Classifications

• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
  • C11D3/00—Other compounding ingredients of detergent compositions covered in group C11D1/00
  • C11D3/0005—Other compounding ingredients characterised by their effect
  • C11D3/0047—Other compounding ingredients characterised by their effect pH regulated compositions
• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
  • C11D17/00—Detergent materials or soaps characterised by their shape or physical properties
  • C11D17/04—Detergent materials or soaps characterised by their shape or physical properties combined with or containing other objects
  • C11D17/041—Compositions releasably affixed on a substrate or incorporated into a dispensing means
  • C11D17/046—Insoluble free body dispenser
• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
  • C11D17/00—Detergent materials or soaps characterised by their shape or physical properties
  • C11D17/04—Detergent materials or soaps characterised by their shape or physical properties combined with or containing other objects
  • C11D17/049—Cleaning or scouring pads; Wipes
• • C—CHEMISTRY; METALLURGY
  • C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
  • C11D—DETERGENT COMPOSITIONS; USE OF SINGLE SUBSTANCES AS DETERGENTS; SOAP OR SOAP-MAKING; RESIN SOAPS; RECOVERY OF GLYCEROL
  • C11D3/00—Other compounding ingredients of detergent compositions covered in group C11D1/00
  • C11D3/02—Inorganic compounds ; Elemental compounds
  • C11D3/04—Water-soluble compounds
  • C11D3/046—Salts

Definitions

• the present invention relates to a substrate (e.g. a sponge) comprising a blocking composition.
• the invention has particular, but not necessarily exclusive, application in blocking a cleaning composition to facilitate detection of microorganisms (such as bacteria) on a surface.
• microorganisms such as bacteria
• BACKGROUND Microorganisms such as bacteria, typically require metal species (e.g. iron), for a number of biological processes.
• Bacteria may extract iron from the ambient environment by means of iron- chelating compounds known as siderophores (e.g. enterobactin and deferoxamine). Siderophores secreted by bacteria have high iron (Fe 3+ ) binding affinity and are thereby able to sequester ambient iron.
• the iron bound to the siderophore can be taken up by the bacteria using active transport mechanisms.
• Chromeazurol S is a colour changing agent which provides a change in colour upon binding and/or release of iron (blue or purple on binding, orange on release/in unbound form). Due to a high iron binding affinity, when CAS with bound iron is exposed to siderophores, the iron is released from the CAS and instead binds preferentially to the siderophores, thereby resulting in a colour change in the CAS.
• Levels of bacteria present in a sample are related to the amount of siderophore present in that sample, and the amount of siderophore present is related to the amount of iron that can be sequestered.
• the colour change resulting from binding and/or release of iron is related to the amount of iron bound and/or released. Therefore, the colour change resulting from release of iron bound to CAS may be used to determine the presence or absence of bacteria present in a sample and various colour changing compositions have been produced which rely on these principles. Other microorganisms adopt similar techniques for iron uptake, and other metals can be sequestered in the same manner. Similarly, other colour changing agents can be used to monitor this. In many environments, it is desirable to reduce or eliminate microorganisms, and thereby to reduce or eliminate risk of infection to living organisms. Suitably, this is undertaken using detergent compositions, disinfecting compositions, or the like.
• compositions operate by virtue of a variety of chemical species, such as surfactants, chelators, pH-altering agents, etc..
• these chemical species can undesirably interact (i.e. interfere) with colour changing agents/compositions configured to detect microorganisms.
• chelators can themselves chelate iron and prevent uptake by CAS and siderophores. This affects various properties of the colour changing compositions, such as sensitivity, selectivity, stability, etc. and may ultimately lead to an ineffective colour changing composition (e.g. eliciting false positives or false negatives).
• a substrate (optionally a sponge) comprising a blocking composition
• said blocking composition comprises: one or more metals; one or more blocking composition surfactants; and one or more pH buffers; wherein 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at least around 1.372 x 10 -8 moles of n-alkyl dimethyl benzyl ammonium chloride; and at least around 3.106 x 10 -8 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• a substrate (optionally a sponge) comprising a blocking composition (optionally around 0.15 ml thereof), wherein said blocking composition comprises: one or more metals; one or more blocking composition surfactants; and one or more pH buffers, optionally wherein around 0.15 ml of the blocking composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: (a) at least around 1.937 x 10 -8 moles of citrate, optionally at least around 4.029x10- 7 , optionally at least around 7.864x10 -7 , optionally at least around 1.17x10 -6 , optionally at least around 1.553x10 -6 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) at least around 2.959 x 10 -8 moles of ethylenediaminetetraacetic acid (EDTA), optionally at least around 6.155x10 -7 , optionally at least around 1.201x10
• EDTA ethylenediamine
• a composition comprising the blocking composition as defined in the first or second aspect.
• a kit comprising: the substrate according to the first or second aspect and/or the composition according to the third aspect; and (a) a colour changing composition configured to detect the presence of microorganisms; and/or (b) a cleaning composition configured to reduce and/or eliminate microorganisms.
• a swab comprising a rod having a substrate according to the first or second aspect at an end thereof (optionally wherein the substrate comprises 0.15 ml of said composition), optionally further comprising a reservoir of a colour changing composition.
• a substrate according to the first or second aspect, the composition according to third aspect or a swab according to the fifth aspect to block a cleaning composition.
• a method comprising contacting a surface, or a sample therefrom, that has been pre-treated with a cleaning composition, with the composition of the third aspect.
• this modifier may indicate a deviation from the quality given of less than or equal to about 20%, such as less than or equal to about 15%, such as less than or equal to about 10%, such as less than or equal to about 5%, such as less than or equal to about 1%, such as about 0%.
• aliphatic means a straight-chain, branched or cyclic hydrocarbon, which is completely saturated, or which contains one or more units of unsaturation (e.g. alkenyl or alkynyl), but which is not aromatic. Where the aliphatic group refers to a range, such as C 1 to C 10 , it is to be understood that it includes each member of the range, i.e.
• (C 1-6 )aliphatic or “C 1 to C 6 ” aliphatic as used herein means an aliphatic group having 1 to 6 carbon atoms, which may be branched or unbranched, saturated or unsaturated and optionally contains a ring.
• alkyl refers to a saturated (no double or triple bonds) aliphatic hydrocarbon radical, including straight-chain, and, where possible, branched-chain and cyclic groups and hybrids thereof, such as (cycloalkyl)alkyl.
• alkyl group refers to a range, such as C 1 to C 10 , it is to be understood that it includes each member of the range, i.e. C 1 , C 2 , C 3 , C 4 , C 5 , C 6 , etc.
• (C1-6)alkyl as used herein means an alkyl group having 1-6 carbon atoms, which may be branched or unbranched and optionally contains a ring.
• cycloalkyl refers to a cyclic alkyl group, for example cycloheptyl, cyclohexyl, cyclopentyl, cyclobutyl or cyclopropyl. Cycloalkyl may be substituted as defined herein.
• alkenyl refers to an alkyl group that contains in the straight or branched hydrocarbon chain one or more double bonds. Examples of alkenyl groups include allenyl, vinylmethyl and ethenyl. An alkenyl group may be unsubstituted or substituted.
• alkynyl refers to an alkyl group that contains in the straight or branched hydrocarbon chain one or more triple bonds. Examples of alkynyls include ethynyl and propynyl. An alkynyl group may be unsubstituted or substituted.
• alkanoate means -C(O)O-alkyl wherein alkyl has the meaning as defined above (e.g. being C 1 to C 10 ).
• Examples of (C 1 - 4 )alkanoate include methanoate, ethanoate, propanoate, isopropanoate, butanoate, isobutanoate and tertiary butanoate.
• halide or “halogen” are used interchangeably and, as used herein mean an ion derived from IUPAC group number 17 of the periodic table.
• the halide/halogen may be a fluoride, a chloride, a bromide, an iodide and the like.
• aryl refers to a carbocyclic (all carbon) monocyclic or multicyclic aromatic ring system (including fused ring systems where two carbocyclic rings share a chemical bond) that has a fully delocalized pi-electron system throughout all the rings.
• the number of carbon atoms in an aryl group can vary.
• the aryl group can be a C 6 -C 14 aryl group, a C 6 -C 10 aryl group, or a C 6 aryl group.
• Examples of aryl groups include, but are not limited to, benzene, naphthalene and azulene.
• heteroaryl refers to a monocyclic or multicyclic aromatic ring system (a ring system with fully delocalized pi-electron system) that contain(s) one or more heteroatoms (for example, 1 to 5 heteroatoms), that is, an element other than carbon, including but not limited to, nitrogen, oxygen and sulfur.
• the number of atoms in the ring(s) of a heteroaryl group can vary.
• the heteroaryl group can contain 4 to 14 atoms in the ring(s), 5 to 10 atoms in the ring(s) or 5 to 6 atoms in the ring(s).
• heteroaryl includes fused ring systems where two rings, such as at least one aryl ring and at least one heteroaryl ring, or at least two heteroaryl rings, share at least one chemical bond.
• heteroaryl rings include, but are not limited to, furan, furazan, thiophene, benzothiophene, phthalazine, pyrrole, oxazole, benzoxazole, 1,2,3-oxadiazole, 1,2,4-oxadiazole, thiazole, 1,2,3-thiadiazole, 1,2,4-thiadiazole, benzothiazole, imidazole, benzimidazole, indole, indazole, pyrazole, benzopyrazole, isoxazole, benzoisoxazole, isothiazole, triazole, benzotriazole, thiadiazole, tetrazole, pyridine, pyridazine, pyrimidine
• Ethylenediaminetetraacetic acid can be present as a salt, such as a tetrasalt (e.g. tetrasodium) thereof.
• EDTA Ethylenediaminetetraacetic acid
• the phrase “colour changing composition” is intended to refer to a composition that comprises an agent (e.g. a dye) configured to change colour upon detection of microbes.
• the colour changing composition may comprise a dye and a metal, wherein the metal is bindable to the colour changing agent to provide a change in colour on binding and/or release thereof.
• Such colour changing compositions can be used to detect the presence of microorganisms by means of a colour change arising from an interaction between the colour changing agent, metal and siderophores as released by microorganisms (e.g. bacteria).
• siderophores can sequester said metal, releasing this from the colour changing agent and thereby causing said colour change.
• colour changing complex refers to a complex formed between the metal and the colour changing agent.
• Suitable colour changing compositions are described in international patent application WO2018185486, the entire content of which is hereby incorporated in its entirety.
• the colour changing composition defined herein may be a sprayable composition in accordance with the first aspect defined in WO2018185486, and related implementations thereof.
• the colour changing composition defined herein may be a composition in accordance with the second aspect defined in WO2018185486, and related implementations thereof.
• the term “metal” is intended to include elemental forms or compounds of metals.
• the phrase “cleaning composition” as used herein is intended to refer to an aqueous composition comprising one or more cleaning chemicals, such as surfactants, emulsifiers, chelators, etc.
• the cleaning composition may be a “detergent composition” and/or a “disinfectant composition”.
• blocking composition refers to a composition which is able to substantially prevent and/or mitigate the ability of cleaning chemicals in a cleaning composition to interfere with activity of a colour changing composition as discussed herein (in particular a metal-dye based colour changing composition, such as an iron-CAS based composition).
• a colour changing composition as discussed herein (in particular a metal-dye based colour changing composition, such as an iron-CAS based composition).
• Such cleaning chemicals may be left as residue on a surface after cleaning with a cleaning composition.
