JPH0671440B2 - Dry cholesterol analytical element - Google Patents

Description

TECHNICAL FIELD The present invention relates to a dry analytical element suitable for analyzing cholesterol in body fluids such as blood, particularly total cholesterol including protein-bound cholesterol.

[Prior Art] A method of utilizing an enzyme such as cholesterol esterase to hydrolyze a cholesterol ester is well known. However, it is well known that this method requires a means other than an enzyme to break the protein-cholesterol ester bond and dissociate the cholesterol ester.

It is known to use proteases with lipases in US Pat. No. 3,869,349. Also US Patent 3 925 16
In No. 4, it is known to use a surfactant such as polyethylene glycol alkyl ether with cholesterol esterase. Also, U.S. Pat.
Nos. 1 and 4 274 152 had 20 ethylene glycol units.
Less than polyethylene glycol alkyl phenyl ethers (alkylphenoxypolyethoxyethanols) have been described as useful.

However, when a dry analytical element having a porous liquid developing layer (hereinafter referred to as a porous developing layer or a developing layer) contains a polyethylene glycol alkylphenyl ether having an ethylene glycol unit of less than 20 together with an enzyme in the developing layer,
This surfactant has a too large liquid development area within a certain period of time, which reduces the sensitivity of colorimetric analysis, and does not have sufficient quantitative development of liquid (proportionality of supply liquid amount and development area).
There is a drawback that the wettability of the coating liquid with respect to the spreading layer is insufficient. Further, when this surfactant is contained in a layer composed of a hydrophilic binder such as a water-absorbing agent, a reagent layer, a light-shielding layer or an adhesive layer, which is present between the spreading layer and the support, it is applied to a certain layer and a next layer. In some cases, the coating liquid of the coating layer was not sufficiently wet, resulting in uneven coating.

[Problems to be Solved by the Invention] An object of the present invention is to have a suitable liquid development area within a certain period of time,
Another object of the present invention is to provide a dry cholesterol analysis element having high sensitivity of colorimetric analysis with respect to total cholesterol including protein-bound cholesterol.

Another object of the present invention is to provide a dry cholesterol analysis element which has a sufficient quantitative property of liquid development in the developing layer (proportionality of the amount of the supply liquid and the developing area) and enables uniform application of the coating liquid.

[Structure of the Invention] The above object of the present invention is a dry analytical element having at least one water-permeable layer and a porous liquid developing layer in a state in which they can be in liquid contact with each other, wherein the developing layer has a cholesterol ester hydrolyzing activity ( Cholesterol ester hydrolase activity)
By an alkylphenoxypolyglycidol in the spreading layer or the at least one water-permeable layer.

[Detailed Description of Structure of the Invention] The enzyme having a cholesterol ester hydrolyzing activity used in the present invention may be either lipase or cholesterol esterase. The lipase may be derived from a plant (eg wheat embryo), derived from an animal organ (eg pancreas) or derived from a microorganism (eg Candida rugosa). Especially Chromobaterium viscosu
The one obtained from m is useful.

The cholesterol esterase may be derived from animal organs or microorganisms, but is preferably microorganisms. For example, US Pat.
Those described in No. 5 164 can be used (Candital gussa, Actinomyces genus, Streptomyces genus,
Penicillium and the like).

The alkylphenoxypolyglycidol used in the present invention preferably has an alkyl group having 4 to 20 carbon atoms.
Two or more alkyl groups may be substituted on the benzene ring (such as dialkylphenoxypolyglycidol). The benzene ring has a substituent other than an alkyl group, such as an alkoxy group,
You may have a halogen atom etc. The glycidol unit in the molecule is appropriately in the range of 7 to 20, but the coating characteristics,
The number of glycidol units is selected according to the purpose such as the development characteristics. The alkylphenoxypolyglycidol is preferably contained in the porous spreading layer together with the enzyme having a cholesterol ester hydrolyzing activity.

In order to detect and quantify cholesterol, a known reagent composition for detecting a reaction can be used. There are the following two types of known detection reactions.

Cholesterol is oxidized by the enzymatic activity of cholesterol oxidase, and the produced hydrogen peroxide or cholestenone is detected and quantified by a known color development reaction or fluorescence of cholestenone.

