CN116583586A

CN116583586A - Use of lactams

Info

Publication number: CN116583586A
Application number: CN202180082476.2A
Authority: CN
Inventors: P·科萨基斯; S·V·梅德帕里; N·J·帕里
Original assignee: Unilever IP Holdings BV
Current assignee: Unilever IP Holdings BV
Priority date: 2020-12-09
Filing date: 2021-11-05
Publication date: 2023-08-11

Abstract

The present invention relates to the use of a lactam composition to improve toilet hygiene, the use comprising treating a toilet surface, wherein the composition comprises 0.0001 to 5 wt% of a lactam; the use thereof relates to reducing off-flavors; or wherein the use involves reducing undesirable urinary scale formation in a toilet; wherein the lactam has the formula (I) or (II) wherein R ₁ And R is ₂ Each independently selected from the group consisting of hydrogen, halogen, alkyl, cycloalkyl, alkoxy, oxoalkyl, alkenyl, heterocyclyl, heteroaryl, aryl, and aralkyl; and R is ₃ Selected from the group consisting of hydrogen, hydroxy, alkyl, cycloalkyl, alkoxy, oxyalkyl, alkenyl, heterocyclyl, and heteroarylRadicals, cycloalkyl, aryl, aralkyl, -C (O) CR ₆ ＝CH ₂ And (CH) ₂ ) _n N ⁺ (R ^a ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8, and wherein each R ^a Independently H or C _1‑4 An alkyl group; r is R ₄ And R is ₅ Independently selected from the group consisting of hydrogen, aryl, heterocyclyl, heteroaryl, and arylalkyl; and R is ₆ Selected from hydrogen and methyl; and R is ₇ Selected from hydrogen and-C (O) CR ₆ ＝CH ₂ The method comprises the steps of carrying out a first treatment on the surface of the And preferably R ₄ And R is ₅ At least one of which is hydrogen. (I)Or (II)

Description

Use of lactams

Technical Field

The present invention relates to improvements in the field of toilet hygiene, and in particular to the use of lactams to improve toilet hygiene.

Background

Toilet hygiene is important to consumers. Consumers still want to maintain a sanitary toilet environment when human waste is disposed of in the toilet.

One way consumers recognize poor toilet hygiene is for the toilet to emit undesirable odors, which are classified as undesirable odors.

There is a need to improve toilet hygiene, for example by reducing the malodour of the toilet, or by reducing unwanted crystal formation in the toilet.

Disclosure of Invention

We have found that toilet hygiene is improved by treating the toilet surface with a lactam composition.

In a first aspect, the present invention relates to the use of a lactam composition to improve toilet hygiene, said use comprising treating a toilet surface, wherein said composition comprises 0.0001-5 wt% of a lactam;

wherein the use relates to reducing off-flavors; or wherein said use involves reducing undesirable urinary scale formation in a toilet;

wherein the lactam has formula (I) or (II):

wherein:

R ₁ and R is ₂ Each independently selected from the group consisting of hydrogen, halogen, alkyl, cycloalkyl, alkoxy, oxoalkyl, alkenyl, heterocyclyl, heteroaryl, aryl, and aralkyl; and

R ₃ selected from the group consisting of hydrogen, hydroxy, alkyl, cycloalkyl, alkoxy, oxoalkyl, alkenyl, heterocyclyl, heteroaryl, cycloalkyl, aryl, aralkyl, -C (O) CR ₆ ＝CH ₂ And (CH) ₂ ) _n N ⁺ (R ^a ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8, and wherein each R ^a Independently H or C _1-4 An alkyl group;

R ₄ and R is ₅ Independently selected from the group consisting of hydrogen, aryl, heterocyclyl, heteroaryl, and arylalkyl; and

R ₆ selected from hydrogen and methyl; and

R ₇ selected from hydrogen and-C (O) CR ₆ ＝CH ₂ The method comprises the steps of carrying out a first treatment on the surface of the And is also provided with

Preferably, R ₄ And R is ₅ At least one of which is hydrogen.

Preferably, the lactam is present at a level of 0.0001 to 2.5 wt%, preferably 0.0001 to 1 wt%, more preferably 0.001 to 1 wt%.

