Urine neutralizing solution composition for preserving cells in urine and urine neutralization treatment method using same

Abstract

The present invention relates to a urine neutralizing solution composition for preserving cells in urine and a urine neutralization treatment method using same. More specifically, the present invention relates to: a urine neutralizing solution composition comprising buffered saline, vitamin C (ascorbic acid), glucose, and optionally polyethylene glycol (PEG); and a urine neutralization treatment method comprising a step of adding the urine neutralizing solution composition to urine. When urine is preserved under refrigeration using the urine neutralizing solution composition according to the present invention, not only can colony forming ability and cell proliferation ability be maintained, but the cell preservation period can also be extended to 24 hours or longer, preferably 48 hours or longer, and most preferably 72 hours or longer.

Description

Urine neutralizing solution composition for preserving cells in urine and urine neutralizing treatment method using the same

[0001] The present invention relates to a urine neutralizing composition for preserving cells in urine and a urine neutralizing treatment method using the same. More specifically, the present invention relates to a urine neutralizing composition comprising buffered saline, vitamin C (ascorbic acid), glucose, and optionally polyethylene glycol (PEG), and a urine neutralizing treatment method comprising a step of adding the urine neutralizing composition to urine.

[0002] When urine is refrigerated and preserved using the urine neutralizing composition according to the present invention, not only can the cell colony formation ability and cell proliferation ability be maintained, but the cell preservation period can be extended to 24 hours or more, preferably 48 hours or more, and most preferably 72 hours or more.

[0003]

[0004] Urine is produced by blood filtration in the kidneys, a process that involves three stages: filtration, reabsorption, and secretion. During the filtration stage, dissolved substances in the blood are filtered through the glomeruli of the kidneys. Then, during the reabsorption stage, necessary molecules, ions, and water are reabsorbed into the blood. Finally, during the secretion stage, hydrogen ions, creatinine, drugs, and urea move from the blood to the collecting ducts, ultimately being excreted as urine.

[0005] Urine contains a variety of cells from the kidneys, bladder, and urinary tract, and recent research on these cells has led to diverse and innovative directions. Representative examples include analyzing urine cells to develop biomarkers for the detection, monitoring, and treatment of bladder cancer, and investigating the effects of traditional and novel treatments on bladder epithelial cells. Furthermore, research using urine-derived stem cells (UDSCs) is opening up new possibilities in various medical fields, including neurological diseases. Urinary stem cells can be easily collected non-invasively and possess stem cell properties, attracting attention in diverse applications such as disease modeling, drug screening, and tissue repair.

[0006] As the potential applications of urine cells continue to grow, the need for their isolation and culture is increasing. However, after urine is excreted from the body, the environment is not conducive to cell survival. Urine contains a high concentration of waste products and electrolytes, and under these conditions, cells struggle to survive for long periods. Due to the cytotoxicity of urine, cells in urine become damaged over time after collection. This leads to reduced cell lifespan, delayed growth, accelerated aging, and cell death, posing significant limitations in analyzing urine cells or culturing them in vitro.

[0007] Although there is little research evidence in the scientific literature to date on the direct cause of urine cytotoxicity, the following main reasons can be speculated as to why urine is toxic to cells. First, osmotic stress: Since urine components are generally highly concentrated, they can cause osmotic imbalances inside and outside the cells. Second, pH imbalance: Urine can be slightly acidic or slightly basic depending on physiological conditions such as health status, food intake, and medication intake, which can affect intracellular enzyme activity and metabolic processes. Third, cytotoxic components: Urea, uric acid, ammonia, and reactive oxygen species contained in urine can accelerate cell death by causing osmotic pressure and pH imbalances, cell membrane damage, cell cycle disruption, cellular DNA damage, and oxidative stress promotion.

[0008] While cell isolation from fresh urine has generally been considered the most efficient, this is only feasible when the urine donor has immediate access to a research facility or cell processing facility. Specifically, when using progenitor or stem cells derived from the kidney, bladder, or other sources isolated from a patient's urine as autologous cell therapy, storage or transportation from the hospital where the urine was collected to a cell processing facility is required. During this process, the cells in the urine can be damaged or killed within hours by the toxicity of the urine.

