Lens disinfectant and production method

Abstract

The invention relates to a lens disinfectant and production method effective against Achantamoeba castellanii.

Description

[0001] DESCRIPTION

[0002] LENS DISINFECTANT AND PRODUCTION METHOD

[0003] Technical Field

[0004] The invention relates to a lens disinfectant and production method effective against Achantamoeba castellanii.

[0005] State of the Art

[0006] Acanthamoeba keratitis is an infection characterized by blurred vision, sensitivity to light, eye redness, which may cause blindness in extreme cases. The risk of transmission of A. castellanii parasite, which can be found in the form of trophozoite and cyst and is the source of this infection, increases with the use of contact lenses, and most of the cases consist of contact lens wearers. [1]

[0007] The disinfecting solutions for contact lenses of the present art are insufficient to effectively neutralize the population of A. castellanii or have toxic effects on human cells. For example, the patent with reference US11331404B2 discloses a system for neutralizing the solution with a metal catalyst after soaking it in a solution prepared with hydrogen peroxide (H2O2). This system includes extra stages such as neutralization with a metal catalyst, as well as the use of toxic hydrogen peroxide. Most importantly, it is unknown whether hydrogen peroxide affects the cyst form of A. castellanii.

[0008] In the article referenced by Abjani et al. [2], it is stated that chlorhexidine used in the previous art is ineffective against the cyst form of A. castellanii, therefore, disinfection can be carried out more effectively with the use of cellulase, an enzyme that helps break down the cyst wall. All the problems mentioned above have made it necessary to make an innovation in the relevant field as a result.

[0009] Objects and Brief Description of the Invention The main object of the invention is to develop an effective lens disinfectant against A. castellanii, an amoeba that causes complications that may lead to blindness in the eyes.

[0010] The object of the invention is to develop a disinfecting solution effective against the trophozoite and cyst forms of A. castellanii and to reduce the transmission of Acanthamoeba keratitis through contact lenses.

[0011] The trophozoite form is the form of the microorganism with active feeding and movement ability. The cyst form is a more durable and resistant resting form compared to the trophozoite form, which occurs under challenging environmental conditions. This cyst wall contains polysaccharides and disaccharides in its structure. The invention, which targets both forms to provide an effective A. castellanii disinfection, preferably uses lactase or combinations of lactase with cellulase to break down the cyst wall, and chlorhexidine to eliminate viability in the trophozoite form. Thanks to the three components used, the toxic effect of chemicals such as hydrogen peroxide is avoided as well as providing the desired disinfecting effect.

[0012] Definitions of Figures Describing the Invention

[0013] Figure 1. Change in the viable Acanthamoeba castellanii number in the phosphate-buffered saline (PBS) medium according to the concentration of lactase and chlorhexidine. (Negative control: cysts only, solvent control: cysts + PBS, positive control: cysts + PBS + chlorhexidine)

[0014] Figure Z.Change in the viable Acanthamoeba castellanii number with the addition of cellulase and chlorhexidine to lens disinfectant (Oxysept (OS), AO SEPT PLUS (AO SP), OPTI-FREE pure moist (OF PM), Renu Fresh (RF), FreshKon CLEAR (FKC) and COMPLETE RevitaLens (CRL), chlorhexidine (CHX))

[0015] Figure 3.Microscope images showing the anti-cystic activity of lactase and chlorhexidine on Acanthamoeba castellanii (a: negative control (cysts), b: solvent control (cysts+PBS), c: 50 pg / mL lactase, d: 75 pg / mL lactase, e: 100 pg / mL lactase, f: positive control (cysts+PBS+chlorhexidine), g: 50 pg / mL lactase+chlorhexidine, h: 75 pg / mL+chlorhexidine, i: 100 pg / mL+chlorhexidine, chlorhexidine concentration in chlorhexidine environments 100 pg / mL)

[0016] Definitions of Components / Pieces / Parts of the Invention Detailed Description of the Invention

[0017] The subject matter of the invention is to develop a lens disinfectant, which is effective against A. castellanii, an eye parasite, and which is non-toxic for human cells, and to develop the production method of this disinfecting solution.

[0018] A. castellanii has two life stages: trophozoite and cyst. There are disinfecting solutions effective against the trophozoite form of the parasite mentioned in the present art. However, although the trophozoite form can be largely eliminated with solutions containing chemicals such as hydrogen peroxide and chlorhexidine, the current state of the art is insufficient to find an effective solution against the cyst form. The parasite, which becomes a cyst and continues to survive as a host, can reactivate and continue its metabolic activities in the host. Therefore, the present invention targets both forms of the parasite for effective disinfection.

[0019] The cyst walls of A. castellanii contain polysaccharide parts that contribute to the formation of a hard shell. Some of these polysaccharides are 1,4 0-glucans (cellulose) and lactose, which is the disaccharide of galactose and glucose. In the event that enzymes and chemicals that can provide the destruction of said molecules are used, the use of substances targeting the membranes of the host cells in the trophozoite form provides effective disinfection in order to neutralize the microorganism, which can only exist in the form of trophozoite. For this purpose, polyhexamethylene biguanide (PHMB) or chlorhexidine, in particular the more effective chlorhexidine (CHX), are preferably used.