• Use of a colour changing composition on a residue-contaminated surface may not elicit the correct colour change in view of unwanted interactions of the residue with the colour changing composition.
• chelators present in the cleaning composition may interact with metal/iron in a metal-dye based colour changing composition and thereby interfere with the colour change otherwise initiated by binding and release of said metal.
• Blocking includes: ⁇ Inhibition of pH-based interference from the cleaning composition (e.g. blocking pH changes that may be caused by residual cleaning compositions, which are typically basic); and/or ⁇ Inhibition of chelation interference commonly elicited by the cleaning composition (e.g. blocking chelators from binding to metal species, such as iron); and/or ⁇ Inhibition of surfactant interference from the cleaning composition (e.g. blocking surfactants from interfering with the colour changing agent, such as CAS).
• pH-based interference from the cleaning composition e.g. blocking pH changes that may be caused by residual cleaning compositions, which are typically basic
• chelation interference commonly elicited by the cleaning composition e.g. blocking chelators from binding to metal species, such as iron
• surfactant interference from the cleaning composition e.g. blocking surfactants from interfering with the colour changing agent, such as CAS.
• ⁇ Inhibition of hypochlorite interference from the cleaning composition e.g. blocking reactive hypochlorites from degrading the colour changing agent, such as CAS.
• ⁇ Inhibition of acid interference e.g. acetic acid interference
• acid interference e.g. acetic acid interference
• peroxide interference from the cleaning composition e.g. blocking reactive peroxides from interfering with the colour change composition by modulating the pH of reactive species
• peracetic acid interference from the cleaning composition e.g. blocking reactive peracetic acid from interfering with the colour change composition by modulating the pH of reactive species.
• the buffer included in the blocking composition can be defined by a buffer capacity and working pH range. These ranges help to ensure that the blocking of undesired surface pH changes can be achieved, whilst also helping to ensure they are not too high to prevent colour change caused by drastic pH changes (e.g. an excessive level of bleach).
• the buffer capacity should preferably have a ⁇ of around -0.7 to 0.7 mol.
• the working pH should preferably be around pH 2.6 to pH 8, optionally pH 3 to pH 8, optionally pH 3.5 to 7, optionally around 5.6 to 7.
• Chelation inhibitors included in the buffers must have an affinity for relevant chelation molecules (e.g. EDTA or Citrate). This ensures that they bind to the relevant chelator to block the chelator from interacting with the colour change composition.
• the affinity can be measured with binding constants (log K d ) and range from 4 - 42. Variation in K d can be modulated via relative abundance.
• Surfactants included in the buffer must be present at such a level as to prevent cleaning chemicals (e.g. quaternary ammonium compounds) from causing an undesired colour change. This can be defined as preventing a specific number of molecules from causing a colour change that would indicate a ‘contaminated’ results and is measured in parts per million (ppm). The range that the surfactant may inhibit is 5 - 2000 ppm.
• the amount of a particular component in a composition or solution is expressed as a “wt%”, this is based on the total weight of composition or solution and may be:
• said blocking composition comprises a certain concentration of said one or more metals, this refers to the amount of free metallic species in the composition (e.g. dissociated metal species from a precursor salt).
• the defined concentration refers to the sum of concentrations of these species.
• binding affinity is intended to refer to the strength of the binding interaction between a molecule to its ligand/binding partner.
• Binding affinities for various chemical species are typically found in textbooks, such as Microchimica Acta 1964, 52, 414–428.
• the term “buffer capacity” is intended to refer to a quantitative measure of the amount a buffer can resist changes in pH. Capacities for various buffers can be measured in accordance with ISO 23497:2019 (https://www.iso.org/obp/ui#iso:std:iso:23497:ed-1:v1:en, accessible as of 24 January 2022).
• present disclosures refer to different compositions, such as a “blocking composition”, a “colour changing composition”, a “detergent composition” or a “sprayable composition”.
• compositions can comprise similar components, such as surfactants.
• a surfactant present in a blocking composition is referred to herein as a “blocking composition surfactant”
• a surfactant present in a colour changing composition is referred to herein as a “colour changing composition surfactant”
• a metal present in a blocking composition is referred to herein as a “blocking composition metal”.
• a substrate (optionally a sponge) comprising a blocking composition
• said blocking composition comprises: one or more metals; one or more blocking composition surfactants; and one or more pH buffers; wherein 0.375 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at least around 1.372 x 10 -8 moles of n-alkyl dimethyl benzyl ammonium chloride; and at least around 3.106 x 10 -8 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• the blocking composition defined herein has been specifically formulated to block cleaning compositions which are typically found in a variety of settings (e.g.
• the amounts of metal, surfactant and other components mentioned below have been specifically chosen bearing these in mind.
• numerous such cleaning compositions are tested (e.g. the “D10” base formulation which commonly comprises a number of cleaning compositions).
• the substrate is useful in applications in which detection of microorganisms is desirable.
• the substrate can be wiped over an article (e.g. a surface) to block action of residual cleaning chemicals. Thereafter, the article/surface may be ready for sampling and detection of microorganisms by means of a colour change using a colour changing composition (such as a colour changing composition defined herein, e.g.
• the blocking composition may comprise a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at least around 2.854x10 -7 , optionally at least around 5.57x10 -7 , optionally at least around 8.287x10 -7 , optionally at least around 1.1x10 -6 , optionally around 1.3x10 -6 , such as around 1.372x10 -6 moles of n-alkyl dimethyl benzyl ammonium chloride; and at least around 6.46x10 -7 , optionally at least around 1.261x10 -6 , optionally at least around 1.876x10 -6 , optionally at least around 2.491x10 -6 , optionally around 3.1x10 -6 such as around 3.106x10 -6 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• the blocking composition may comprise a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at most around 1.372 x 10 -4 , optionally at most around 1.1x10 -4 , optionally at most around 8.287x10 -5 , optionally at most around 5.57x10 -5 , optionally at most around 2.854x10 -5 , around 1.3x10 -6 , such as around 1.372x10 -6 moles of n-alkyl dimethyl benzyl ammonium chloride; and at most around 3.106 x 10 -4 , optionally at most around 2.491x10 -4 , optionally at most around 1.876x10 -4 , optionally at most around 1.261x10 -4 , optionally at most around 6.46x10 -5 , optionally around 3.1x10 -6 such as around 3.106x10 -6 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• the blocking composition may comprise a combined amount of said metal and said blocking composition surfactant that is sufficient to block: around 1.372x10 -8 to 1.372x10 -4 , optionally around 2.854x10 -7 to 1.1x10 -4 , optionally around 5.57x10 -7 to 8.287x10 -5 , optionally around 8.287x10 -7 to 5.57x10 -5 , optionally around 1.1x10 -6 to 2.854x10 -5 moles of n-alkyl dimethyl benzyl ammonium chloride; and around 3.106x10 -8 to 3.106x10 -4 , optionally around 6.46x10 -7 to 2.491x10 -4 , optionally around 1.261x10 -6 to 1.876x10 -4 , optionally around 1.876x10 -6 to 1.261x10 -4 , optionally around 2.491x10 -6 to 6.46x10 -5 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• the blocking composition may comprise a combined amount of said metal and said blocking composition surfactant that is sufficient to block said amount of n-alkyl dimethyl benzyl ammonium chloride and C 9 -C 11 alkyl alcohol ethoxylate, and at least: (a) around 1.937x 10 -8 moles of citrate, optionally at least around 4.029x10 -7 , optionally at least around 7.864x10 -7 , optionally at least around 1.17x10 -6 , optionally at least around 1.553x10 -6 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) around 2.959 x 10 -8 moles of ethylenediaminetetraacetic acid (EDTA), optionally at least around 6.155x10 -7 , optionally at least around 1.201x10 -6 , optionally at least around 1.787x10 -6 , optionally at least around 2.373x10 -6 , optionally around 3x10 -6 such as
• Citrate and EDTA are commonly-encountered chelators present in a variety of cleaning compositions and it is desirable, in some implementations, to provide a blocking composition which is able to cater for such chelators.
• the blocking composition may comprise a combined amount of said metal and said blocking composition surfactant that is sufficient to block said amount of n-alkyl dimethyl benzyl ammonium chloride and C 9 -C 11 alkyl alcohol ethoxylate, and at most: (a) around 1.937x10 -4 moles of citrate, optionally at most around 1.553x10 -4 , optionally at most around 1.17x10 -4 , optionally at most around 7.864x10 -5 , optionally at most around 4.029x10 -5 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) around 2.959x10 -4 moles of ethylenediaminetetraacetic acid (EDTA), optionally at most around 2.373x10
• the blocking composition may comprise a combined amount of said metal and said blocking composition surfactant that is sufficient to block said amount of n-alkyl dimethyl benzyl ammonium chloride and C 9 -C 11 alkyl alcohol ethoxylate, and: (a) around 1.937 x 10 -8 to 1.937 x 10 -4 , optionally around 4.029x10 -7 to 1.553x10 -4 , optionally around 7.864x10 -7 to 1.17x10 -4 , optionally around 1.17x10 -6 to 7.864x10 -5 , optionally around 1.553x10 -6 to 4.029x10 -5 moles of citrate; and/or (b) around 2.959 x 10 -8 to 2.959 x 10 -4 , optionally around 6.155x10 -7 to 2.373x10 -4 , optionally around 1.201x10 -6 to 1.787x10 -4 , optionally around 1.787x10 -6 to 1.201x10 -4 , optionally around 2.3
• a substrate (optionally a sponge) comprising a blocking composition (optionally around 0.15 ml thereof), wherein said blocking composition comprises: one or more metals; one or more blocking composition surfactants; and one or more pH buffers, optionally wherein: around 0.15 ml of the blocking composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: (a) at least around 1.937 x 10 -8 moles of citrate, optionally at least around 4.029x10- 7 , optionally at least around 7.864x10 -7 , optionally at least around 1.17x10 -6 , optionally at least around 1.553x10 -6 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) at least around 2.959 x 10 -8 moles of ethylenediaminetetraacetic acid (EDTA), optionally at least around 6.155x10 -7 , optionally at least around 1.201x
• EDTA ethylenediamine
• the one or more metals may comprise one or more metal salts selected from the group consisting of metal halides (optionally metal chlorides, metal fluorides, metal iodides and metal bromides), metal acetates, metal sulphates, metal carbonates, metal nitrates, metal phosphates, metal gluconates, metal oxides, metal hydroxides, metal citrates, metal lactates, metal glubionates, metal hydrates, metal peroxides, metal hypochlorides, metal dioxides and metal fumarates.
