Diabetic wound healing ointment with hypnum curvifolium bryophyte extract and chlorella vulgaris microalgae enriched with tuna waste oil

Abstract

This invention involves a bioactive ointment for promoting diabetic wound healing which speeds up treatment process significantly. The ointment utilizes Hypnum Curvifolium bryophyte extract, Chlorella Vulgaris microalgae and fatty acids from tuna waste oil. Initially the hypnum bryophyte extract and microalgae are prepared via ultrasound. Next, the oil is extracted from the tuna waste using the Blythe-Dyre method. After separating the unsaturated fatty acids from the tuna waste oil, the hydrogel base is prepared. The proposed solution includes the step by step preparation method of the ingredients and the finalized obtained formula.

Description

Diabetic Wound Healing Ointment with Hypnum Curvifolium Bryophyte Extract and Chlorella Vulgaris Microalgae Enriched with Tuna Waste Oil

[0001] The development of healing ointment for diabetic wounds using Chlorella microalgae agents, fatty acids and hypnum curvifolium bryophyte is a groundbreaking innovation in the medical field. By harnessing the potent healing properties of microalgae fatty acids and hypnum bryophyte extracts, this proposed composition can create a powerful ointment that accelerates the healing process of diabetic wounds. These natural ingredients are rich in antioxidants and anti-inflammatory compounds, which help reduce inflammation, promote tissue regeneration, and prevent infections. The ointment provides a safe and effective alternative to traditional wound care methods, offering diabetic patients a faster and more efficient way to heal their wounds. This innovative approach has the potential to revolutionize diabetic wound care and improve the quality of life for millions of patients worldwide.

[0002] A61K 36 / 10 – A61L 15 / 36 – C11C 3 / 08

[0003] US9668966B2

[0004] Cosmetic compositions comprising microalgal components

[0005] The invention provides cosmetic compositions comprising microalgal biomass, whole microalgal cells, and / or microalgal oil in combination with one or more other cosmetic ingredients, and methods of making such compositions. In preferred embodiments, the microalgal components of the cosmetic compositions are derived from microalgal cultures grown heterotrophically and which comprise at least 10% oil by dry weight.

[0006] The mentioned patent shares certain qualities with our claimed one in terms of ingredients and components such as chlorella vulgaris microalgae and fatty acids. However, the method of production is different in the two patents and they feature other ingredients that set them apart. Moreover, this mentioned composition does not specify a final product while ours focuses primarily on diabetic wound healing via topical application.

[0007] US9511034B1

[0008] Method for applying a skin treatment

[0009] The invention is generally a skin treatment for managing skin conditions ranging from scars, including keloids, to burns and wrinkles. The skin treatment may comprise of several components including a cleansing solution, a silicone elastomer, and a mineral powder that may be utilized to treat and camouflage the affected skin. In one embodiment, a cleansing component is used to prime a skin area, cleansing and treating the area so that a silicone component can heal the skin.

[0010] This mentioned invention addresses skin conditions including scars and offers a treatment solution using ingredients that resemble ours. However, the scope of this patent is wide and the developed solution can be used for a variety of conditions. On the other hand, our solution is carefully designed to speed up the healing process of diabetic wounds and features a special formula.

[0011] US20160256480

[0012] THERAPEUTIC COMPOSITION FOR WOUND HEALING

[0013] A therapeutic composition comprising a beta-glucan, at least one fatty acid, and at least one secondary polysaccharide, which may increase a rate of wound healing. A method for healing a wound in a subject, by administering to a subject in need thereof, the therapeutic composition as described.

[0014] This invention involves a wound healing composition which uses fatty acids and shares a general purpose with ours. But when It comes to specifics of the production process and comprising elements, the two patents are different. For instance, this design relies on bacterial agents while our solution includes microalgae and bryophytes extract in its formulation which is enriched with fish waste oil.

[0015] United States Patent 8691298

[0016] Stabilized formulation comprising omega-3 fatty acids and use of the fatty acids for skin care and / or wound care

[0017] A stabilized formulation for skin care, wound care and / or other tissue healing applications and methods for making the same is described. The stabilized formulation stabilizes omega-3 polyunsaturated fatty acids and is constituted of the omega-3 polyunsaturated fatty acids in combination with tocopherol (Vitamin E), ascorbic acid (Vitamin C), herb extract, and a fat-soluble antioxidant. Methods for making and using the stabilized formulation are also described.