[0009]

[0010] Prior art

[0011] 1. Republic of Korea Publication Patent No. KR 10-2023-0165334 A

[0012]

[0013] The present inventors have made extensive efforts to develop a urine neutralizing solution capable of extending the preservation and transportation period of urine by preventing damage and death of cells present in urine over time, and as a result, they have confirmed that when urine is refrigerated and preserved using a composition containing buffered saline, vitamin C (ascorbic acid), and glucose, the influence of cytotoxic components can be minimized, and cell damage in urine can be prevented and the cell preservation period can be extended to 24 hours or more, preferably 48 hours or more, thereby completing the present invention.

[0014] In addition, the inventors of the present invention confirmed that when urine is refrigerated and preserved using a composition containing buffered saline, vitamin C (ascorbic acid), glucose, and polyethylene glycol (PEG), cell colony formation ability and cell proliferation ability can be maintained (see Example 6), and the cell preservation period can be extended to 72 hours (see Example 7).

[0015]

[0016] Accordingly, the present invention aims to provide a urine neutralizing composition comprising buffered saline, vitamin C (ascorbic acid), and glucose.

[0017] The present invention also aims to provide a urine neutralization treatment method using a urine neutralization composition containing buffered saline, vitamin C (ascorbic acid), and glucose.

[0018]

[0019] However, the problems that the present invention seeks to solve are not limited to the problems mentioned above, and other problems not mentioned will be clearly understood by those skilled in the art from the description below.

[0020]

[0021] The present invention provides a urine neutralizing composition comprising buffered saline, vitamin C (ascorbic acid), and glucose, and a urine neutralizing treatment method using the urine neutralizing composition comprising buffered saline, vitamin C (ascorbic acid), and glucose.

[0022]

[0023] According to the first implementation example,

[0024] A urine neutralizing composition comprising buffered saline, vitamin C (ascorbic acid), and glucose is disclosed.

[0025] In the present invention, the buffered saline solution can be added in an amount of 50 to 200% based on the volume of urine.

[0026] In the present invention, the vitamin C can be added at a concentration of 50 to 600 μg / mL.

[0027] In the present invention, the glucose may be added at a concentration of 5 to 60 mM.

[0028] In the present invention, the urine neutralizing composition may further include polyethylene glycol (PEG).

[0029] In the present invention, the polyethylene glycol may be added at a concentration of 10 to 60 mg / mL.

[0030] In the present invention, the urine neutralizing composition may be composed of buffered saline, vitamin C (ascorbic acid), glucose, and polyethylene glycol (PEG).

[0031] In the present invention, the urine neutralizing composition may be composed of 100% buffered saline, 600 μg / mL of vitamin C (ascorbic acid), 60 mM of glucose, and 60 mg / mL of polyethylene glycol (PEG) based on the volume of urine.

[0032]

[0033] According to the second implementation example,

[0034] A urine neutralization treatment method is disclosed, which comprises a step of adding a urine neutralizing composition according to the first embodiment to urine.

[0035] In the present invention, the method may further include a step of storing urine with the urine neutralizing composition added thereto at 2 to 8°C for 1 to 5 days.

[0036]

[0037] When urine is refrigerated and preserved using the urine neutralizing composition according to the present invention, not only can the cell colony formation ability and cell proliferation ability be maintained, but the cell preservation period can be extended to 24 hours or more, preferably 48 hours or more, and most preferably 72 hours or more.

[0038]

[0039] Figure 1 shows changes in cell viability in urine according to Example 1 of the present invention.

[0040] Figure 2 shows the change in cell viability in urine according to the added dose and concentration of buffered saline solution according to Example 2 of the present invention.

[0041] Figure 3 shows the change in cell viability in urine depending on the concentration of polyethylene glycol added according to Example 3 of the present invention.

[0042] Figure 4 shows the change in cell viability in urine depending on the concentration of vitamin C added according to Example 4 of the present invention.

[0043] Figure 5 shows the change in cell viability in urine according to the concentration of added glucose according to Example 5 of the present invention.

[0044] Figure 6 shows the cell preservation effect of the urine neutralizing solution composition according to Example 6 of the present invention.

[0045] Figure 7 shows the cell preservation effect of the urine neutralizing solution composition according to Example 7 of the present invention.