• metal halides optionally metal chlorides, metal fluorides, metal iodides and metal bromides
• metal acetates metal sulphates
• metal carbonates metal nitrates
• metal phosphates metal gluconates
• metal oxides metal hydroxides
• metal citrates metal lactates
• metal glubionates metal hydrates
• metal peroxides metal hypochlorides
• the one or more metals may be selected from the group consisting of zinc, calcium, magnesium, copper, bismuth, indium, manganese, nickel, titanium, chromium, aluminum, lithium, sodium, potassium, beryllium, radium, scandium, yttrium, lanthanum, vanadium, iron, cobalt, iridium, hafnium, silver, thallium, palladium, cadmium, tin, gallium, cerium, praseodymium, neodymium, promethium, samarium, europium, gadolinium, terbium, dysprosium, holmium, erbium, thulium, ytterbium, lutetium, americium, curium, berkelium, californium, thorium, uranium and neptunium, optionally from the group consisting of magnesium, calcium, zinc, copper, bismuth and manganese, or metal ions
• the one or more metals may be selected from the group consisting of Zn 2+ , Ca 2+ , Mg 2+ , Cu 2+ , Cu 3+ , Zn 3+ , Bi 3+ , In 3+ , Mn 2+ , Mn 3+ , Ni 2+ , Ti 3+ , Cr 3+ , Al 3+ , Li + , Na + , K + , Be 2+ , Sr 2+ , Ba 2+ , Ra 2+ , Sc 3+ , Y 3+ , La 3+ , V 2+ , Cr 2+ , Fe 2+ , Co 2+ , V 3+ , Ir 4+ , Hf 4+ , VO 2+ , Ag + , Tl + , Pd 2+ , Cd 2+ , Sn 2+ , Ga 3+ , Tl 3+ , Ce 3+ , Pr 3+ , Nd 3+ , Pm 3+ , Sm 3+ , Eu 3+ , G
• the one or more metals may comprise magnesium, optionally Mg 2+ .
• the one or more metals may comprise calcium and zinc (optionally Ca 2+ and Zn 2+ ).
• the one or more metals may comprise zinc and/or calcium, optionally Zn 2+ and/or Ca 2+ , optionally ZnCl2 and/or CaCl2.
• Said blocking composition may comprise around 0.0001 to 4 M of said one or more metals, optionally around .001 to 2 M; optionally 0.01 to 1 M, optionally around 0.05 to 0.25 M, optionally around 0.1 to 0.15 M.
• the one or more metals comprise ZnCl 2 and CaCl 2 .
• said blocking composition comprises: around 0.00005 to 2 M of ZnCl 2 , optionally around 0.001 to 1 M; optionally 0.005 to 0.05 M, optionally around 0.025 to 0.125 M, optionally around 0.05 to 0.1 M, optionally around 0.06 to 0.08 M; and around 0.00005 to 2 M of CaCl 2 , optionally around 0.001 to 1 M; optionally 0.005 to 0.05 M, optionally around 0.025 to 0.125 M, optionally around 0.03 to 0.07 M, optionally around 0.04 to 0.06 M.
• the one or more metals may have a binding affinity, log K f , for EDTA of at least around 4, optionally at least around 8, 10, 15, 20, 25, 30, 35, 40 or 45. Said metal may have a binding affinity, log Kf, for EDTA of at most around 30, optionally at most around 25, 20, 15, 10 or 8. Said metal may have a binding affinity, log K f , for EDTA of around 4 to 45, optionally around 8 to 30, optionally around 10 to 30, optionally around 15 to 25, optionally around 20 to 25.
• the metal may have a binding affinity for Chromeazurol S of at most 15, optionally at most 13, optionally at most 10 (e.g. per Microchimica Acta 1964, 52, 414–428, log K by the method of Dey et al.).
• the one or more pH buffers may be selected from the group consisting of: glycine, acetate, citrate, phosphate, ethanesulfonic acid and bis-tris methane.
• the one or more pH buffers may be selected from the group consisting of: glycine-hydrochloric acid, sodium acetate (counter-substrate acetic acid), piperazine-N,N′-bis(2-ethanesulfonic acid), citrate, phosphate, phosphate-citrate, 2-(N-morpholino)ethanesulfonic acid, 3-(N- morpholino)propanesulfonic acid and bis-tris methane.
• the one or more pH buffers may provide a pH working range of about 2.6 to 7, optionally 3.5 – 5.8, optionally around 5.6 – 7.
• the one or more pH buffers may provide a buffer capacity ( ⁇ , beta) of about -0.7 to 0.7 mol, optionally about -0.4 to 0.7 mol, optionally about -0.1 to 0.7 mol, optionally about 0.075 to 0.7 mol, optionally about 0.04 to 0.7 mol; and/or optionally about -0.7 to 0.4 mol, optionally about -0.7 to 0.1 mol, optionally about -0.7 to 0.04 mol, optionally about -0.7 to 0.01 mol; and/or optionally about -0.06 to 0.04, optionally about -0.05 to 0.02, optionally about -0.04 to 0.015 mol.
• pH was determined to influence the formation of distinct colour changing complexes and their activity.
• Glycine buffer was tested due to its ability to keep a strong acidic pH (3.5) preventing the rise of pH after the addition of the highly basic detergents. The red colour change was achieved by the release of the protonated dye molecule upon reaction. It was also observed that glycine might have had a catalysing effect on the more stable colour changing complexes. Although the low pH was beneficial in blocking chelator-iron interactions because of protonation, it simultaneously lowered the favoured siderophore-iron interactions. Acetate buffer was investigated as an alternative buffering system due to its greater buffering range of pH 3.6 - 5.8.
• the blocking composition may comprise around 10 mmol/L to 1 mol/L of said one or more pH buffers, optionally around 10 mmol/L to 750 mmol/L, optionally around 50 mmol/L to 500 mmol/L, optionally around 100 mmol/L to 300 mmol/L, optionally around 100 to 200 mmol/L, optionally around 130 to 170 mmol/L.
• the one or more pH buffers may comprise acetate, such as sodium acetate.
• the blocking composition surfactant may comprise: a polysorbate (optionally a polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or optionally a Tween TM ) optionally at a level of around 1 mM to 200 mM; an aliphatic phenol ethoxylate (such as Triton X-100 TM ; optionally wherein the aliphatic group is an alkyl group; optionally wherein the aliphatic phenol ethoxylate is octyl phenol ethoxylate) optionally at a level of around 0.1 mM to 200 mM; a cyclodextrin (optionally ⁇ -cyclodextrin or ⁇ -cyclodextrin); an aliphatic sulfate (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has a straight chain length of 8 to 16 carbon atoms; optionally wherein the aliphatic
• Polysorbate may be helpful to block action of QATs, which are present in a variety of cleaning compositions. Its large structural composition can incorporate the QAT molecule and prevent it from interacting with the colour changing complex. Inclusion of polysorbates (e.g. polysorbate-80) may be helpful to reduce friction of absorbent materials (e.g. sponges/swabs/tips) with target surfaces. Polysorbate surfactants are not believed to interact with the colour changing agent (such as the chrome azurol family, e.g. CAS or CAB).
• An additional advantage of including surfactant-like molecules in the blocking composition is the minimisation of surface tension between the analysed surface and substrate (e.g. the swab/a cotton tip thereof).
• cyclodextrin substrates were tested due to their ability to incorporate QAT molecules and thus to inhibit their interaction with the colour changing system.
• Triton was included in the blocking composition due to its reported property to lyse bacterial membrane. It was hypothesised that the triton molecules would lyse bacteria and release internal siderophores leading to an increase in siderophore sample size and thus enhancing the sensitivity of the colour changing system. It was also observed that triton molecules influenced colour. SDS is known in the literature to have a higher efficiency in lysing cellular membranes. Due to its structural differences to the glycol-based polysorbate and triton substrates, SDS was studied to also have a higher stability in acidic media.
• SDS has a negatively charged head allowing it to interact with the positively charged surfactant molecules of certain dye complexes (e.g. CAS) and thus catalysing the desired reaction, resulting in increased overall sensitivity .
• CAS dye complexes
• Lecithin was found to be extremely effective in limiting QAT interaction.
• Pyruvate substrates were used to deactivate the active component of the bleach cleaners. Pyruvate chemicals react with peroxides and hypochlorites leading to decarboxylation and thus their decomposition.
• the blocking composition surfactant may comprise an aliphatic sulfate (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has a chain length of 8 to 16 carbon atoms (optionally straight chain); optionally wherein the aliphatic sulfate is sodium dodecyl sulfate).
• the blocking composition surfactant (selected from the options above, optionally sodium dodecyl sulfate) may be present from an amount of about 1 x 10 -5 mol/L to about 3.5 x 10 -3 mol/L, optionally about 1 x 10 -4 mol/L to about 3.5 x 10 -3 mol/L, optionally about 2.059x10 -4 to 2.926x10 -3 , optionally about 3.119x10 -4 to 2.352x10 -3 , optionally about 4.178x10 -4 to 1.778x10 -3 , optionally about 5.238x10 -4 to 1.204x10 -3 .
• the blocking composition may further comprise 5-sulfosalicyclic acid.
• 5-sulfosalicyclic acid may be present from an amount of about 5x10 -3 mol / L, optionally 1 x10 -6 mol / L to 2.5x10 -3 mol / L, optionally, 1x10 -4 mol / L to 1x10 -3 mol / L.
• the blocking composition may comprise: ⁇ around 12.5-400 mM acetate buffer at a pH of around 5.6 – 5.8 ⁇ around 0.05-3.5 mM SDS ⁇ around 10-200 mM CaCl 2 ⁇ around 15-250 mM ZnCl 2
• Around 0.05 ml to 0.5 ml of said blocking composition e.g. around 0.1 ml to 0.4 ml, optionally around 0.125 ml to 0.3 ml
• the substrate carrier may be an absorbent material, such as a sponge or wipe.
• the substrate may be able to absorb at least about 5 times its weight (based on the total weight of the substrate) in liquid, such as at least about 7 its weight, such as at least about 10 times its weight, such as at least about 15 times its weight in liquid, such at least about 20 times its weight in liquid.
• the substrate may be able to absorb between about 4-40 times its weight (based on the total weight of the substrate) in liquid, such as between about 4-30 times its weight, such as between about 4-20 times its weight in liquid.
• the substrate may be a non-woven material, such as a fibrous material (optionally paper or a fabric).
• the substrate carrier may comprise cellulose (e.g.
• the substrate carrier may comprise fibres of cellulose, polyester, lignin and/or mixtures thereof (such as lignocellulosic fibres), optionally wherein the substrate carrier comprises cellulose.
• the substrate carrier may comprise pulp, wool, silk, jute, linen, ramie, sisal, bagasse, banana fibres, hemp, flax, camel hair, kenaf and/or mixtures thereof.
• the substrate may be a wipe, such as a surface wipe or a personal (e.g.
• the substrate may be a surface wipe.
• a composition comprising the blocking composition as defined in the first or second aspect. In other words, the third aspect is not limited to the presence of a substrate.
• the composition may comprise: around 0.001 to 3 mol / L of one or more metals selected from the group consisting of: Zn 2+ , optionally wherein said one or more metals are selected from the group consisting of: Zn 2+ , Ca 2+ , Mg 2+ , Cu 2+ , Cu 3+ , Bi 3+ , Mn 2+ , Mn 3+ , Cr 2+ , Cr 3+ , Li + , K + , Na + , Be 2+ and Fe 2+ ; optionally wherein said one or more metals are selected from the group consisting of: Zn 2+ , Ca 2+ , Mg 2+ , Fe 2+ , Cr 3+ , Mn 2+ and Bi 3+ .
• the one or more metals may be present as salts selected from the group consisting of: metal halides, metal acetates, metal carbonates and metal sulfates.
• the composition may comprise around 10 to 750 mmol / L of buffer selected from the group consisting of: acetate, glycine, ethanesulfonic acid, bis-tris methane and phosphate.
• the composition may comprise: (a) around 0.01 to 10 mmol / L of an aliphatic sulfate; or (b) around 1 to 100 mmol / L of an aliphatic phenol ethoxylate; or (c) around 10 to 200 mmol / L of a polysorbate.
• the composition may comprise around 0.001 to 2 mol / L of the one or more metals, optionally around 0.001 to 1.5 mol / L, optionally around 0.001 to 1 mol / L.
