Lutein-containing water-soluble topical ophthalmic preparation and method for producing same

Abstract

A method for the preparation of a topical ophthalmic formulation useful as a tear substitute and a treatment for computer vision syndrome and related products are disclosed. In addition to the water-soluble components typical of aqueous eye drops or gels, the formulation also contains a stable micellar solution (or microemulsion) of lutein and Vitamin E TPGS. The method includes four steps performed sequentially, of which the second step of preparing an oily component consisting of Vitamin E TPGS and lutein is performed by liquidizing Vitamin E TPGS and adding lutein thereto at a temperature of 90-170°C under constant stirring. The third step of forming a stably suspended lutein / TPGS micellar solution in aqueous solution is performed by gradually adding an aqueous solution previously heated to 80-90°C to the stirred oily solution and continuing the stirring at high temperature until an optically uniform and transparent orange micellar solution is formed.

Description

[Technical field]

[0001] The present invention relates to a lutein-containing water-soluble topical ophthalmic formulation and a method for its preparation. More specifically, the present invention relates to a specific method for the preparation of a tear substitute containing lutein, a lipid-soluble active ingredient with high antioxidant and photoprotective power, in combination with Vitamin E TPGS, a well-known emulsifier, in addition to the essential thickening and mucosal mimetic agents of tear substitutes and the common additives typically contained in aqueous eye drops, such as buffers, osmoprotectants or tonicity agents, chelating agents, and possibly antibacterial agents. The specific continuous preparation method makes it possible to obtain a stable micellar solution (or microemulsion) suitable for topical ocular administration. [Background technology]

[0002] The organ of vision, whose function is to capture light radiation from the outside world and convert it into nerve impulses used for sensory purposes such as vision and reflexes, is separated from the external environment by the tear film, a layer of liquid composed of several layers. The tear film is continuously and evenly distributed over the ocular surface by the closing of the eyelids and has the crucial function of maintaining the homeostasis of the ocular surface. It allows proper lubrication of the corneal epithelium by reducing friction with the eyelids and allows the transport and diffusion of molecules important for the survival of the epithelium and the cornea (oxygen, carbon dioxide, ions, mucins, lipids). It also has a pronounced antibacterial activity due to the presence of several enzymes, which ultimately guarantees the exchange and keeps the ocular surface clean, removing impurities from the environment, metabolic wastes, and exfoliated cells.

[0003] From outside to inside, the layers of the tear film are: 1) A thin superficial lipid layer produced by the meibomian glands that slows evaporation of the underlying aqueous layer, prevents leakage of tears from the eyelid margins, and maintains hydration during sleep; 2) The intermediate aqueous layer contains various molecules (electrolytes, proteins, amino acids, organic acids) with enzymatic and antibacterial functions, and not only protects, nourishes, and replaces the epithelium, but also reduces blinking friction and washes away many impurities that have accumulated on the ocular surface. 3) The innermost mucosal layer, which is composed of mucin and has a pronounced hydrophilicity, and whose presence allows an aqueous layer to spread over the otherwise highly hydrophobic corneal epithelium.

[0004] Thus, the stability of the tear film is the result of a balance of its various components, the alteration of which can lead to the development of lacrimal dysfunction syndrome, also known as dry eye syndrome, and other ocular diseases.

[0005] The aqueous layer of the tear film is constantly evaporating, which is constantly being replenished by lacrimal secretion. If this compensatory secretion is reduced or insufficient for any reason, the symptoms of dry eye disease appear, consisting of dehydration with persistent ocular discomfort accompanied by a foreign body sensation in the eye, itching, burning, difficulty in opening the eyes, the desire to wash or rub the eyes, discomfort when looking at light, and daytime eye fatigue. Eye redness and reduced light intensity are most often signs of poor hydration of the ocular surface.

[0006] Dry eye caused by reduced tear production is more precisely divided into two main subclasses: dry eye associated with Sjögren's syndrome (SSDE), an autoimmune chronic inflammatory disease, and dry eye not caused by Sjögren's syndrome, which represents a group of cases of lacrimal gland dysfunction in which the systemic autoimmune pathology characteristic of SSDE has been excluded.

[0007] Also, some ocular surface diseases can alter the quantity and composition of tears. The most obvious case is blepharitis, where inflammation of the eyelids alters meibomian gland secretions, resulting in altered tear film. Misuse of any type of contact lens, such as extended daytime wear, can also cause changes in the tear film.

[0008] Another problem that has only recently appeared, some of which share symptoms with those associated with dry eye syndrome, is caused by the extensive use of smartphones, computers and all screens that emit light radiation that interacts with various parts of the eye. The latter must "filter" these radiations, of which the artificial component called "blue light" is the most harmful. Blue light is more or less clearly emitted by all light sources, especially cold sources such as LED lights, which are widely used. Exposure to radiation corresponding to wavelengths in the range of 380-500 nanometers, which also includes parts of the near ultraviolet, is indeed harmful to the eye and is associated with increased ocular inflammation, since it leads to an increased production of oxygen free radicals that contribute to the progressive degeneration of the retina.

[0009] Additionally, blinking frequency is significantly reduced during computer use, resulting in increased exposure of the ocular surface to greater tear evaporation.

