Synthesis and applications of multifunctional biopolymers from coconut husk and / or coiapis powder

Abstract

We provide novel, naturally biocompatible, multifunctional biopolymers that can be used in pharmaceuticals, nutritional supplements, cosmetics, and / or leather industry applications. [Solution] A natural biocompatible multifunctional biopolymer comprising: a) lignin extracted from coconut husk and / or koiapis powder obtained from the fruit of Cocos nucifera, a tropical plant belonging to the family Palmae; b) copolymer; c) crosslinking agent; d) plasticizer; and e) at least one activator.

Description

[Technical Field]

[0001] This invention relates to the extraction of lignin from coconut husk and / or coiapis powder. The extracted lignin is purified, and the physicochemical properties of the purified lignin are studied. This lignin is further processed with copolymers, crosslinking agents, and plasticizers to produce biopolymer sheets / films. These sheets / films exhibit multifunctional properties and have been found to be effective for wound dressing in severe burn injuries. These sheets / films have also been used in the development of leather substitute materials. The developed biopolymer sheets / films have been evaluated for their properties and have been found to be promising for a variety of applications. [Background technology]

[0002] Coconut husk and / or coiapis powder is a versatile and environmentally friendly material that is gaining popularity for a variety of applications. These natural lignocellulose fibers are obtained from the fruit of Cocos nucifera, a tropical plant of the palm family (Palmae). Its unique properties make it ideal for a wide range of applications, and its sustainable sourcing makes it an attractive option for those who prioritize environmental responsibility. Coconut husk and / or coiapis powder is produced globally, primarily in Indonesia, the Philippines, India, and Sri Lanka. Coconut fiber stands out as the most prominent fibrous by-product of coconut cultivation, with an annual global production of at least 30 million tons. While most parts of the coconut are used in food, textiles, and other industries, the husk portion is mainly used for crafts and decorative items, and much of it remains unused, with its plant components largely untapped in the pharmaceutical field. According to Panyakaew and Fotios (2011), coconut husk consists of 30% fiber and 70% pith, and boasts high lignin and phenol content. Van Dam et al. (2006) concluded that coconut husk contains 35% cellulose, 35% lignin, and 17% hemicellulose (see Figure 1). Due to its high lignin content, coconut fiber is highly elastic, durable, and rot-resistant.

[0003] Lignin is a highly beneficial complex organic polymer that plays a crucial role in the structure of cell walls, particularly wood and bark. Lignin provides essential rigidity and resistance to decay, making it an excellent building material. In fact, among lignocellulosic biomass, lignin is the second most abundant biopolymer after cellulose. Furthermore, lignin forms strong and durable composites with cellulose and hemicellulose, resulting in outstanding strength, flexibility, and high abrasion resistance, making it extremely useful in various industrial applications. Lignin is primarily composed of phenolic monomers such as coniferyl, cinapyr, and p-coumaryl alcohol, which are interconnected via various carbon-carbon and carbon-oxygen bonds, forming a highly branched and irregular structure. This complex network of aromatic rings and hydrocarbon chains contributes to lignin's strength and elasticity, making it essential for plant growth and a valuable resource for many industrial applications.

[0004] Lignin, a highly complex organic polymer found in plant cell walls, plays a crucial role in many industries due to its outstanding properties. Lignin is an impressive biopolymer that plays a vital role in improving the structural support, rigidity, and durability of plant tissues. While cellulose and hemicellulose have traditionally received more attention in biomass utilization, lignin's unique and complex properties have recently attracted interest due to its potential applications in sustainable materials, bioenergy, and various industrial sectors. In the paper and pulp sector, lignin, previously considered a by-product of waste, is now used to produce valuable chemicals such as vanillin. It also serves as a sustainable adhesive for wood products, thereby reducing reliance on synthetic resins. In the field of bioplastics and composite materials, lignin enhances mechanical properties and contributes to biodegradability, making it particularly beneficial in the production of automotive parts. Furthermore, in agriculture, lignin derivatives are used as soil conditioners and controlled-release fertilizers, improving utility while minimizing environmental impact. In the pharmaceutical industry, lignin's antioxidant and antimicrobial properties contribute to improved drug stability and delivery. Furthermore, in the cosmetics industry, lignin is valued for its UV protection properties in sunscreens and anti-aging products. Additionally, lignin can be converted into biofuels, thus contributing to renewable energy efforts. The inventors' previously filed patent PCT / KR2022 / 019441 expands its applications to include nutritional supplements and cosmetics. Finally, its unique chemical structure and properties present increasingly important untapped potential in various new fields, particularly in the development of biopolymers.