• the composition may comprise: around 0.001 to 3 mol / L of one or more metals are selected from the group consisting of: Zn 2+ , Ca 2+ , Mg 2+ , Fe 2+ , Cr 3+ , Mn 2+ and Bi 3+ ; around 10 to 750 mmol / L buffer (optionally according to claim 25, optionally acetate buffer); around 0.01 to 10 mmol / L surfactant (optionally an aliphatic sulfate, optionally sodium dodecyl sulfate).
• the buffer may be selected from the group consisting of: acetate, glycine, ethanesulfonic acid, bis-tris methane and phosphate; optionally wherein the buffer is acetate.
• the surfactant may be an aliphatic sulfate (optionally sodium dodecyl sulfate).
• the one or more metals may be Zn 2+ and Ca 2+ .
• the composition may comprise: around 0.001 to 1.5 mol / L of Zn 2+ ; around 0.001 to 1.5 mol / L of Ca 2+ .
• the composition may comprise: around 10 to 750 mmol / L of acetate buffer.
• the composition may comprise: around 0.01 to 10 mmol / L of an aliphatic sulfate.
• the composition may comprise: around 0.001 to 1 mol / L of Zn 2+ ; around 0.001 to 1 mol / L of Ca 2+ ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• the composition may comprise: around 15 to 250 mmol / L of Zn 2+ ; around 10 to 200 mmol / L of Ca 2+ ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• the one or more metals may comprise CaCl 2 and ZnCl 2 ; and the blocking composition may comprise a sodium dodecyl sulfate blocking composition surfactant.
• the composition may comprise: around 0.001 to 1.5 mol / L of ZnCl 2 ; around 0.001 to 1.5 mol / L of CaCl 2 ; around 10 to 750 mmol / L of acetate buffer; around 0.01 to 10 mmol / L sodium dodecyl sulfate.
• the composition may comprise: around 0.001 to 1 mol / L of ZnCl 2 ; around 0.001 to 1 mol / L of CaCl 2 .
• the composition may comprise around 12.5 to 400 mmol / L of acetate buffer.
• the composition may comprise around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• the composition may comprise: around 12.5 to 400 mmol / L of acetate buffer; and around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• the composition may comprise: around 0.001 to 1 mol / L of ZnCl 2 ; around 0.001 to 1 mol / L of CaCl 2 ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• the blocking composition may comprise: ⁇ around 15 mmol/L to 250 mmol/L of ZnCl 2 ; ⁇ around 10 mmol/L to 200 mmol/L of CaCl 2 ; ⁇ around 12.5 mmol/L to 400 mmol/L acetate buffer; ⁇ around 0.05-3.5 mM SDS.
• a kit comprising: the substrate according to the first or second aspect and/or the composition according to the third aspect; and (a) a colour changing composition configured to detect the presence of microorganisms; and/or (b) a cleaning composition configured to reduce and/or eliminate microorganisms.
• the colour changing agent may be a chromeazurol (such as chromeazurol S or chromeazurol B) or a tannin.
• the colour changing agent may be chromeazurol S (CAS).
• Chromeazurol compounds form a blue colour upon binding with iron and are coloured orange in the absence of iron. It will be appreciated that chromeazurol compounds may exist in different isomeric forms and/or salt forms and/or de/protonated forms, particularly as a result of conjugation in the compounds. All such forms are envisaged herein.
• Tannins are a class of polyphenolic compounds, such as vescalagin, castalagin [(33beta)-isomer of vescalagin], penta-O-galloyl-beta-D-glucose.
• Said substrate may comprise around 0.05 ml to 0.75 ml of said blocking composition, optionally around 0.1 ml to 0.6 ml, optionally around 0.125 ml to 0.5 ml, optionally around 0.15 ml.
• Said colour changing composition may be present in an amount around 0.1 ml to 1 ml, optionally around 0.2 ml to 0.7 ml, optionally around 0.3 ml to 0.5 ml.
• Said colour changing composition may comprise: a colour changing composition metal, and a colour changing agent, wherein said colour changing composition metal is bindable to said colour changing agent to provide a change in colour on binding and/or release thereof.
• the kit may further comprise a colour changing composition surfactant, optionally wherein said colour changing composition metal and colour changing composition surfactant are present at a molar ratio of about 1:0.25 to 30, optionally about 1:0.5 to 7; optionally about 1:0.75 to 6; optionally about 1:0.75 to 5; optionally about 1:0.75 to 3; optionally about 1:1 to 3; optionally about 1:1 to 2.5.
• Said colour changing composition surfactant may be selected from the group consisting of: a polysorbate (optionally a polysorbate 80 or polysorbate 20, optionally a Tween TM ), an aliphatic phenol ethoxylate (such as Triton X-100; optionally wherein the aliphatic group is an alkyl group; optionally wherein the aliphatic phenol ethoxylate is octyl phenol ethoxylate), an aliphatic sulfobetaine (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has a chain length of 8 to 16 carbon atoms (optionally straight chain); optionally wherein the aliphatic sulfobetaine is lauryl sulfobetaine), an aliphatic quaternary ammonium halide such as aliphatic trimethylammonium halide (optionally wherein the aliphatic group is an alkyl group, optional
• Said colour changing composition metal and colour changing agent may be present at a molar ratio of about 1:0.25 to 25; optionally about 1:0.25 to 20, optionally 1:0.5 to 15; optionally about 1:1 to 13; optionally about 1:1.5 to 11.5; optionally about 1:1.5 to about 8; optionally about 1:1.5 to 5; optionally about 1:1.5 to 4; optionally about 1:1.5.
• Said colour changing composition metal may be iron; optionally iron (III); optionally FeCl 3 , optionally hydrated FeCl 3 (FeCl 3 ⁇ 6 H2O).
• Said colour changing agent may be chromeazurol (such as chromeazurol S or chromeazurol B) or a tannin, optionally wherein said colour changing agent is a chromeazurol, optionally chromeazurol S (CAS).
• the colour changing agent such as CAS (e.g. CAS S) may be tetrabasic, having 4 ionizable hydrogen moieties. Ionization of one or more of said moieties may lead to a colour change. A chemical capable of achieving said ionization may therefore be detectable upon exposure to such a colour changing agent.
• the skilled person will understand the nature of chemicals for which the composition is suitable for detecting.
• the colour changing agent may provide a change in colour in the visible spectrum (e.g.
• the colour changing composition may comprise: ⁇ Fe : CAS : (C 10-16 )-trimethylammonium Bromide/Chloride ⁇ Equivalent molar ratios around 1 : 1-3 : 0-4 (e.g.1-4) Around 0.1 ml to 1 ml (e.g. around 0.2 ml to 0.7 ml, optionally around 0.3 ml to 0.5 ml) of this solution may be used as the colour change liquid (e.g. in the swab reservoir).
• Said cleaning composition may comprise: an n-alkyl dialkyl triamine (e.g.
• n-alkyl is at least C 10 , optionally C 12 ; and/or optionally wherein the trialkyl is dipropylene, optionally wherein the n-alkyl dialkyl triamine is dodecyl dipropylene triamine), n-alkyl dimethyl ammonium halide (e.g.
• n-alkyl dialkyl aryl ammonium halide is n-alkyl dimethyl benzyl ammonium halide, optionally n-alkyl dimethyl benzyl ammonium chloride, optionally wherein the n-alkyl is at least C 8 (e.g.
• Aliphatic alcohol alkanoate is an alkyl alcohol alkanoate, optionally wherein the aliphatic group has a chain length of 8 to 18 carbon atoms (optionally straight chain), optionally wherein the aliphatic group has a chain length of 9 to 11 carbon atoms (optionally straight chain).
• Aliphatic alcohol alkanoate may be an alkyl alcohol ethoxylate, optionally C 8 -C 18 alkyl alcohol ethoxylate, optionally C 9 -C 11 alkyl alcohol ethoxylate.
• Said cleaning composition may comprise n-alkyl dialkyl aryl ammonium halide present from about 0.5 wt% to 15 wt%; and aliphatic alcohol alkanoate present from about 0.5 wt% to 15 wt%.
• Said cleaning composition may further compromise sodium hypochlorite present from about 1.8 wt% to 7 wt%, optionally 2.1 wt% to 6.5 wt%, optionally 2.4 wt% to 6 wt%, optionally 2.7 wt% to 5.5 wt%, optionally about 3 wt% to 5 wt%; and hydrogen peroxide present from about 10 wt% to 30 wt%; and peracetic acid present from about 10 wt% to 30 wt%; and acetic acid present from about 10 wt% to 30 wt%.
• Said cleaning composition may comprise: n-alkyl dialkyl aryl ammonium halide present from about 0.6 wt% to 14 wt%, optionally 0.7 wt% to 13 wt%, optionally 0.8 wt% to 12 wt%, optionally 0.9 wt% to 11 wt%, optionally about 1 wt% to 10 wt%; and aliphatic alcohol alkanoate present from about 0.6 wt% to 14 wt%, optionally 0.7 wt% to 13 wt%, optionally 0.8 wt% to 12 wt%, optionally 0.9 wt% to 11 wt%, optionally about 1 wt% to 10 wt%.
• Said cleaning composition may further comprise citrate present from about 0.5 wt% to 10 wt%; optionally about 0.6 wt% to 9 wt%, optionally 0.7 wt% to 8 wt%, optionally 0.8 wt% to 7 wt%, optionally 0.9 wt% to 6 wt%, optionally about 1 wt% to 5 wt%.
• Said cleaning composition may further comprise EDTA present from about 0.5 wt% to 30 wt%; optionally about 0.6 wt% to 25 wt%, optionally 0.7 wt% to 20 wt%, optionally 0.8 wt% to 15 wt%, optionally 0.9 wt% to 12.5 wt%, optionally, about 1 wt% to 10 wt%.
• EDTA present from about 0.5 wt% to 30 wt%; optionally about 0.6 wt% to 25 wt%, optionally 0.7 wt% to 20 wt%, optionally 0.8 wt% to 15 wt%, optionally 0.9 wt% to 12.5 wt%, optionally, about 1 wt% to 10 wt%.
• Said cleaning composition may further comprise: sodium hypochlorite present from about 1.8 wt% to 7 wt%, optionally 2.1 wt% to 6.5 wt%, optionally 2.4 wt% to 6 wt%, optionally 2.7 wt% to 5.5 wt%, optionally about 3 wt% to 5 wt%; and hydrogen peroxide present from about 6 wt% to 42 wt%, optionally 7 wt% to 39 wt%, optionally 8 wt% to 36 wt%, optionally 9 wt% to 33 wt%, optionally, about 10 wt% to 30 wt%; and peracetic acid present from about 6 wt% to 42 wt%, optionally 7 wt% to 39 wt%, optionally 8 wt% to 36 wt%, optionally 9 wt% to 33 wt%, optionally about 10 wt% to 30 wt%; and acetic acid present from
• Said colour changing composition may comprise: a colour changing composition metal, wherein said colour changing composition is iron; a colour changing agent, wherein said colour changing agent is chromeazurol S; and optionally a colour changing composition surfactant, wherein said colour changing composition surfactant is dodecyltrimethylammonium bromide or dodecyltrimethylammonium chloride; wherein said iron is bindable to chromeazurol S to provide a change in colour on binding and/or release thereof, optionally wherein said iron and said aliphatic quaternary ammonium halide are present at a molar ratio of 1:0.25 - 1:5; and wherein said iron and said chromeazurol S are present at a molar ratio of 1:0.5 - 1:5.