The use includes reducing off-flavors. In particular, it is preferred that the reduction of off-flavors relates to the reduction of off-flavors associated with urine, in particular to the reduction of off-flavors associated with phenol and indole; or the use relates to reducing undesirable urine scale formation in toilets. In particular, it is preferred that the reduction of undesired urinary scale formation in toilets involves inhibition of Proteus mirabilis (P.Mirabilis).

The lactam has the formula (I) or (II):

wherein:

R ₆ selected from hydrogen and methyl; and

R ₇ selected from hydrogen and-C (O) CR ₆ ＝CH ₂ The method comprises the steps of carrying out a first treatment on the surface of the And

preferably R ₄ And R is ₅ At least one of which is hydrogen.

Preferably a lactam of formula (I) or (II), R ₁ 、R ₄ And R is ₅ Is H; r is R ₃ Is H or (CH) ₂ ) _n N ⁺ (CH ₃ ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8; and R is ₂ Phenyl or monosubstituted phenyl; preferably R ₂ Selected from phenyl, 4-fluorophenyl, 2-fluorophenyl, 4-chlorophenyl, 3-chlorophenyl, 4-bromophenyl and 4-methylphenyl.

Preferably the lactam is a lactam selected from the group consisting of:

preferably, the lactam is selected from:

most preferably, the lactam is

4- (4-chlorophenyl) -5-methylene-pyrrol-2-one.

Preferably, the lactam is delivered from a water-based composition or an organic solvent-based composition.

Preferably, the toilet surface to be treated is selected from ceramics, such as porcelain.

Drawings

Figure 1 relates to example 2 and shows the effect of lactam on odor control (phenol).

Figure 2 relates to example 2 and shows the effect of lactam on odor control (indole).

FIG. 3 relates to example 3 and depicts an image showing the effect of lactam on reducing Proteus mirabilis.

FIG. 4 relates to example 3 and depicts a graph showing the effect of lactam on reducing Proteus mirabilis.

Detailed Description

The indefinite articles "a" or "an" as used herein and the corresponding definite article "the" mean at least one or more unless specified otherwise.

It is to be understood that all preferences are combinable unless explicitly stated otherwise.

Use for improving toilet hygiene

The use relates to reducing malodour and/or the use relates to reducing undesirable formation of urinary scale in a toilet.

In particular, it is preferred that the reduction of off-flavors relates to the reduction of off-flavors associated with urine, in particular to the reduction of off-flavors associated with phenol and indole.

Where the use relates to reducing undesirable urinary scale formation in a toilet, it is preferred that the undesirable reduction in urinary scale formation in a toilet involves inhibition of Proteus mirabilis.

The formation of urinary scale is at least partly due to the formation of crystals. Proteus mirabilis (Proteus mirabilis) produces urease which hydrolyzes urea to release carbon dioxide and ammonia, thereby increasing the pH of the urine and resulting in magnesium ammonium phosphate and apatite crystal deposits. Inhibition of Proteus mirabilis is accompanied by inhibition of crystal formation.

Lactam

The lactam is a cyclic amide. The preferred lactam is a gamma-lactam having 5 ring atoms.

The lactam has the formula (I) or (II):

wherein:

R ₆ selected from hydrogen and methyl; and

preferably, R ₄ And R is ₅ At least one of which is hydrogen.

It will be appreciated that the groups may be optionally substituted where appropriate. Optional substituents mayComprising halogen, C _1-4 Alkyl, C _1-4 Haloalkyl (e.g., CF) ₃ ) And C _1-4 An alkoxy group.

The alkyl group may be, for example, C _1-12 Alkyl radicals, e.g. C _1-6 An alkyl group. Aryl groups can be, for example, C _6-10 Aryl groups such as phenyl.

Preferably, R ₁ And R is ₂ Is selected from the group consisting of heterocyclyl, heteroaryl, aryl, and arylalkyl.

Preferably, R ₁ Is hydrogen. Preferably, R ₃ Is hydrogen or (CH) ₂ ) _n N ⁺ (R ^a ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8, and wherein each R ^a Independently H or C _1-4 Alkyl, more preferably R ^a Is CH ₃ The method comprises the steps of carrying out a first treatment on the surface of the Preferably, R ₄ Is hydrogen. Preferably, R ₅ Is hydrogen. Preferably, R ₆ Is hydrogen. Preferably, R ₇ Is hydrogen. Preferably, R ₂ Is aryl or aralkyl. More preferably, R ₂ Is phenyl or substituted phenyl, for example mono-substituted phenyl. The substitution may be ortho, meta or para. Preferred substituents include halogen and methyl. For example, but not limited to, R ₂ Can be selected from phenyl, 4-fluorophenyl, 2-fluorophenyl, 4-chlorophenyl, 3-chlorophenyl, 4-bromophenyl and 4-methylphenyl.