[0010] All this has led the American Optometric Association to define the so-called "Computer Vision Syndrome" (CVS), which is currently a widespread problem, affecting a huge number of people worldwide and with a very large potential patient population. The symptoms are varied and include visual, neurological and musculoskeletal symptoms, which do not necessarily occur together, vary greatly from person to person and are classified as external and internal symptoms. The former include burning, irritation, tearing and dryness, which seem to be related to poor tearing problems, while the latter include eye fatigue, headaches, eye pain, double vision and blurred vision, which are generally related to vision problems in the strict sense (refraction, accommodation or binocular).

[0011] In order to contribute to the improvement of dry eye and provide an appropriate therapeutic means to alleviate symptoms, there are many commercially available tear substitute (artificial tear) formulations that are applied periodically by instillation onto the cornea or into the conjunctival fornix. Specifically, the main goal of dry eye treatment is to improve ocular comfort and quality of life, and to restore the normal homeostatic balance between the ocular surface and the tear film. Although it is almost impossible to completely eliminate symptoms, they can often be improved.

[0012] Conventional tear substitutes, consisting of artificial tears, ointments, and gels, generally function as lubricants for the ocular surface, and many substances used for this purpose as thickeners have no real pharmacological action; that is, the only activity reliably attributed to them is lubrication. As for computer vision syndrome, in any case, it is not recommended to use electronic device screens for more than 4-5 hours a day. Here too, it is recommended to instill ophthalmic lubricants as needed to reduce burning and dryness.

[0013] Lutein (β,ε-carotene-3,3'-diol), a member of the xanthophyll family of carotenoids, and its stereoisomer zeaxanthin are antioxidant molecules that cannot be synthesized by the human body. Therefore, their absorption is known to depend on the intake of certain fruits and vegetables, or animal foods such as eggs. As reported in the literature, these antioxidant molecules are highly concentrated in the retina and macula of the human eye (Roberts JE, Dennison J., The Photobiology of Lutein and Zeaxanthin in the Eye, J. Ophthalmol., 2015: 687173). Lutein absorption has also been reported to improve vision and slow the progression of age-related macular degeneration (AMD) (Moschos, MM et al., Effect of carotenoids dietary supplementation on macular function in diabetic patients. Eye Vis. 2017, 4, 23; Gong, X. et al. Effects of the Macular Carotenoid Lutein in Human Retinal Pigment Epithelial Cells, Antioxidants 2017, 6, 100).

[0014] Lutein exerts a barrier effect that protects the photoreceptor structures of the eye and retina from photodamage caused by sunlight and light from computer screens, smartphones and other electronic devices, due to its physical property of absorbing blue light and specific wavelengths of UVA / UVB / UVC radiation. Its characteristic structure contains nine double bonds: [ka] are in fact involved in the absorption of some wavelengths of light and the emission of others. In particular, lutein and zeaxanthin absorb visible blue light (400-500 nm) with an absorption peak at 440 nm. Cited literature has shown that age- or diet-related loss of lutein and zeaxanthin increases UV-induced eye damage, and that dietary supplementation with these carotenoids exerts a protective effect against UV radiation-induced retinal damage.

[0015] Although lutein is widely used in the ophthalmic field in the formulation of certain dietary supplements, it is not commercially available in topical ophthalmic preparations, i.e. in the form of eye drops or aqueous gels, or in the form of ointments or eye ointments, which are administered directly to the eye. To formulate a lipophilic molecule such as lutein in a topical ophthalmic product, it is necessary in fact to transport it in a long-term stable oil-in-water emulsion, in order to guarantee the preservation of the active ingredient without compromising its bioavailability during the period envisaged for this type of commercial product.

[0016] U.S. Patent No. 6,573,299 (Petrus) discloses a topical ophthalmic composition for administration to the eyelid and orbital field that may include lutein and tocopherol acetate along with a gelling agent, carboxyvinyl polymer (carbomer). The composition utilizes a penetration enhancer to allow the active ingredient to penetrate the underlying tissue and vascular network of the orbit.

[0017] WO 2009 / 084069 (Lio Farmaceutici Srl) discloses an ophthalmic composition in the form of an aqueous solution or gel, in which lutein is encapsulated in liposomes / cerasomes, which may further comprise hyaluronic acid (salts) and a buffering agent.

[0018] Widely used emulsifiers also known in the ophthalmic field are amphiphilic derivatives of vitamin E (especially β-tocopherol, the most widespread component of vitamin E), which consist of a hydrophilic polar head formed by a polyethylene glycol chain connected to the tocopherol structure by a diester bridge of succinic acid.

[0019] [ka]

[0020] This compound has the chemical name d-alpha-tocopheryl polyethylene glycol 1000 succinate (where 1000 represents the chain length of the PEG) and is also known as Vitamin E TPGS, or TPGS (English acronym) and tocophersolan (INN).

[0021] Vitamin E TPGS exhibits low critical micelle concentration and large surface area, characteristics that are characteristic of a good emulsifier. For this reason, the compound is considered a multifunctional excipient, capable of emulsifying or aiding in the solubilization of a wide range of immiscible water-oil compounds and other poorly soluble compounds, and is widely used in dietary supplements (Maya et al., Pharmaceutical Profile of Alpha-tocopherol - A Brief Review, Int. J. Pharma. Chem. Sci. 2012 1 (3): 1023-1039).