In the dehydrogenation reaction due to the enzymatic activity of cholesterol dehydrogenase, the produced cholestenone is detected and quantified by a known reaction, or NADH or NADPH generated by the reduction of NAD or NADP coupled with the dehydrogenation reaction is directly optically detected and quantified. Or the colored substance produced through a chemical reaction is optically detected and quantified.

A chromogenic reagent composition for detecting hydrogen peroxide produced by cholesterol oxidase contains [A] chromogen and a coupler. In the presence of peroxidase, hydrogen peroxide oxidatively couples the chromogen and coupler to form a quinoneimine dye. Or a compound [B] which is a leuco dye or an auto-oxidative chromogen that is oxidized by hydrogen peroxide in the presence of peroxidase to form a dye.

As a chromogen, Ann. Clin. Biochem., 6 , 24-27 ((1
969), 4-aminoantipyrine (also known as 4-aminophenazone, that is, 1-phenyl-2,3-dimethyl-4-amino-3-pyrarin-5-one), JP-A-59-
1- (2,4,6-trichlorophenyl) described in 54962 and the like
2,3-Dimethyl-4-amino-3-pyrrolidin-5-one, 1- (3,5-1 (3,5-dichlorophenyl) -2,3-
Tri-substituted-4-amino-3-pyrazolin-5-one such as dimethyl-4-amino-3-pyrazolin-5-one, 1-phenyl-2,3-dimethyl-4 described in JP-B-55-25840 4-Aminoantipyrine analogues such as dimethylamino-3-pyrazolin-5-one can be used.
Among these compounds, 4-aminoantipyrine,
1- (2,4,6-trichlorophenyl) -2,3-dimethyl-
4-amino-3-pyrazolin-5-one, 1- (3,5-
Dichlorophenyl) -2,3-dimethyl-4-aminopyrazolin-5-one and the like are preferable.

As a coupler, Ann. Clin. Biochem., 6 , 24-27 (1
969), Japanese Patent Sho 55-25840, Japanese Patent Sho 58-45599, Japanese Patent Sho 58
-1628, JP-A-55-164356, JP-A-56-124398, JP-A-56-155852, and the like, phenol, 2-hydroxy-
1-benzenesulfonic acid, 4-hydroxy-1-benzenesulfonic acid, 3,5-dichloro-2-hydroxy-1
-Benzenesulfonic acid, 2-hydroxy-3-methoxy-1-benzenesulfonic acid and other phenolsulfonic acids (including alkali metal salts and alkaline earth metal salts); 1-naphthol, 2-naphthol; 1,7-dihydroxy Dihydroxynaphthalene such as naphthalene; Naphtholsulfonic acid such as 1-hydroxy-2-naphthalenesulfonic acid and 1-hydroxy-4-naphthalenesulfonic acid (including alkali metal salt and alkaline earth metal salt); Other phenol or naphthol derivative There is. Among these compounds, 1,7-dihydroxynaphthalene, 1-hydroxy-
2-naphthalenesulfonic acid (including Na salt, K salt, Li salt),
3,5-Dichloro-2-hydroxy-1-benzenesulfonic acid (including Na salt, K salt, Li salt) is preferable.

As a leuco dye, 2- (4-) described in JP-B-57-5519
Triarylimidazole leuco dyes such as hydroxy-3,5-dimethoxyphenyl) -4,5-bis [4- (dimethylamino) phenyl] imidazole, JP-A-59-1933
52- (3,5-dimethoxy-4-hydroxyphenyl) -4- [4- (dimethylamino) phenyl]-
Diaryl imidazole leuco dyes such as 5-phenethyl imidazole and diaryls such as 2- (2-phenyl-3-indolyl) -4,5-di [4- (dimethylamino) phenyl] imidazole described in JP-A-61-1960. Indolyl imidazole leuco dye, 2- (4-hydroxy-3,5-dimethoxyphenyl)-described in JP-A-61-229868
Triarylmonoacylimidazole leuco dyes such as 3-acetyl-4,5-bis [4- (dimethylamino) phenyl] imidazole, 2- (4-hydroxy-3,5-dimethoxyphenyl) -3-methyl-4, 5-bis [4-
Examples include triarylmonoalkylimidazole leuco dyes such as (diethylamino) phenyl] imidazole.