More preferably in the lactam of formula (I) or (II), R ₁ 、R ₄ And R is ₅ Is H; r is R ₃ Is H or (CH) ₂ ) _n N ⁺ (CH ₃ ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8; and R is ₂ Phenyl or monosubstituted phenyl; preferably R ₂ Selected from phenyl, 4-fluorophenyl, 2-fluorophenyl, 4-chlorophenyl, 3-chlorophenyl, 4-bromophenyl and 4-methylphenyl.

Even more preferably the lactam has formula (I), R ₁ 、R ₄ And R is ₅ Is H; r is R ₃ Is H or (CH) ₂ ) _n N ⁺ (CH ₃ ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8; and R is ₂ Phenyl or monosubstituted phenyl; preferably R ₂ Selected from phenyl, 4-fluorophenyl, 2-fluorophenyl, 4-chlorophenyl, 3-chlorophenyl, 4-bromophenyl and 4-methylphenyl.

Where the lactam is cationic in nature, it may be used as such or suitably with a counter ion (e.g. iodide).

Preferably the lactam is a lactam selected from the group consisting of:

more preferably the lactam is selected from:

most preferably the lactams are:

4- (4-chlorophenyl) -5-methylene-pyrrol-2-one.

Level of lactam

Preferably the lactam is present at a level of 0.0001 to 2.5 wt%, preferably 0.0001 to 1 wt%. For example, the lactam may suitably be present at a level of 0.001 to 1 wt% or even 0.01 to 0.5 wt%.

Composition and method for producing the same

Preferably, the lactam is delivered from an aqueous or organic solvent based composition, most preferably an aqueous based composition.

The composition comprises 0.0002 to 0.1 wt%, preferably 0.001 to 0.1 wt% of lactam.

Toilet surface

Preferably, the toilet surface to be treated is selected from ceramics (e.g. porcelain).

Further ingredients

The composition may comprise further ingredients such as surfactants, chelating agents, thickeners, pH modifiers.

The invention will be further described by the following non-limiting examples.

Examples

Example 1 preparation of preferred lactam examples

Preparation of 4- (4-chlorophenyl) -5-hydroxy-5-methylfuran-2 (5H) -one

1- (4-chlorophenyl) propan-2-one (40.00 g,34.75mL,237.2 mmol), glyoxylate monohydrate (32.75 g,355.8 mmol) and phosphoric acid (69.74 g,711.7 mmol) were combined at room temperature and then heated to 85℃overnight. After cooling to room temperature, the mixture was poured into a mixture of water (500 mL) and ethyl acetate (500 mL). The layers were separated and the aqueous phase was extracted with ethyl acetate (500 mL). The combined organic layers were washed with a 1:1 mixture of water and brine (2×500 mL), dried (MgSO ₄ ) And concentrated under reduced pressure to give 4- (4-chlorophenyl) -5-hydroxy-5-methylfuran-2 (5H) -one (66.00 g,>100% yield) as a brown oil. The material was used in the next step without further purification.

Preparation of 4- (4-chlorophenyl) -5-hydroxy-5-methyl-1H-pyrrol-2 (5H) -one

4- (4-chlorophenyl) -5-hydroxy-5-methylfuran-2 (5H) -one (66.00 g,293.8 mmol) was dissolved in thionyl chloride (196.8 g,120.0mL,1654 mmol) and heated at 40℃for 1 hour and then at 80℃for 2 hours. Concentrating the mixture under reduced pressure, and mixing with 2-Methyl tetrahydrofuran (200 mL) azeotropes. The residue was diluted with 2-methyltetrahydrofuran (160 mL) and the solution was added to a cooled stirred mixture of 28% aqueous ammonia (180 mL) in 2-methyltetrahydrofuran (20 mL) at 0 ℃. The mixture was warmed to room temperature and stirred overnight. Water (100 mL) and ethyl acetate (200 mL) were added and the layers separated. The aqueous phase was extracted with ethyl acetate (200 mL) and the combined organic extracts were dried (MgSO ₄ ) And concentrated under reduced pressure. Purification by dry flash column chromatography (5-60% ethyl acetate in heptane) afforded 4- (4-chlorophenyl) -5-hydroxy-5-methyl-1H-pyrrol-2 (5H) -one (23.18 g,35% yield) as a cream solid.