[0046]

[0047] Hereinafter, a urine neutralizing composition for preserving urine cells according to specific embodiments of the present invention and a urine neutralizing treatment method using the same will be described in detail. However, this is presented as one example of the invention, and the scope of the invention is not limited thereby, and it is obvious to those skilled in the art that various modifications to the embodiments are possible within the scope of the invention. Unless otherwise specified, throughout this specification, "include" or "containing" refers to including a certain component (or component) without any particular limitation, and cannot be interpreted as excluding the addition of other components (or components).

[0048]

[0049] 1. Urine neutralizing solution composition

[0050] The present invention provides a urine neutralizing composition comprising buffered saline, vitamin C (ascorbic acid), and glucose.

[0051] In the urine neutralizing composition according to the present invention, the buffered saline solution may be added at a concentration of 50 to 200%, preferably 100 to 200%, and most preferably 100%, based on the volume of urine.

[0052] In the urine neutralizing composition according to the present invention, the vitamin C may be added at a concentration of 50 to 600 μg / mL, preferably 400 to 600 μg / mL, and most preferably 600 μg / mL.

[0053] In the urine neutralizing composition according to the present invention, the glucose may be added at a concentration of 5 to 60 mM, preferably 40 to 60 mM, and most preferably 60 mM.

[0054] In the urine neutralizing composition according to the present invention, the urine neutralizing composition may further include polyethylene glycol (PEG).

[0055] In the urine neutralizing composition according to the present invention, the polyethylene glycol may be added at a concentration of 10 to 60 mg / mL, preferably 40 to 60 mg / mL, and most preferably 60 mg / mL.

[0056] In the urine neutralizing composition according to the present invention, the urine neutralizing composition may be composed of buffered saline, vitamin C (ascorbic acid), glucose, and polyethylene glycol (PEG).

[0057] In the urine neutralizing composition according to the present invention, the urine neutralizing composition may be composed of 100% buffered saline, 600 μg / mL of vitamin C (ascorbic acid), 60 mM of glucose, and 60 mg / mL of polyethylene glycol (PEG) based on the volume of urine.

[0058] In the urine neutralizing composition according to the present invention, the urine neutralizing composition may be added in an amount of 5 to 30%, preferably 10 to 20%, and most preferably 10% of the urine volume.

[0059] In the urine neutralizing composition according to the present invention, the urine may be derived from a mammal. For example, the mammal may include, but is not limited to, a human, a non-human primate, a mouse, a rat, a dog, a cat, a horse, or a cow.

[0060]

[0061] 2. Urine neutralization treatment method

[0062] The present invention provides a urine neutralization treatment method comprising a step of adding a urine neutralization composition containing buffered saline, vitamin C (ascorbic acid), and glucose to urine.

[0063] In the urine neutralization treatment method according to the present invention, the method may further include a step of storing urine to which a urine neutralizing composition has been added at 2 to 8°C for 1 to 5 days.

[0064] In the urine neutralization treatment method according to the present invention, the buffered saline solution may be added at a concentration of 50 to 200%, preferably 100 to 200%, and most preferably 100% based on the volume of urine.

[0065] In the urine neutralization treatment method according to the present invention, the vitamin C may be added at a concentration of 50 to 600 μg / mL, preferably 400 to 600 μg / mL, and most preferably 600 μg / mL.

[0066] In the urine neutralization treatment method according to the present invention, the glucose may be added at a concentration of 5 to 60 mM, preferably 40 to 60 mM, and most preferably 60 mM.

[0067] In the urine neutralization treatment method according to the present invention, the urine neutralization composition may further include polyethylene glycol (PEG).

[0068] In the urine neutralizing composition according to the present invention, the polyethylene glycol may be added at a concentration of 10 to 60 mg / mL, preferably 40 to 60 mg / mL, and most preferably 60 mg / mL.

[0069] In the urine neutralization treatment method according to the present invention, the urine neutralization composition may be composed of buffered saline, vitamin C (ascorbic acid), glucose, and polyethylene glycol (PEG).

[0070] In the urine neutralization treatment method according to the present invention, the urine neutralization composition may be composed of 100% buffered saline, 600 μg / mL of vitamin C (ascorbic acid), 60 mM of glucose, and 60 mg / mL of polyethylene glycol (PEG) based on the volume of urine.

[0071] In the urine neutralization treatment method according to the present invention, the urine neutralizing liquid composition may be added in an amount of 5 to 30%, preferably 10 to 20%, and most preferably 10% of the urine volume.