[0022] The use of TPGS in the ophthalmic field is described, for example, in U.S. Pat. No. 5,886,030 (Alcon Laboratories Inc.), which proposes its use in ophthalmic formulations containing nonsteroidal anti-inflammatory active ingredients (NSAIDs), where TPGS functions to reduce the irritant effect of the NSAID when applied directly to the cornea and to improve its solubility in the aqueous phase.

[0023] Furthermore, regarding the issue of lipophilic active ingredients, the use of TPGS as a specific emulsifier has been proposed, for example, in EP 2197461 (Aurinia Pharmaceuticals), where it is formulated together with another surfactant (PEG alkyl-aryl ether, Octoxynol-40) to obtain a micellar solution for the delivery of active ingredients that inhibit calcineurin in topical ophthalmic formulations, in particular cyclosporine A. The preparation method involves mixing the lipophilic active ingredient with the two surfactants in a common solvent, evaporating the solvent to obtain a substantially solid substance, hydrating this substance with an aqueous solution and dissolving the product to obtain an optically clear micellar solution.

[0024] In the same field, EP 25798451 (Medivis) discloses a topical ophthalmic formulation comprising: one or more lipophilic active ingredients (or none if artificial tears are desired), an emulsifier consisting of d-α-tocopheryl polyethylene glycol 1000 succinate (TPGS), an oily component consisting of medium chain triglycerides (MCT), and an ophthalmologically acceptable aqueous phase. The concentration of TPGS in the product is 0.1-5% by weight, and the weight ratio of MCT to TPGS is 1:2.8-1:3.6. Furthermore, no co-surfactant is present, and the average size of the particles of the oil phase dispersed in the aqueous phase is less than 100 nm, usually less than 30 nm. The above specification reports that the preparation of the microemulsion occurs spontaneously at room temperature by first mixing the oil phase ingredients (i.e., any fat-soluble active ingredients, MCT, and Vitamin E TPGS), then mixing together the aqueous phase ingredients, and then taking care to combine the two ingredients with constant stirring at room temperature.

[0025] The presence of vitamin E (α-tocopherol) in topical ocular products is certainly beneficial and well tolerated, firstly because α-tocopherol has been detected in tears at physiological concentrations of 0.033-0.95 μM (Khaksari M. et al., Determination of water-soluble and fat-soluble vitamins in tears and blood serum of infants and parents by liquid chromatography / mass spectrometry, Experimental Eye Res. 2017 155: 54-63). Furthermore, the antioxidant properties of vitamin E have been confirmed to be useful against glaucoma, cataracts, dry eye syndrome, and other ocular pathologies (Ribeiro A. et al., Poloxamine micellar solubilization of alpha-tocopherol for topical ocular treatment, Colloids Surf. B ​​Biointerfaces 2017 103: 550-5572013). Vitamin E also has a photoprotective function due to its antioxidant properties. Indeed, literature data shows that vitamin E can filter ultraviolet radiation (range 300-400 nm, with a peak at 350 nm) and has a protective effect against cell damage caused by ultraviolet radiation (Ayala MN and Soderberg PG, Vitamin E can protect against ultraviolet radiation-induced cataract in albino rats, Ophthalmic Res. 2004, 36 (5): 264-269).

[0026] Vitamin E and lutein are also known to act together as powerful antioxidants that neutralize reactive oxygen species (ROS) induced by UV radiation. The combination of these two compounds is even more useful for UV ocular photoprotection, as it protects the eye from lipid peroxidation, which leads to oxidative degradation of lipids and the formation of free radicals. Thus, lutein and vitamin E have been shown to act synergistically to neutralize free radicals. In particular, they investigated the effect of lutein and vitamin E on lipid peroxidation in human lens epithelial cells after UVB irradiation and found that pretreatment of cell cultures with lutein and vitamin E prior to UVB radiation exposure reduced lipid peroxidation by 47-57% compared to control epithelial cells treated with UVB alone (Chitchumroonchokchai C. et al., Xanthophylls and alpha-tocopherol decrease UVB-induced lipid peroxidation and stress signaling in human lens epithelial cells, J. Nutr. 2004, 134 (12): 3225-32322004).

[0027] In view of the described prior art, it is clear that the existence of an ophthalmic formulation for topical use that makes lutein easily administrable and bioavailable together with other ingredients, mostly water-soluble, commonly used in the treatment of disorders and conditions requiring protection and lubrication of the ocular surface and internal structures of the eye would be highly advantageous. Summary of the Invention

[0028] The object of the present invention is therefore to provide a tear substitute comprising a lipid phase consisting only of a combination of Vitamin E-TPGS and lutein. Such a combination has never been used in a topical ocular formulation. The tear substitute must be biocompatible and suitable for use with contact lenses and must be suitable for the management and treatment of the aforementioned conditions, due to its lubricating, antioxidant and photoprotective properties, to reduce the symptoms common to dry eye syndrome and CVS (irritation, burning, foreign body sensation, photophobia).

