TECHNICAL FIELD

[0001] The present disclosure relates to a composition which is used as an environmental purification material. The present disclosure further relates to a process for producing the composition.

BACKGROUND

[0002] An absorbent or an acid or alkali solution has been widely used in many fields for preventing or removing malodors or for removing harmful substances in the air. One major problem associated with this method is the treatment of the waste solution or the adsorbent utilized, which may cause secondary pollution. A method to hide unpleasant odors by using a perfume may also be used, but there is a possibility that the odor of the perfume gets transferred to the food materials or substances and hence resulting in damage to the food by the smell of the perfume itself.

[0003] Further, organic chemical substances, particularly based on alcohols, phenols, aldehydes, carboxylic acid, esters, ethers, nitriles, peroxide-epoxides, halogens, and organometals have long been used to inhibit the proliferation of microbes. Although these chemicals provide good antibacterial and bactericidal strength, they are essentially toxic, irritate the skin, and may lead to allergies, "sickhouse syndrome", "hypersensitivity to chemical substances, etc. Therefore, care must be taken in the use of these chemicals.

[0004] US7375069 describes photocatalytic methods, compositions and kits for removal of stains such as oil, grease, paint, mold, rust and the like from solid surfaces.

[0005] EP 1437397 describes a cleaning agent comprising diamond-like carbon, or a covered component produced by partially covering the surface of diamond-like carbon with a ceramic, a thickener and an oxidant.

[0006] EP 1064999 discloses a photocatalytic powder for environmental clarification, comprising finely divided titanium dioxide having a coating of porous calcium phosphate, further calcium phosphate precipitated form pseudo substance solution consisting of sodium chloride, sodium phosphate, potassium chloride and so on.

[0007] US6180548 discloses an environment purifying material wherein porous apatite film is clotted on the surface of titanium oxide which comprises a carrier selected from alumina, silica gel, zeolite, clay sintered. The apatite is obtained from pseudo-body fluid having composition ratio: Na ⁺ 120 to 160 mM, HP0 ₄ ^2" 0.5 to 30 mM.

[0008] Therefore, there is a need of a composition for environmental clarification, and an economical process for its preparation which are safe and easy to use.

SUMMARY

[0009] The present disclosure relates to a composition comprising: a photocatalyst; a redox agent; an absorbent; a stabilizing agent, and water; wherein the photocatalyst is coated with apatite.

[0010] The present disclosure further relates to a process for producing a composition, the process comprising heating 2%-8% w/w of a photocatalyst coated with apatite in a slurry comprising at least one redox agent in the range of 40%-65% w/w, an absorbent in the range of 0.5%- 1% w/w, a stabilizing agent in the range of 20%-25% w/w, and water in the range of 0.5%- 1% v/w to obtain the composition.

[0011] The composition disclosed in the present disclosure is used as an environmental purification material by dissolving toxic materials into non-hazardous materials.

[0012] These and other features, aspects, and advantages of the present disclosure will become better understood with reference to the following description. This statement is provided to introduce a selection of concepts in simplified form. This statement is not intended to identify key features or essential features of the subject matter, nor is it intended to be used to limit the scope of the subject matter.

BRIEF DESCRIPTION OF THE DRAWINGS

[0013] The above and other features, aspects, and advantages of the subject matter will be better understood with regard to the following description, appended claims, and accompanying drawings where:

[0014] Figure 1 shows a plot of gas concentration (ppm) vs time period (h), where the testing gas is formaldehyde, in accordance with an embodiment of the present disclosure.

[0015] Figure 2 shows a plot of gas concentration (ppm) vs time period (h), where the testing gas is acetaldehyde, in accordance with an embodiment of the present disclosure. [0016] Figure 3 shows a plot of gas concentration (ppm) vs time period (h), where the testing gas is ammonia, in accordance with an embodiment of the present disclosure. DETAILED DESCRIPTION

[0017] The present invention now will be described more fully hereinafter. Indeed, the invention may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements. As used in the specification, and in the appended claims, the singular forms "a", "an", "the", include plural referents unless the context clearly dictates otherwise.

[0018] The present disclosure is to provide a composition for environmental clarification that has high dissolving capability, high absorption efficiency, improved safety and stability, and react with visible ray having low intensity.

[0019] The present disclosure provides a composition comprising: a photocatalyst; a redox agent; an absorbent; a stabilizing agent; and water; wherein the photocatalyst is coated with apatite.

[0020] The present disclosure provides a composition comprising: a photocatalyst in the range of 2%-8% w/w; a redox agent in the range of 40%-65% w/w; an absorbent in the range of 0.5%-l% w/w; a stabilizing agent in the range of 20%-25% w/w; and water in the range of 0.5%-l% v/w, wherein the photocatalyst is coated with apatite.

[0021] The present disclosure provides a composition for environmental clarification comprising: 8% w/w of a photocatalyst; 65% w/w of at least one redox agent; 1 % w/w of an absorbent; 25% w/w of a stabilizing agent; and 1% v/w of water, wherein photocatalyst is coated with apatite.

[0022] The photocatalyst used in the present disclosure is a substance that absorbs light and, in doing so, catalyzes a reaction. The photocatalyst of the present disclosure is selected from the group consisting of titanium dioxide, zinc oxide, tungsten oxide, silver oxide, copper oxide, and magnesium oxide.

[0023] The preferred photocatalyst used in the present disclosure is titanium dioxide which is further coated with apatite. [0024] Photocatalytic action results when titanium oxide is irradiated with light, an electron having a strong reducing action and a positive hole having a strong oxidizing action are generated and a molecular seed coming into contact therewith is decomposed by the oxidation-reduction action.

[0025] In accordance with the present disclosure, the average particle diameter of titanium dioxide is in the range of 20 nanometer to 500 nanometer.

[0026] The preferred average diameter of titanium dioxide is in the range of 100 nanometer to 300 nanometer.

[0027] If the average particle diameter is smaller than 20 nanometer, the efficient production is difficult to attain, whereas if the average particle diameter exceeds 500 nanometer, the performance as a photocatalyst greatly decreases.

[0028] In an embodiment, the composition is mixed with water in a range of 1 :70 to 1 : 100 w/v. The aqueous composition of the photocatalyst composition is used in the application quantity range of 0.1 -1.5 % (w/v) with water.

[0029] The apatite used in the present disclosure is selected from the group consisting of hydroxyapatite, fluorapatite, chlorapatite, and combinations thereof. The apatite of the present disclosure is a phosphate mineral used to coat the photocatalyst. The apatite of the present disclosure is in crystalline form. In general, apatite is used as a material for medical products and cosmetics, such as treatment for bone fracture and cavity.

[0030] The preferred apatite used in the present disclosure is hydroxyapatite. Hydroxyapatite is used because of safety to human body and has excellent absorbance of ammonia and aldehyde.

[0031] The photocatalyst coated with apatite can be produced by methods known in the art or can be purchased separately.

[0032] The redox reagent of the present disclosure is selected from the group consisting of citric acid, sodium percarbonate and combinations thereof. The redox reagent used in the present disclosure is a reagent that can induce an oxidation or a reduction.

[0033] In yet an embodiment of the present disclosure, the absorbent is selected from the group consisting of alumina, silica gel, zeolite, clay sintered compacts, glass and foam ceramics. The absorbent of the present disclosure is a substance that is capable of absorbing heat, light, moisture, etc.