[0072] In the urine neutralization treatment method according to the present invention, the urine may be derived from a mammal. For example, the mammal may include, but is not limited to, a human, a non-human primate, a mouse, a rat, a dog, a cat, a horse, or a cow.

[0073]

[0074] Hereinafter, the present invention will be described in detail through examples, but the following examples and experimental examples are merely illustrative of one form of the present invention, and the scope of the present invention is not limited by the following examples and experimental examples.

[0075]

[0076] <Example>

[0077] Example 1. Changes in cell viability in urine over time.

[0078] Because the number of cells present in urine is extremely small, it is difficult to accurately assess cytotoxicity over time for actual cells in urine. Therefore, by utilizing urinary-derived stem cells (UDSCs), which possess superior cell adhesion and proliferation capabilities and stable biological properties, among the cells that can exist in urine, the level of cell death in urine cells can be reproduced in the laboratory, as follows:

[0079] 3 X10 Urinary Stem Cells Isolated from Fresh Urine of Healthy Donors 5Dogs were suspended in 1 mL of urine from 10 different male and female donors, and samples were collected at 0 h (immediately after urine suspension), 3 h, 6 h, and 24 h. Cell viability (%) was measured using an automated cell counter (ADAM-MC, NanoEntek) and an automated cell counter kit (ADAM™ AccuChip Kit, NanoEntek). The reliable measurement range of the automated cell counter used in this example was 5X10 4 cells / mL 4X10 inland 6 cell / mL was appropriate, and the total cell number was the initial suspended cell concentration (3X10 5 Only values ​​measured within approximately 20% of the error range of cells / mL were judged to be valid.

[0080] As a result, all cells suspended in urine showed a tendency for their viability to decrease over time. Viability varied significantly among samples for the first 3 to 6 hours, and after that, cell viability in all samples decreased significantly, averaging 18.11±10.55% after 24 hours (Fig. 1).

[0081]

[0082] Example 2. Cell protection effect of buffered saline in urine

[0083] 3X10 of stem cells derived from this 5 After suspending the dog in 1 mL of urine, buffered saline was added and mixed at 10, 50, 100, and 200% of the urine volume, respectively, to evaluate the cell protection effect.

[0084] As a result, compared to the cell viability of the negative control group suspended in urine, which was less than about 20% after 24 hours, a significant increase in cell viability was observed when HBSS was added in an amount of 50% to 200% of the urine volume, and it was shown to be dose-dependent on the added HBSS. In addition, in order to confirm the difference in cell protection efficacy according to the concentration of HBSS, the concentration (1X) generally used in cell culture was diluted to 0.5X and 0.1X and the change was observed, and it was shown that there was no statistically significant difference in the concentration range of 0.1X to 1X of HBSS (Fig. 2).

[0085]

[0086] Example 3. Cytoprotective effect of polyethylene glycol (PEG) in urine

[0087] 3X10 of stem cells derived from this 5 After suspending the dog in 1 mL of urine, the cytoprotective effect was evaluated by adding PEG solutions at concentrations of 0, 10, 20, 40, and 60 mg / mL, respectively. PEG was dissolved in HBSS, and the amount of PEG solution added for each concentration was 10% of the urine volume to properly confirm the cytoprotective effect of PEG while minimizing the influence of HBSS.

[0088] As a result, a significant increase in cell viability was observed when PEG was added at concentrations of 10 mg / mL to 60 mg / mL compared to the negative control and HBSS control group (PEG 0 mg / mL), and it was shown to be concentration-dependent on the added PEG. In particular, it was confirmed that while there was almost no cell protection effect when HBSS was added at 10% of the urine volume, cell viability was improved to a level comparable to the positive control when PEG was added at a concentration of 60 mg / mL. This cell protection effect of 60 mg / mL of PEG was maintained for up to 72 hours, confirming that the survival period of cells in urine was significantly extended. In addition, when the addition volume and concentration of the PEG solution were increased and the same analysis was performed, it was shown that a cell protection effect comparable to the positive control was maintained up to 100% of the urine volume and a concentration of 300 mg / mL of PEG (Fig. 3).

[0089] Example 4: Urinary cell protective effect of vitamin C (ascorbic acid)

[0090] 3X10 of stem cells derived from this 5 After suspending the dog in 1 mL of urine, the cell protective effect was evaluated by adding Vit. C solution at concentrations of 0, 50, 200, 400, and 600 μg / mL, respectively. Vit. C was dissolved in HBSS, and the amount of Vit. C solution added for each concentration was added at 50% of the urine volume to appropriately confirm the cell protective effect of Vit. C while minimizing the influence of HBSS.