[0029] As mentioned, lutein is a lipophilic molecule and is not soluble in water. During the initial stages of experimentation related to the present invention, preliminary tests were performed which confirmed that most lutein added to an aqueous solution quickly settles to the bottom, even if the mixture is vigorously stirred for a long period of time.

[0030] According to the present invention, it has been found that it is possible to obtain lutein-containing ophthalmic formulations that are biocompatible and at the same time have lubricating, antioxidant and photoprotective activity and keep their physicochemical properties stable over time, and that a specific manufacturing method results in a micellar solution (or microemulsion) of lutein emulsified with Vitamin E TPGS. This lutein-based micellar solution is produced according to the present invention without the use of additional lipid excipients, except for a minimal amount of Vitamin E-TPGS.

[0031] The aforementioned micellar solution is dispersed in an aqueous phase which contains one or more thickening agents (rheology modifiers) and / or mucoadhesives in addition to the usual tonicity agents (osmolarity agents, osmolality regulators), buffers, and any further water-soluble components. The ophthalmic formulations of the present invention can be obtained by carrying out a specific, specially developed four-step preparation process.

[0032] In fact, according to the present invention, it has been found that under certain temperature and stirring conditions, Vitamin E-TPGS, an amphiphilic molecule with a polar hydrophilic head and a lipophilic tail, can bind to lutein and subsequently transport it to the interior of an aqueous solution in a stable micellar form. Due to its molecular properties, Vitamin E-TPGS is an excellent solubilizer of lutein, and under the conditions specified according to the present invention, its hydrophilic head faces the external aqueous phase, while the hydrophobic tail bound to lutein faces the interior of the micelle, spontaneously forming a bilayer without the need for an additional oily component in the lipid phase.

[0033] Thanks to the thermal / mechanical energy supplied to the system during preparation, it is possible to obtain dispersed particle sizes below 30 nm, usually in the range of 5-10 nm. Indeed, the formulation has a transparent appearance with a bright orange color.

[0034] The topical ophthalmic product of the present invention can be obtained by a four-step preparation method. In this method, in the first step, a predetermined amount of deionized water is introduced into a first container, to which one or more other water-soluble components of the formulation, such as a buffer system, are added. In the second step, Vitamin E TPGS is heated in the second container under stirring, first melting and then bringing it to a temperature of 90-170°C. Lutein is then added, while maintaining the high temperature and stirring speed, until a homogeneous oily solution is formed. In the third step, a portion of the aqueous solution from the first container is heated to a temperature of 80-90°C and then slowly added under stirring to the oily solution, and once the addition is complete, the stirring speed is increased while maintaining the temperature at 90-170°C and continuing heating, in order to obtain a stable micellar solution of Vitamin E TPGS and lutein in the aqueous phase. Finally, in the fourth step, the resulting micellar solution is added to the solution remaining in the first container under stirring. Once the solution has cooled to a temperature not exceeding 30° C., the remaining water soluble ingredients, including one or more thickening or gelling agents, are added, one at a time. The ingredients are then completely dissolved and the product is sterilized.

[0035] According to the present invention, by proceeding rigorously according to the methods presented, it is possible to ensure the successful formation of micellar aggregates Vitamin E-TPGS-lutein, rendering the lutein substantially "water soluble" and stable in solution.

[0036] Due to the antioxidant activity exhibited by lutein and the β-tocopherol contained in TPGS, and the light absorption characteristics towards wavelengths typical of blue light and UVA / UVB / UVC rays, the combination of Vitamin E TPGS and lutein according to the present invention can provide an enhanced barrier effect to protect the eye and retinal photoreceptor structures from photodamage caused by light from sources such as sunlight, computer screens, smart phones, and other electronic devices.

[0037] In addition to the innovative combination of the aforementioned components, the ophthalmic formulations of the present invention comprise in the aqueous phase one or more useful conventional water-soluble components already known as ocular surface lubricants, such as viscosity modifiers, mucoadhesives, and corneal surface moisturizers, preferably those based on polysaccharides, e.g., hyaluronic acid (HA) or its salts, and cellulose derivatives such as carboxymethylcellulose (CMC) and hydroxypropylmethylcellulose (HPMC, hypromellose), optionally modified by crosslinking.

[0038] According to another embodiment of the present invention, the formulation containing lutein and Vitamin E TPGS can be formulated in the form of eye drops or in the form of an ophthalmic gel by including a gel forming polymer such as polyacrylic acid (carbomer) along with an associated neutralizing agent in the aqueous component of the fourth step of the preparation process.

[0039] The ophthalmic formulations of the present invention have been developed in a variety of both preservative-free single dose and preservative-containing multi-dose formulations, both of which can be formulated with different buffer systems, such as borate, citrate, or phosphate buffers, as needed.