[0034] The preferred absorbent is silica gel. Silica gel is suitable for the coating by its transparency.

[0035] The stabilizing agent of the present disclosure is sodium acid phosphate. Stabilizing agent is used in a very small amount to avoid the deterioration of its performance.

[0036] The composition of the present disclosure has an application in form of a paste, powder or slurry, preferably in form of a powder.

[0037] The shelf life of the composition for environmental clarification of the present disclosure is 2-3 years. However, the storage conditions to maintain the performance of the composition of the present disclosure during shelf life is by avoiding moisture and direct sunlight from the composition, further avoid storage at room temperature or higher temperature.

[0038] In accordance with the present disclosure, the temperature to store the composition is less than 10°C. The preferred temperature to store the composition is less than 5°C.

[0039] The present disclosure provides a composition for environmental clarification comprising: titanium dioxide in the range of 6%-8% w/w; citric acid in the range of 20%- 30% w/w; sodium percarbonate in the range of 30%-35% w/w; silica gel in the range of 0.5%-l% w/w; sodium acid phosphate in the range of 20%-25% w/w; and water in the range of 0.5%- 1% v/w, wherein titanium dioxide is coated with apatite.

[0040] The present disclosure provides a composition for environmental clarification comprising: 8% w/w of titanium dioxide; 30% w/w of citric acid; 35% w/w of sodium percarbonate; 1% w/w of silica gel; 25% w/w of sodium acid phosphate; and 1 % w/v of water, wherein titanium dioxide is coated with apatite.

[0041] In accordance to the present disclosure, the composition for environmental clarification is active at an intensity of light in a range from 1/10 to 1/100 candela. The composition of the present disclosure reacts with visible ray having low intensity like light emitted from electric bulbs, room light to high energy ray like sunlight and generate high dissolving capability.

[0042] A process for producing a composition, comprising heating 2%-8% of a photocatalyst coated with apatite in a slurry containing at least one redox agent, an absorbent, a stabilizing agent, and water to obtain a composition.

[0043] In an embodiment of the present disclosure, the process comprising heating 2%- 8% w/w of a photocatalyst coated with apatite in a slurry containing at least one redox agent in the range of 40%-65% w/w, an absorbent in the range of 0.5%- 1 % w/w, a stabilizing agent in the range of 20%-25% w/w; and water in the range of 0.5%-l% v/w to obtain a composition.

[0044] In an embodiment of the present disclosure, the process comprising heating 6%- 8% w/w of a photocatalyst coated with apatite in a slurry containing at least one redox agent in the range of 40%-65% w/w, an absorbent in the range of 0.5%-l % w/w, a stabilizing agent in the range of 20%-25% w/w; and water in the range of 0.5%- 1 % v/w in the temperature at the range of 30-40 °C for 20-26 hours to obtain a composition.

[0045] In another embodiment of the present disclosure, the process further comprises diluting the composition with water in a range of 1 : 70 to 1 : 100 w/v.

[0046] The composition disclosed in the present disclosure is used as an environmental purification material by dissolving toxic materials into non-hazardous materials like water and carbon dioxide gas. The composition for environmental clarification of the present disclosure is used to inactivate or eliminate micro-organisms, sterilize, deodorize sludge,

[0047] The advantageous effect of the composition of the present disclosure includes (1) anti bacteria and anti fungus effect: dissolve and eliminate E. coli, Legionella, influenza, viruses, fungus, and other toxicity causing microorganisms in foods, (2) deodorization: dissolve viruses, bacteria, and toxic organic materials that cause bad odor into odor-free and non-hazardous water and carbon dioxide gas, and (3) sanitization of soil and water quality: sterilize, deodorize sludge, water pollutant with anaerobes and blue-green algae. [0048] The composition for environmental clarification of the present disclosure is applied by spray coating a 100-fold diluted solution by plating machine or by ultra sonic nebulizer.

[0049] The applicable fields for the use of composition for environmental clarification of the present disclosure include hospital/operation room, hospital/examination room, waiting room, care home, kitchen/food processing plant, hotel, restaurant, vehicle, ship airplane, residence, chicken farm, dairy farm, risk management, and contingency planning.

[0050] The composition of the present disclosure is irradiated with light to generate hydroxyl radical (OH radical), whereby hydroxyl radical (OH radical) generated results in the conversion of toxic organic materials into non-hazardous materials.

[0051] In accordance to the present disclosure, the mechanism of the composition for the purification of environment includes; irradiating the light on the composition for environmental clarification to generate hydroxyl radical (OH radical), due to the polarity of the .OH radical the dissolving capability of the composition for environmental clarification increases that results in the conversion of toxic organic materials into non- hazardous materials such as water and carbon dioxide.

EXAMPLES:

[0052] The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the disclosed methods and compositions, the exemplary methods, devices and materials are described herein.

Example 1:

[0053] The composition for environmental clarification comprises the following active ingredients: • titanium dioxide coated with apatite in the range of 2%-8% w/w;

• citric acid and sodium percarbonate in the range of 40%-65% w/w;

• silica gel in the range of 0.5%-l% w/w;

• sodium acid phosphate in the range of 20%-25% w/w; and

• water in the range of 0.5%- 1 % v/w.

Example 2:

[0054] The composition for environmental clarification comprises the following active ingredients:

• titanium dioxide coated with apatite in the range of 6% -8% w/w;

• citric acid in the range of 20% -30% w/w;

• sodium percarbonate in the range of 30%-35% w/w;

• silica gel in the range of 0.5%- 1 % w/w;

• sodium acid phosphate in the range of 20%-25% w/w; and

• water in the range of 0.5%-l% v/w.

Example 3:

[0055] The composition for environmental clarification comprises the following active ingredients:

. · 8% w/w of titanium dioxide coated with apatite;

• 30% w/w of citric acid;

• 35% w/w of sodium percarbonate;

• 1% w/w of silica gel;

• 25% w/w of sodium acid phosphate; and

• 1 % v/w of water.

Example 4:

Preparation and storage of the composition:

[0056] 2%-8% w/w of titanium dioxide coated with apatite was heated in a slurry containing citric acid and sodium percarbonate in the range of 40%-65% w/w, silica gel in the range of 0.5%- 1 % w/w, sodium acid phosphate in the range of 20%-25% w/w and water in the range of 0.5%-l % v/w in the temperature at the range of 36-37 °C for 24 hours to obtain a composition.

[0057] The composition was stored at temperature less than 10°C.

Example 5:

Preparation and storage of the composition:

[0058] 6%-8% w/w of titanium dioxide coated with apatite was heated in a slurry containing citric acid and sodium percarbonate in the range of 50%-65% w/w, silica gel in the range of 0.5%-l% w/w, sodium acid phosphate in the range of 20%-25% w/w and water in the range of 0.5%-l% v/w in the temperature at the range of 36-37 °C for 24 hours to obtain a composition.

[0059] The composition was stored at temperature less than 10°C.

Example 6:

Preparation and storage of the composition:

[0060] 8% w/w of titanium dioxide coated with apatite was heated in a slurry containing 65% w/w of citric acid and sodium percarbonate, 1 % w/w of silica gel, 25% w/w of sodium acid phosphate and 1% v/w of water in the temperature at the range of 36- 37 °C for 24 hours to obtain a composition

Example 7:

Comparison of activity of the composition for environmental clarification with other methods:

[0061] Table 1 describes the comparison disclosed in the present disclosure with other methods such as ozone, hypochlorous acid, chrorine dioxide. It was observed from Table 1 that the composition disclosed in the present disclosure have higher anti-bacterial activity than other known methods.

