JPS60237360A - Inspection apparatus - Google Patents

Abstract

PURPOSE:To improve the gas barrier property along with a higher transparency by making a specified silicon compound react with the inner or outer of a synthetic resin container to form a film with a continuous gas barrier. CONSTITUTION:The mixture of silicon compounds expressed by the formulas 1 and 2 is made to react with at least one of the inner and outer surfaces of a synthetic resin container body 2 to form a film with a continuous gas barrier property. Then, an inspection chemical agent 5a is housed into the body 2 and the opening is closed with a stickable plug member 4.

Description

DETAILED DESCRIPTION OF THE INVENTION 10. BACKGROUND TECHNICAL FIELD OF THE INVENTION The present invention relates to testing instruments. Specifically, the present invention relates to a testing instrument that has extremely high gas barrier properties, good transparency, and does not cause the internal testing agent to deteriorate over a long period of time.

PRIOR ART Conventionally, in order to culture microorganisms in blood or other body fluids for diagnosis, or to detect abnormal components in these blood or body fluids, there have been test instruments that contain test agents in containers. It is used. - For example, there are test instruments that contain body fluids, gel-like media, anticoagulants, etc. in a container.

As such testing instruments, glass containers have conventionally been used because the container main body is not gas permeable and has good transparency. However, glass containers have the disadvantage that they are easily damaged during storage, transportation, or use, and are heavy. For this reason, we investigated the use of lightweight and transparent containers made of synthetic resin, but since synthetic resins are more or less gas permeable, during long-term storage they may be exposed to surrounding atmospheric gases, such as air (oxygen gas). ) is transmitted through the test tube, resulting in the deterioration of the test chemicals inside. For example, a chemical that is easily oxidized, such as a liquid or gel medium, inside a synthetic resin container.
If an anticoagulant or the like is stored, the oxygen contained in the surrounding air that passes through the container will oxidize the drug and cause deterioration, so it is difficult to use a synthetic resin container. For example, it was necessary to store it in a vacuum packaging container. However, storage in a reduced pressure packaging container has the disadvantage that, since the packaging container is a reduced pressure container, it is extremely expensive and requires considerable time and effort to seal and open the can, resulting in high costs. On the other hand, the aqueous solutions of the test chemicals have the disadvantage that the moisture inside evaporates and passes through the synthetic resin container as water vapor, resulting in a change in concentration.

■Object of the Invention Accordingly, an object of the present invention is to provide a novel testing instrument. Another object of the present invention is to provide a testing device that contains a liquid or gel medium, an anticoagulant, and other chemical solutions, has a high gas barrier property, has good transparency, and does not deteriorate the test chemical solution over a long period of time. It is about providing. Still another object of the present invention is to provide an inspection instrument that is free from damage and has extremely high gas barrier properties.

The purpose of these openings is to form a synthetic resin container main body having a sealable opening in at least one place, on at least one of the inner and outer surfaces of the container body with the general formula I (where m is an integer of 1 to 5). ) and a silicon compound having the general formula ■ (where n is an integer of 1 to 5, and R+ and R2 are an alkyl or alkoxy group having 1 to 4 carbon atoms, a phenyl group, or a hydroxyl group). (However, R'l and R2 are not hydroxyl groups at the same time.) A test agent is stored in a synthetic resin container made by reacting a mixture of silicon compounds having R'l and R2 to form a continuous film with gas barrier properties. However, this is achieved by a testing instrument in which the opening is closed with a plug member that can be pierced to form a sealed space inside.

'Furthermore, the present invention is an inspection instrument in which the synthetic resin container body is made of transparent synthetic resin. Furthermore, the present invention is an inspection instrument in which the coating is a transparent coating. The present invention is an inspection instrument in which the coating has a thickness of 0.01 to 2.0 μm. Further, the present invention provides a method in which m in the general formula (1) is an integer of 1 to 5, n in the general formula (2) is an integer of 1 to 5, and R1 and R2 are alkyl having 1 to 4 carbon atoms. It is an instrument for testing whether it is a group or a hydroxyl group. Furthermore, the present invention is an inspection instrument in which R1 in the general formula (2) is an alkyl group having 1 to 2 carbon atoms, and R2 is a hydroxyl group. The present invention is a testing instrument in which the testing agent is an oxidizing substance. Further, the present invention is a test instrument in which the oxidizing substance is a liquid or gel-like medium. Furthermore, the present invention is a test instrument in which the oxidizing substance is an anticoagulant. Further, the present invention is an inspection instrument in which the closed space has a pressure lower than atmospheric pressure.

