mannitol; arabitol; xylitol; or any combination of two or more thereof;

- the one or more organic alcohol is a sugar alcohol compound comprising, for example consisting of, a chain of 1 , 2 or 3 monosaccharide units terminated with one acyclic alcohol unit, optionally where. 1 , 2, 3 or each monosaccharide unit is a Cs or Ce monosaccharide unit and/or the acyclic alcohol unit is a Cs or Ce sugar alcohol unit; for example, isomalt, maltitol, lactitol, maltotriitol, maltotetraitol; and

- The one or more organic alcohol is present in about 5 % to about 15 % by weight/volume of the e-liquid composition;

- The method of prevention or treatment includes at least one treatment session including five, six or seven operations of the vaping device in succession; Each operation of the vaping device may occur for an operation time period in the range of about four to about six seconds, such as about five seconds;

Each operation of the vaping device may be separated for a separation time period be in the range of about ten seconds to about two minutes, or about thirty seconds to about 90 seconds;

- the disease or disorder being treated or prevented is associated with or caused by a microbe, for example without limitation influenza virus, SARS-CoV, SARS- CoV-2, Mycobacterium tuberculosis, Mycobacterium abscessus, Pseudomonas aeruginosa including antibiotic-resistant strains thereof.

The present invention will be described in more detail below, with particular reference to the Examples and the following Figures:

Figure 1 shows a cumulative plot of NOx generated using 1.0M Sodium Nitrite in 0% glycerol solution.

Figure 2 shows a plot of NOx release in ppb over time. 1mL of 1.0M Sodium Nitrite in 0% Glycerol solution vaporised through IBIZA vape device at 30W and NOx measured through SIFT-MS.

Figure 3 shows a cumulative plot of NOx generated using 0.5M Sodium Nitrite in 10% glycerol solution

Figure 4 shows a plot of NOx release in ppb over time. 1mL of 0.5M Sodium Nitrite in 10% glycerol solution vaporised through IBIZA vape device at 30W and NOx measured through SIFT-MS.

Figure 5 shows a cumulative plot of NOx generated using 0.5M Sodium Nitrite in 0% glycerol solution.

Figure 6 shows a cumulative plot of NOx generated using 0.5M Sodium Nitrite in 10% glycerol solution. Figure 7 shows a NOx release in ppb over time plot. 1 mL of 0.5M Sodium Nitrite in 0% glycerol solution vaporised through IBIZA vape device at 30W and NOx measured through SIFT-MS

Figure 8 shows a cumulative plot of NOx generated using 0.5M Sodium Nitrite in 10% glycerol solution.

Figure 9 shows an example of a SIFT-MS - vape device setup for NOx generation using silicone tubes and a parafilm.

Examples

The following non-limiting Examples are provided for further illustration of the present invention.

Materials, Apparatus, and Methods Used

Solutions

Sodium Nitrite Solution preparation

Stock solutions of 2.0M sodium nitrite were prepared using sodium nitrite (Honeywell Fluka™ 99-100.5%) by dissolving the appropriate mass in deionised water. Deionised water (11.2 MQcm) was obtained from Millipore ELIX water systems (Merck Group, France). Final concentrations of sodium nitrite solutions shown in table 1 were prepared using the stock solution of 2.0M and dissolved in deionised water.

Alcohol/Polyol Solution Preparation

Stock solution of glycerol (Honeywell Riedl-de-haen™ >99.5%) was used to make various concentrations of glycerol dissolved in deionised water (See table 2). Deionised water was obtained from Millipore ELIX water systems (Merck Group, France).

Example 1: Various Sodium Nitrite Solution Preparation

For analysing sodium nitrite in the absence of glycerol, 0.1M, 0.5M and 1.0M of sodium nitrite were prepared. 2.0 M stock solution of sodium nitrite was used to prepare the various formulations of sodium nitrite, mixed with deionised water. Final concentration of 0.1M, 0.5M and 1.0M were prepared in 0% glycerol (no glycerol was added). See