¹ H NMR(400MHz,d ₆ -DMSO)8.55(brs,1H),7.88-7.83(m,2H),7.51-7.46(m,2H),6.37(d,1H),6.32(s,1H),1.45(s,3H)

UPLC (alkaline) 1.51/5.00min,100% purity, M+H ⁺ 224

MP 177℃

Preparation of 4- (4-chlorophenyl) -5-methylene-1H-pyrrol-2 (5H) -one

To a cooled solution of 4- (4-chlorophenyl) -5-hydroxy-5-methyl-1H-pyrrol-2 (5H) -one (10.00 g,44.51 mmol) in anhydrous dichloromethane (100 mL) was added a solution of boron trifluoride etherate (8.213 g,7.142mL,57.87 mmol) in anhydrous dichloromethane (45 mL) at 0deg.C over 15 min. The mixture was stirred at 0 ℃ and then slowly warmed to room temperature and stirred for 2 hours. The reaction was quenched with ice water (100 mL) and the layers separated. The aqueous layer was extracted with dichloromethane (100 mL) and the combined organic layers were washed with a 1:1 mixture of water and saturated aqueous sodium bicarbonate (100 mL), dried (MgSO ₄ ) And filtered. Silica was added to the filtrate and the mixture was stirred for 10 minutes, then filtered through a plug of silica, washed thoroughly with dichloromethane, then dichloromethane: the 3:1 mixture of diethyl ether was washed. Fractions containing the desired product were combined and concentrated under reduced pressure. After concentration a precipitate forms which passesCollected by filtration and washed with diethyl ether to give 4- (4-chlorophenyl) -5-methylene-1H-pyrrol-2 (5H) -one (5.25 g,57% yield) as a cream solid.

¹ H NMR(400MHz,d ₆ -DMSO)10.10(s,1H),7.54-7.47(m,4H),6.36(s,1H),5.04(t,1H),4.85(s,1H)

UPLC (alkaline) 1.87/5.00min,100% purity, M+H ⁺ 206

MP 182℃

Preparation of 5-hydroxy-5-methyl-4- (p-tolyl) furan-2 (5H) -one

1- (p-tolyl) propan-2-one (25.00 g,24.00mL,168.7 mmol), glyoxylate monohydrate (23.29 g,253.0 mmol) and phosphoric acid (49.60 g,506.1 mmol) were combined at room temperature and then heated overnight at 90 ℃. After cooling to room temperature, the mixture was poured into a stirred mixture of ice water (400 mL) and ethyl acetate (400 mL). The layers were separated and the organic phase was washed with water (100 mL), dried (MgSO ₄ ) And concentrated under reduced pressure. Azeotropes of the mixture with 2-methyltetrahydrofuran (50 mL) gave 5-hydroxy-5-methyl-4- (p-tolyl) furan-2 (5H) -one (16.50 g,48% yield) as a brown solid.

¹ H NMR(400MHz,d ₆ -DMSO)7.86(s,1H),7.75(d,2H),7.28(d,2H),6.59(s,1H),2.32(s,3H),1.61(s,3H)

Preparation of 5-hydroxy-5-methyl-4- (p-tolyl) -1H-pyrrol-2 (5H) -one

5-hydroxy-5-methyl-4- (p-tolyl) furan-2 (5H) -one (16.50 g,80.80 mmol) was dissolved in thionyl chloride (48.06 g,29.47mL,404.0 mmol) and heated at 50℃for 1 hour, then at reflux for 1 hour. After cooling to room temperature, the mixture was concentrated under reduced pressure and azeotroped with 2-methyltetrahydrofuran (2×50 mL). The residue was diluted with 2-methyltetrahydrofuran (60 mL) and the solution was added to a cooled stirred mixture of 28% aqueous ammonia (55 mL,808.0 mol) in 2-methyltetrahydrofuran (10 mL) at 0 ℃. The mixture was warmed to room temperature and stirred overnight. 1, 2-methyltetrahydrofuran was removed under reduced pressure, the residue was diluted with (200 mL) and diethyl ether (100 mL), and the mixture was stirred at room temperature for 20 min. The solid was collected by filtration and stirred at room temperature in water (100 mL) and diethyl ether (50 mL) for 10 min. The solid was collected by filtration, washed with water, diethyl ether, and dried under vacuum at 50 ℃ to give 5-hydroxy-5-methyl-4- (p-tolyl) -1H-pyrrol-2 (5H) -one (10.49 g,31% yield) as a pale beige solid.