As a method for quantifying NADH or NADPH, a colorimetric method for forming a formazan dye having an absorption spectrum in the visible region from an electron-accepting dye precursor in the presence of an electron transfer agent is disclosed in JP-A-49-11395. Is known for. Using this reaction system, NADH or NA can be obtained by spectrophotometry in the visible range.
Can quantify DPH.

As an electron transfer agent, diaphorase (EC 1.6.4.3),
N-methylphenazonium, methosulfate, etc. can be used.

As the formazan dye precursor, INT, that is, 2-
(P-Iodophenyl) -3- (p-nitrophenyl)
-5-phenyltetrazolium chloride, BT i.e. 3,
3 '-(3,3'-dimethoxy-4,4'-biphenylene) bis [2,5-diphenyltetrazolium chloride], 3,3'
-(4,4'-biphenylene) bis [2,5-diphenyltetrazolium chloride] and the like can be used, but nitrotetrazolium blue (NTB or NBT), that is, 3,3 '
-(3,3'-dimethoxy-4,4'-diphenylene) bis [2- (p-nitrophenyl) -5-phenylotetrazolium chloride] is preferred.

In the present invention, deoxycholic acid or a salt of deoxycholic acid can be contained in a layer containing an enzyme having a cholesterol ester hydrolase activity or a layer (adjacent layer etc.) which can be in liquid contact with the layer. As the salt of deoxycholic acid, sodium salt, potassium salt, lithium salt and the like can be used. Instead of deoxycholic acid, cholic acid, lithocholic acid, chenodeoxycholic acid, taurocholic acid, glycolic acid, taurodeoxycholic acid, glycodeoxycholic acid, and any one or a combination of two or more of these sodium salts and lithium salts. Combinations can be used.

The content (coating amount per 1 m ² of element) of each component of the reagent composition used in the element of the present invention in the element is as follows.

Cholesterol esterase: about 500 IU to about 50,000 IU, preferably about 1000 IU to about 30,000 IU lipase: about 5000 IU to about 300,000 IU, preferably 10,000 IU to 200,000 IU alkylphenoxypolyglycidol: about 200 mg to about 10 g, Preferably about 500 mg to about 5.0 g cholesterol oxidase: about 1000 IU to about 30,000 IU, preferably about 1500 IU to about 20,000 IU peroxidase: about 1000 IU to about 100,000, preferably about 2000 IU to about 60,000 IU chromogen or Leuco dye: about 0.5 mmol to about 10 mmol, preferably about 1 mmol to about 5 mmol Coupler: about 2.5 mmol to about 50 mmol, preferably about 5 mmol to about 25 mmol Deoxycholic acid or its salt: about 200 mg to about 10 g, preferably about 500 mg to about 5.0 g (same amount in case of acid or salt used instead of deoxycholic acid) Cholesterol dehydrogenase: about 200 IU to about 30,000 IU, preferably About 300 IU to about 20,000 IU NAD or NADP: about 10 mg to about 50 g, preferably about 30 mg to about 20 g Electron transfer agent: about 1 mg to about 1000 mg, preferably about 5 mg to about 500 mg About 500 IU for diaphorase 20,000 IU, preferably about 700 IU to about 10,000 IU formazan dye precursor: about 5 mg to about 10 g, preferably about 25 mg to about 5 g The present invention can be applied to various known dry analytical elements. In particular, it can be applied to an element including a solid carrier through which both the detection reagent system and the test liquid can pass. The element is provided on a light-transmissive, water-impermeable support on which a submerged layer, a water-absorbing layer, an adhesive layer, a coloring (or coloring) reagent layer, a reaction reagent layer, a light-shielding layer, and a transparent layer are provided in addition to the porous liquid spreading layer. It may be a multilayer structure having layers and other known layers. As such an analysis element, U.S. Pat. No. 3,992,158, JP-A-55-16435
No. 6, and JP-A No. 60-222769.

When a support is used, the dry analytical element of the present invention can be practically adopted as (1) having a porous liquid developing layer on the support. Those having a water absorbing layer between the support and the spreading layer are preferable.

(2) A support having a reagent layer and a spreading layer in this order on the support. A water absorption layer may be provided between the support and the reagent layer.

(3) A support having a second reagent layer, a first reagent layer, and a spreading layer in this order on a support. A water absorbing layer may be provided between the support and the second reagent layer.

(4) A support having a reagent layer, a light shielding layer, and a spreading layer in this order on a support. A water absorption layer may be provided between the support and the reagent layer.