• Said cleaning composition may comprise: n-alkyl dialkyl aryl ammonium halide, wherein said n-alkyl dialkyl aryl ammonium halide is n-alkyl dimethyl benzyl ammonium chloride present at an amount from about 1 wt% to 10 wt%; and aliphatic alcohol alkanoate, wherein said aliphatic alcohol alkanoate is C 9 -C 11 alkyl alcohol ethoxylate present at an amount from about 1 wt% to 10 wt%, optionally wherein said cleaning composition further comprises citrate, optionally present at an amount from about 1 wt% to 5 wt% and/or EDTA, optionally present at an amount from about 1 wt% to 10 wt%, optionally wherein said cleaning composition further compromises sodium hypochlorite, optionally present at an amount from about 3 wt% to 5 wt%; and/or hydrogen peroxide, optionally present at an amount from about 10 w
• a swab comprising a rod having a substrate according to the first or second aspect at an end thereof (e.g. at a tip thereof, optionally wherein the substrate comprises 0.15 ml of said composition), optionally further comprising a reservoir of a colour changing composition (e.g. at an opposite end to the tip end).
• the reservoir may be rupturable to release colour changing composition (e.g. to enable it to interact with the swab substrate/tip).
• the swab may be provided with a collecting vial, to facilitate mixture of the colour changing composition with the blocking composition on/around the substrate.
• a method comprising contacting a surface, or a sample therefrom, that has been pre-treated with a cleaning composition with the composition of the third aspect. Also provided herein is a method comprising contacting a surface that has been pre-treated with a cleaning composition with a substrate according to the first or second aspect or a swab as defined in the fifth aspect, to collect a sample therefrom. The method may further comprise contacting said surface, or sample therefrom (e.g.
• first, second, third, fourth, fifth, sixth and seventh aspects are interrelated.
• features described above in relation to one or more of the first, second, third, fourth, fifth, sixth and seventh aspects apply mutatis mutandis to the subject matter of any one or more of the other aspects as will be readily understood by a person of skill in the art.
• BRIEF DESCRIPTION OF FIGURES Figs.1 to 10 absorption spectroscopy traces.
• Figs.11 to 24 photographs of vials of compositions.
• Fig.25(A&B) absorption spectroscopy traces.
• Fig.26 pH vs absorbance measurements.
• stock colour changing complex solutions including the following: ⁇ FreshCheck Complexes: o Complexes are given in the ratio of Iron : dye : surfactant. The 3 letters preceding the ratios (e.g. FCH) indicate the metal ion : dye : surfactant/aggregation inhibitor.
• an analogue colour changing complex (based on iron-CAS-S colour changing dye as discussed above) was mixed with a range of chemicals (D10, BioK and bleach) typically found in commonly-available cleaning compositions, as well as siderophore-based composition Dfx. Light absorption spectroscopy measurements were conducted, and the results are shown in Fig.1. In the absence of cleaning chemicals, the colour changing complex adopts two distinct curves depending on the presence or absence of siderophores.
• the solid curve depicts a metal-bound colour changing complex (absence of siderophores) and the dash-dot-dot curve (peak ⁇ 495 nm) depicts the protonated colour changing complex (presence of siderophores), wherein metal has been released from the colour changing complex through chelation by the siderophores.
• a colour change shift can therefore clearly be seen, depending on the presence or absence of siderophores, which in turn indicates the presence or absence of microbes. Interaction with common cleaning chemicals interferes with this process, leading to several new peaks/curves in the light spectrum and consequent contamination of the colour-change shift as can be seen in Fig.1.
• Example 1 – glycine buffer blocking of bleach Glycine buffer at pH 3.0 – 4.0 (150 ⁇ l, 50 - 150 mM), bleach (100 ⁇ l), BioKlenz (100 ⁇ l) and D10 (100 ⁇ l) were added to the stock colour changing complex solution (400 ⁇ l).
• the buffer blocked bleach as shown with the long dashes curve which no longer appears with a peak in the ⁇ 427 nm region and instead appears approximately aligned with solid metal-bound dye complex ⁇ 576 nm).
• blocking here occurred by inhibiting the increase of pH after the addition of bleach.
• Example 2(B) – metal blocking of TEAs (combination effect with glycine buffer)
• glycine buffer solution 150 ⁇ l containing 50 - 150 mM glycine and 0.006 – 0.008 mol/L magnesium chloride
• the stock colour changing complex solution 400 ⁇ l
• the common cleaning chemicals bleach 100 ⁇ l, BioK 100 ⁇ l and D10100 ⁇ l; see Fig.3B).
• the undesired release of free dye as result of TEA interaction was blocked (see short dashes curve, which no longer appears with a peak in the ⁇ 430 nm region).
• the QAT and TEA components of the common cleaning chemicals still undesirably interact with the dye complex, despite the combination of glycine buffer and magnesium chloride (both eliciting peaks ⁇ 656 nm), e.g. forming a dye-QAT complex.
• Fig.4 shows the decay of the dye-QAT complex as the result of the interaction of Polysorbate-80 (long dash, with peak spread between ⁇ 490 and 690 nm).
• Example 4 5-sulfosaliyclic acid effect on colour changing speed
• 5-sulfosalicyclic acid was investigated as a potential catalyst. Without wishing to be bound by theory, it is believed that 5-sulfosalicyclic acid can enhance the reactivity of metal- dye (e.g. CAS)-surfactant complexes by competing with the colour changing dye and coordinating to metal (e.g. iron). It will be appreciated that a balance is needed, between inducing a quicker colour change on the one hand and degrading the complex on the other.
• metal- dye e.g. CAS
• metal e.g. iron
• Fig.5 shows the interaction of 5-sulfosalicyclic acid with the dye complex in the stock colour changing complex solution (400 ⁇ l).
• Low levels of 5-sulfosalicyclic acid show no change to the peak, but medium to high levels show degradation of the peak. This suggests a limit for 5-sulfosalicyclic acid based on the degradation of the formulation.
• glycine was used as a buffering agent, enabling the pH of the stock colour changing complex solution to be kept at low acidic pH ( ⁇ 3.5).
• the low pH enabled a number of factors: (i) pH blocking of basic detergents found in common cleaning compositions, (ii) regulation of the binding affinity (K d ) of chelators to metal/iron in the presence of the dye; and (iii) influencing the colour of bound and unbound dye. It was determined that the low pH of the solution simultaneously lowered the binding affinity of siderophore molecules to metal/iron by protonating siderophore binding sites.
• acetate buffer was next investigated as a potential buffer to stabilise the pH of the stock colour changing complex solution after the addition of a detergent.
• Fig.6 breaks down the interaction of 400 ⁇ l of the stock colour changing complex solution with a range of common cleaning chemicals (upper graph is with 100 ⁇ l D10; middle is with 100 ⁇ l BioK,; and lower is with 100 ⁇ l bleach-based cleaning formulation) in both the absence (milli-Q purified water, MQ, dashed curve) and the presence (solid curve) of an acetate buffer (150 ⁇ l, 100-200 mM at pH 5.4-5.8).
• the acetate buffer stabilises the colour changing complex against the tested common cleaning chemicals (i.e. blocks these) and prevents a following colour change to bright green or yellow.
• vial colouring vial colouring: upper graph: upper teal, lower blue; middle graph: upper yellow, lower green; lower graph: upper yellow, lower teal).
• Fig.7 depicts the results of experiments conducted to demonstrate the effect of acetate buffer (150 ⁇ l, 100 - 200 mM) on 400 ⁇ l of the specific colour changing composition described in the general exemplary section above having a molar ratio Fe : CAS : HDTMA of 1:2-3:2- (based on 4.99 * 10 ⁇ -5 mol/L Fe in 400 ⁇ l) and the results are shown in Fig.7.
• acetate buffer 150 ⁇ l, 100 - 200 mM
• acetate buffer 150 ⁇ l, 25 - 125 mM
• polysorbate 80 5-10 molar equivalents to Fe
• magnesium chloride 1-5 molar equivalents to EDTA
• Example 7 pH effect on colour change
• the stability of the dye complex can be influenced by the pH of the buffer and thus its interaction with common cleaning chemicals can vary in the presence of the blocking composition.
• the following solution was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : dodecyl pyridinium bromide of 1:2 - 3:1 - 3 (based on 2.50 * 10 ⁇ -5 mol/L Fe in 200 ⁇ l).
• a series of vials was prepared with the solution above, with D10 or Dfx at various dilution levels as indicated in the tables below. In the tables below, the layout of cells (upper left to lower right) matches the layout of vials in Figs.11(A) and (B).
• Fig.11(A) – vial layout In general terms, the D101x vial had the deepest blue colour, extending progressively towards a green/teal colour at 0.1x with a notable change towards green/teal at 0.5x and below. Vials 0.9x to 0x remained approximately consistent with a green/teal colour similar to that of 0.1x. For Dfx, the 1x vial had a yellow colour, transitioning through green between 0.6 to 0.4x and on to teal at 0.1x.
• Fig.11(B) – vial layout In general terms, the D101x vial had a blue-purple colour, extending progressively towards a blue colour at 0.5x, transitioning towards purple again at 0.4x with a notable change towards red at 0.3x and below.
• the 1x vial had an orange colour, transitioning progressively towards purple at 0.1x.
• a vial of D10 at 1x had a blue colour and a vial of Dfx at 1x had a yellow colour.
• the following solution composition was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : dodecyltrimethyl ammonium chloride of 1:2-3:1-3 (based on 2.50 * 10 ⁇ -5mol/L Fe in 200 ⁇ l).
• a series of vials was prepared with the composition above, with EDTA or citric acid at various dilution levels as indicated in the tables below.
• the layout of cells (upper left to lower right) matches the layout of vials in Figs.12 and 13.
• the ZnCl 2 vials had the best performance with CaCl 2 next, followed by MgCl 2 . Deeper blues were generally encountered with lower equivalent levels of EDTA. By changing the metal ion from magnesium to calcium or zinc, the efficiency of chelator blocking was increased maintaining the blue colour changing composition colour at higher EDTA concentrations. Zinc chloride was the preferred chelator inhibitor as it blocked EDTA (1x to 0.001x) and was the most successful in blocking citrate. The middle row shows that CaCl 2 can block EDTA up to around 0.5x (which is representative of around 0.5x a standard level of cleaning chemicals typically found in real-world applications).
• the bottom row shows that ZnCl 2 can block EDTA (except when EDTA is encountered in very high concentrations).
• Fig.13 – vial layout Vials labelled with bold underlined text above had a blue colour (i.e. were not substantially affected by citric acid presence/citric acid was blocked) while the remaining vials were green.
• the top row shows that MgCl 2 is able to block/stabilise the colour change complex up to around 0.01x dilution.
• the middle row shows that CaCl 2 can block citric acid to approximately the level as MgCl 2 .
• the bottom row shows that ZnCl 2 can block citric acid up to around 0.1x – 0.5x concentration.
• Example 10 metal effect on stability and sensitivity Copper chloride amongst others has been reported in the literature to have a higher binding efficacy to EDTA than zinc chloride. Consequently, it was incorporated into the blocking composition and investigated for chelator inhibition at lower concentrations.
• the following solution composition was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : dodecyltrimethyl ammonium chloride of 1:2-3:0.5 - 3 (based on 2.50 * 10 ⁇ -5 mol/L Fe in 200 ⁇ l).
• a series of vials was prepared with the composition above, with citric acid at 0.5x or 0.05x dilution levels as indicated in the tables below.