Table 1 : Comparison of the activity of the composition with other methods.

Compositi 2.80V Very Good Good Safe Long No on for

environm

ental

clarificati

on

Ozone 2.07V Good Good Hazardo Short Yes (Sterilizat us

ion and

Deodoriza

tion)

Hypochlo 1.49V Fair Fair Hazardo Short Yes rous Acid us

(Bleach &

Sterilizati

on)

Chrorine 1.36V Fair Good Rather Short Yes Dioxide Safe

(Antisepti

c)

Results based on the experiments carried out in table 2:

Table 2:

Example 8:

Preparation of Suspension Solution of the composition for environmental clarification: [0062] The suspending solution was prepared by homogenous mixing of composition for environmental clarification in water for 1-2 hours with constant stirring. The above mixture was degassed for 1 day under an inert atmosphere. The resulting mixture was kept under constant stirring for 2-3 minutes to obtain the suspending solution of the composition for environmental clarification in water.

Example 9:

Acute Oral Toxicity Test in Female Mice:

• Preparation of test solution:

[0063] The test sample was dissolved in water to make 1 w/v% test solution. 1 w/v% test solution was diluted with water for injection to make 100 mg/mL test dilution.

• Experimental animals:

[0064] Female mice, at an age of 5 weeks were tested. Before test, they are acclimated to laboratory conditions for about 1 week to verify that no abnormalities are shown in general conditions. They were hosted in plastic cages (five animals per cage) under standard laboratory conditions (temperature 23 °C ± 2 °C, light dark cycle: 12/12 hours). Feed (Labo MR Stock diet, Nihon Nosankogyo K.K) and tap water were provided ad libitum throughout the experiment.

• Procedure:

[0065] Female mice are allocated into experimental and control groups each consisting of five mice. The mice are starved for about 4 hours before administration. After measurement of body weight, the animals in the experimental group are administered orally the test solution in a single dose using a stomach tube. The animals in the control group are administered solvent as vehicle control in the same manner. The administered volume did not exceeded 20 mL/kg of body weight (b.w). The clinical observation is carried out frequently on the day of the administration and once a day for the following 13 days. The body weight is measured after 7 and 14 days of the administration, and the mean body weight values of the experimental and control groups are statistically analyzed by t-test (a = 0.05). At the completion of the test, all of the mice are sacrificed for necropsy. • Results:

[0066] (1) Death of animals: None of the mice died during the experimental period.

[0067] (2) Clinical observations: No abnormalities were observed in any of the mice during the experimental period.

[0068] (3) Body weight changes: The body weight changes were shown in Table 3. From the Table 3, it was observed that no significant differences in body weight were detected between the experimental and control groups.

Table 3

The values are mean ± SD.

The values in parentheses represent the number of animals.

[0069] (4) Necropsy: No remarkable changes were found in any of the mice.

• Conclusion:

[0070] The acute oral toxicity in female mice of 1 w/v% test solution was determined. Oral administration of 2000 mg/kg b.w. of the test sample caused neither anbormalities nor death in any of the mice during the observation period. Consequently, the LD50 value (single dose, oral administration) of 1 w/v% test solution is considered to be more than 2000 mg/kg b.w. in female mice.

Example 10:

Eye Irritation Test in Rabbits:

• Experimental animals

[0071] Three male Japanese white rabbits were tested. They are acclimated to laboratory conditions for more than 1 week to confirm that there are no abnormalities in general conditions. Then, they were individually housed in fiber reinforced plastic (FRP) cages under standard laboratory conditions (Temperature; 22 °C ± 2 °C, Light-dark cycle: 12/12 hours). Tap water was provided ad libitum, and a certain amount of LRC4 diet (Oriental Yeast Co., Ltd.) was provided in proportion to each animal's body weight.

• Procedure:

[0072] Both the eyes of each animal are examined on the first day of the test to ensure the absence of any ocular abnormalities. After the animals are weighed, 0.1 g / 0.1 mL of the test sample is instilled into the conjunctival sac of one eye of each animal. The lids are then gently held together for about 1 second. Similarly, water for injection, as vehicle control, was instilled into another eye. Then, the cornea, iris, and conjunctivae are observed clinically using a hand slit-lamp at 1, 24, 48 and 72 hours after the instillation. The ocular reactions are observed and scored according to the criteria of the Draize method, shown in Table 4. The corneal epithelium is examined further in detail with the use of fluorescein sodium at each observation time except for 1 hour after the instillation. Other eyes, which are not treated with the test sample, are used as controls. The total score was calculated for each animal according to the formula shown in Table 5, and the mean total score is obtained at each observation time. Eye irritation potential of 1 w/v% test sample was graded by means of the maximum mean total score on the basis of Table 6.

• Results:

[0073] After 1 hour of instillation, the test eyes of one animal (Rabbit No. 3) showed redness of palpebral and bulbar conjunctiva (both score 1). These responses, however, disappeared by the 24-hour observation. No abnormalities were found during the subsequent period. In the remaining animals, no abnormalities were found throughout the observation period.

[0074] In contrast, the control eyes of all of the animals showed no abnormalities throughout the observation period.

[0075] In addition, fluorescein staining was not seen in the test and control eyes of all of the animals.

[0076] The maximum mean total scores were 0.7 (at 1 hour after instillation) in the test eyes and 0 in the control eyes. • Conclusion:

[0077] The test sample was tested for eye irritation potential in rabbits in accordance with OECD Guidelines for the Testing of Chemicals 405 (201 2).

[0078] A 0.1 -mL portion of 1 w/v% test solution was applied to one eye of three rabbits, and water for injection, as vehicle control, was similarly applied to another eye. As a result, in the test eyes, redness of palpebral and bulbar conjunctiva were seen in one animal at I hour after instillation. These responses, however, disappeared by the 24-hour observation. In the control eyes, no eye irritation was observed in any of the rabbits throughout the observation period. The maximum mean total scores, calculated by Draize method, were 0.7 (at 1 hour after instillation) in the test eyes and 0 in the control eyes. Consequently, 1 w/v% test solution is classified as a non-irritant to rabbit eyes.