[0040] All embodiments of the product obtainable according to the invention can be used even with contact lenses on. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0041] Detailed Description of the Invention According to a first aspect of the invention, a specific object of the invention is a method for the preparation of a water-soluble topical ophthalmic formulation containing lutein, comprising the following steps in sequence: Step 1: a) introducing a quantity of deionized water, optionally components of a buffer system, and optionally a quantity of an ophthalmic thickener into a first stirred vessel and waiting for each of the optional components to completely solubilize; Step 2: a) heating a predetermined amount of Vitamin E TPGS in a second stirring vessel to a temperature of 80-90°C, preferably 80-85°C, and waiting for it to completely melt; b) bringing Vitamin E TPGS to a temperature of 90-170°C, preferably 150-160°C, and adding a predetermined amount of lutein while continuing stirring until the Vitamin E TPGS and lutein are completely dissolved to form a homogeneous orange oily solution consisting of only Vitamin E TPGS and lutein; Step 3: a) heating the water or aqueous solution obtained in operation a) of step 1 to a temperature of 80-90°C, preferably 80-85°C, and adding an aliquot of it gradually to a second vessel while keeping it under stirring (the aliquot added gradually should correspond to about 1 / 10 of the total volume of the final product, e.g. if the total volume of the product is 1000 ml, a volume equivalent to about 100 ml should be added to the second vessel), during which the product transitions from a water-in-oil emulsion to an oil-in-water emulsion by phase inversion, b) Once the addition is complete, increase the stirring speed while maintaining the temperature in the range of 90-170°C until a completely clear, optically homogeneous orange liquid is obtained; Step 4: a) adding the solution of step 3 to the remainder of the water or aqueous solution obtained in operation a) of step 1; b) After confirming that the temperature has dropped below 30°C, add the other water-soluble ingredients of the topical ophthalmic formulation to be produced, including one or more thickeners and / or mucoadhesives, one at a time in a predetermined amount to the stirring vessel, and wait until each is completely solubilized; c) The product thus obtained is sterilized.

[0042] According to some specific embodiments of the present invention, the topical ophthalmic formulation is liquid, i.e., eye drops administered dropwise. In this case, in step 1, operation a), in addition to deionized water, also introduce components of a buffer system into the first container, said components being preferably, but not necessarily limited to, boric acid and sodium tetraborate, or citric acid and sodium citrate, or phosphate buffer. If the ophthalmic thickener is crosslinked hyaluronic acid (HA-CK), in step 1, add this component after the two components of the buffer system.

[0043] Additionally or alternatively, one or more thickening agents and / or mucoadhesives, preferably selected from hyaluronic acid or one of its salts (HA), carboxymethylcellulose (CMC) and cross-linked carboxymethylcellulose (CMC-CK), are incorporated into the one prepared during operation b) of step 4.

[0044] Sodium hyaluronate or hyaluronic acid (HA) is a natural polysaccharide consisting of a specific structure formed by repeating units of D-glucuronic acid and N-acetylglucosamine that enhances the lubrication, hydration and protection of the ocular surface due to its well-known hygroscopicity. HA stabilizes both the aqueous layer of the tear film and, in the presence of contact lenses, the aqueous layer of the prelens tear film (PrLTF). Furthermore, the mucoadhesive properties and lubricating action of hyaluronic acid support the physiological repair process of the corneal epithelium.

[0045] In fact, in a healthy eye, the presence of an intact mucous layer is essential, since it transforms the corneal epithelium from a hydrophobic to a hydrophilic surface, protects the epithelium from eyelid friction, prevents bacterial adhesion thanks to its viscoelastic properties, and allows a regular and uniform distribution of the lipid layer of the tear film on the aqueous layer, thus reducing the surface tension. For example, if mucus production is reduced due to damage to the corneal goblet cells, the distribution of mucus on the ocular surface is impaired, resulting in a reduced contact of the tear film with the ocular surface, loss of film stability and the formation of dry areas.

[0046] It follows that the symptomatic effectiveness of tear substitutes is closely related to both their ability to remain on the ocular surface and their mucosomimetic and mucoadhesive properties. For these reasons, polymeric compounds that act as viscosity regulators are added to tear substitutes, which, in addition to washing away and diluting toxic or irritating substances present in the tear film, also hydrate the gelatinous mucin formation. These compounds also extend the retention time, providing comfort to the user for longer periods of time. Furthermore, thickeners protect the epithelium of the ocular surface.

[0047] With particular reference to hyaluronic acid, numerous experiments have been carried out with the aim of chemically modifying the molecule in order to further improve its properties. The most used chemical strategy to modify HA is cross-linking, either by direct addition of side chains or by addition of spacer arms, which then form stabilizing bonds between the HA molecules. In this way, it becomes more viscoelastic, which extends the contact time of HA with the ocular surface.

[0048] Carboxymethylcellulose (CMC) is one of the most widely used viscosity modifiers in the ophthalmic field. This polymer is often added to medical devices due to its properties of forming a transparent, viscoelastic protective shield that can thicken, stabilize, and hydrate the tear film on the corneal surface and extend the life of the tear film on the cornea between blinks (break-up time). Typically, the break-up time is shortened in dry eye conditions. CMC also exhibits cytoprotective activity on the ocular surface when applied before contact lenses. Moreover, subacute and chronic toxicity tests have reported no adverse effects caused by CMC, confirming its safety.

[0049] Carboxymethylcellulose is also commercially available in cross-linked form (CMC-CK, cross-linked type), and its toxicological profile has been determined by the European Commission to be equivalent to that of CMC.

[0050] According to the first embodiment of the invention described above, the ophthalmic solution obtained in operation b) of step 4 of the method is sterilized by 0.2 μm filtration and sent for packaging in suitable containers.