• Fig.15 – vial layout Vials labelled with bold underlined text above had a blue colour (i.e. were not substantially affected) while the remaining vials were green (albeit 0.05x citrate dilution for the 0.001x and 0x vials had a blue tinge).
• the addition of CuCl 2 caused the colour changing composition to become a much darker blue than the control (not shown) but prevent the interference of citrate with the dye complex, until there was less than 0.01 mol of CuCl 2 compared to Fe.
• Example 11 metal effect on stability and sensitivity
• the following solution composition was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : HDTMA of 1:2 - 3:0.5 - 3 (based on 2.50 * 10 ⁇ -5 mol/L Fe in 400 ⁇ l).
• a series of vials was prepared with the composition above, with D10 or Dfx at various dilution levels as indicated in the tables below. Each dilution was conducted in pairs (i.e. each cell below represents a pair of vials), with a left hand (0 equivalents CaCl 2 ) and a right hand (50 equivalents CaCl 2 ) vial in each pair.
• Fig.16 – vial layout All upper vials were teal coloured. The 0.4x Dfx vials were yellow, while the remainder were green-blue (with the deepest blues in the 0.1x and 0x control vials). The level of CaCl 2 had limited effect on the presence of D10, except where D10 was at high levels (0.1x standard cleaning solution) where it prevented the solution turning yellow. The CaCl 2 also had limited effect on the potential for the siderophore DFX to bind to the Fe centre of the colour changing composition. The experiment was repeated, with 100 (left hand vial in each pair) or 150 (right hand vial in each pair) equivalents of CaCl 2 .
• Fig.17(A) which matches the vial layout in Fig.16
• Fig.17(B) which shows a specific comparison of the D10 vials at 0.06x and 0.05x against a MilliQ control and varying concentrations of calcium chloride. All upper vials were teal coloured. The 0.4x and 0.3x Dfx vials were yellow, while the remainder were green-blue (with the deepest blues in the 0.1x and 0x control vials).
• the level of CaCl 2 again had limited effect on the presence of D10, except where D10 was at high levels (0.2x standard cleaning solution), where it prevented the solution turning yellow.
• the CaCl 2 also had limited effect on the potential for the siderophore DFX to bind to the Fe centre of the colour changing composition.
• the following solution composition was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : surfactant of 1:2-3:0.5-3 (based on 0.0499 mol/mL Fe in 400 ⁇ l).
• a series of vials was prepared with the composition above, with D10 or Dfx at various dilution levels as indicated in the tables below. Each dilution was conducted in pairs (i.e.
• each cell below represents a pair of vials), with a left hand (of FCP) and a right hand (a comparator formulation of FCD) vial in each pair.
• Fig.18 – vial layout All upper vials were teal coloured. The 0.4x and 0.3x Dfx vials were yellow, while the remainder were green-blue (with the deepest blues in the 0.1x and 0x control vials).
• FCD showed good stability to D10-mediated colour change, with sensitivity to siderophore (deferoxamine) at 0.2x equivalence to iron and above. FCD showed some colour change, but the colour change with Dfx occurred at a lower concentration and was more distinct.
• Example 13 Interaction of FCD complex with D10 or Dfx with (Fig.19) or without (Fig. 20) a blocking composition
• the following solution composition was used:
• FCD becomes discoloured and a brighter blue than the control.
• Fig.20 shows that the blocking composition still allows FCD to interact with a siderophore (deferoxamine) as the colour change become more intense as more Dfx is added (right to left). The effect of Dfx on colour is more apparent on the left than on the right, showing that the blocking composition prevents unwanted D10 interaction, whilst encouraging sensitivity to Dfx.
• siderophore deferoxamine
• the following solution composition was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : HDTMA of 1:2-3:0.5 - 3 (based on 0.0499 mol/mL Fe in 400 ⁇ l).
• a series of vials was prepared with the composition above with D10 or Dfx at various dilution levels as indicated in the tables below.
• Fig.21 – vial layout All upper and middle vials were blue or teal coloured, with the deepest blues being at 1x D10 dilution.
• the 1x to 0.6x Dfx vials were yellow, 0.5x to 0.3x green-teal and 0.2x to 0.1x blue.
• the test was repeated with the same vial layout, but with 3 equivalents MgCl 2 (relative to EDTA) instead of ZnCl 2 and CaCl 2 .
• the results are shown in Fig.22.
• the top two lines of both figures show a reducing level of D10 from left to right and top to bottom.
• the bottom line shows a reducing level of siderophore reducing in concentration.
• the blocking composition that includes MgCl 2 (Fig.22) showed some discolouration with D10 from below 0.3x – 0.04x dilution compared to stock concentration.
• the effect of Dfx on the MgCl 2 blocking composition is apparent from 0.1x and up.
• the blocking composition that includes the ZnCl 2 and CaCl 2 (Fig.20) showed a smaller sensitivity range to D10 (0.09x – 0.07x) and still had sensitivity to DFX from 0.2x and up. This shows a clear trade-off between metal ions that prevent D10 interference, and the effect of the metal ions on the sensitivity to DFX.
• Example 15 – metal effect on stability and sensitivity the following solution composition was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : HDTMA of 1:2-3:0.5-3 (based on 0.0499 mol/L Fe in 400 ⁇ l).
• a series of vials was prepared with the composition above with D10 or Dfx at various dilution levels as indicated in the tables below.
• Fig.23 – vial layout All upper vials were blue with the exception of 0.3-0.1x, which progressively became more teal coloured. The middle row was teal. The 1x to 0.6x Dfx vials were yellow, 0.5x to 0.1x green- teal. The experiment was repeated, but with 50 equivalents CaCl 2 (relative to EDTA) and no ZnCl 2 .
• Fig.24 – vial layout All upper vials were blue with the exception. The middle two rows were teal.
• the bottom row was yellow-teal, with deeper yellows towards the 1x side and deeper teal towards 0.1x.
• the range of sensitivity to D10 was increased compared to ZnCl 2 at 50 equivalents, but the sensitivity to Dfx was increased. This further shows that a lower amount of ZnCl 2 has a direct effect on sensitivity to both cleaning chemicals and bacterial residues.
• Example 16 the effect of detergent on colour change sensitivity
• the stability & sensitivity of the dye complex can be influenced by the amount of surfactant (e.g. HDTMA) that is present.
• surfactant e.g. HDTMA
• the following solution was used: *described in the general exemplary section above, here having a molar ratio Fe : CAS : (C12-16) trimethyl ammonium chloride of 1:2 - 3:0 - 4 (based on 4.99 * 10 ⁇ -5 mol/L Fe in 400 ⁇ l).
• An increased level of surfactant led to preservation of the peak from 600 – 710 nm, and reduced the increase of the peak from 450 – 460 nm when DFX was introduced.
• Example 17 Stock solutions of compositions were prepared by mixing the following solutions: Stock solution A: ⁇ 50 ml of a solution comprising 0.06 g CAS in 50 ml H 2 O; ⁇ 9 ml of a solution comprising 0.0027 g hydrated FeCl 3 (FeCl 3 ⁇ 6H 2 O) in 10 ml 10 mM HCl; ⁇ 8 ml of a solution comprising 0.0146 g HDTMA in 8 ml H 2 O; and ⁇ 33 ml of a solution comprising 2 g Tween ® 80 in 33 ml H 2 O
• Stock solution B ⁇ 50 ml of a solution comprising 0.06 g CAS in 50 ml H 2 O; ⁇ 9 ml of a solution comprising 0.0081 g hydrated FeCl 3 (FeCl 3 ⁇ 6H 2 O) in 10 ml 10 mM HCl; ⁇ 8 ml of a solution comprising 0.0146 g HDTM
• compositions for use in a label.
• Compositions may be added to the label by mixing with the carrier (e.g. agar) at a temperature of about 40 °C.
• the carrier e.g. agar
• Example 18 A range of tests were performed to investigate the colouring effect of increasing molar ratio of iron relative to CAS S (hydrated FeCl 3 (FeCl 3 ⁇ 6H 2 O) to CAS S).
• Example 20 Membrane testing was performed with 10 kDa MWCO dialysis tubing. Dialysis tubing was cut to 10cm in length to form a cylinder of tubing open and both ends. After tying one end of the dialysis tubing, the formulation was added to 5ml total volume and then the second end was tied off to prevent any leaking of the formulation from the ends of the tubing.
• Example 21 An exemplary composition suitable for inclusion in a substrate, such as a surface wipe, is given below. 0.0008475 g of the composition was mixed with 12.25 mL water and then doped into a 10x10 cm cellulose wipe.
• Example 22 An experiment was conducted to demonstrate the effect of pH on the colour of compositions.
• compositions having pH between 0.8 and 12.8 were prepared in accordance with the following procedure.
• Preparatory solutions with a pH between 0.8 and 6.8 were prepared from a stock solution comprising hydrochloric acid (1 mL, 37%) in distilled water (50 mL) and then diluted with sufficient further distilled water to yield solutions having a pH level 0.3 units lower than that intended for the final compositions for testing (e.g. where the final composition for testing was intended to have a pH of 1.8, then the preparatory solution was prepared by diluting the stock solution with further distilled water to a pH of 1.5).
• Preparatory solutions with a pH between 7.8 and 12.8 were prepared from a stock solution comprising sodium hydroxide (0.4 g) in distilled water (50 mL) and then diluted with further distilled water to yield solutions having a pH level 0.3 units higher than that intended for the final compositions for testing. Preparatory solutions were then diluted 1:1 (volume) with a water-mixed composition prepared in accordance with Example 21, to yield final compositions for testing having the desired pH. The (unbuffered) final compositions for testing were observed to have colouring as set out in the table below.
• Example 23 validation of blocking effectiveness
• a method adapted from BS EN 1276:2019 was used to evaluate the blocking effectiveness of the blocking composition and any false negatives.
• a culture of Escherichia coli was produced and adjusted to 1.5-5.0x10 8 CFU*/mL according to BS EN 1276:2019 to form a cell suspension. This organism was chosen due to its sensitivity to quaternary ammonium compounds.
• the cell suspension was then further diluted to achieve a cell count of 3.0x10 2 to 1.6x10 3 CFU/mL to form a validation suspension.
• This validation suspension was then used in the blocking test.
• 100 ⁇ L of a commercial quaternary ammonium compound (QAC)-based disinfectant (HycolinTM hospital disinfectant, diluted to 2% by volume) was added to 550 ⁇ L of blocking composition according to the present disclosure (including 150 ⁇ L of buffer) and incubated at ambient temperature for 5 minutes to allow blocking of the disinfectant.
• 50 ⁇ L of the E. coli validation suspension was added to the tube and mixed thoroughly. Samples were incubated at ambient temperature for 5 minutes contact time, allowing exposure of the test organism to any residual un-blocked disinfectant.