(5) The second reagent layer, the light shielding layer, the first reagent layer, on the support,
Having a development layer in this order. A water absorbing layer may be provided between the support and the second reagent layer.

In the above (1) to (5), between the water absorption layer and the reagent layer or the spreading layer, between the reagent layer and the liquid spreading layer, between the second reagent layer and the first reagent layer, or between the light shielding layer and the reagent layer or spreading. Overlayers may be provided between the layers.

The multilayer analytical element of the present invention may be provided with a water-absorbing layer (optionally via another layer such as a subbing layer) on the light-transmitting water-impermeable support. The water absorption layer provided in the dry analytical element of the present invention is preferably a layer made of a hydrophilic binder, that is, a layer having a hydrophilic polymer that absorbs and swells water as a layer forming component.

Hydrophilic polymers that can be used in the water-absorbing layer are natural or synthetic hydrophilic polymers having a swelling ratio at water absorption of 30 ° C. of about 1.5 to about 20, preferably about 2.5 to about 15.
Examples of such hydrophilic polymers include gelatin (eg, alkali-treated gelatin, acid-treated gelatin, deionized gelatin, etc.), gelatin derivatives (eg, phthalated gelatin), agarose, polyacrylamide, polyvinyl alcohol, polyvinylpyrrolidone. Etc. can be given.

The dry thickness of the water-absorbent layer is in the range of about 1 μm to about 100 μm, preferably in the range of about 3 μm to about 30 μm. Further, the water absorbing layer may contain a surfactant (cationic, amphoteric or nonionic) and a pH buffering agent, if necessary.

When the water absorbing layer also serves as the detection layer, the water absorbing layer may contain light shielding (reflecting or light absorbing) fine particles as necessary.

An adhesive layer for adhering and laminating the spreading layer may be provided on the water absorbing layer, the light shielding layer, the overlayer, the reagent layer and the like. The adhesive layer is preferably made of a hydrophilic polymer capable of adhering the spreading layer when wet with water or when swollen with water. Examples of hydrophilic polymers that can be used in the adhesive layer include the same hydrophilic polymers that are used in the water absorbing layer. Of these, gelatin, gelatin derivatives, polyacrylamide and the like are preferable. The dry thickness of the adhesive layer is generally about 0.5 μm to about 2
It is in the range of 0 μm, preferably about 1 μm to about 10 μm. The adhesive layer may be provided between the desired two layers in order to improve the adhesive force between the other layers, in addition to between the reagent layer and the spreading layer. The adhesive layer can be provided by a known method, such as a method of applying an aqueous solution containing a hydrophilic polymer and a surfactant and the like, which is added, on the reagent layer and the like.

The light shielding layer is preferably a water permeable layer in which light reflecting fine particles are dispersed with a hydrophilic polymer as a binder. The light-reflecting fine particles were spotted and supplied to the developing layer when the detectable change (color change, color development, etc.) that occurred in the reagent layer (or the water-absorbing layer) was reflected and measured from the light-transmissive support side. It also functions as a light-reflecting layer or a background layer while blocking the color of the aqueous liquid, particularly hemoglobin and red when the sample is whole blood. Preferred examples of the light-reflecting fine particles are titanium dioxide fine particles and barium sulfate fine particles. If necessary, the light-shielding fine particles as described above may also be contained in the spreading layer.

The porous liquid spreading layer is preferably a spreading layer having a liquid metering action. Having a liquid metering action means that the liquid sample spot-supplied to the surface of the liquid sample is distributed in a unit area in the lateral direction (direction along the plane of the layer) without substantially unevenly distributing the components contained therein. That is, it has the action of spreading the liquid sample to the lower layer by spreading it at a substantially constant rate.

The material forming the matrix of the spreading layer is paper,
Non-woven fabrics, woven fabrics (eg, plain weaves such as broad and poplin), knitted fabrics (eg, tricot knit, double tricot knit, Milanese knit, etc.), glass fiber paper, membrane heilter (blushed polymer layer), or polymer micro A three-dimensional lattice-like structure made of beads or the like can be used. Among these, it is preferable to use a fibrous layer represented by a woven fabric and a knitted fabric.
For details of these, see JP-A-55-164356 and JP-A-57.
-66359 and JP-A-60-222769 may be referred to.