[0005] The leather, pharmaceutical, and cosmetics industries are seeing increased demand for novel biopolymer materials based on natural resources due to their sustainability, regulatory compliance, and improved functional properties. Consumers and governments are increasingly calling for environmentally friendly, biodegradable materials as alternatives to conventional synthetic polymers to mitigate environmental impact and health risks. Naturally derived biopolymers, such as lignin, offer outstanding advantages such as biocompatibility, antimicrobial properties, and customizability, making them suitable for a variety of applications, including sustainable leather alternatives, advanced drug delivery systems, and natural skincare products. This shift towards natural biopolymers aligns with a global trend toward market diversification, technological innovation, and more sustainable industrial practices.

[0006] Lignin-based biopolymers offer innovative and sustainable solutions across various industries. In the leather industry, they can serve as environmentally friendly alternatives while maintaining high quality and durability. The pharmaceutical industry can benefit from lignin's biocompatibility and functional properties, making it ideal for drug delivery systems, wound dressings, and other medical applications. In the cosmetics industry, incorporating lignin-derived biopolymers into skincare products can provide natural ingredients with antioxidant and UV protection properties, enhancing product efficacy and sustainability.

[0007] In the leather industry, animal hides present numerous disadvantages, including the slaughtering process carried out to obtain the hides. Furthermore, there are concerns about environmental pollution due to the large amounts of waste and carbon dioxide emitted by the livestock industry for leather production. The use of harmful substances such as chromium in the leather manufacturing process not only harms the health of manufacturers but also contributes to environmental pollution when discharged with wastewater. On the other hand, lignin can improve leather processing and finishing by enhancing texture, appearance, and durability, and providing natural antioxidant protection. Lignin also acts as a natural dye and color stabilizer, as well as a biodegradable adhesive, potentially contributing to more environmentally friendly manufacturing processes. Overall, the use of lignin in leather production has the potential to promote sustainability and high-quality leather products.

[0008] Burns are one of the most common types of injuries, after traffic accidents, falls, and physical violence. In the early 20th century, treatment options for burn patients were extremely limited, and deaths due to hypovolemic shock on the first day after injury were frequent. However, the latter half of the 20th century saw remarkable progress in regenerative medicine, burn treatment, and drug therapy. Despite these advances, effective management of burns remains challenging. Burn recovery is a complex and lengthy process that relies on diverse repair mechanisms mediated by the immune system. It is necessary to restore the continuity of tissue damaged by unexpected events such as fires. To achieve complete recovery, burns can progress through three stages: inflammation, granulation tissue formation (proliferation), and remodeling (reshaping), before scar formation. The patient's immune system plays a crucial role in regulating the entire wound healing process. In the initial stages of wound healing, injuries are most susceptible to bacterial infection. This infection can drastically reduce healing and, if left untreated, can be fatal. To overcome this problem, many types of wound dressings have been tried, including surface dressings, hydrogels, hydrocolloids, foams, fibers, and films. However, each has been found to have its own set of drawbacks, such as issues with biocompatibility, dressing properties, ease of application, dressing abrasion, shape and size, activity against pathogens, and drug delivery capacity. To address these problems, the development of innovative biocompatible polymers was necessary.