• a substrate (optionally a sponge) comprising a blocking composition
• said blocking composition comprises: one or more metals; one or more blocking composition surfactants; and one or more pH buffers; wherein 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at least around 1.372 x 10 -8 moles of n-alkyl dimethyl benzyl ammonium chloride; and at least around 3.106 x 10 -8 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at least around 2.854x10 -7 , optionally at least around 5.57x10 -7 , optionally at least around 8.287x10 -7 , optionally at least around 1.1x10 -6 , optionally around 1.3x10 -6 , such as around 1.372x10 -6 moles of n-alkyl dimethyl benzyl ammonium chloride; and at least around 6.46x10 -7 , optionally at least around 1.261x10 -6 , optionally at least around 1.876x10 -6 , optionally at least around 2.491x10 -6 , optionally around 3.1x10 -6 such as around 3.106x10 -6 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: at most around 1.372 x 10 -4 , optionally at most around 1.1x10 -4 , optionally at most around 8.287x10 -5 , optionally at most around 5.57x10 -5 , optionally at most around 2.854x10- 5 , around 1.3x10 -6 , such as around 1.372x10 -6 moles of n-alkyl dimethyl benzyl ammonium chloride; and at most around 3.106 x 10 -4 , optionally at most around 2.491x10 -4 , optionally at most around 1.876x10 -4 , optionally at most around 1.261x10 -4 , optionally at most around 6.46x10- 5 , optionally around 3.1x10 -6 such as around 3.106x10 -6 moles of C 9 -C 11 alkyl alcohol ethoxylate.
• 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: around 1.372x10 -8 to 1.372x10 -4 , optionally around 2.854x10 -7 to 1.1x10 -4 , optionally around 5.57x10 -7 to 8.287x10 -5 , optionally around 8.287x10 -7 to 5.57x10 -5 , optionally around 1.1x10 -6 to 2.854x10 -5 moles of n-alkyl dimethyl benzyl ammonium chloride; and around 3.106x10 -8 to 3.106x10 -4 , optionally around 6.46x10 -7 to 2.491x10 -4 , optionally around 1.261x10 -6 to 1.876x10 -4 , optionally around 1.876x10 -6 to 1.261x10 -4 , optionally around 2.491x10 -6 to 6.46x10 -5 moles of C 9 -C
• 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block said amount of n-alkyl dimethyl benzyl ammonium chloride and C 9 -C 11 alkyl alcohol ethoxylate, and at least: (a) around 1.937x 10 -8 moles of citrate, optionally at least around 4.029x10 -7 , optionally at least around 7.864x10 -7 , optionally at least around 1.17x10 -6 , optionally at least around 1.553x10 -6 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) around 2.959 x 10 -8 , optionally at least around 6.155x10 -7 , optionally at least around 1.201x10 -6 , optionally at least around 1.787x10 -6 , optionally at least around 2.373x10 -6 , optionally around 3x10 -6 such as around
• 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block said amount of n-alkyl dimethyl benzyl ammonium chloride and C 9 -C 11 alkyl alcohol ethoxylate, and at most: (a) around 1.937x10 -4 moles of citrate, optionally at most around 1.553x10 -4 , optionally at most around 1.17x10 -4 , optionally at most around 7.864x10 -5 , optionally at most around 4.029x10 -5 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) around 2.959x10 -4 moles of ethylenediaminetetraacetic acid (EDTA), optionally at most around 2.373x10 -4 , optionally at most around 1.787x10 -4 , optionally at most around 1.201x10 -4 , optionally at most around 6.155x10
• EDTA ethylenediaminet
• 0.15 ml of said composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block said amount of n-alkyl dimethyl benzyl ammonium chloride and C 9 -C 11 alkyl alcohol ethoxylate, and: (a) around 1.937 x 10 -8 to 1.937 x 10 -4 , optionally around 4.029x10 -7 to 1.553x10 -4 , optionally around 7.864x10 -7 to 1.17x10 -4 , optionally around 1.17x10 -6 to 7.864x10 -5 , optionally around 1.553x10 -6 to 4.029x10 -5 moles of citrate; and/or (b) around 2.959 x 10 -8 to 2.959 x 10 -4 , optionally around 6.155x10 -7 to 2.373x10 -4 , optionally around 1.201x10 -6 to 1.787x10 -4 , optionally around 1.787x10
• a substrate comprising a blocking composition (optionally around 0.15 ml thereof), wherein said blocking composition comprises: one or more metals; one or more blocking composition surfactants; and one or more pH buffers, optionally wherein: around 0.15 ml of the blocking composition comprises a combined amount of said metal and said blocking composition surfactant that is sufficient to block: (a) at least around 1.937 x 10 -8 moles of citrate, optionally at least around 4.029x10- 7 , optionally at least around 7.864x10 -7 , optionally at least around 1.17x10 -6 , optionally at least around 1.553x10 -6 , optionally around 1.9x10 -6 such as around 1.937x10 -6 ; and/or (b) at least around 2.959 x 10 -8 moles of ethylenediaminetetraacetic acid (EDTA), optionally at least around 6.155x10 -7 , optionally at least around 1.201x10 -6 , optionally at least around EDTA
• the one or more metals comprise one or more metal salts selected from the group consisting of metal halides (optionally metal chlorides, metal fluorides, metal iodides and metal bromides), metal acetates, metal sulphates, metal carbonates, metal nitrates, metal phosphates, metal gluconates, metal oxides, metal hydroxides, metal citrates, metal lactates, metal glubionates, metal hydrates, metal peroxides, metal hypochlorides, metal dioxides and metal fumarates.
• metal halides optionally metal chlorides, metal fluorides, metal iodides and metal bromides
• metal acetates metal sulphates
• metal carbonates metal nitrates
• metal phosphates metal gluconates
• metal oxides metal hydroxides
• metal citrates metal lactates
• metal glubionates metal hydrates
• metal peroxides metal hypochlorides
• the one or more metals comprise one or more metals, selected from the group consisting of zinc, calcium, magnesium, copper, bismuth, indium, manganese, nickel, titanium, chromium, aluminum, lithium, sodium, potassium, beryllium, radium, scandium, yttrium, lanthanum, vanadium, iron, cobalt, iridium, hafnium, silver, thallium, palladium, cadmium, tin, gallium, cerium, praseodymium, neodymium, promethium, samarium, europium, gadolinium, terbium, dysprosium, holmium, erbium, thulium, ytterbium, lutetium, americium, curium, berkelium, californium, thorium, uranium and neptunium, optionally from the group consisting of magnesium, calcium, zinc,
• the one or more metals comprise zinc and/or calcium, optionally Zn 2+ and/or Ca 2+ , optionally ZnCl2 and/or CaCl2.
• said blocking composition comprises around 0.0001 to 4 M of said one or more metals, optionally around 0.001 to 2 M; optionally 0.01 to 0.1 M, optionally around 0.05 to 0.25 M, optionally around 0.1 to 0.15 M. 14.
• the one or more metals comprise ZnCl 2 and CaCl 2 , optionally wherein said blocking composition comprises: around 0.00005 to 2 M of ZnCl 2 , optionally around 0.001 to 1 M; optionally 0.005 to 0.05 M, optionally around 0.025 to 0.125 M, optionally around 0.05 to 0.1 M, optionally around 0.06 to 0.09 M, and around 0.00005 to 2 M of CaCl 2 , optionally around 0.001 to 1 M; optionally 0.005 to 0.05 M, optionally around 0.025 to 0.125 M, optionally around 0.03 to 0.07 M, optionally around 0.04 to 0.06 M. 15.
• the one or more metals has a binding affinity, log K f , for EDTA of at least around 4, optionally at least around 8, 10, 15, 20, 25, 30, 35, 40 or 45. 16.
• said metal has a binding affinity, log K f, for EDTA of at most around 30, optionally at most around 25, 20, 15, 10 or 8.
• said metal has a binding affinity, log K f , for EDTA of around 4 to 45, optionally around 8 to 30, optionally around 10 to 30, optionally around 15 to 25, optionally around 20 to 25. 18.
• the substrate according to any preceding clause wherein said metal has a binding affinity for Chromeazurol S of at most 15, optionally at most 13, optionally at most 10. 19.
• the one or more pH buffers are selected from the group consisting of: glycine, acetate, citrate, phosphate, ethanesulfonic acid and bis-tris methane. 20.
• the one or more pH buffers are selected from the group consisting of: glycine-hydrochloric acid, sodium acetate, piperazine- N,N′-bis(2-ethanesulfonic acid), citrate, phosphate, phosphate-citrate, 2-(N- morpholino)ethanesulfonic acid, 3-(N-morpholino)propanesulfonic acid and bis-tris methane. 21. The substrate according to any preceding clause, wherein the one or more pH buffers provide a pH working range of about 2.6 to 8, optionally 3.5 – 7, optionally around 5.6 – 7. 22.
• the one or more pH buffers provide a buffer capacity ⁇ of about -0.7 to 0.7 mol, optionally about -0.4 to 0.7 mol, optionally about -0.1 to 0.7 mol, optionally about 0.075 to 0.7 mol, optionally about 0.04 to 0.7 mol; and/or optionally about -0.7 to 0.4 mol, optionally about -0.7 to 0.1 mol, optionally about -0.7 to 0.04 mol, optionally about -0.7 to 0.01 mol; and/or optionally about -0.06 to 0.04, optionally about -0.05 to 0.02, optionally about -0.04 to 0.015 mol. 23.
• said blocking composition comprises around 10 mmol/L to 1 mol/L of said one or more pH buffers, optionally around 10 mmol/L to 750 mmol/L, optionally around 50 mmol/L to 500 mmol/L, optionally around 100 mmol/L to 300 mmol/L, optionally around 100 to 200 mmol/L, optionally around 130 to 170 mmol/L.
• the one or more pH buffers comprise acetate, such as sodium acetate. 25.
• said blocking composition surfactant comprises a polysorbate (optionally a polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or optionally a Tween TM ); optionally at a level of around 1 mM to 200 mM; an aliphatic phenol ethoxylate (such as Triton X-100 TM ; optionally wherein the aliphatic group is an alkyl group; optionally wherein the aliphatic phenol ethoxylate is octyl phenol ethoxylate); optionally at a level of around 0.1 mM to 200 mM; a cyclodextrin (optionally ⁇ -cyclodextrin or ⁇ -cyclodextrin); of a range between 1mM to 200mM; an aliphatic sulfate (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has a
• said blocking composition surfactant comprises a polysorbate (optionally a polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80 or optionally a Tween TM ); optionally at a level of around 1 mM to 200 mM; an aliphatic phenol ethoxylate (such as Triton X-100 TM ; optionally wherein the aliphatic group is an alkyl group; optionally wherein the aliphatic phenol ethoxylate is octyl phenol ethoxylate); optionally at a level of around 0.1 mM to 200 mM; a cyclodextrin (optionally ⁇ -cyclodextrin or ⁇ -cyclodextrin); of a range between 1mM to 200mM; an aliphatic sulfate (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has
• said blocking composition surfactant comprises an aliphatic sulfate (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has a chain length of 8 to 16 carbon atoms (optionally straight chain); optionally wherein the aliphatic sulfate is sodium dodecyl sulfate).
• said blocking composition surfactant is as defined in any one of clauses 25 to 27 (optionally wherein said blocking composition surfactant is sodium dodecyl sulfate) present from an amount of about 1 x 10 -5 mol/L to about 3.5 x 10 -3 mol/L, optionally about 1 x 10 -4 mol/L to about 3.5 x 10 -3 mol/L, optionally about 2.059x10 -4 to 2.926x10 -3 , optionally about 3.119x10 -4 to 2.352x10 -3 , optionally about 4.178x10 -4 to 1.778x10 -3 , optionally about 5.238x10 -4 to 1.204x10 -3 . 29.
• said blocking composition further comprises 5-sulfosalicyclic acid.
• said 5-sulfosalicyclic acid is present from an amount of about 1 x 10 -6 mol / L to about 5x10 -3 mol / L, optionally 1 x10 -5 mol / L to 2.5x10 -3 mol / L, optionally, 1x10 -4 mol / L to 1x10 -3 mol / L.