Table 4 (1): Grades for ocular lesions

Cornea

(A) Opacity-degree of density (area most dense taken for reading)

No opacity · · · ^■ · 0

Scattered or diffuse area,

details of iris clearly visible 1

Easily discernible translucent areas,

details of iris slightly obscured 2

Opalescent areas, no details of iris visible,

size of pupil barely discernible 3

Opaque, iris invisible · · 4

(B) Area of cornea involved

One quarter (or less) but not zero ^■ · ^■ · 1

Greater than one quarter, but less than half · 2

Greater than half, but less than three quarters-- 3

Greater than three quarters, up to whole area 4

[Score = A x B 5 Maximum possible—— 80] Iris

(A) Normal ^■ ···· ··· ^■ · 0

Folds above normal, congestion, swelling,

circumcorneal injection (any or all of these

or combination of any thereof) iris still reacting

to light (sluggish reaction is positive) 1

No reaction to light, hemorrhage, gross destruction

(any or all of these) · ^■ 2

[Score = A x 5 Maximum possible 10]

Table 4 (1): Grades for ocular lesions (continued)

Conjunctivae

(A) Redness (refers to palpebral and bulbar conjunctivae excluding cornea and iris)

Vessels normal · 0

Vessels definitely injected above normal 1

^• More diffuse, deeper crimson red, individual vessels not

easily discernible · 2

Diffuse beefy red 3

(B) Chemosis

No swelling · 0

Any swelling above normal (includes nictitating membrane) 1

Obvious swelling with partial eversion of lids 2

Swelling with lids about half closed 3

Swelling with lids about half closed to completely closed 4

(C) Discharge

No discharge · · 0

Any amount different from normal (does not include small

amounts observed in inner canthus of normal animals) 1

Discharge with moistening of the lids and hairs

just adjacent to lids 2 Discharge with moistening of the lids and hairs,

and considerable area around the eye

[Score = (A + B + C) x 2 Maximum po -20]

Table 5: Calculation method of the total score

A, B, and C show the scores of (A), (B), and (C) given in Table 5, respectively.

* Calculation at each observation time.

Table 6: Evaluation of eye irritation

Table 7: Body weight of the lest animals at the start of the test

Table 8: Mean total scores

Rabbit No. Total score 1 hour 24 hours 48 hours 72 hours

1 0(0) 0(0) 0(0) 0(0)

2 0(0) 0(0) 0(0) 0(0)

3 2 (0) 0(0) 0(0) 0(0)

Mean total score 0.7(0) 0 (0) 0(0) . 0(0)

The values in parentheses show the scores of the control eyes.

Table 9: Scores of irritant response in the rabbit No. I

The values in parentheses show the scores of the control eye.

— : Not recorded

Table 10: Scores of irritant response in the rabbit No. 2

(2) Iris (A) 0(0) 0(0) 0(0) 0(0)

Redness (A) 0(0) 0 (0) 0(0) 0(0)

(3) Conjunctivae Chemosis (B) 0(0) 0(0) 0(0) 0(0)

Discharge (C) 0(0) 0(0) 0(0) 0(0)

Score (l) = A x B x 5 0(0) 0(0) 0(0) 0(0)

Score (2) = A x 5 0(0) 0(0) 0(0) 0(0)

Score (3) = (A + B + C) * 2 0(0) 0(0) 0(0) 0(0)

Total score [(1) + (2) + (3)] 0(0) 0(0) 0(0) 0(0)

The values in parentheses show the scores of the control eye.

— : Not recorded

Table 1 1 : Scores of irritant response in the rabbit No. 3

The values in parentheses show the scores of the control eye.

— : Not recorded

Example 11:

Primary Skin Irritation Test in Rabbits:

• Preparation of test solution [0079] The test sample is dissolved / suspended / diluted in water for injection to make 1 w/v% test solution.

• Experimental animals

[0080] Three, male Japanese white rabbits were tested. They were acclimated to laboratory conditions for more than 1 week to confirm that there are no abnormalities in general conditions. Then, they were individually housed in FRP cages under standard laboratory conditions (Temperature: 22 °C ± 2 °C, Light-dark cycle: 12/12 hours). Tap water was provided ad libitum, and a certain amount of LRC4 diet (Oriental Yeast Co., Ltd.) was provided in proportion to each animal's body weight.

• Procedure:

[0081] The back of each animal is completely shaved about 24 hours before the test, and the shaved area was divided into six sections (about 6 cm ² ). Two of the test sections were abraded. Abrasion was made by epidermal incision through the stratum corneum; two incisions perpendicular to the other two were made in a manner resembling a "#" pattern by use of a sterile 18-gage needle. The other two sections are left intact. A 0.5- mL portion of 1 w/v% test solutions was applied to each gauze patch (about 2 cm ^χ 3 cm) and held in one intact and one abraded sections on each animal with MULTI FIX-ROLL. To ensure the contact of the test solutions with skin, the patches were covered with Blenderm surgical tape. The other intact and abraded sections served as controls. After 4 hours of exposure, all the test sections are wiped with water for injection to remove the test solutions. Then, the skin reactions are read at 1 , 24, 48 and 72 hours after removal of 1 w/v% test solution according to the criteria shown in Table 12. The P.I.I, was calculated according to ISO 10993-10 Biological Evaluation of Medical Devices-Part 10 (2010). The scores of the readings at 24, 48 and 72 hours were added together and divided by 6. The mean value of three animals is regarded as P.I.I. The degree of the primary skin irritation was evaluated in accordance with the criteria shown in Table 13. The animals are weighed at the start and completion of the test.

• Results: [0082] At 1 hour after removal, very slight erythema (score 1) was found on the intact and abraded 'skin of all of the animals. But the erythema in one animal (Rabbit No. 3) disappeared at the 24-hour observation, and the erythema in the remaining two animals (Rabbits No. 1 and 2) disappeared by the 48-hour observation. No abnormalities were found during the subsequent period. At the non-treatment intact and abraded sections, no irritation response was observed throughout the observation period. The P.1.1., calculated usinu the scores of the readings, was 0.2.

• Conclusion

[0083] The test sample was tested for primary skin irritation in rabbits in accordance with OECD Guidelines for the Testing of Chemicals 404 (2002). 1 w/v% test solution was applied occlusively for 4 hours to the intact and abraded skin. As a result, very slight erythema was found in all of the animals at 1 hour after removal, but all of the erythema disappeared within 48 hours. The primary irritation index (P.I.I.) calculated according to ISO 10993-10 Biological Evaluation of Medical Devices-Part 10 (2010) was 0.2. Consequently, the primary skin irritation of 1 w/v% test solution is considered to be negligible.

Table 12: Evaluation of skin reactions

Erythema and eschar formation

No erythema : 0

Very slight erythema (barely perceptible) · 1

Well-defined erythema 2

Moderate to severe erythema 3

Severe erythema (beet redness) to slight eschar formation

(injuries in depth) · 4*

[Total possible erythema score 4]

* Necrosis, ulcer, alopecia, scars, etc. are classified into injuries in depth, score 4.

Edema formation

No edema · 0

Very slight edema (barely perceptible) 1 Slight edema

(edges of area well-defined by definite raising) · 2

Moderate edema (raised approximately 1 mm) · 3

Severe edema (raised more than 1 mm

and extending beyond area of exposure) · 4

[Total possible edema score 4]

Irritation response categories in rabbit

Table 14: Body- weight changes (kg)

Table 15: Scores of the irritation response

Example 12:

Virus lnactivation test • Outline of methods

[0084] The sample solution was mixed with Poliovirus suspension and stored at room temperature. After 30 minutes and also after 6 hours, virus infectivity titers were determined. The method of infectivity liters was validated by a preliminary test.

• , Results:

(1) Preliminary test

[0085] The results of the preliminary test indicated that the effects of the sample were removed by diluting the mixture of the sample solution and the virus suspension to 100- fold with a cell support medium.

(2) Virus infectivity assay

[0086] Table 16 shows the results.

Table 16: Virus infectivity of the mixture

Initial: TCID ₅₀ of the control at inoculation

Control: Purified water

Storage temperature: room temperature

* Logarithm of TCID ₅ o per 1 ml, of the mixture

• Method in detail:

[0087] (1) Test virus: Poliovirus 3

10088] (2) Test cell: HEp-2 cells: HEp-2 ATCC CCL-23 (Dainippon Pharmaceutical Co. Ltd.)