The woven cloth material or knitted cloth material that can be used in the dry analysis element of the present invention is preferably substantially free from at least oils and fats attached at the time of manufacturing a thread, a woven material or a knitted material by a degreasing treatment such as washing with water.

When a liquid sample is applied to the development layer, the color-developing reagent layer, the reaction reagent layer or the water-absorption layer and the analysis operation is performed, the pH in the element is about 4.
It can be contained by appropriately selecting from known pH buffering agents that can be maintained at an appropriate value within the range of 5 to about 9.0. As a buffering agent that can be used, "Chemical Handbook Basic Edition" edited by The Chemical Society of Japan (Tokyo, Maruzen Co., Ltd., 1966) 1312-1320
Page, RMCDawson et al. Eds. “Data for Biochemical R
esearch "Second Edition (Oxford at the Clarendon Press, 19
1969) pp. 476-508, "Biochemistry" Vol. 5, 467-4
77 pages (1966), "Analytical Biochemistry" Vol. 104
There is a pH buffering agent described in 300 to 310 (1980).

The multilayer analytical element of the present invention can be prepared by known methods described in the above-mentioned patent specifications.

The multilayer analysis element of the present invention is cut into small pieces such as a square or a circle of approximately the same size having a side of about 15 mm to about 30 mm.
-28331, S.A. 56-142454, JP-A-57-63452, S.S. 58
-32350, Tokushusho Sho-501501144, etc. can be used as a chemical analysis slide by placing it in a slide frame,
It is preferable from various viewpoints such as transportation, storage, and measurement operation. Depending on the purpose of use, it can be used as a long tape in a cassette or magazine, or a small piece can be attached or stored in a card with an opening.

The multilayer analytical element of the present invention can carry out a quantitative analysis of cholesterol as an analyte in a liquid sample by the operations described in the above-mentioned patent specifications and the like. That is, about 5 μL to about 30
1 μL, preferably 8 μL to 15 μL, of the whole liquid, plasma, serum, urine, etc.
Incubate at a substantially constant temperature in the range of about 20 ° C. to about 40 ° C. for a period of about 10 minutes, preferably at a substantially constant temperature of about 37 ° C. to visualize color development or discoloration within the element. The total cholesterol content in a liquid sample is measured by reflection measurement from the side of the light-transmissive support using light having a maximum absorption wavelength of light or ultraviolet light or a wavelength in the vicinity of the absorption maximum wavelength and using a calibration curve prepared in advance according to the principle of colorimetric measurement method. The quantity can be calculated. Alternatively, the intensity of fluorescence in the element can be measured, and the total cholesterol content in the liquid sample can be determined using a calibration curve prepared in advance. The quantitative analysis of the analyte can be carried out with high accuracy by keeping the amount of the liquid sample to be spotted, the incubation time and the temperature constant. Measurement operation is JP-A-60-12
5543, JP-A-60-220862, JP-A-61-294367, JP-A-58
With the chemical analyzer described in -161867 and the like, highly accurate quantitative analysis can be performed with extremely easy operation.

Example 1 A 180 μm thick polyethylene terephthalate (PET) colorless transparent smooth film (support) provided with a gelatin subbing was coated with an aqueous solution for forming a color forming layer having the following composition so that the dry film thickness was 15 μm. It was applied and dried to provide a coloring layer.

Components of aqueous solution for forming color layer Alkali-treated gelatin 12g Nonifphenoxypolyglycidol (containing 10 units of glycidol units) 0.3g NAD 0.8g Diaphorase 2800IU NTB (*) 0.2g Water 98ml * 3,3 '-(3,3'- Dimethoxy-4,4'-biphenylene) bis [2- (p-nitrophenyl) -5-phenyl-2H-tetrazolium chloride] Next, an aqueous solution for forming a light-shielding layer having the following composition is dried on the coloring layer. It was applied to a thickness of 7 μm and dried to provide a light shielding layer.

Aqueous solution for forming light shielding layer Alkali-treated gelatin 14 g Rutile-type titanium oxide 70 g Water 100 ml A gelatin aqueous solution having the following composition was coated on the light shielding layer to a dry film thickness of 2 μm, and dried to form an adhesive layer. .