■Specific structure of 0 invention Next, the present invention will be described in detail with reference to the drawings.

That is, FIG. 1 shows an example of a test instrument using a reduced pressure sampling method. This is a plug member 4 that seals an opening 3 of a main body 2, and a test agent 5a is stored in a space 5 within the container main body 2. Therefore, it is preferable that this space 5 is depressurized and further maintained under an atmosphere of gas inert to the testing agent. Further, a transparent film is formed on at least one of the inner and outer surfaces of the transparent synthetic resin container body 2 by reacting the silicon compounds of the general formulas I and (2) with plasma treatment. For example, as shown in FIG. 2A, a transparent coating 2a is formed on the entire outer surface of the container body 2 by reacting the silicon compound. Further, as shown in FIG. 2(B), the transparent coating 2b is formed on the entire inner surface of the container body 2.

Furthermore, as shown in FIG. 2(C), the transparent coating 2a is formed on the outer surface of the container body 2, and the transparent coating 2b is formed on the inner surface thereof. In addition, in FIGS. 2(A) to 2(C), the thicknesses of the transparent coatings 2a and 2b are exaggerated.

The synthetic resin constituting the tubular member used in the present invention is not particularly limited, but transparent synthetic resin is preferable;
Particularly preferred are styrene homopolymers or copolymers, methyl methacrylate homopolymers or copolymers, polycarbonates, and the like.

In addition to polystyrene, styrene polymers include copolymers of styrene and other copolymerizable monomers, such as butazine, methyl methacrylate, maleic anhydride, etc. There is at least one kind of thing.

One of the film-forming components is a silicon compound having the general formula (1) (where m is an integer of 1 to 5, preferably 1 to 2).

The other film-forming component is a silicon compound having the general formula (2). In the general formula (■), n
is an integer of 1 to 5, preferably 1 to 2.

R1 and R2 are alkyl or alkoxy groups having 1 to 4 carbon atoms, phenyl groups or hydroxyl groups, but R
1 and R2 are never hydroxyl groups at the same time. R1 and R2 are preferably an alkyl group having 1 to 4 carbon atoms or a hydroxyl group, but both are not hydroxyl groups, and in particular R1 is an alkyl group having 1 to 2 carbon atoms and R2 is a hydroxyl group. The case is preferred.

Therefore, the blending ratio of the silicon compound of the general formula (2) to 1 mole of the silicon compound of the general formula (I) is 0.5 to 3 moles, preferably 1 to 2 moles. Such mixtures of silicon compounds are used as solutions in organic solvents such as methanol, ethanol, isopropanol, etc. Its concentration is 3-50% by weight, preferably 5-35% by weight.

The transparent film is formed as follows.

For example, after the opening @ 13 of the tubular member 2 is closed using the plug member 4 or other closing member or occluder (not shown), the tubular member 2 is immersed in the mixed solution of the silicon compound. adhere to the outer surface of the The immersion time is usually 0.01 to 10 minutes, preferably 1 to 5 minutes. In this case, applying ultrasonic waves during dipping will promote the removal of air from the micropores on the surface of the synthetic resin tubular member and the infiltration of solutes into the micropores, thereby improving the gas barrier properties of the resulting film. improves. Note that the application of the mixed solution is not limited to dipping, but can also be performed by spraying or other methods.

The temperature of the immersion treatment is usually 0 to 50°C, preferably 10-50°C.
The temperature is 30°C. In addition, when carrying out under the action of ultrasonic waves, the liquid temperature is 0 to 50°C, preferably 10 to 30°C, and 20 to 20°C.
2'0OK)-1z, preferably 25-50 K Hz
It is carried out at a frequency of 0.1 to 10 minutes, preferably 0.5 to 5 minutes.

The tubular member coated with the 8-solution solution in this way is
3 to 3 at a temperature of 50 to 120°C, preferably 60 to 70°C.
After being dried for 30 minutes, preferably 5 to 15 minutes, it is subjected to plasma treatment.