[0089] (3) Culture media

a) Cell culture medium: Eagle's MEM "Nissui"(l) (Nissui Pharmaceutical Co., Ltd.) including 10 % of fetal bovine serum

b) Cell support medium: Eagle's MEM "Nissui"(l ) including 2 % of fetal bovine serum.

[0090] (4) Preparation of virus suspension

a) Cell incubation: The test cell was monolayer-cultured in a tissue culture flask using the cell culture medium.

b) Virus inoculation: After monolayer culture, the culture medium was removed from the flask, and the test virus was inoculated. Next, the cell support medium was added, and the virus was cultured in a C0 ₂ incubator (C0 ₂ : 5%) at 37 °C ± 1 °C for 1 to 5 days.

c) Preparation of virus suspension: The cell form was observed with an inverted phase-contrast microscope. After morphological changes (cytopathic effects) were confirmed, the culture solution was centrifuged (3000 r/min, 10 minutes). The supernatant was used as a virus suspension.

[0091] 5) Test procedure: The sample was dissolved in purified water to make 0.3 % solution. A 0.1 -mL portion of the virus suspension was inoculated into 1 ml, of 0.3 % sample solution and stored at room temperature. After 30 minutes and also after 6 hours, it was diluted to 100-fold with the cell support medium, and virus infectivity was determined. As a control, purified water was prepared in the same manner as the sample. Virus infectivity was determined at the initial point, after 30 minutes, and after 6 hours.

[0092] (6) Virus infectivity assay: The test cell was monolayer-cultured in a tissue culture microplate (96-well) using the cell culture medium. After removal of the culture medium, 0.1 mL of the cell support medium was added. Next, 100-fold dilution of the mixture of the sample and the virus suspension, and the control were diluted with the cell support medium to obtain serial 10-fold dilutions. Then, 0.1 mL of each dilution was poured into four wells each and cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 4 to 7 days. After culture, the cells were observed by inverted phase-contrast microscope to observe if any morphological changes (cytopathic effects) had occurred. Then, median tissue culture infectious dose (TCID ₅₀ ) was calculated by Reed-Muench method and converted into virus infectivity liter per 1 mL of the mixture.

Example 13: Virus Inactivation test

• Outline of methods

[0093] The sample solution was mixed with Poliovinis suspension and stored at room temperature. After 30 minutes and also after 6 hours, virus infectivity titers were determined. The method of infectivity liters was validated by a preliminary test.

• Results:

(1 ) Preliminary test

[0094] The results of the preliminary test indicated that the effects of the sample were removed by diluting the mixture of the sample solution and the virus suspension to 100- fold with a cell support medium.

(2) Virus infectivity assay

[0095] Table 17 shows the results.

Table 17: Virus infectivity of the mixture

Initial: TCID50 of the control at inoculation

Control: Purified water

Storage temperature: room temperature

* Logarithm of TCIDso per 1 ml, of the mixture

• Method in detail:

[0096] (1 ) Test virus: Poliovirus 3

[0097] (2) Test cell: HEp-2 cells: HEp-2 ATCC CCL-23 (Dainippon Pharmaceutical Co. Ltd.)

[0098] (3) Culture media

a) Cell culture medium: Eagle's MEM "Nissui"(l) (Nissui Pharmaceutical Co., Ltd.) including 10 % of fetal bovine serum b) Cell support medium: Eagle's MEM "Nissui"(l) including 2 % of fetal bovine serum.

[0099] (4) Preparation of virus suspension

a) Cell incubation: The test cell was monolayer-cultured in a tissue culture flask using the cell culture medium.

b) Virus inoculation: After monolayer culture, the culture medium was removed from the flask, and the test virus was inoculated. Next, the cell support medium was added, and the virus was cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 1 to 5 days.

c) Preparation of virus suspension: The cell form was observed with an inverted phase-contrast microscope. After morphological changes (cytopathic effects) were confirmed, the culture solution was centrifuged (3000 r/min, 10 minutes). The supernatant was used as a virus suspension.

[00100] 5) Test procedure: The sample was dissolved in purified water to make 0.1 % solution. A 0.1 -mL portion of the virus suspension was inoculated into 1 ml, of 0.1 % sample solution and stored at room temperature. After 30 minutes and also after 6 hours, it was diluted to 100-fold with the cell support medium, and virus infectivity was determined. As a control, purified water was prepared in the same manner as the sample. Virus infectivity was determined at the initial point, after 30 minutes, and after 6 hours.

[00101] (6) Virus infectivity assay: The test cell was monolayer-cultured in a tissue culture microplate (96-well) using the cell culture medium. After removal of the culture medium, 0.1 mL of the cell support medium was added. Next, 100-fold dilution of the mixture of the sample and the virus suspension, and the control were diluted with the cell support medium to obtain serial 10-fold dilutions. Then, 0.1 mL of each dilution was poured into four wells each and cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 4 to 7 days. After culture, the cells were observed by inverted phase-contrast microscope to observe if any morphological changes (cytopathic effects) had occurred. Then, median tissue culture infectious dose (TCID ₅₀ ) was calculated by Reed-Muench method and converted into virus infectivity liter per 1 mL of the mixture. Example 14:

Virus Inactivation test

• Outline of methods

[00102] The sample solution was mixed with Feline calicivirus suspension and stored at room temperature. After 30 minutes and also after 6 hours, virus infectivity titers were determined. The method of infectivity liters was validated by a preliminary test. Feline calicivirus is a commonly used as a surrogate for Norovirus, which can not been cultured routinely.

• Results:

(1) Preliminary test

[00103] The results of the preliminary test indicated that the effects of the sample were removed by diluting the mixture of the sample solution and the virus suspension to 10000-fold with a cell support medium.

(2) Virus infectivity assay

{00104] Table 18 shows the results.

Table 18: Virus infectivity of the mixture

Initial: TCID ₅₀ of the control at inoculation

Control: Purified water

Storage temperature: room temperature

* 1 Logarithm of TCID ₅₀ per 1 ml, of the mixture

*2 Surrogate virus for Norovirus

• Method in detail:

[00105] (1) Test virus: Feline calicivims F-9 ATCC VR-782

[00106] (2) Test cell: CRFK cells (Dainippon Pharmaceutical Co., Ltd.) [00107] (3) Culture media

a) Cell culture medium: Eagle's MEM "Nissui"(l) (Nissui Pharmaceutical Co., Ltd.) including 10 % of fetal bovine serum

b) Cell support medium: Eagle's MEM "Nissui "(l) added 2 % of fetal bovine serum

[00108] (4) Preparation of virus suspension

a) Cell incubation: The test cell was monolayer-cultured in a tissue culture flask using the cell culture medium.

b) Virus inoculation: After monolayer culture, the culture medium was removed from the flask, and the test virus was inoculated. Next, the cell support medium was added, and the virus was cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 1 to 5 days.

c) Preparation of virus suspension: The cell form was observed w i th an inverted phase-contrast microscope. After morphological changes (cytopathic effects) were confirmed, the culture solution was centrifuged (3000 r/min, 10 minutes). The supernatant was used as a virus suspension.