Table 1 for all relevant nitrite formulations

Table 1: Sodium Nitrite formulations in 0% Glycerol vaporised at 30W

Example 2: Glycerol-Sodium Nitrite Formulations Preparation

Stock solution of glycerol (Honeywell Riedl-de-haen™ >99.5%), stock solution of 2.0 M sodium nitrite and deionised water was used to make 0.5 M sodium nitrite concentrations, in 0%, 25% and 50% glycerol (purity >99.5%). See Table 2 for all relevant nitrite-glycerol formulations

Table 2: 0.5M Sodium Nitrite and varying glycerol formulations

Vape Device Setup

A Sub-Ohm vape device (IBIZA Vape Club model: 115521 , 2020) was used to generate

NOx. Full anatomy of the vape is illustrated on Figure 1. The vape contained an atomiser (capacity 2 mL), a 22 x 11 mm coil embedded inside the atomiser, a 1500 mAh battery capacity, and a power range between 20-50 W. Vape was connected to the SIFT-MS inlet by using silicone tubes (SLS, 2020) that were complementary to the mouthpiece of the vape device and to the SIFT-MS inlet (See Figure 2). Parafilm (Bemis™, SLS 2020) was used to coat the inlet and mouthpiece tubing areas to prevent any gas escape

Selected Ion Flow Tube Mass Spectrometry (SIFT-MS) Start up and Validation

A voice200 Ultra Selected ion flow Tube Mass Spectrometer (SIFT-MS) (Syft Technologies Ltd, New Zealand) was used for all the gas analyses described in this report. This instrument uses helium (Air Products, Surrey, UK) as a carrier gas. Prior to analysis, the SIFT-MS was prepared for use with a simple start up procedure. The instrument was taken out of standby mode and a series of pressure checks were made to ensure that capillary flow is within the acceptable range for operation. This was followed by automated validation procedure using the manufacturer’s calibrant gas standard (Syft Technologies Ltd, New Zealand) containing ethylene, hexafluorobenzene, isobutane, octafluorotoluene, tetrafluorobenzene, and toluene.

Finally, an in-house performance check was undertaken using 10-ppm nitric oxide and nitrogen dioxide standards (Air Products PLC, Surrey, UK).

Table 3 shows the parameters used to preform SIFT-MS analysis using the IBIZA vape device.

Table 3: Parameters of Sampling Using SIFT-MS and IBIZA vape device

The glass chamber of the vape was attached to the battery and rotated clockwise. 0.5mL of each sodium nitrite-glycerol formulation (see table 1 and 2) was added into the glass chamber of the vape device and the mouthpiece was sealed tight to close the chamber. The power on the vape device was set to 30W for all formulations.

After a successful SIFT-MS validation, the heated transfer line with 1/8^th PTFE tubing, heated to 150°C, was attached to the vape reaction chamber outlet allowing the SIFT- MS to sample the gases flowing out from the vape chamber in real-time.

The SIFT-MS equipment, vape device, and gas pathway were set up as illustrated in Figure 9.

A continuous SIFT-MS scan was initiated by firstly observing a background for 5 minutes prior to the run. Once a stable baseline background was observed, the fire button of the vape was pressed to prime the coil (priming the coil is a crucial step to allow the formulation to enter the coil).

The fire button on the vape was pressed for 5 seconds to allow NOx measurement through SIFT-MS for 60 seconds. After the 1 -minute interval the fire button was pressed for 5 seconds again and this was repeated 6 times (generating 6 peaks in total). Between each analysis of the formulations, an air purge was performed using a dry filtered compressed air to allow the clearance of any NOx that might be present in the vape chamber. Vape coil was replaced with a new one, when example 2 nitriteglycerol formulations were analysed. So separate coils were used for example 1 and 2, and they were flushed with air purge in between the formulations of each example

Example 1 Results: 0.5M Sodium Nitrite in Varying Glycerol formulations Table 4: Table 4: Various Sodium Nitrite formulations in 0% Glycerol vaporised through IBIZA Vape and measured using SIFT-MS.