The coating film on the surface of the tubular member is plasma-treated to form a transparent film, for example, in the following manner. That is, as shown in FIG. 3, an electrode 14 is provided in a reactor 13 equipped with a gas inlet 11 and a gas outlet 12, and a tubular member support 15 is installed on the electrode 14. The support 15 supports the tubular member 2 coated with the mixed solution.

For example, the tubular member support 15 is inserted and supported inside the tubular member 2 after the seal is removed. Note that the cooling fA device 16, 1 should not be brought into contact with the electrode 14, and the cooling device 16 should not be brought into contact with the electrode 14.
a is connected to a temperature controller 16b to circulate a cooling medium such as water. Further, the electrode 14 is connected to a ground 17 . On the other hand, a counter electrode 18 is provided on the opposite side of the electrode 14,
The counter electrode 18 is connected to a high frequency power source 20 through one matching box. An oxygen container 21 and a flow meter 22 are connected to the gas inlet 11 . On the other hand, pressure reducing devices such as an oil diffusion pump 24 and an oil rotary pump 25 are connected to the gas discharge port 12 via a trap 23 .

In FIG. 3, reference numeral 26 is a pressure sensor, 27 is a vacuum gauge, and 28 is a thermometer.

Then, a pressure reducing device such as an oil rotary pump is operated to exhaust the atmospheric gas in the reactor 13 from the gas exhaust port 12, and while maintaining a predetermined degree of pressure reduction, the oxygen gas is transferred from the oxygen container 21 to the flow meter 22. While supplying the gas through the gas inlet 11 to the reactor 13, the electrodes are energized to irradiate plasma onto the coated surface 8e for treatment.

In this case, 1
For example, it is desirable to perform irradiation while rotating the container body.

The pressure inside the reactor during plasma treatment is o, oi~2, □
Torr, preferably 0.1 to 0.5 Torr. The electrode 14, which is the base, is cooled by a cooling medium circulated in the cooling device 16a, but the temperature inside the reactor is between 0 and 1.
The temperature is 50°C, preferably 30-70°C. Further, the high frequency power amount is 0.05 to 2°0 W/cm2, preferably 0.05 to 2°0 W/cm2.
2 to 1,5 W/cm2. Furthermore, gases containing oxygen atoms include monomolecular oxygen, ozone, carbon oxide, carbon dioxide, nitrogen oxide, dinitrogen oxide, and mixed gases of these and other gases (e.g., argon, nitrogen helium, etc.). However, it is preferably molecular oxygen (hereinafter referred to as i! elementary gas), and the oxygen content is preferably 20 to 100%.The plasma irradiation is performed for 0.1 to 60 minutes, preferably 0. The irradiation is carried out for 3 to 5 minutes.This is preferable since it is possible to prevent the temperature of the tubular member, which is the base material, from increasing due to the plasma irradiation.

The thickness of the transparent film formed under these reaction conditions is 0.
．． 01~2.0μm, preferably 0.03~0.2μm
It is.

In the method of the present invention, better results can be obtained if the tubular member is subjected to a cleaning treatment before or after closing the opening before applying the mixed solution of the silicon compound.

Cleaning can be done with water, acid aqueous solution, alkaline aqueous solution, alcohol,
This is carried out using an aqueous surfactant solution, but an alkaline aqueous solution is preferred. Examples of alkaline aqueous solutions include sodium carbonate, potassium carbonate, lithium carbonate, sodium hydrogen carbonate, sodium hydroxide, potassium hydroxide, etc.
．． It is used as an aqueous solution of 1 to 20% by weight, preferably 3 to 10% by weight. The processing time is usually 0.1 to 30
minutes, preferably 5 to 10 minutes, and the liquid temperature is 0 to 10 minutes.
The temperature is 150°C, preferably 10-40°C. Furthermore, even better results can be obtained if the cleaning treatment is performed using ultrasonic waves. Ultrasonic treatment is 20-200KHz
, preferably 0.1-10 at a frequency of 25-50 KHz
This is carried out for 0.5 to 5 minutes, preferably 0.5 to 5 minutes. It goes without saying that if cleaning treatment is carried out using ultrasonic waves, fairly good results can be obtained even with treatment agents other than aqueous alkaline solutions. Furthermore, the best results can be obtained by performing ultrasonic cleaning using an alkaline aqueous solution.