[00109] (5) Test procedure: The sample was dissolved in purified water to make 3 % solution. A 0.1 -mL portion of the virus suspension was inoculated into 1 mL of 3 % sample solution and stored at room temperature. After 30 minutes and also after 6 hours, it was diluted to 10000-fold with the cell support medium, and virus infectivity was determined. As a control, purified water was prepared in the same manner as the sample. Virus infectivity was determined at the initial point, after 30 minutes, and after 6 hours.

[00110] (6) Virus infectivity assay: The test cell was monolayer-cultured in a tissue culture microplate (96-well) using the cell culture medium. After the culture medium was removed, 0.1 mL of the cell support medium was added. Next, 10000-fold dilution of the mixture of the sample and the virus suspension, and the control were diluted with the cell support medium to obtain serial 10-fold dilutions. Then 0.1 mL of each dilution was poured into four wells each and cultured in a CO; incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 4 to 7 days. After culture, the cells were observed by inverted phase- contrast microscope to observe if any morphological changes (cylopathic effects) had occurred. Then, median tissue culture infectious dose (TCID ₅₀ ) was calculated by Reed- Muench method and converted into virus infectivity liter per 1 mL of the mixture.

Example 15:

Virus Inactivation test

• Outline of methods

[00111] The sample solution was mixed with Feline calicivirus suspension and stored at room temperature. After 30 minutes and also after 6 hours, virus infectivity titers were determined. The method of infectivity titers was validated by a preliminary test. Feline calicivirus is a commonly used as a surrogate for Norovirus, which can not been cultured routinely.

• Results:

( 1 ) Preliminary test

[00112] The results of the preliminary test indicated that the effects of the sample were removed by diluting the mixture of the sample solution and the virus suspension to 10000-fold with a cell support medium.

(2) Virus infectivity assay

[00113] Table 19 shows the results.

Table 19: Virus infectivity of the mixture

Initial: TCID ₅ o of the control at inoculation

Control: Purified water

Storage temperature: room temperature

* 1 Logarithm of TCID ₅₀ per 1 ml, of the mixture

*2 Surrogate virus for Norovirus • Method in detail:

[00114] (1) Test virus: Feline calicivims F-9 ATCC VR-782

[00115] (2) Test cell: CRFK cells (Dainippon Pharmaceutical Co., Ltd.)

[00116] (3) Culture media

a) Cell culture medium: Eagle's MEM "Nissui "(l) (Nissui Pharmaceutical Co., Ltd.) including 10 % of fetal bovine serum

b) Cell support medium: Eagle's MEM "Nissui "(1) added 2 % of fetal bovine serum

[00117] (4) Preparation of virus suspension

a) Cell incubation: The test cell was monolayer-cultured in a tissue culture flask using the cell culture medium.

b) Virus inoculation: After monolayer culture, the culture medium was removed from the flask, and the test virus was inoculated. Next, the cell support medium was added, and the virus was cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 1 to 5 days.

c) Preparation of virus suspension: The cell form was observed with an inverted phase-contrast microscope. After morphological changes (cytopathic effects) were confirmed, the culture solution was centrifuged (3000 r/min, 10 minutes). The supernatant was used as a virus suspension.

[00118] (5) Test procedure: The sample was dissolved in purified water to make 1 % solution. A 0.1 -mL portion of the virus suspension was inoculated into 1 mL of 1 % sample solution and stored at room temperature. After 30 minutes and also after 6 hours, it was diluted to 10000- fold with the cell support medium, and virus infectivity was determined. As a control, purified water was prepared in the same manner as the sample. Virus infectivity was determined at the initial point, after 30 minutes, and after 6 hours.

[00119] (6) Virus infectivity assay: The test cell was monolayer-cultured in a tissue culture microplate (96-well) using the cell culture medium. After the culture medium was removed, 0.1 mL of the cell support medium was added. Next, 10000-fold diluti on of the mixture of the sample and the virus suspension, and the control were diluted with the cell support medium to obtain serial 10-fold dilutions. Then 0.1 mL of each dilution was poured into four wells each and cultured in a CO; incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 4 to 7 days. After culture, the cells were observed by inverted phase-contrast microscope to observe if any morphological changes (cylopathic effects) had occurred. Then, median tissue culture infectious dose (TCID ₅₀ ) was calculated by Reed-Muench method and converted into virus infectivity liter per 1 mL of the mixture.

Example 16:

Virus Inactivation test

• Outline of methods

[00120] The sample solution was mixed with Influenza virus suspension and stored at room temperature. After 30 minutes and also after 6 hours, virus infectivity titers were determined. The method of infectivity titers was validated by a preliminary test.

• Results:

( 1 ) Preliminary test

[00121] The results of the preliminary test indicated that the effects of the sample were removed by diluting the mixture of the sample solution and the virus suspension to 10000-fold with a cell support medium.

(2) Virus infectivity assay

[00122] Table 20 shows the results.

Table 20: Virus infectivity of the mixture

TCID50: Median tissue culture infectious dose

initial: TCIDso of the control at inoculation

Control: Purified water

Storage temperature: room temperature

<4.5: Not detected

* Logarithm of TCID ₅ o per 1 ml, of the mixture • Method in detail:

[00123] (1) Test virus: Influenza A virus (H1N1)

[00124] (2) Test cell: MDCK (NBL-2) cells ATCC CCL-34 strain (Dainippon Pharmaceutical Co., Ltd.)

[00125] (3) Culture media

a) Cell culture medium: . Eagle's MEM "Nissui " (l) (Nissui Pharmaceutical Co., Ltd.) including 10 % of fetal bovine serum

b) Cell support medium: Eagle's MEM "Nissui " (l) l OOOmL

1 0% NaHCO ₃ 1 4mL

L-glutamine (30g/L) 9.8mL l OOxvitamins for MEM 30mL

1 0% albumin 20mL

0.25 % trypsin 20mL

(4) Preparation of virus suspension

a) Cell incubation: The test cell was monolayer-cultured in a tissue culture flask using the cell culture medium.

b) Virus inoculation: After monolayer culture, the culture medium was removed from the flask, and the test virus was inoculated. Next, the cell support medium was added, and the virus was cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 1 to 5 days.

c) Preparation of virus suspension: The cell form was observed with an inverted phase-contrast microscope. After morphological changes (cytopathic effects) were confirmed, the culture solution was centrifuged (3000 r/min, 10 minutes). The supernatant was used as a virus suspension.

[00126] (5) Test procedure: The sample was dissolved in purified water to make 3 % solution. A 0.1 -mL portion of the virus suspension was inoculated into 1 mL of 3 % sample solution and stored at room temperature. After 30 minutes and also after 6 hours, it was diluted to 10000- fold with the cell support medium, and virus infectivity was determined. As a control, purified water was prepared in the same manner as the sample. Virus infectivity was determined at the initial point, after 30 minutes, and after 6 hours.

[00127] (6) Virus infectivity assay: The test cell was monolayer-cultured in a tissue culture microplate (96-well) using the cell culture medium. After the culture medium was removed, 0.1 mL of the cell support medium was added. Next, 10000-fold dilution of the mixture of the sample and the virus suspension, and the control were diluted with the cell support medium to obtain serial 10-fold dilutions. Then 0.1 mL of each di lution was poured into four wells each and cultured in a CO; incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 4 to 7 days. After culture, the cells were observed by inverted phase-contrast microscope to observe if any morphological changes (cylopathic effects) had occurred. Then, median tissue culture infectious dose (TCID5 ₀ ) was calculated by Reed-Muench method and converted into virus infectivity liter per 1 mL of the mixture.