¹ H NMR(400MHz,d ₆ -DMSO)8.44(brs,1H),7.73(d,2H),7.21(d,2H),6.24(s,2H),2.29(s,3H),1.45(s,3H)

¹³ C NMR(400MHz,d ₆ -DMSO)170.4(s,1C),161.1(s,1C),139.8(s,1C),129.7(s,2C),128.9(s,1C),128.2(s,2C),119.1(s,1C),87.8(s,1C),26.7(s,1C),21.5(s,1C)

UPLC (alkaline) 1.41/5.00min,100% purity, M+H ⁺ 204

MP 178 ℃ decomposition

Preparation of 5-methylene-4- (p-tolyl) -1H-pyrrol-2 (5H) -one

To a cooled solution of 5-hydroxy-5-methyl-4- (p-tolyl) -1H-pyrrol-2 (5H) -one (8.68 g,42.7 mmol) in anhydrous dichloromethane (87 mL) was added a solution of boron trifluoride etherate (6.85 g,5.96mL,55.5 mol) in anhydrous dichloromethane (40 mL) at 0deg.C over 15 min. After 1 hour, the mixture was allowed to slowly warm to room temperature. After a further 3 hours, the reaction was diluted with dichloromethane (50 mL) and ice water (100 mL) and stirred for 10 minutes. The layers were separated and the organic layer was washed with water (100 mL), water and 1% saturated aqueous sodium bicarbonate: 1 mixture (100 mL) and brine (100 mL), and the organic layer was filtered through celite, eluting with twoWashing with chloromethane. Any excess water was removed by pipette and the filtrate was dried (MgSO ₄ ) And concentrated under reduced pressure to a brown solid. The solid was stirred in hot dichloromethane (120 mL) for 15 min, then slowly cooled to room temperature, then cooled to 0 ℃. The solid was collected by filtration to give 5-methylene-4- (p-tolyl) -1H-pyrrol-2 (5H) -one (3.87 g,49% yield) as a yellow solid. Silica was added to the filtrate and the mixture was stirred for 10 minutes, then filtered through a plug of silica, washed thoroughly with dichloromethane, then dichloromethane: the 4:1 mixture of diethyl ether was washed. The filtrate was concentrated under reduced pressure to give 5-methylene-4- (p-tolyl) -1H-pyrrol-2 (5H) -one (0.58 g, 7%) as a yellow solid. Total yield of 5-methylene-4- (p-tolyl) -1H-pyrrol-2 (5H) -one (4.45 g,56% yield).

¹ H NMR(400MHz,d ₆ -DMSO)10.11(brs,1H),7.35(d,2H),7.25(d,2H),6.25(s,1H),5.01(s,1H),4.85(s,1H),2.31(s,3H)

UPLC (alkaline) 1.83/5.00min,100% purity, M+H ⁺ 186

MP 200 ℃ decomposition

EXAMPLE 2 odor control

Artificial urine media was prepared using the composition described below and filter sterilized using a 0.22 μm filter.

The lactam used is denoted "488" and is 4- (4-chlorophenyl) -5-methylene-pyrrol-2-one.

The artificial urine medium composition is shown in table 1.

TABLE 1

By mixing equal volumes of 10 ⁸ Cell/ml Morganii Morganella morganii (M. Morganii), streptococcus agalactiae (S. Agalactiae), citrobacter k (C. Koseri), enterococcus faecalis (E. Faecalis), proteus mirabilis (P. Mirabilis), proteus vulgaris (P. Vulgaris), E.coli (E. Coli) to prepare a consortium of bacterial cultures.