• said substrate is an absorbent material, such as a sponge or wipe.
• a composition comprising the blocking composition as defined in any preceding clause. 33.
• composition according to clause 32 comprising: around 0.001 to 3 mol / L of one or more metals selected from the group consisting of: Zn 2+ , Ca 2+ , Mg 2+ , Cu 2+ , Cu 3+ , Bi 3+ , Mn 2+ , Mn 3+ , Ag + , Fe 2+ , Cr 2+ , Cr 3+ , Ce 4+ , Co 2+ , Bi 3+ , Ti 3+ , Al 3+ , Li + , K + , Na + and Be 2+ ; optionally wherein said one or more metals are selected from the group consisting of: Zn 2+ , Ca 2+ , Mg 2+ , Cu 2+ , Cu 3+ , Bi 3+ , Mn 2+ , Mn 3+ , Cr 2+ , Cr 3+ , Li + , K + , Na + , Be 2+ and Fe 2+ ; optionally wherein said one or more metals are selected from the group consisting of: Zn 2+
• composition according to clause 32 or 33 wherein the one or more metals are present as salts selected from the group consisting of: metal halides, metal acetates, metal carbonates and metal sulfates.
• 35 The composition according to any one of clauses 32 to 34, comprising around 10 to 750 mmol / L of buffer selected from the group consisting of: acetate, glycine, ethanesulfonic acid, bis-tris methane and phosphate. 36.
• composition according to any one of clauses 32 to 35 comprising: (a) around 0.01 to 10 mmol / L of an aliphatic sulfate; or (b) around 1 to 100 mmol / L of an aliphatic phenol ethoxylate; or (c) around 10 to 200 mmol / L of a polysorbate. 37.
• the composition according to any one of clauses 32 to 36 comprising around 0.001 to 2 mol / L of the one or more metals, optionally around 0.001 to 1.5 mol / L, optionally around 0.001 to 1 mol / L. 38.
• composition according to clause 32 comprising: around 0.001 to 3 mol / L of one or more metals selected from the group consisting of: Zn 2+ , Ca 2+ , Mg 2+ , Fe 2+ , Cr 3+ , Mn 2+ and Bi 3+ ; around 10 to 750 mmol / L buffer (optionally according to clause 35, optionally acetate buffer); around 0.01 to 10 mmol / L surfactant (optionally an aliphatic sulfate, optionally sodium dodecyl sulfate). 39.
• composition according to clause 38 or 39 (optionally according to clause 39), wherein the surfactant is an aliphatic sulfate (optionally sodium dodecyl sulfate).
• the surfactant is an aliphatic sulfate (optionally sodium dodecyl sulfate).
• 41. The composition according to any one of clauses 32 to 40, wherein the one or more metals are Zn 2+ and Ca 2+ .
• 42. The composition according to any one of clauses 38 to 41 (optionally according to any one of clauses 39 to 41, optionally according to clause 40 or 41, optionally according to clause 41), comprising: around 0.001 to 1.5 mol / L of Zn 2+ ; around 0.001 to 1.5 mol / L of Ca 2+ . 43.
• composition according to any one of clauses 32 to 42 (optionally according to any one of clauses 39 to 42), comprising around 10 to 750 mmol / L of acetate buffer.
• the composition according to any one of clauses 32 to 43 (optionally according to any one of clauses 39 to 43), comprising around 0.01 to 10 mmol / L of an aliphatic sulfate.
• the composition according to clause 41 comprising: around 0.001 to 1 mol / L of Zn 2+ ; around 0.001 to 1 mol / L of Ca 2+ ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate. 46.
• composition according to clause 45 comprising: around 15 to 250 mmol / L of Zn 2+ ; around 10 to 200 mmol / L of Ca 2+ ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• said one or more metal comprises CaCl 2 and ZnCl 2 ; the composition comprises a sodium dodecyl sulfate blocking composition surfactant.
• composition according to clause 48 comprising: around 0.001 to 1.5 mol / L of ZnCl 2 ; around 0.001 to 1.5 mol / L of CaCl 2 ; around 10 to 750 mmol / L of acetate buffer; around 0.01 to 10 mmol / L sodium dodecyl sulfate. 50.
• the composition according to clause 49 comprising: around 0.001 to 1 mol / L of ZnCl 2 ; around 0.001 to 1 mol / L of CaCl 2 .
• the composition according to clause 49 or 50 comprising around 12.5 to 400 mmol / L of acetate buffer. 52.
• composition according to any one of clauses 49 to 51 comprising around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• the composition according to clause 49 comprising: around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• 54. The composition according to clause 49, comprising: around 0.001 to 1 mol / L of ZnCl 2 ; around 0.001 to 1 mol / L of CaCl 2 ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate. 55.
• composition according to clause 49 comprising: around 15 to 250 mmol / L of ZnCl 2 ; around 10 to 200 mmol / L of CaCl 2 ; around 12.5 to 400 mmol / L of acetate buffer; around 0.05 to 3.5 mmol / L sodium dodecyl sulfate.
• a kit comprising: a substrate according to any one of clauses 1-31 and/or a composition according to any one of clauses 32-55; and (a) a colour changing composition, wherein said colour changing composition is configured to detect the presence of microorganisms; and/or (b) a cleaning composition, wherein said cleaning composition is configured to reduce and/or eliminate microorganisms. 57.
• said substrate comprises around 0.05 ml to 0.75 ml of said blocking composition, optionally around 0.1 ml to 0.6 ml, optionally around 0.125 ml to 0.5 ml, optionally around 0.15 ml.
• said colour changing composition is present in an amount around 0.1 ml to 1 ml, optionally around 0.2 ml to 0.7 ml, optionally around 0.3 ml to 0.5 ml. 59.
• said colour changing composition comprises: a colour changing composition metal, and a colour changing agent, wherein said colour changing composition metal is bindable to said colour changing agent to provide a change in colour on binding and/or release thereof.
• a colour changing composition surfactant optionally wherein said colour changing composition metal and colour changing composition surfactant are present at a molar ratio of about 1:0.25 to 30, optionally about 1:0.5 to 7; optionally about 1:0.75 to 6; optionally about 1:0.75 to 5; optionally about 1:0.75 to 3; optionally about 1:1 to 3; optionally about 1:1 to 2.5.
• said colour changing composition surfactant is selected from the group consisting of: a polysorbate (optionally a polysorbate 80 or polysorbate 20, optionally a Tween TM ), an aliphatic phenol ethoxylate (such as Triton X-100; optionally wherein the aliphatic group is an alkyl group; optionally wherein the aliphatic phenol ethoxylate is octyl phenol ethoxylate), an aliphatic sulfobetaine (optionally wherein the aliphatic group is an alkyl group, optionally wherein the aliphatic group has a chain length of 8 to 16 carbon atoms (optionally straight chain); optionally wherein the aliphatic sulfobetaine is lauryl sulfobetaine), an aliphatic quaternary ammonium halide such as aliphatic trimethylammonium halide (optionally wherein the aliphatic group is an
• said colour changing agent is a chromeazurol (such as chromeazurol S or chromeazurol B) or a tannin, optionally wherein said colour changing agent is a chromeazurol, optionally chromeazurol S (CAS).
• said cleaning composition comprises: an n-alkyl dialkyl triamine (e.g.
• n-alkyl is at least C 10 , optionally C 12 ; and/or optionally wherein the trialkyl is dipropylene, optionally wherein the n-alkyl dialkyl triamine is dodecyl dipropylene triamine), n-alkyl dimethyl ammonium halide (e.g.
• n-alkyl dialkyl aryl ammonium halide is n-alkyl dimethyl benzyl ammonium halide, optionally n- alkyl dimethyl benzyl ammonium chloride, optionally wherein the n-alkyl is at least C 8 (e.g.
• said cleaning composition comprises: n-alkyl dialkyl aryl ammonium halide present from about about 0.6 wt% to 14 wt%, optionally 0.7 wt% to 13 wt%, optionally 0.8 wt% to 12 wt%, optionally 0.9 wt% to 11 wt%, optionally about 1 wt% to 10 wt%; and aliphatic alcohol alkanoate present from about 0.6 wt% to 14 wt%, optionally 0.7 wt% to 13 wt%, optionally 0.8 wt% to 12 wt%, optionally 0.9 wt% to 11 wt%, optionally about 1 wt% to 10 wt% 69.
• said cleaning composition further comprises citrate present from about 0.5 wt% to 10 wt%; optionally about 0.6 wt% to 9 wt%, optionally 0.7 wt% to 8 wt%, optionally 0.8 wt% to 7 wt%, optionally 0.9 wt% to 6 wt%, optionally about 1 wt% to 5 wt%. 70.
• said cleaning composition further comprises EDTA present from about 0.5 wt% to 30 wt%; optionally about 0.6 wt% to 25 wt%, optionally 0.7 wt% to 20 wt%, optionally 0.8 wt% to 15 wt%, optionally 0.9 wt% to 12.5 wt%, optionally about 1 wt% to 10 wt%. 71.
• said cleaning composition further comprises sodium hypochlorite present from about 1.8 wt% to 7 wt%, optionally 2.1 wt% to 6.5 wt%, optionally 2.4 wt% to 6 wt%, optionally 2.7 wt% to 5.5 wt%, optionally about 3 wt% to 5 wt%; and hydrogen peroxide present from about 6 wt% to 42 wt%, optionally 7 wt% to 39 wt%, optionally 8 wt% to 36 wt%, optionally 9 wt% to 33 wt%, optionally about 10 wt% to 30 wt%.
• said cleaning composition further comprises peracetic acid present from about 6 wt% to 42 wt%, optionally 7 wt% to 39 wt%, optionally 8 wt% to 36 wt%, optionally 9 wt% to 33 wt%, optionally about 10 wt% to 30 wt%; and acetic acid present from about 6 wt% to 42 wt%, optionally 7 wt% to 39 wt%, optionally 8 wt% to 36 wt%, optionally 9 wt% to 33 wt%, optionally about 10 wt% to 30 wt%. 73.
• said blocking composition is as defined in any one of clauses 33 to 55; and wherein said colour changing composition comprises: a colour changing composition metal, wherein said colour changing composition is iron; a colour changing agent, wherein said colour changing agent is chromeazurol S; and optionally a colour changing composition surfactant, wherein said colour changing composition surfactant is an aliphatic quaternary ammonium halide; wherein said iron is bindable to chromeazurol S to provide a change in colour on binding and/or release thereof, optionally wherein said iron and said aliphatic quaternary ammonium halide are present at a molar ratio of 1:0.25 - 1:5; and wherein said iron and said chromeazurol S are present at a molar ratio of 1:0.5 - 1:5.
• a swab comprising a rod having a substrate according to clauses 1-31 at an end thereof (optionally wherein the substrate comprises 0.15 ml of said composition), optionally further comprising a reservoir of a colour changing composition, optionally wherein said colour changing composition is as defined in any one of clauses 56 to 74, optionally wherein said reservoir is rupturable to enable release of said colour changing composition.
• a method comprising contacting a surface, or a sample therefrom, that has been pre- treated with a cleaning composition, with a blocking composition as defined in any one of clauses 32 to 55, optionally wherein said cleaning composition is as defined in any one of clauses 56 to 74. 79. The method according to clause 78, further comprising contacting said surface, or sample therefrom (e.g.

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• 2023
  • 2023-02-07 WO PCT/EP2023/053022 patent/WO2023148409A1/en not_active Ceased
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