Example 17:

Virus Inactivation test

• Outline of methods

[00128] The sample solution was mixed with Influenza virus suspension and stored at room temperature. After 30 minutes and also after 6 hours, virus infectivity titers were determined. The method of infectivity titers was validated by a preliminary test.

• Results:

( 1 ) Preliminary test

[00129] The results of the preliminary test indicated that the effects of the sample were removed by diluting the mixture of the sample solution and the virus suspension to 10000-fold with a cell support medium.

(2) Virus infectivity assay

[00130] Table 21 shows the results.

Table 21 : Virus infectivity of the mixture

Influenza Sample 1 % solution 7.5 7.5 <3.5 virus Control - 7.5 7.3 7.5

TCID ₅₀ : IV edian tissue culture infectious dose

Initial: TCID ₅₀ of the control at inoculation

Control: Purified water

Storage temperature: room temperature

<3.5: Not detected

* Logarithm of TCID ₅₀ per 1 ml, of the mixture

• Method in detail:

[00131 J (1) Test virus: Influenza A virus (H1N1)

[00132] (2) Test cell: MDCK (NBL-2) cells ATCC CCL-34 strain (Dainippon Pharmaceutical Co., Ltd.)

[00133J (3) Culture media

a) Cell culture medium: Eagle's MEM "Nissui "(l) (Nissui Pharmaceutical Co., Ltd.) including 10 % of fetal bovine serum

b) Cell support medium: Eagle's MEM "Nissui " (l) l OOOmL

1 0% .NaHCO ₃ 14mL

L-glutamine (30g/L) 9.8mL 1 00 x vitamins for MEM 30mL

10% albumin 20mL

0.25 % trypsin 20mL

(4) Preparation of virus suspension

a) Cell incubation: The test cell was monolayer-cultured in a tissue culture flask using the cell culture medium.

b) Virus inoculation: After monolayer culture, the culture medium was removed from the flask, and the test virus was inoculated. Next, the cell support medium was added, and the virus was cultured in a C0 ₂ incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 1 to 5 days.

c) Preparation of virus suspension: The cell form was observed with an inverted phase-contrast microscope. After morphological changes (cytopathic effects) were confirmed, the culture solution was centrifuged (3000 r/min, 10 minutes). The supernatant was used as a virus suspension.

[00134] (5) Test procedure: The sample was dissolved in purified water to make 1 % solution. A 0.1 -mL portion of the virus suspension was inoculated into 1 mL of 1 % sample solution and stored at room temperature. After 30 minutes and also after 6 hours, it was diluted to 10000-fold with the cell support medium, and virus infectivity was determined. As a control, purified water was prepared in the same manner as the sample. Virus infectivity was determined at the initial point, after 30 minutes, and after 6 hours.

[00135] (6) Virus infectivity assay: The test cell was monolayer-cultured in a tissue culture microplate (96-well) using the cell culture medium. After the culture medium was removed, 0.1 mL of the cell support medium was added. Next, 10000-fold dilution of the mixture of the sample and the virus suspension, and the control were diluted with the cell support medium to obtain serial 10-fold dilutions. Then 0.1 mL of each dilution was poured into four wells each and cultured in a CO; incubator (C0 ₂ : 5 %) at 37 °C ± 1 °C for 4 to 7 days. After culture, the cells were observed by inverted phase-contrast microscope to observe if any morphological changes (cylopathic effects) had occurred. Then, median tissue culture infectious dose (TCID ₅ 0) was calculated by Reed-Muench method and converted into virus infectivity liter per 1 mL of the mixture.

Example 18:

Antibacterial Activity test

• Outline of methods

[00136] Suspensions of Escherichia coli, Escherichia coli (Serovar 0157:H7, Shiga toxin I & Il-producing Escherichia coli), Listeria monocytogenes, Salmonella enterica, and Methicillin-resistant Staphylococcus aureus (MRSA) were separately inoculated to the sample (hereafter called "the test solutions") and stored at room temperature. After 30 minutes and also after 6 hours, the viable cell count of each test solution was determined. The test method for determining viable cell count was validated by a preliminary test.

• Results:

[00137] Table 22 shows the results. [00138] The results of the preliminary test revealed that the test solution should be diluted by 10-fold [for Escherichia coli (Serovar 0157:H7)], by 100-fold (for Escherichia coli) and by 1000-fold (for Listeria monocytogenes, Salmonella enterica, and MRSA) with SCDLP broth to remove the effects of the sample.

Table 22: Viable counts of the lest solutions

<10, <100 and <1000: Not detected

Control: Purified water and physiological saline (only for MRSA)

Storage temperature: room temperature

* The viable count of the control immediately after inoculation was defined as the viable count at the initial point.

• Method in detail:

[00139] (1) Test strains: (a) Escherichia coli NBRC 3972, (b) Escherichia coli ATCC 43895 (Serovar 0157:H7, Shiga toxin I & 11-producing Escherichia coli), (c) Listeria monocytogenes VTU 206, (d) Salmonella enterica subsp. enterica NBRC 3313, (e) Staphylococcus aureus IID 1 77 (MRSA)

[00140] (2) Medium for determination of viable count and incubation conditions: SCDLP Agar (Nihon Pharmaceutical Co., Ltd.). Pour plate method 35 °C ± 1 °C, 2 days. [00141] (3) Preparation of cell suspensions: The test organism was grown on Nutrient Agar (Liken Chemical Co., Ltd.) for 18 to 24 hours al 35 °C ± 1 °C. After incubation, the culture was suspended in purified water [for the test strains (a) to (d)] and in physiological saline [for the test strain (e)[ to contain about 10 ⁷ to 10 ⁸ cells/mL. The obtained suspensions were used as cell suspensions.

[00142] (4) Test procedure: A 0.1 -mL portion of the cell suspension was inoculated to 10 mL of 3 % sample solution (prepared with purified water), and this solution was called the test solution. Then, the test solution was stored at room temperature. After 30 minutes, it was immediately diluted by 10-fold [for the test strains (b)], by 100-fold [for the test strain (a)] and by 1000-fold [for the test strains (c) to (e)] with SCDLP broth (Nihon Pharmaceutical Co. Ltd.). After 6 hours, it was diluted in a similar way. Then, the viable cell counts of the test solutions were measured at each time. As controls, purified water and physiological saline [only for the test strain (e)] were prepared in the same manner. The viable cell counts of the controls were determined at the initial point, after 30 minutes and also after 6 hours.

Example 19:

Antibacterial Activity test

• Outline of methods

[00143] Suspensions of Escherichia coli, Escherichia coli (Serovar 0157.Ή7, Shiga toxin I & Il-producing Escherichia coli), Listeria monocytogenes, Salmonella enterica, and Methicillin-resistant Staphylococcus aureus (MRSA) were separately inoculated to the sample (hereafter called "the test solutions") and stored at room temperature. After 30 minutes and also after 6 hours, the viable cell count of each test solution was determined. The test method for determining viable cell count was validated by a preliminary test.

• Results:

[00144] Table 23 shows the results.