[0051] According to another specific embodiment of the present invention, where the topical formulation is in the form of an ophthalmic gel, in step 4, operation b), a predetermined amount of gelling agent, preferably carboxyvinyl polymer (carbomer), is added to the stirring vessel through a sieve. The gelling agent is allowed to soak and then dispersed while stirring is continued until a lump-free dispersion is obtained. Then, with continued gentle stirring, it is neutralized with a neutralizing agent.

[0052] In the case of the manufacture of a gel formulation according to the invention, the sterilization of step 4, operation c) is carried out by subjecting the product obtained to autoclaving or treatment with gamma radiation.

[0053] The basic steps of the proposed method are to heat the solution obtained in step 1 to 80-90°C and then add a portion of it to the vitamin E TPGS-lutein aggregates in step 2. If this operation is carried out at a lower temperature, thermal shock will occur in the vitamin E TPGS-lutein aggregates, and a cloudy liquid will be obtained instead of a transparent solution. If the manufacturing method is continued and the solution is filtered through 0.2 μm, a transparent solution will be obtained, but in any case, such a solution will not meet the specifications, since the finished product may be colorless. This result means that the vitamin E TPGS-lutein aggregates have been lost.

[0054] In Step 2, the key points are: · We need to make sure that a total phase change occurs in Vitamin E TPGS, which indicates that the molecule is ready to combine with lutein in the best way possible. After all the lutein has been added, the temperature must be held between 80-170°C by increasing the stirring speed for at least 1 minute.

[0055] If the following operations are carried out without ensuring compliance with the two conditions indicated, a product is obtained which does not have a transparent appearance and contains a precipitate.

[0056] In step 3, if all previous steps of the preparation method have been followed, it is important not to slow down the stirring speed and to maintain the temperature at 80-170° C. until the micellar solution becomes clear.

[0057] Compliance with all specified operating conditions is necessary to obtain a product that is not only compatible but also stable over time. In this regard, it is important to note that it is not sufficient to simply add all declared ingredients to the solvent in the stated order, since this will result in a final product in which lutein settles to the bottom as an insoluble precipitate, despite the use of Vitamin E-TPGS as an emulsifier.

[0058] The preparation method according to the invention does not result in the loss of lutein in solution, as shown by the fact that the orange color of the product continues to be present despite the treatments it undergoes during some steps of the preparation, in particular during the steps at high temperature and during the filtration of the semi-finished product. In fact, if the molecular structure of lutein were somehow damaged by temperature by breaking the chain of double bonds that contribute to the absorption of visible blue light at 400-500 nm (Roberts et al., 2015, supra) and the resulting emission of the corresponding wavelengths (ranging from yellow, through orange, in fact, to red), it would not be possible to obtain the color characteristic of lutein already in the semi-finished product (i.e. before filtration), even if a micelle solution of vitamin E TPGS-lutein aggregates was mechanically blocked by filtration at 0.2 μm.

[0059] The products obtained by applying the manufacturing method of the present invention have the following characteristics:

[0060] [Table 1]

[0061] According to a further aspect, the present invention relates to a topical ophthalmic formulation consisting of a stable micellar solution (microemulsion) of Vitamin E TPGS and lutein suspended in an aqueous phase containing a thickening and / or mucoadhesive or gelling agent, and other pharmacologically acceptable excipients.

[0062] Preferred embodiments of the described formulations are those in which the thickening agent and / or mucoadhesive agent is selected from hyaluronic acid or its salts (HA), cross-linked hyaluronic acid (HA-CK), carboxymethylcellulose (CMC), cross-linked carboxymethylcellulose (CMC-CK) and mixtures thereof. Other preferred embodiments are those in which the gelling agent is, for example, in an aqueous gel based on carboxyvinyl polymers (carbomers).

[0063] A typical formulation of an ophthalmic solution obtainable according to the present invention may have the composition shown in the table below.

[0064] [Table 2]

[0065] Above is an example of an ophthalmic product formulation based on the stable micellar solution of Vitamin E TPGS and lutein according to the present invention, which is an eye drop. A similar gel product example can be obtained by removing the buffer system and polysaccharide from the previous formulation, and instead adding a hydrophilic gelling polymer along with any associated neutralizing agents. EXAMPLES

[0066] Some specific embodiments of the formulation methods and related products of the present invention are described below by way of example and not by way of limitation, together with the results of experiments that have been performed.

[0067] It should be emphasized that the preparation methods proposed for liquid products always follow the same steps, only differing in the formulation with or without preservatives or with HA-CK. For this type of formulation, the difference relates to one of the steps in step 1, where it is also possible to add HA-CK immediately after the addition of the components that determine the buffer. As suggested by the manufacturer, doing this can improve and speed up the transition into solution.

[0068] Example 1 and Example 2 The compositions of two eye drop formulations, with or without a preservative, containing lutein solubilized with Vitamin E TPGS in a citrate buffer and having the viscosity enhancer sodium hyaluronate (HA), are described below.

[0069] [Table 3]

[0070] The specific procedures for obtaining the two formulations to be tested are as follows: Step 1 Weigh out deionized water, place it in a suitable container and keep it under stirring. The buffer components, citric acid and sodium citrate, are weighed out one at a time and introduced into the stirred vessel in the order shown. Continue stirring until complete solubilization.