A1:40 dilution mixture of the bacterial culture mixture and sterile artificial urine medium was prepared in whole and 5ml of this mixture was transferred to a sterile GC vial. A 1% stock of lactam 488 was prepared in DMSO. Lactam 488 at various concentrations corresponding to 50, 200 and 400ppm was added in duplicate to GC vials prepared as described above. Positive controls were also established without lactam 488. The vials were incubated at 37℃for 48 hours. After 48 hours, the vials were subjected to GC analysis. Sample vials were placed in CombiPAL vial holders. The analysis was performed using an Auto SPME accessory.

GC-MS (SIM) condition

Instrument: perkin Elmer Clarus 500 gas chromatograph-mass spectrometer. The GC conditions were: GC column: CP WAX (30m x 0.25mm ID x 0.25 μm thickness); carrier gas: helium (1 mL/min); column box procedure: the initial temperature is kept at 35 ℃ for 5min; heating: maintaining at 10 deg.c/min to 230 deg.c for 5min; run time: 29.5min; injection port temperature: 230 ℃. The MS conditions were: inlet line temperature: 200 ℃; electron energy: 70eV; source temperature: 180 ℃; multiplier voltage: 400V

combiPAL Condition

Sample vials were equilibrated at 60 degrees for 5 minutes. The sampling time was 2 minutes. The desorption time was 2 minutes. Post fiber conditioning time (post fiber conditioning time) was 38 minutes. The total run time was 40 minutes.

Detection of off-flavor molecules using GC/MS-SIM mode

Operating the GC/MS in SIM mode allows detection of specific analytes with increased sensitivity. In this mode, the MS gathers data of the quality of interest, rather than seeking all the quality over a wide range. Because the instrument is configured to seek only the mass of interest, it may be specific to the particular analyte of interest. Thus, they are analyzed using SIM patterns. Typically, more than two ions are monitored per compound, and the ratio of these ions is unique to the analyte of interest. The ions monitored for each molecule are recorded in table 2. To increase sensitivity, the mass scan rate and dwell time (the time it takes to observe at each mass) are adjusted. Using this method we can detect phenol (retention time-18.5) and indole (retention time-22.4).

TABLE 2

The odor generated from the sample without lactam was considered to be 100%. The amount of odor generated was recorded from the peak area of the MO peak. In comparison to this peak, the odor produced in the presence of lactam was calculated. The peak areas of phenol and indole from the defect-only (bugs) samples were recorded and considered to be 100%. Peak areas of these two peaks in the sample with lactam were recorded and the% reduction relative to the sample without lactam was compared and plotted in the graphs shown in fig. 1 and 2, the data also being shown in tables 3 and 4.

TABLE 3 odor control of lactams (phenol)

Composition and method for producing the same	% phenol
		Positive control	100
Positive control+lactam 400ppm	28

TABLE 4 odor control of lactams (indoles)

Composition and method for producing the same	% indole
		Positive control	100
Positive control+lactam 400ppm	68

EXAMPLE 3 Proteus mirabilis inhibition and Urea Crystal control

A multi-strain biofilm experiment was performed with ds-red labeled Proteus mirabilis Hauser 1885, m-turquoise labeled Pseudomonas aeruginosa PAO1-uw and eGFP labeled Staphylococcus aureus SH 1000. The OD600 of each strain was adjusted to 0.05 inoculum with artificial urine medium in polystyrene 24 well plates (Greiner), kept static at 37 ℃ for 2 hours and incubated at 70rpm for a further 22 hours for 72 hours. Firstly, the Proteus mirabilis grows, then the Pseudomonas aeruginosa grows, and finally the Staphylococcus aureus grows for 72 hours. The medium was carefully removed and a new bacterial suspension was gently added before the addition of the new inoculum. Cells were cultured in artificial urine. Crystals are the product of the urease activity of Proteus mirabilis and when the pH increases, the natural salt precipitate comes from the medium.

Artificial urine medium

According toBrooks and Keevil (1997) for artificial urine media. 1g peptone L37,0.005g yeast extract, 0.1g lactic acid, 0.4g citric acid, 2.1g NaHCO ₃ 10g of urea, 0.07g of uric acid, 0.8g of creatinine and 0.37g of CaCl ₂ ·2H ₂ O,5.2g sodium chloride, 0.0012g FeSO ₄ ·7H ₂ O，3.2g Na ₂ O ₄ S·10H ₂ O，0.95g KH ₂ PO ₄ ，1.2g HK ₂ O ₄ P，1.3g NH ₄ Cl was made up to 1L with distilled water. The pH was adjusted to 6.5 using HCl (molar, e.g., 2.5M). Finally, the media was filter sterilized by a 0.22 μm membrane filter (Millipore).