[0018] The above mentioned invention involves a wound healing agent with a similar purpose to our patent which also addresses diabetic ulcers. While both patents utilize fatty acids from fish, their composition includes a variety of ingredients that make them different. For instance, ours prepares fatty acids from fish waste oil, whereas, this one includes omega-3 polyunsaturated fatty acids derived from various sources. Moreover, our formula has a primary focus on diabetic wound treatment, unlike this patent which has a wider scope in the field of skin care.

[0019] This invention introduces a bioactive ointment specifically designed to accelerate the healing process of diabetic wounds, offering a solution to a common medical challenge. The key components of this advanced ointment include extracts from the Hypnum Curvifolium bryophyte, Chlorella Vulgaris microalgae, and fatty acids sourced from tuna waste oil.

[0020] The preparation process begins by subjecting the hypnum bryophyte extract and microalgae to ultrasound, effectively enhancing their bioactive properties. Subsequently, the oil is extracted from tuna waste using the Blythe-Dyre method, ensuring the purity and potency of the fatty acids. Through a meticulous process, the unsaturated fatty acids are carefully separated from the tuna waste oil.

[0021] Then a hydrogel base is meticulously prepared, serving as a stable and effective carrier for the potent extracts and fatty acids. The step-by-step preparation method outlined in this invention provides clarity and guidance for creating the final formula, ensuring consistency and efficacy in every batch produced. Overall, this innovative bioactive ointment represents a breakthrough in diabetic wound treatment, offering a safe, efficient, and cost-effective solution.

[0022] The importance of diabetes is due to its prevalence, long course and complications. The prevalence of this disease is increasing in developed and industrialized countries as well as in developing countries. According to a documented report, approximately 10% of the world has diabetes (Kar et al., 1999).

[0023] Chronic lower limb ulcers are a major clinical challenge that become more prevalent with increasing age and increasing risk factors, including smoking, obesity and diabetes (Hardwicke et al., 2011). In this regard, diabetes is of particular importance due to its severe complications such as diabetic nephropathy, neuropathy, retinopathy and diabetic ulcers (Gabir et al., 2000).

[0024] Although insulin and other chemical treatments can control the disease to some extent, the development of wounds and burns is associated with delayed healing due to the persistence of tissue inflammation and the presence of inflammatory cells at the wound site, the disorder of the extracellular matrix, and the delay in epithelial regeneration (Hardwicke et al., 2011). For this reason, delayed wound healing has become one of the greatest therapeutic challenges for physicians and patients, with approximately 15% of patients with diabetes, due to prolonged hospitalization or poor healthcare, eventually leading to amputation (Guo and DiPietro, 2010).

[0025] The treatment of wounds, especially for long-term care, can impose significant costs on healthcare systems. For example, diabetic wounds cost the US healthcare system more than $9 billion annually to treat. Conflict-related injuries, such as amputations, can lead to lifelong rehabilitation costs and impose a significant economic burden on governments, so treating diabetic wounds is of utmost importance.Solution of Problem

[0026] Aquatic animals are rich in nutrients such as proteins, minerals, vitamins, lipids, and especially a significant amount of unsaturated omega-3 and -6 fatty acids, which can be very effective in maintaining the health, strength, and stability of skin and hair.

[0027] Extraction of fatty acids, beta-carotene, and carotenoids from bryophyte and microalgae has made them a rich and accessible source for the production of cosmetic and health products. β-carotene is the most important pigment in the carotenoid family, which prevents the formation of active oxygen and is used as provitamin A in the formulation of anti-aging products.

[0028] Polysaccharides, fatty acids, collagen, hyaluronic acid, alguronic acid are bioactive compounds from marine sources that lead to the synthesis of elastin in the skin, in other words, skin repair. Since the microalgae Chlorella is rich in bioactive compounds and considering the effectiveness of fatty acids, which are abundant in tuna waste, it seems that the simultaneous use of these two can be very effective in wound treatment. So far, efforts to accelerate wound healing have not led to the introduction of an effective drug, and research is still ongoing. Since the microalgae Chlorella contains fatty acids, especially omega, it can be very effective in wound treatment.