[0071] Step 2 Weigh out Vitamin E TPGS and place in a suitable borosilicate glass container and place on a magnetic stirrer with a heating plate. Insert the magnet into the vessel and start the stirring, not exceeding 500 rpm. Heat Vitamin E-TPGS to 80°C and wait until it changes from a waxy solid to a liquid. Lutein is weighed out in a special container. - Heat the Vitamin E TPGS to 150-160℃, add lutein to the container containing Vitamin E TPGS, and confirm that it is completely dissolved in the Vitamin E TPGS, that the mixture turns deep orange, and that there are no undissolved dark-colored particles. Once all the lutein has been added, increase the stirring speed to 1000 rpm and wait an additional minute to ensure all the lutein has bound to the Vitamin E TPGS and formed stable aggregates.

[0072] Step 3 Add an aliquot of the aqueous solution obtained in step 1, heated to 80°C, in small portions to the oil-phase Vitamin E TPGS-Lutein, previously checked to ensure that its temperature was not lower than 80°C (when the aqueous solution is added, the oil phase changes first, and the translucent orange gel partially turns back into a solid), After adding the entire amount of the solution from step 1, increase the stirring speed from 1000 rpm to 1500 rpm while maintaining the temperature at 150-160°C until there is no undissolved Vitamin E TPGS-Lutein aggregates. Stop stirring and heating only when the orange solution becomes completely clear.

[0073] Step 4 Pour the solution obtained in step 3 into the solution obtained in step 1, and make sure that the solution temperature does not exceed 30℃ before carrying out the following steps. Weigh out the following ingredients: edetate disodium, sodium chloride, sodium hyaluronate, (optionally Microglicin 50), Add freshly weighed ingredients, one at a time, to the mixing vessel in the order listed above. Keep the solution stirred until the newly added ingredient is completely dissolved. · The finished product is 0.2μm filtered and sterile prior to filling into suitable primary packaging.

[0074] Examples 3 and 4 The compositions of two eye drop formulations, with or without a preservative, containing lutein solubilized with Vitamin E TPGS in a citrate buffer and with cross-linked sodium hyaluronate (HA-CK) as a viscosity enhancer, are described below.

[0075] [Table 4]

[0076] The procedures to obtain the two formulations tested are the same as those in Examples 1 and 2, respectively, except for the aforementioned fact that sodium hyaluronate CK is added in step 1, rather than as the final step.

[0077] Examples 5 and 6 The compositions of two eye drop formulations, with or without a preservative, containing lutein solubilized with Vitamin E TPGS in a citrate buffer and having carboxymethylcellulose (CMC) as a viscosity enhancer, are described below.

[0078] [Table 5]

[0079] The procedures for obtaining the two formulations tested were the same as in Examples 1 and 2, respectively.

[0080] Examples 7 and 8 The compositions of two eye drop formulations, with or without a preservative, containing lutein solubilized by Vitamin E TPGS in a citrate buffer and having cross-linked carboxymethylcellulose (CMC-CK) as a viscosity enhancer, are described below.

[0081] [Table 6]

[0082] The procedures for obtaining the two formulations tested were the same as in Examples 1 and 2, respectively.

[0083] Examples 9 and 10 The compositions of two eye drop formulations, with or without a preservative, containing lutein solubilized with Vitamin E TPGS in a citrate buffer and having a carboxyvinyl polymer as a gelling agent, are described below.

[0084] [Table 7]

[0085] The specific procedures for obtaining the two gel formulations are similar to those in Examples 1 and 2, respectively, except for some procedures in steps 1 and 4 described below.

[0086] Step 1 Deionized water is weighed out and introduced into a suitable container and placed under stirring.

[0087] Steps 2 and 3 -Same as Example 1.

[0088] Step 4 Pour the solution obtained in step 3 into the deionized water remaining in step 1, making sure that the temperature of the solution does not exceed 30°C. Weigh out the following ingredients: Disodium edetate, Sorbitol, Sodium hyaluronate, (optionally Microglicin 50), Add freshly weighed ingredients, one at a time, to the mixing vessel in the order listed above. Continue stirring the solution until the newly added ingredient is completely dissolved. Add the carbomer through the sieve in small increments, expected to be wetted and thus dispersed in the monitored solution, increase the stirring speed to about 800-1200 rpm and continue stirring for about another 20 minutes or until a lump-free dispersion is obtained. Add 32% sodium hydroxide (neutralizer) little by little with gentle stirring until a uniform viscous system is obtained. · Subject the finished product to gamma irradiation or autoclaving prior to filling into suitable primary packaging to render it sterile.

[0089] Product Experiments To experimentally prove the presence of lutein in the final product, concentration data for total carotenoids (meaning lutein and zeaxanthin) present in the finished product was obtained from several product samples prepared according to the method of the present invention using a spectrophotometric method utilizing the absorption peak at 446 nm.

[0090] The primary objective of the analysis is to provide experimental data to prove the presence or absence of lutein in the finished product, and a secondary objective is to quantify the lutein so that it can be determined what initial concentration to use to proportionally obtain the desired final concentration in the product.