Confocal laser scanning microscopy

Confocal Laser Scanning Microscopy (CLSM) was used to visualize and generate Z-stack representations of mixed species biofilms. Display was performed with a Zeiss LSM 700 confocal laser scanning microscope (Carl Zeiss, ltd, welwyn, UK) controlled by a ZEN 2009 software platform (Carl Zeiss, ltd, welwyn, UK) using either a 5x or 10x objective lens.

Acquisition settings depend on fluorescence: pinhole 1.0, laser intensity 5% -15%, and gain of bioluminescence staining (bio-fluorescent strains) between 450-700. However, once adjusted to optimize the visualization, the settings were never modified in the experiment to facilitate unbiased comparison. The Z-stack height is set for each acquisition in a new well by starting from the first point where no emission through the biofilm itself was detected (just before the start of the biofilm) until the point where no emission was reached again (which is set as the last slice). Once set, the spacing was kept constant throughout the experiment.

Biofilm quantification and analysis

Using TECAN ^TM The biofilm was analyzed and quantified by an infinite F200 PRO plate reader. To be able to analyze the Z-stack images generated with CLSM, the 3D pictures were processed in Zen 2.3SP1 (black) software (Carl Zeiss, jena, germany) using the maximum intensity projection function. The tool generates a two-dimensional (2D) image for each channel, the image being made up of pixels containing a maximum value over all images of the Z-stack at a single pixel location. Each new picture is output in tiff format (64 bits) without compression to Zen 2.3 (blue version).The whole image was analyzed and quantified using the Measure function in ImageJ 1.52i software (Rueden et al, 2017) implemented with Fiji plugin bundle (Schindelin et al, 2012). The data generated per single image is defined as the sum of the gray values of all pixels divided by the average gray value of the number of pixels and the associated standard deviation. The final data represent the average of samples grown under specific conditions as the average of the average gray values generated in the images collected per well. The resulting pattern is shown in figures 3 and 4.

Claims

1. Use of a lactam composition to improve toilet hygiene, the use comprising treating a toilet surface, wherein the composition comprises 0.0001 to 5 wt% of a lactam;

wherein the use relates to reducing off-flavors; or wherein the use involves reducing undesirable urinary scale formation in the toilet;

wherein the lactam has formula (I) or (II):

(I)or (II)>

Wherein:

R ₄ and R is ₅ Independently selected from hydrogen, aryl, heterocyclyl, heteroaryl, and arylAn alkyl group; and

R ₆ selected from hydrogen and methyl; and

Preferably, R ₄ And R is ₅ At least one of which is hydrogen.

2. Use according to claim 1, wherein the lactam is present at a level of 0.0001-2.5 wt%, preferably 0.0001-1 wt%, more preferably 0.001-1 wt%.

3. Use according to claim 1 or claim 2, wherein the reduction of off-flavors comprises a reduction of off-flavors associated with urine, in particular with respect to reduction of off-flavors associated with phenol and indole.

4. The use according to claim 1 or claim 2, wherein reducing undesirable urinary scale in the toilet involves inhibition of proteus mirabilis.

5. The use according to any one of the preceding claims, wherein in the lactam of formula (I) or (II), R ₁ 、R ₄ And R is ₅ Is H; r is R ₃ Is H or (CH) ₂ ) _n N ⁺ (CH ₃ ) ₃ Wherein n is an integer from 1 to 16, preferably from 2 to 8; and R is ₂ Is phenyl or monosubstituted phenyl; preferably R ₂ Selected from phenyl, 4-fluorophenyl, 2-fluorophenyl, 4-chlorophenyl, 3-chlorophenyl, 4-bromophenyl and 4-methylphenyl.

6. Use according to any one of the preceding claims, wherein the lactam is a lactam selected from the group consisting of:

7. use according to any one of the preceding claims, wherein the lactam is selected from:

and/or +.>

8. The use according to any one of the preceding claims, wherein the lactam is:

4- (4-chlorophenyl) -5-methylene-pyrrol-2-one.