[00145] The results of the preliminary test revealed that the test solution should be diluted by 10-fold [for Escherichia coli (Serovar 0157:H7) and Listeria monocytogenes], and by 100-fold (for Escherichia coli, Salmonella enterica, and MRSA,) with SCDLP broth to remove the effects of the sample.

Table 23: Viable counts of the lest solutions

<10, and <100: Not detected

Control: Purified water and physiological saline (only for MRSA)

Storage temperature: room temperature

* The viable count of the control immediately after inoculation was defined as the viable count at the initial point.

• Method in detail:

[00146] (1) Test strains: (a) Escherichia coli NBRC 3972, (b) Escherichia coli ATCC 43895 (Serovar 0157:H7, Shiga toxin I & Il-producing Escherichia coli), (c) Listeria monocytogenes VTU 206, (d) Salmonella enterica subsp. enterica NBRC 3313, (e) Staphylococcus aureus IID 1677 (MRSA)

[00147] (2) Medium for determination of viable count and incubation conditions: SCDLP Agar (Nihon Pharmaceutical Co., Ltd.). Pour plate method 35 °C ± 1°C, 2 days.

[00148] (3) Preparation of cell suspensions: The test organism was grown on Nutrient Agar (Liken Chemical Co., Ltd.) for 18 to 24 hours al 35 °C ± 1 °C. After incubation, the culture was suspended in purified water [for the test strains (a) to (d)] and in physiological saline [for the test strain (e)] to contain about 10 ⁷ to 10 ⁸ cells/mL. The obtained suspensions were used as cell suspensions.

[00149] (4) Test procedure: A 0.1 -mL portion of the cell suspension was inoculated to 10 mL of 1 % sample solution (prepared with purified water), and this solution was called the test solution. Then, the test solution was stored at room temperature. After 30 minutes, it was immediately diluted by 10-fold [for the test strains (b) and (c)], and by 100-fold [for the test strain (a), (d) and (e)] with SCDLP broth (Nihon Pharmaceutical Co.. Ltd.). After 6 hours, it was diluted in a similar way. Then, the viable cell counts of the test solutions were measured at each time. As controls, purified water and physiological saline [only for the test strain (e)] were prepared in the same manner. The viable cell counts of the controls were determined at the initial point, after 30 minutes and also after 6 hours.

Example 20:

Gas removal effect test

• Outline of methods

[00150] A 1 % water solution of the sample was used as the test solution (1) and a 3 % water solution of the sample was used as the test solution (2). The gas removal effects of the test solutions and control (water alone) were evaluated using formaldehyde by gas detector tube method.

• Results:

[00151] Table 24 and figure 1 shows the test results.

Table 24 Test results (Testing gas: formaldehyde)

Unit: ppm

Initial gas concentration: about 20 ppm • Method in detail:

[00152] (1) Reagents and apparatus: Film bag (25 cm x 40 cm. ARAM Corporation), Formaldehyde: generated from formaldehyde (36 %, special grade. Kanio Chemical Co., Inc.) and Gas detector tube (Gastec Corporation).

[00153] (2) Preparation of test solutions: The sample weighing 0.5 g was dissolved and filled up to 50 niL with water to make I % water solution. The solution obtained was used as the test solution (1). Similarly 1.5 g of the sample was dissolved and filled up to 50 mL to make 3 % water solution. The solution obtained was used as the test solution

(2)·

[00154] (3) Procedures: The test solutions and control (water alone) were put in film bags. After the bags were heat-sealed, 3 L of air was injected into the bags. Next, the testing gas was added to a specified concentration, and the bags were allowed to stand at room temperature. Then, the gas concentrations in the bags were determined at each determination time. In addition, the testing gas alone (without the test solutions or water) was prepared in the same manner as a blank test.

[00155] Table 25 shows the test conditions

Table 25: Test conditions

Example 21:

Odor removal effect test

• Outline of methods [00156] A 1 % water solution of the sample was used as the test solution (1), and a 3 % water solution of the sample was used as the test solution (2). The odor removal effects of the test solutions and control (water alone) were evaluated using acetaldehyde by gas detector tube method.

• Results:

[00157] Table 26 and figure 2 shows the test results.

Table 26: Test results (testing gas: acetaldehyde)

Unit: ppm

Initial gas concentration: about 20 ppm • Method in detail:

[00158] (1) Reagents and apparatus: Film bag (25 cm x 40 cm. ARAM Corporation), Acetaldehyde: generated from acetaldehyde (first grade. Wako Pure Chemical Industries, Ltd.) and Gas detector tube (Gastec Corporation).

[00159] (2) Preparation of test solutions: The sample weighing 0.5 g was dissolved and filled up to 50 mL with water to make 1 % water solution. The solution obtained was used as the test solution (1). Similarly, 1.5 g of the sample was dissolved and filled up to 50 mL to make 3 % water solution. The solution obtained was used as the test solution (2).

[00160] (3) Procedures: The test solutions and control (water alone) were put in film bags. After the bags were heat-sealed, 3 L of air was injected into the bags. Next, the testing gas was added to a specified concentration, and the bags were allowed to stand at room temperature. Then, the gas concentrations in the bags were determined at each determination time. In addition, the testing gas alone (without the test solutions or water) was prepared in the same manner as a blank test.

[00161] Table 27 shows the test conditions.

Table 27: Test conditions

Example 22:

Odor removal effect test

• Outline of methods

[00162] A 1 % water solution of the sample was used as the test solution (1), and a 3 % water solution of the sample was used as the test solution (2). The odor removal effects of the test solutions and control (water alone) were evaluated using ammonia by gas detector tube method.

• Results:

[00163] Table 28 and figure 3 show the test results.

Table 28: Test results (testing gas: ammonia)

Unit: ppm

Initial gas concentration: about 100 ppm • Method in detail: [00164] (1) Reagents and apparatus: Film bag (25 cm x 40 cm, ARAM Corporation), Ammonia: generated from ammonia water (28 %, special grade. Koso Chemical Co.. Ltd.) and Gas detector tube (Gastec Corporation).

[00165] (2) Preparation of test solutions: The sample weighing 0.5 g was dissolved and filled up to 50 mL with water to make 1 % water solution. The solution obtained was used as the test solution (1). Similarly, 1.5 g of the sample was dissolved and filled up to 50 mL to make 3 % water solution. The solution obtained was used as the test solution ^• (2)·

[00166] (3) Procedures: The test solutions and control (water alone) were put in film bags. After the bags were heat-sealed, 3 L of air was injected into the bags. Next, the testing gas was added to a specified concentration, and the bags were allowed to stand at room temperature. Then, the gas concentrations in the bags were determined at each determination time. In addition, the testing gas alone (without the test solutions or water) was prepared in the same manner as a blank test.

[00167] Table 29 shows the test conditions.

Table 29: Test conditions

Advantages:

[00168] The previously described versions of the subject matter and its equivalent thereof have many advantages, including those which are described below:

1. The present disclosure provides a composition for environmental clarification which has high dissolving capability.

2. The present disclosure enhances the improved safety and stability. 3. The present disclosure also provides a composition for environmental clarification with high absorption efficiency in a cost effective way.

4. The present disclosure further reacts even with slightly visible ray.

[00169] Although the subject matter has been described in considerable detail with reference to certain preferred embodiments thereof, other embodiments are possible. As such, the spirit and scope of the disclosure should not be limited to the description of the preferred embodiment contained therein.