[0091] As a result, a significant increase in cell viability was observed at 72 hours in the test groups to which Vit. C was added at concentrations of 50 μg / mL to 600 μg / mL compared to the negative control group and the HBSS control group (Vit. C 0 μg / mL), and it was shown to be concentration-dependent on the added Vit. C. In particular, it was confirmed that when HBSS was added at 50% of the volume of urine, the cell viability was about 40%, whereas when Vit. C was added at a concentration of 600 μg / mL, the cell viability could be improved to about 80%, which was equivalent to the positive control group (Fig. 4).

[0092]

[0093] Example 5: Cellular protective effect of glucose in urine

[0094] 3X10 of stem cells derived from this 5 After suspending the dog in 1 mL of urine, glucose solutions were added at concentrations of 0, 5, 20, 40, and 60 mM, respectively, to evaluate the cytoprotective effect. Glucose solutions were diluted in HBSS, and the amount of glucose solution added for each concentration was 50% of the urine volume to appropriately confirm the cytoprotective effect of glucose while minimizing the influence of HBSS.

[0095] As a result, a significant increase in cell viability was observed at 72 hours in the test groups to which Glucose was added at concentrations of 5 mM to 60 mM compared to the negative control group and the HBSS control group (Glucose 0 mM), and it was shown to be concentration-dependent on the added Glucose. In particular, it was confirmed that when HBSS was added at 50% of the volume of urine, the cell viability was about 40%, whereas when Glucose was added at a concentration of 60 mM, the cell viability could be improved to about 80%, which was equivalent to the positive control group (Fig. 5).

[0096]

[0097] Example 6: Urine cell preservation effect of urine neutralizing solution composition

[0098] The preservation effect on actual urine cells was evaluated using an optimal urine neutralizing solution containing a mixture of buffered saline, polyethylene glycol, vitamin C, and glucose. The research team of Reference 1 demonstrated the best cell preservation efficiency by adding cell culture medium (a mixture of keratinocyte-serum free medium [KSFM] and embryonic fibroblast medium [EFM], containing 10% FBS) to 10% of the urine volume. For comparative analysis, the urine neutralizing solution of this example was prepared by mixing 1X buffered saline, polyethylene glycol (60 mg / mL), vitamin C (600 μg / mL), and glucose (60 mM), and added to 10% of the urine volume in the same amount as the solution of the research team above.

[0099] Each test group was added to 100 mL of fresh urine collected from 12 healthy donors. Cells were isolated and cultured after 0, 6, 24, and 48 hours to compare the number of colonies, cell culture success rate, and cell proliferation ability of the total urine-derived cells. The number of cell colonies was counted by observing only the number of colonies composed of single cells under a microscope. The cell culture success rate was expressed as a percentage of the 12 specimens in which attachment and culture of urine-derived cells were confirmed with the naked eye. Cell proliferation ability was evaluated using the formula for calculating the population doubling level (PDL) after culturing the cells for 4 days.

[0100] PDL = ln(Nf / Ni) ln2 (Ni = number of cells at initial inoculation, Nf = final number of cells).

[0101] As a result of comparing the number of cell colonies according to the urine preservation time, fresh urine (G1) did not culture cell colonies after 24 hours. The urine neutralizing solution (G3) of the present invention showed excellent cell colony culture up to 48 hours, showing a high cell culture success rate. On the other hand, although some cell colonies were cultured in the cell culture medium (G2) up to 24 hours, it was shown that almost no cell colonies were formed at 48 hours. As a result of comparing the cell proliferation ability according to the urine preservation time, the urine-derived cells isolated at 48 hours in the cell culture medium (G2) showed a significantly reduced cell proliferation ability compared to the cells isolated immediately after urine collection. On the other hand, the urine neutralizing solution (G3) of the present invention was confirmed to maintain a cell proliferation ability at a level equivalent to that of the cells isolated immediately after urine collection even in the cells isolated at 48 hours (Fig. 6).

[0102] Accordingly, it was demonstrated that the urine neutralizing composition according to the present invention can significantly extend the maximum cell preservation period in urine by more than 100% under the same processing conditions as a cell culture medium containing conventional fetal bovine serum (FBS).