Aqueous solution for forming adhesive layer Alcali-treated gelatin 4g Nonylphenoxypolyethoxyethanol (containing 10 oxyethylene units on average) 0.1g Water 90g Water is supplied almost uniformly over the entire surface at a rate of about 30g / m ² on the adhesive layer. DET spun yarn tricot knitted cloth material (equivalent to 50 denier yarn thickness,
A cloth developing layer was provided by laminating a cloth having an average thickness of 250 μm while lightly applying pressure, drying and adhering.

An aqueous suspension of the reagent composition for cholesterol analysis having the following composition was applied onto the cloth spreading layer almost uniformly at a rate of 200 ml / m ² , and dried to prepare an integrated multilayer analytical element for cholesterol determination.

Ethanol 500 ml Polyvinylpyrrolidone (average molecular weight 360,000) (10% ethanol solution) 18 g 2-Amino-2-ethyl-1,3-propadiol 11 g Nonylphenyloxypolyethoxyethanol (containing 40 oxyethylene units on average) 6 g Nonylphenoxypoly Glycidol (containing 10 glycidol units on average) 3 g sodium deoxycholate 7 g The following enzyme solution was added to the above ethanol solution and suspended.

Lipase (*) 15000IU Cholesterol dehydrogenase 5000IU Water 5ml * Integrated multi-layer analytical element for quantifying cholesterol produced from Chromobacterium viscosum was cut into a 15mm x 15mm square and placed in a plastic mount to complete an analytical slide for quantifying cholesterol. did.

10 μl of human plasma having different cholesterol concentrations was spotted on the development layer of the analytical slide and incubated at 37 ° C. for 6 minutes, and then the reflection optical density was measured from the PET support side with visible light having a central wavelength of 600 nm. The results are shown in Table 1.

[Example 2] On a PET film similar to that in Example 1, an aqueous solution having the following composition was applied to a dry film thickness of 15 µm and dried to form a color-developing layer.

Alkali-treated gelatin 12g Nonylphenoxypolyglycidol (containing 10 units of glycidol units) 0.1g Diaphorase 3000U NTB 0.35g Bis (vinylsulfonylmethylcarbonylamino) methane 0.12g Water 105g Next, dry film thickness of 7μm with water suspension of the following composition Was applied and dried to provide a light shielding layer.

Alkali-treated gelatin 7 g Rutile-type titanium oxide 35 g Water 50 ml Next, an adhesive layer containing cholesterol dehydrogenase was applied by the following formulation to a dry film thickness of 4 μm and dried.

Cholesterol dehydrogenase 39000U NAD 0.9g Alkali-treated gelatin 20g Nonylphenoxypolyglycidol (containing an average of 10 glycidol units) 0.2g Water 220g Dampening water was supplied at a rate of 30g / m ² on the adhesive layer, and then Example 1 A PET spun yarn knitted cloth material similar to that was laminated and adhered while pressing it to form a cloth development layer.

Next, an aqueous suspension of the reagent composition for cholesterol separation having the following composition was applied almost uniformly on this cloth-developed layer at a rate of 200 ml / m ² , and dried to obtain an integrated multilayer analytical element for cholesterol determination. Was prepared.

Ethanol 500 ml Polyvinylpyrrolidone (average molecular weight 360,000) (10% ethanol solution) 18 g 2-Amino-2-methyl-1,3-propanediol 11 g Sodium deoxycholate 7 g Nonylphenoxypolyethoxyethanol (containing 40 oxyethylene units on average) 6 g Nonylphenoxypolyglycidol (containing an average of 10 glycidol units) 3 g The following enzyme solution was added to the above ethanol solution.

Lipase (*) 15000U Water 3g * The element produced from Chromobacterium viscosum was processed in the same manner as in Example 1 to complete a chemical analysis slide for quantifying cholesterol. Next, a calibration curve was prepared in the same manner as in Example 1 and the values shown in Table 2 were obtained.

[Example 3] The same result as in Example 2 was obtained in the same manner as in Example 2 except that 12000 IU of cholesterol esterase was used instead of 15000 IU of lipase.

Claims (1)

[Claims] 1. A dry analytical element having at least one water-permeable layer and a porous liquid developing layer in a state in which they can be in liquid contact, wherein the developing layer contains an enzyme having a cholesterol ester hydrolyzing activity. A dry analytical element comprising an alkylphenoxypolyglycidol in a layer or at least one of the water-permeable layers.