In addition to butyl rubber, the material constituting the plug member 4 may include materials with sufficient elasticity to allow the blood collection needle to be inserted during use and to prevent the gap between the blood collection needle and the plug member from loosening due to the puncture of the blood collection needle, as will be described later. It is desirable that the material has a low gas permeability. Typical examples thereof include, for example, a blend of a thermoplastic elastomer, polyisobutylene, and partially crosslinked butyl rubber, and preferably a blend of a thermoplastic elastomer, polyisobutylene, and partially crosslinked butyl rubber.

The composition of each component in the formulation is 100 to 200 parts by weight, preferably 120 to 150 parts by weight, of polyisobutylene per 100 parts by weight of thermoplastic elastomer, and 100 to 200 parts by weight, preferably 120 parts by weight, of partially crosslinked butyl rubber.
~150 parts by weight.

Examples of thermoplastic elastomers include ethylene-propylene rubber, polyester elastomer, nylon elastomer, styrene-isoprene block copolymer, styrene-butadiene block copolymer, polybutadiene,
Examples include thermoplastic polyurethane, hydrogenated styrene-butadiene block copolymer, and the like. Polyisobutylene has a molecular weight of 15°000 to 200,000. Preferably 80,0
00 to 150.000. Partially crosslinked butyl rubber is obtained by partially crosslinking butyl rubber obtained by copolymerizing isobutylene with a small amount (for example, 0.3 to 3.0 mol%) of isoprene.

The test agent 6 is stored in the synthetic resin container thus obtained, and the internal space 5 is brought into a reduced pressure state or an inert gas atmosphere such as nitrogen, argon, helium, carbon dioxide, etc., and the opening is opened. The portion 3 is closed with a plug member 4.

Testing agents include various liquid or gel media, anticoagulants such as ethylenediaminetetraacetate, sodium oxalate, CPD solution, ACD-A solution, and the like.

The test instrument thus obtained is sterilized before or after storing the test drug. For sterilization, steam sterilization, radiation sterilization, etc. can be used, but radiation sterilization is preferable. In this case, the radiation used is gamma rays, electron beams, etc. ! There is radiation, preferably gamma rays, the irradiation intensity of which is between 0.1 and 4 Mrad, preferably between 0.5 and 2.5 Mrad.

(3) Specific effects of the invention The reduced pressure collection tube having the above configuration is used as described below. That is, as shown in FIG. 4, one end is closed and the other end is open, and the opening is inserted into the screw hole 7 of the closed end 6. The collection needle 8 is composed of, for example, a blood vessel piercing part 8a and a plug puncturing part 8b, and the plug puncturing part 8b is wrapped with a luer adapter 10 made of synthetic resin. Next, the blood vessel piercing portion 8a of the sampling needle 8 is pierced into a blood vessel, for example, a vein, and the reduced pressure blood sampling whistle 1 is inserted under pressure into the occluded portion 6 of the blood sampling tube holter 9. As shown in FIG. The plug puncturing part 8b punctures the Luer adapter 10 and the plug member 4, and the tip thereof reaches the internal space 5 of the collection tube 1, so that the blood vessel and the internal space 5 communicate with each other, and the internal space 5
Due to the negative pressure within the blood vessel, blood within the blood vessel flows into the internal space 5 of the collection tube 1 in an amount corresponding to the degree of reduced pressure. Next, the blood vessel piercing portion 8a of the sampling needle 8 is removed from the blood vessel, thereby completing the surface sampling. In this case, if the subject or cultured material is anaerobic, the test or culture is performed as is, or if it is aerobic, the test or culture is performed by opening the cap or introducing oxygen or air.

Next, the present invention will be explained in more detail by giving examples.

Example 1 A bottomed tube made of polyethylene terephthalate was prepared at a concentration of 2.
After immersing in a 5% Na2CO3 solution for 5 minutes (with ultrasonication), the above silicon compound'a concentration was 12% by weight.
After being immersed in an isopropanol solution at 20° C. for 1 minute, it was pulled up at a pulling speed of 16 cm for 77 minutes. Then,
Dry at 70'C for 30 minutes. Next, the synthetic resin tube was inserted into the low-temperature plasma reactor (Fig. 3, 13) and degassed (0.05 Torr).
With bare cuff, under pressure of about 0.3 Torr,
Plasma output power 100W, plasma treatment for 2 minutes using parallel plate electrodes (50mm x 50m flat plate used), distance 5Qm
m, the film thickness was 0.1 on one side of this tube.
A transparent film of μm was deposited. In order to measure the oxygen and carbon dioxide permeability of this coated tube, the same treatment as above was carried out using a film of the above material (film thickness: 12 μm, area: 500 m 2 ). The gas permeability of this coating film was measured using a gas permeation measuring device manufactured by Ridge Co., Ltd. and was 50.
6m/! /m2 ・day-atm and 204 ml
/m2-day-atom.