[0020] In a preferred embodiment of the invention, lactase, an enzyme that can digest the lactose molecule, is used to ensure the destruction of the cyst wall. Preferably, the viability of the parasite is also eliminated by targeting the cell membrane in the trophozoite form by using chlorhexidine or another disinfecting agent used in the formulation of the lens disinfectant.

[0021] The effect of lactase on the viability of A. castellanii is shown in Figure 1. Three different lactase concentrations of 50 pg / mL, 75 pg / mL and 100 pg / mL were tried in phosphate buffered saline solution, and a decrease in the amount of viable A. castellanii was observed with the increase in the amount of lactase. The viable A. castellanii number decreased further with the addition of chlorhexidine to this solution in addition to lactase. Comparison of the positive control and the viable microorganism number corresponding to the solution containing lactase and chlorhexidine proves that the disinfecting solution becomes more effective with the use of lactase with chlorhexidine. Microscope images of these solutions are presented in Figure 3. An alternative embodiment of the invention is the formulation comprising both lactase and cellulase enzymes or other chemical combinations targeting these molecules. Thus, more effective destruction of the cyst wall is provided, and then the parasite in the form of trophozoite is neutralized by the use of chlorhexidine or another disinfecting agent.

[0022] The effect of cellulase on the viability of A. castellanii is shown in Figure 2. It is seen that these solutions become more effective against A. castellanii as a result of the addition of chlorhexidine (CHX) and cellulase to the currently used contact lens solutions.

[0023] The production method of the disinfecting solutions developed and mentioned within the scope of the invention includes the following steps: i. Preparing phosphate buffered saline solution with pH 7.0-7.4, ii. Adding lactase and cellulase to this solution, which provides the destruction of the cyst wall, iii. Adding disinfecting agents to the solution obtained.

[0024] The dimensions used in the production method of the formulation in which cellulase and lactase, which is a preferred embodiment of the invention, are used as a destructive agent of the cyst wall and chlorhexidine is used as a disinfecting agent against the trophozoite form of A. castellanii are given in the table below:

[0025] Table 1 Contents of the lens disinfectant subject to the invention In a preferred embodiment of the invention, PBS solution with a pH of 7.4 is prepared first. 200 mg of lactase, preferably 200 mg of cellulase and preferably 20 mg of chlorhexidine per liter is preferably added to this solution. REFERENCES

[0026] [1] Simau, F.A., Ahmed, U., Khan, K.M. et al. Lactase can target cellular differentiation of Acanthamoeba castellanii belonging to the T4 genotype. Parasitol Res 123, 117 (2024). https: / / doi.org / 10.1007 / s00436-024-08131-2

[0027] [2] Abjani, F., Khan, N. A., Yousuf, F. A., Siddiqui, R. Targeting cyst wall is an effective strategy in improving the efficacy of marketed contact lens disinfecting solutions against Acanthamoeba castellanii cysts. Contact Lens and Anterior Eye 39 (3) 239-243 (2016). https: / / doi.org / ! 0.1016 / j.clae.2015.11.004

Claims

CLAIMS1. A lens disinfectant, characterized in that it comprises lactase and cellulase to break down the cyst wall in phosphate buffered saline solution with pH 7.0-7.4 and a disinfecting agent targeting the cell membrane.

2. The lens disinfectant according to claim 1, characterized in that it comprises chlorhexidine or polyhexamethylene biguanide as disinfecting agent.

3. The lens disinfectant according to claim 2, characterized in that it comprises chlorhexidine at a concentration of 20 pg / mL.

4. The lens disinfectant according to claim 3, characterized in that it comprises lactase at a concentration of 200 pg / mL.

5. The lens disinfectant according to claim 4, characterized in that it comprises cellulase at a concentration of 200 pg / mL.

6. A production method for a lens disinfectant according to claim 1, characterized in that it comprises the following: i. Preparing phosphate buffered saline solution with pH 7.0-7.4, ii. Adding lactase and cellulase to this solution, which provides the destruction of the cyst wall, iii. Adding disinfecting agents to the solution obtained.

7. The method according to claim 6, characterized in that, when preparing the phosphate buffered saline solution, 0.137 millimoles of sodium chloride (NaCl), 0.0027 millimoles of potassium chloride (KC1), 0.01 millimoles of disodium phosphate (Na2HPC>4) and 0.0018 millimoles of mono potassium phosphate (KH2PO4) are added per milliliter of water.

8. The method according to claim 6, characterized in that chlorhexidine or polyhexamethylene biguanide is added to this solution as a disinfecting agent.

9. The method according to claim 8, characterized in that 20 pg per milliliter of chlorhexidine is added to this solution.

10. The method according to claim 9, characterized in that 200 pg per milliliter of lactase is added to this solution.

11. The method according to claim 10, characterized in that 200 pg per milliliter of cellulase is added to this solution.

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