[0029] This invention aims to introduce a healing ointment, specifically designed to address prior art shortcomings in the field. Although, the use of aquatic sources and bryophytes has been part of the development process for healing agent compositions such as ours, there are factors that set apart this innovative endeavor. By using natural ingredients and relying on the power of microorganisms, this solution is presented in order to speed up the treatment process of skin issues for diabetic patients.

[0030] The claimed patent involves a bioactive wound dressing composition which can be highly effective in topical treatment of diabetic wounds. The process of achieving this unique composition is discussed here step by step.

[0031] The initial phase involves preparation of the microalgae agent which includes the extraction of dried samples were prepared from ground powder using the ultrasonic extraction method as follows.

[0032] To begin, 18.165 g of Tris solution was combined with 762.8 g of Merck sodium chloride and dissolved in 800 ml of distilled water. The acidity of the solution was adjusted to 7 to 7.5 using hydrochloric acid. The prepared solution was autoclaved at 121°C for 15 minutes to prepare the desired buffer. The prepared solution was stored in the refrigerator until use.

[0033] In order to prepare the algal extract, 10 grams of microalgae were added to every 200 cc of buffer prepared. Then, the ultrasonic device was placed in an ice bath at a frequency of 20 kHz, in 10 cycles for 3 minutes with an interval of 1 minute. Finally, the prepared extract was transferred to a centrifuge and placed at 9000 rpm until the remains of fragmented and crushed microalgae cells that had been created in the ultrasonic part of the process were deposited.

[0034] The next phase involves optimal oil extraction from tuna waste which typically consists of minced entrails and viscera. The extraction can be performed using the Blythe and Dyer method as follows: 2000 grams of minced entrails and viscera are mixed well with 1100 ml of chloroform and 2000 ml of methanol for 2-4 minutes, then 1000 ml of distilled water is added and the mixture is stirred for another 30 seconds and kept in the dark at 25°C for 72 hours.

[0035] The filtrate was transferred to a decanter. Separation took place after the two phases were completely separated from each other. The upper layer contained water and methanol and the lower layer contained chloroform, which contained purified fat. The chloroform layer contained oil, which was separated from the oil after evaporation. The concentration of the extracted oil was calculated based on [Formula. 1].

[0036] The next step entails the separation and identification of fatty acids (optimized oil) from waste fish oil. In order to separate unsaturated fatty acids, 80 grams of pure oil prepared from tuna waste was used. 68 ml of distilled water, 20 grams of potash and 108 ml of ethanol were added to the above sample and it was placed in a mantle heater for 2 hours. After the above time, pure hydrochloric acid with a molecular weight of 36.45 g / mol in an amount of 27.6 ml, 200 ml of distilled water, 740 ml of ethanol and 200 grams of urea were added to it and it was placed in a decanter for 20 minutes until the above solution became two phases, then the phase containing the urea complex was removed from it. After that, it was placed at room temperature for 24 hours and another 24 hours at 1°C.

[0037] After the aforementioned time, 17.6 ml of hydrochloric acid and 1440 ml of distilled water were added. The fat was placed on top. Then, 38 ml of ethanol and 0.6 ml of sulfuric acid were added to it and it was left to rest for 2 hours. After the aforementioned time, 80 ml of distilled water was added and it was placed at -30°C for 24 hours. After that, petroleum benzene was added to the upper layer, passed through a filter, and placed in a rotary centrifuge. Finally, what remained were the fatty acids.

[0038] In order to prepare the hydrogel ointment, Carbomer 940 (acrylic polymer), which is a polyvinyl carboxy polymer, was used and the formula was prepared as follows:

[0039] Microalgae extract 15% + bryophyte extract Hypnum curvifolium 15% + fish oil 5% + hyaluronic acid 1% + collagen 1% + elastin 1%.Advantage Effects of the Invention

[0040] • Cost-effective production

[0041] • Speeding up the treatment process

[0042] • Minimizing treatment expenses

[0043] • Minimizing production waste

[0044] • Safe and durable

[0045] • Comprehensive and accessible method

[0046] • Tested and proven effective

[0047] Shows a general flowchart of the production process.