Hereinafter, all the gas permeability tests in the Examples were conducted in the same manner using separate films.

The culture medium shown in Table 1 was placed in this container, and 25 parts by weight of thermoplastic elastomer (1,2-polybutadiene), 25 parts by weight of polyisobutylene (molecular weight 100,000>35 parts by weight, 25 parts by weight of partially crosslinked butyl rubber, and 15 parts by weight of liquid paraffin) were added. It was sterilized with gamma rays of 1.4 Mrad after being sealed under reduced pressure with a stopper made of a compound of There was almost no change in the amount.

Table 1 Tryptone 17 (] Soybean peptone 3g Meat extract 3g, Yeast extract 5g Liver ice melt 19 Glucose 2.5g Potassium hydrogen phosphate 2.5° Sodium chloride 5g L-cysteine hydrochloride 0.45g P-aminobenzoic acid 0 .05Q Borianetholsulfate A nate sodium 0,35 (] Hemin 0,005g Agar 0.01g Gelatin 12g Distilled water 1,000ml pH 7,3±0.1 Example 2 In the same method as Example 1, the concentration When a 5% by weight mixed solution was ultrasonically treated for 5 minutes, dried at 70°C for 3 minutes, and immediately subjected to plasma treatment under the same conditions, a transparent film with a thickness of 0.04 μm was formed. The oxygen and carbon dioxide permeabilities were 28 and 313, respectively.

Comparative Example 1 Oxygen and carbon dioxide gas permeability G and t of the untreated polyethylene terephthalate used in Example 1 were 2, respectively.
31 and 939.

Comparative Example 2 In the method of Example 1, the film was immersed for 5 minutes in a mixed solution containing 32% iron without performing ultrasonic treatment or plasma treatment, and then dried at 60° C. for 180 minutes, resulting in a film thickness of 0.1
A transparent film of 4 μm was formed. The oxygen and carbon dioxide permeabilities were 213 and 854, respectively.

Examples 3 to 12 Same as Example 1, film thickness 12 μm (Examples 8 to 12)
In No. 12, a polyethylene terephthalate film (area: 50 cm 2 ) with a film thickness of 11.5 μm was ultrasonically cleaned using the treatment agent shown in Table 2, and then dried. Next, a mixed solution of silicon compounds similar to that in Example 1 is shown in Table 2. The sample was used at a concentration of 1', immersed in the mixed solution for 5 minutes, subjected to ultrasonication at a frequency of KHz, and then dried at 70° C. for 5 minutes. Next, plasma treatment was performed under the conditions of 0.18 Torr and the output numbers shown in Table 2, and the results shown in Table 2 were obtained.

(The following is a blank space) Example 13 As shown in FIG. 1, a tubular container 2 having a wall thickness of imm and having one end closed and the other end open was made of polyethylene terephthalate. This tubular container was subjected to ultrasonic treatment in a 5% aqueous sodium carbonate solution for 1 hour, and then dried at a temperature of 60° C. for 15 minutes. Then, 1 of the same silicon compound as in Example 1 was added.
45 K by immersion in a mixed solution with a concentration of 2% by weight for 5 minutes.
After ultrasonication at a frequency of Hz, drying was performed at a temperature of 70° C. for 15 minutes. Furthermore, plasma treatment was performed for 5 minutes in the presence of oxygen gas at a power output of 200 W to obtain a transparent zero.
．． A synthetic resin container having a coating of 0.07 μm on the outer surface was obtained. The culture medium shown in Fig. 3 was placed in this container, and 25 parts by weight of thermoplastic elastomer (1,2-polybutadiene) and polyimbutylene (molecular weight 100°000>35 weight a81'5) were added.
The container was sealed under reduced pressure with a stopper member made of a mixture of 25 parts by weight of partially cross-linked butyl rubber and 15 parts by weight of liquid paraffin, and then sterilized with gamma rays at 1.5 Mrad. The results showed that there was no change in the quality of the culture medium and there was almost no change in the amount of pressure reduction even though it was left for three months.