Example 2 Results: Formulations with 0.5M Sodium Nitrite and Varying Glycerol % Table 5: 0.5M Sodium Nitrite formulations in various Glycerol concentrations, vaporised through IBIZA Vape and measured using SIFT-MS

Final nmol nmol % Ratio Peak

Sodium Glycerol Repeat NO NO2 of NO to Range

Nitrite NO₂ NO

Cone. (ppm)

Comparing results for 0.5 Sodium Nitrite in 0% Glycerol with 0.5M Sodium Nitrite in

10% Glycerol (example 1 and 2)

Table 6: 1mL of 0.5M Sodium Nitrite in 0% Glycerol solution vaporised through IBIZA vape device and NOx measured through SIFT-MS

Table 7: 1mL of 0.5M Sodium Nitrite in 10% Glycerol solution vaporised through IBIZA vape device and NOx measured through SIFT-MS

As can be seen by the results above, an increased nitrite concentration results in an increased NO output. In general, the higher the sodium nitrite concentrations result in the greater the NO output. In addition, the NO/NO2 ratio increases as the nitrite concentration increases.

Further, the results above show that increasing the glycerol concentration, decreases the levels of NO generated. There appears to be no significant change in the NO/NO2 ratio with increasing glycerol concentration.

Claims

33 CLAIMS:

1 . An aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder, the method including operating an e-cigarette or vaping device including an aqueous e-liquid composition and wherein the e-liquid composition comprises one or more nitrite salt.

2. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of claim 1 wherein the one or more nitrite salt comprises one or more alkali metal or alkaline earth metal nitrite salt, for example: sodium nitrite; potassium nitrite; or any combination thereof.

3. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of claim 1 or claim 2, wherein the nitrite salt has a concentration in the e-liquid composition in the range of about 0.1 to about 1.2 M.

4. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of any one of claims 1 to 3, wherein the e-liquid composition includes one or more organic alcohol.

5. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of claim 4, wherein the one or more organic alcohol comprises (for example, includes or consists essentially of or consists only of) an acyclic organic alcohol having 2 to 4 carbon atoms and 1 to 3 hydroxy groups that are not a proton source; for example ethanol (ethyl alcohol), glycerol, propylene glycol and combinations thereof.

6. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of claim 4, wherein the one or more organic alcohol comprises (for example, includes or consists essentially of or consists only of) a cyclic organic alcohol having 8 to 12 carbon atoms and 1 to 3 hydroxy groups that are not a proton source; for example menthol.

7. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of claim 4, wherein the one or more organic alcohol comprises (for example, includes or consists essentially of 34 or consists only of) a straight-chain sugar alcohol or alditol having from 4 to 12 carbon atoms and from 4 to 12 OH groups per molecule; for example sorbitol; mannitol; arabitol; xylitol; or any combination of two or more thereof.

8. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of claim 4, wherein the one or more organic alcohol is a sugar alcohol compound comprising, for example consisting of, a chain of 1, 2 or 3 monosaccharide units terminated with one acyclic alcohol unit, optionally where. 1 , 2, 3 or each monosaccharide unit is a C5 or C6 monosaccharide unit and/or the acyclic alcohol unit is a C5 or C6 sugar alcohol unit; for example, isomalt, maltitol, lactitol, maltotriitol, maltotetraitol.

9. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of any one of claims 4 to 8, wherein the one or more organic alcohol is present in about 5 % to about 15 % by weight or by volume based on the total weight or total volume, respectively, of the e-liquid composition.

10. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of any one of claims 1 to 9, wherein the method of prevention or treatment includes at least one treatment session including five, six or seven operations of the vaping device or e cigarette in succession.

11. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of any one of claims 1 to 10, wherein each operation of the vaping device occurs for an operation time period in the range of about four to about six seconds, such as about five seconds.

12. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of any one of claims 1 to 11 , wherein each operation of the vaping device is separated for a separation time period be in the range of about ten seconds to about two minutes, or about thirty seconds to about 90 seconds. The aqueous e-liquid composition for use in a method of treatment or prevention of a respiratory disease or disorder of any one of claims 1 to 12, wherein the disease or disorder being treated or prevented is associated with or caused by a microbe, for example without limitation Influenza virus, SARS-CoV, SARS-CoV-2, Mycobacterium tuberculosis, Mycobacterium abscessus,

Pseudomonas aeruginosa including antibiotic-resistant strains thereof. A method of treatment or prevention of a respiratory disease or disorder, the method including operating a vaping device including an aqueous e-liquid composition and wherein the e-liquid composition comprises a nitrite salt. A vaping device containing an aqueous e-liquid composition, the e-liquid composition comprising a nitrite salt