[0048] Declares a flowchart of the algal extract preparation.

[0049] Displays a flowchart of tuna waste oil preparation.

[0050] Depicts a flowchart of hydrogel preparation.

[0051] Shows a flowchart of the production process for the claimed wound healing ointment. The ingredients and components are prepared in different stages to achieve the desired solution.

[0052] Shows a flowchart of the preparation of microalgae and sheet moss bryophyte extract via ultrasound as one of the main ingredients of the claimed topical ointment.

[0053] Shows a flowchart of the preparation of tuna waste oil extract via the Blythe and Dyre method. The separation of unsaturated fatty acid is an important step in obtaining the final product.

[0054] Shows a flowchart of the hydrogel preparation from Carbopol 940 (acrylic polymer), which is a polyvinyl carboxy polymer.Examples

[0055] The invention was tested in a standard setting and the results were highly effective. Three reports of the treatment process and the progress of the wound healing are included here. Photomicrographs were prepared to show the healing wound area on different days using Masson's trichrome stain.

[0056] In the macroscopic examination on the fourth day of the experiment, no infection was observed in the wound, and the microscopic results of the tissue analysis also confirm it because the amount of polymorphonuclear cells and mononuclear cells in each case is between 5 and 25, and no ulcer (tissue inflammation) is observed and complete destruction of the epithelial tissue has not occurred. In this sample, collagen synthesis is observed moderately in all wounds and densely in some areas, and vascularization is observed moderately everywhere, and epithelial cell migration is less than 50%, which is a sign of wound healing.

[0057] In the macroscopic examination on the eighth day of the experiment, no infection was observed in the wound, and the microscopic results of the tissue analysis also confirm it because the amount of polymorphonuclear cells and mononuclear cells in each case is between 5 and 25, and no ulcer (tissue inflammation) is observed and complete destruction of the epithelial tissue has not occurred. In this sample, collagen synthesis is observed in an average manner in all wounds and in some places in a dense manner. Vascularization is observed in cases and the migration of epithelial cells is more than 50%, which is a sign of wound healing.

[0058] In the macroscopic examination on the 12th day of the experiment, the effects of the wound were observed carefully, the amount of polymorphonuclear cells, mononuclear cells in each case was between 5 and 25, and no ulcer (tissue inflammation) was observed and complete destruction of the epithelial tissue was not done. In this sample, collagen synthesis was observed in an average manner in all the wounds and in some places in a dense manner. Vascularization was seen mildly and the epithelial tissue epithelium was formed in a unified and bridged manner.

[0059] In conclusion, a sample was successfully treated with the developed hydrogel ointment.

[0060] [Formula. 1] The formula used to determine the extracted oil percentage is listed here:

[0061] [Formula. 1]

[0062] This invention will be very widely used in the field of pharmaceuticals by companies that value cost-effective and tested solutions. Health and hygiene facilities, clinics and institutes could all benefit from the production and implementation of this wound healing ointment. It could also be manufactured as an OTC topical gel and become a household item for those dealing with the impact of diabetes.

Claims

1.A bioactive topical ointment is developed for the purpose of promoting diabetic wound healing which utilizes Hypnum Curvifolium bryophyte extract and Chlorella Vulgaris microalgae enriched with unsaturated fatty acids as the main components.2.According to claim 1, the claimed ointment preparation method includes obtaining the hypnum curvifolium bryophyte extract and chlorella vulgaris microalgae via ultrasound.3.According to claim 1, the claimed ointment preparation process includes obtaining oil from fish waste via the Blythe and Dyre method.4.According to claim 3, the obtained composition is further processed in order to identify and separate the unsaturated fatty acids from the tuna waste oil.5.According to claim 1, the claimed ointment preparation method includes making the hydrogel base with acrylic polymer.6.According to claim 1, the claimed wound healing ointment is prepared via this developed formula: Microalgae extract 15% + bryophyte extract Hypnum curvifolium 15% + fish oil 5% + hyaluronic acid 1% + collagen 1% + elastin 1%.

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