(Left below) Table 3 Tryptone 179 Soybean peptone 3Q Meat extract 3Q Yeast extract 5g Liver lysate 1g Glucose 2.5g Potassium hydrogen phosphate 2.5g Sodium chloride 5g Cystine hydrochloride 0.45g P-aminobenzoic acid o , 059 Sodium polyanetholesulfonate 0,35g Hemin o,oosg Agar 0.01g Gelatin i2g Distilled water i,ooomλ DH7,3±0.1 ■0Specific effects of the invention As described above, the present invention has the following effects: A gas barrier is formed by reacting a mixture of a silicon compound having the general formula (■) and a silicon compound having the general formula (■) on at least one of the inner and outer surfaces of a synthetic resin container body having at least one sealable opening. This is a testing instrument in which a testing agent is stored in a synthetic resin container formed with a continuous film, and the opening is closed with a punctureable plug member to form a sealed space inside. The gas permeability, particularly the oxygen gas permeability coefficient, is extremely low, so that gas permeation from the outside of the testing instrument into the instrument and from the inside of the instrument to the outside of the instrument is substantially eliminated. Therefore, when storing and using testing agents that are susceptible to oxygen deterioration, such as culture media and anticoagulants, the permeation of gas, especially oxygen, from outside the device into the device is virtually eliminated, resulting in a long period of time. There will be no deterioration of the testing agent over time. Furthermore, in the case of a test agent stored as an aqueous solution, even if water evaporates, the water vapor does not permeate outside the instrument, and therefore a predetermined concentration can be maintained for a long period of time. Moreover, since the testing instrument according to the present invention is made of synthetic resin, there is no fear of damage even if it is subjected to impact during transportation, storage, or use. In addition, since it has good gas barrier properties, anaerobic culture is also possible if used as is.

[BRIEF DESCRIPTION OF THE DRAWINGS] FIG. 1 is a sectional view showing an example of an inspection instrument according to the present invention;
Figures 2(A) to 2(C) are partially enlarged views of the inspection equipment shown in Figure 1, Figure 3 is a schematic sectional view showing an example of the plasma processing apparatus used in the present invention, and Figures 4 to 5. The figure is a sectional view showing the state in which the vacuum collection tube is used. 1... Inspection equipment, 2... Container body, 2a, 2b.
...Transparent film, 3...Opening, 4...Plug member, 5
...Space, 5a...Test drug. Figure 1 (C-Figure 3)

Claims (9)

[Claims] (1) At least one sealable opening on at least one surface of the inner or outer surface of the synthetic resin container body having a diameter of 1° with the general formula I (where m is 1 to 5 is an integer of 1 to 4 carbon atoms, and a silicon compound having the general formula Although it is a hydroxyl group, R1 and R
A test agent is stored in a synthetic resin container made by reacting a mixture of silicon compounds having a hydroxyl group (2 and 2 are not hydroxyl groups at the same time to form a continuous film with gas barrier properties), and the opening is punctured. An inspection instrument that is closed with a plug member to form a sealed space inside. (2) The testing instrument according to claim 1, wherein the synthetic resin container body is made of transparent synthetic resin. (3) The second claim is that the coating is a transparent coating.
Inspection equipment described in Section. (4) The testing instrument according to any one of claims 1 to 3, wherein the coating has a thickness of 0.01 to 2.0 μm. (5) m in the general formula I is an integer of 1 to 5, and n in the general formula π is an integer of 1 to 5, and R1 and R2 are an alkyl group or a hydroxyl group having 1 to 4 carbon atoms. An inspection instrument according to claim 1. (6) The testing instrument according to claim 5, wherein R1 in the general formula π is an alkyl group having 1 to 2 carbon atoms, and R2 is a hydroxyl group. (7) The testing device according to any one of claims 1 to 6, wherein the testing agent is an oxidizing substance. (8) The testing instrument according to claim 7, wherein the oxidant is a liquid or gel-like medium. (9) Claim 7 in which the oxidant is an anticoagulant
Inspection equipment described in Section. (10) The testing instrument according to claim 1, wherein the closed space is in a reduced pressure state due to atmospheric pressure.