[0091] The analysis was carried out on three different solutions obtained according to the invention, containing the same initial concentration of lutein equal to 0.005% w / w, but with different excipients.

[0092] The data obtained is shown below.

[0093] [Table 8]

[0094] As previously hypothesized, the results support the assertion that lutein is not removed during the manufacturing process.

[0095] In the finished product, the lutein intended for the raw material, expressed as total carotenoids, averaged about 0.0033%. This result should not be considered relative to the starting amount added to the solution, which was 0.005% w / w, but rather the fact that the total carotenoids in the raw material was equal to 43.92% and not 50%. Thus, an average product loss of about 25% is likely to be observed. The data obtained therefore makes it possible in the present invention to establish upstream the initial percentage of lutein to be used in order to obtain the desired amount in the finished product.

[0096] In conclusion, the present invention makes it possible to add to the performance of conventional topical ophthalmic preparations for artificial tears or products for the treatment of computer vision syndrome the benefits of lutein and vitamin E contained in the corresponding emulsifiers. The resulting composition, in addition to its normal lubricating effect, also exerts a photoprotective effect, acting as a filter for light entering the eye and at the same time exerting an antioxidant effect, since it is also able to neutralize reactive oxygen species (ROS) induced by ultraviolet light.

[0097] Although the present invention has been described with reference to certain specific embodiments thereof, it is understood that a person skilled in the art may make variations or modifications without departing from the scope of protection defined in the appended claims.

Claims

1. A method for producing a water-soluble topical ophthalmic formulation containing lutein, comprising the steps of: Step 1: a) introducing a quantity of deionized water, optionally components of a buffer system, and optionally a quantity of an ophthalmic thickener into a first stirred vessel and waiting for the optional components to completely solubilize; Step 2: a) heating a quantity of Vitamin E TPGS in a second stirred vessel to a temperature of 80-90° C. and waiting for it to completely melt; b) bringing the Vitamin E TPGS to a temperature of 90-170° C. and adding a predetermined amount of lutein while continuing to stir until the Vitamin E TPGS and lutein are completely dissolved to form a homogeneous orange oily solution consisting of only the Vitamin E TPGS and lutein; Step 3: a) heating the water or aqueous solution obtained in step 1 operation a) to a temperature of 80-90° C. and slowly adding an aliquot thereof equal to 10% by volume of the final product to the second stirred vessel while keeping it under stirring; b) once the addition is complete, increase the stirring speed while maintaining the temperature in the range of 90-170°C until a completely clear, optically homogeneous orange liquid is obtained; Step 4: a) adding the solution of step 3 to the remainder of the water or aqueous solution obtained in operation a) of step 1; b) after confirming that the temperature has dropped below 30°C, add other water-soluble ingredients of the topical ophthalmic formulation to be produced, including one or more thickeners and / or mucoadhesives, one at a time in a predetermined amount to the stirring vessel and wait for complete solubilization; c) Sterilizing the product thus obtained.

2. 2. The method of claim 1, wherein the topical ophthalmic formulation is in liquid form and in step 1, operation a), in addition to the deionized water, the components of the buffer system are also introduced into the first stirring vessel.

3. The method of claim 2, wherein a component of the buffer system is one of a borate, a citrate, and a phosphate buffer.

4. The method according to any one of claims 1 to 3, wherein in operation a) of step 1, a predetermined amount of cross-linked hyaluronic acid (HA-CK) is also introduced into the first stirred vessel.

5. The method according to any one of claims 1 to 4, wherein the one or more thickening agents and / or mucoadhesives of operation b) of step 4 comprise hyaluronic acid or its salts (HA), or carboxymethylcellulose (CMC) or cross-linked carboxymethylcellulose (CMC-CK).

6. The method according to any one of claims 1 to 5, wherein the sterilization in operation c) of step 4 is carried out by filtering the obtained product through a 0.2 μm filter.

7. 2. The method of claim 1, wherein the topical ophthalmic formulation is in gel form, and in step 4, operation b), a predetermined amount of gelling agent is added to the stirring vessel through a sieve, waiting for the gelling agent to soak and thus disperse, continuing stirring until a lump-free dispersion is obtained, and then neutralizing the dispersion with a neutralizing agent while continuing the stirring gently.

8. The method of claim 7, wherein the gelling agent is a carboxyvinyl polymer (carbomer).

9. 9. The method according to claim 7 or 8, wherein the sterilization in operation c) of step 4 is carried out by subjecting the obtained product to autoclaving or treatment with gamma radiation.

10. A topical ophthalmic formulation consisting of a stable micellar solution (microemulsion) of Vitamin E TPGS and lutein suspended in an aqueous phase containing a thickening and / or mucoadhesive or gelling agent, and other ophthalmically acceptable excipients.

11. 11. The topical ophthalmic formulation of claim 10, wherein the thickening agent and / or mucoadhesive agent is selected from the group consisting of hyaluronic acid or its salts (HA), cross-linked hyaluronic acid (HA-CK), carboxymethylcellulose (CMC), cross-linked carboxymethylcellulose (CMC)-CK) and mixtures thereof.

12. 11. The topical ophthalmic formulation of claim 10, wherein the gelling agent is based on a carboxyvinyl polymer (carbomer).

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