[0103]

[0104] Example 7: Effect of cell preservation in urine according to changes in the detailed composition of urine neutralizing solution

[0105] The cell preservation effect in urine of solutions mixed with different combinations of buffered saline, polyethylene glycol, vitamin C, and glucose, which are the detailed compositions of urine neutralizing solutions, was compared. The concentrations of each component of the urine neutralizing solutions in this example were composed of different combinations of 1X buffered saline, polyethylene glycol (60 mg / mL), vitamin C (600 μg / mL), and glucose (60 mM), and were added at 10% of the urine volume. After adding each test group to 100 mL of fresh urine collected from three healthy donors, cells were isolated and cultured after 0, 6, 24, 48, and 72 hours, and the number of single colonies of urine-derived cells was counted.

[0106] As a result of comparing the number of cell colonies according to urine preservation time, in the test group (G1) with only 10% HBSS added, colonies were formed in 2 out of 3 specimens at a maximum of 6 hours, but the number of colonies formed was reduced by approximately 85% compared to 0 hours. In the test group (G2) with 10% added HBSS and Glucose mixture solution, the number of colonies formed at 6 hours was improved compared to the G1 test group, but the colony formation probability was very low in specimens at 24 hours. On the other hand, in the test group (G3) to which 10% of a mixed solution of HBSS, Glucose, and Vit. C was added, the number of colonies formed at 24 hours was significantly improved compared to the G2 test group, and colonies were formed up to 48 hours. In particular, in the test group (G4) to which 10% of a mixed solution of HBSS, Glucose, Vit. C, and PEG was added, the number of colonies formed at 48 hours was significantly improved compared to the G3 test group, and it was confirmed that colonies of urine-derived cells could be stably formed in all specimens even up to 72 hours (Fig. 7).

[0107]

[0108] While specific aspects of the present invention have been described in detail above, it should be apparent to those skilled in the art that these specific descriptions are merely preferred implementation examples and do not limit the scope of the present invention. Therefore, the substantial scope of the present invention is defined by the appended claims and their equivalents.

[0109]

[0110] [References]

[0111] 1. Lang, Ren, Guihua Liu, Yingai Shi, Shantaram Bharadwaj, Xiaoyan Leng, Xiaobo Zhou, Hong Liu, Anthony Atala, and Yuanyuan Zhang. “Self-renewal and differentiation capacity of urine-derived stem cells after urine preservation for 24 hours.” PloS one 8, no. 1 (2013): e53980.

[0112]

[0113] When urine is refrigerated and preserved using the urine neutralizing composition according to the present invention, not only can the cell colony formation ability and cell proliferation ability be maintained, but the cell preservation period can be extended to 24 hours or more, preferably 48 hours or more, and most preferably 72 hours or more.

Claims

1. A urine neutralizing solution composition containing buffered saline, vitamin C (ascorbic acid), and glucose.

2. In paragraph 1, A urine neutralizing composition, characterized in that the buffered saline solution is added in an amount of 50 to 200% based on the volume of urine.

3. In paragraph 1, A urine neutralizing composition characterized in that the above vitamin C is added at a concentration of 50 to 600 μg / mL.

4. In paragraph 1, A urine neutralizing solution composition characterized in that the glucose is added at a concentration of 5 to 60 mM.

5. In paragraph 1, A urine neutralizing composition, characterized in that the urine neutralizing composition further comprises polyethylene glycol (PEG).

6. In paragraph 5, A urine neutralizing composition, characterized in that the polyethylene glycol is added at a concentration of 10 to 60 mg / mL.

7. In paragraph 1, A urine neutralizing composition characterized in that the urine neutralizing composition comprises buffered saline, vitamin C (ascorbic acid), glucose, and polyethylene glycol (PEG).

8. In paragraph 1, A urine neutralizing composition characterized in that the above urine neutralizing composition is composed of 100% buffered saline, 600 μg / mL of vitamin C (ascorbic acid), 60 mM of glucose, and 60 mg / mL of polyethylene glycol (PEG) based on the volume of urine.

9. A step of adding a urine neutralizing composition according to any one of claims 1 to 8 to urine; A method for neutralizing urine comprising:

10. A method for neutralizing urine, characterized in that the method further includes a step of storing urine with a urine neutralizing composition added thereto at 2 to 8°C for 1 to 5 days.

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