[0009] Biopolymer sheets and films primarily composed of lignin have the potential to offer significant advantages over conventional materials for burn and wound dressings. Their natural antimicrobial and antioxidant properties help prevent infection and reduce oxidative stress, promoting faster healing. These biopolymers can form hydrogels, creating a moist healing environment while managing excess exudate, facilitating cell migration, tissue regeneration, and necrotic tissue removal. These processes are crucial for optimal wound healing outcomes. Their biocompatibility also reduces the risk of allergic reactions and minimizes potential inflammation. Furthermore, lignin can be used as a flexible and durable dressing, protecting wounds from contaminants and enabling controlled drug delivery. In other words, lignin-based biopolymers have the potential to deliver therapeutic agents such as antimicrobials and growth factors directly to wound sites in a controlled manner, enabling sustained treatment. This targeted drug delivery approach aims to enhance therapeutic efficacy while minimizing systemic side effects. Additionally, lignin is biodegradable and cost-effective, making it environmentally friendly and suitable for sustainable healthcare. Overall, lignin-based biopolymer dressings offer an attractive and versatile solution for advanced wound care, contributing to improved patient outcomes and reduced environmental impact.

[0010] This invention discloses the development of various lignin-based biopolymer blends using lignin extracted from coconut husk and / or coiapis powder to prepare biopolymer sheets and / or films for burn healing / wound dressings and as leather substitute materials. The lignin polymers are produced by oxidative radical polymerization of three main p-hydroxycinnamyl alcohols known as monolignols, namely p-coumaryl alcohol, coniferyl alcohol, and synapyl alcohol. These alcohols are involved in the formation of p-hydroxyphenyl (H), guaiacyl (G), and syringyl (S) units and are incorporated into the lignin polymers.

[0011] Broad-leaved trees consist of syringyl (S) and guaiacyl (G) units, while coniferous trees consist mainly of guaiacyl (G) units and have a small amount of p-hydroxyphenyl (H) units (Figure 2). [Overview of the Initiative] [Problems that the invention aims to solve]

[0012] The main objective of the present invention is to extract lignin from coconut husk and / or coiapis powder obtained from the fruit of Cocos nucifera, a tropical plant belonging to the palm family, and to develop a novel natural biocompatible multifunctional biopolymer that can be used in pharmaceutical, nutritional supplement, cosmetic, and / or leather industry applications.

[0013] Another object of the present invention is to extract and purify lignin components from coconut husk and / or coiapis powder, investigate their physicochemical properties, and process them into various biopolymer sheets / films / gels using other additional copolymers, crosslinking agents, and plasticizers.

[0014] Another object of the present invention is to develop a biopolymer sheet / film / gel for victims of severe burn accidents, the protective wound dressing layer comprising 1) lignin extracted from coconut husk and / or coiapis powder, and 2) α-lactalbumin extracted from whey protein isolate, and 3) a combination of additional other copolymers, crosslinkers, and plasticizers. Due to its antimicrobial, antioxidant, and moisturizing properties, the lignin biopolymer may be usable for wound dressing in severe burn accidents, particularly when combined with APIs such as recombinant human growth factors, as it restores new growth to replace damaged burn tissue. The lignin biopolymer can be further utilized for the healing and repair of damaged tissue through diverse immune system-mediated repair mechanisms and the regeneration and replacement of damaged burn tissue with new growth.

[0015] Another object of the present invention is to provide a leather substitute material that can be converted to leather (non-animal source) using a leather reinforcing material obtained from coconut husk and / or coiapis powder fibers, particularly as a leather substitute, to exhibit excellent abrasion resistance and flexibility. [Means for solving the problem]

[0016] As an agricultural waste, coconut husk and / or coiapis powder has proven to be a rich source of lignin, as well as cellulose. The solubility of lignin can be improved by adjusting the extraction and drying methods, pH, and temperature. Lignin has been successfully converted into multifunctional biopolymer sheets / films / gels by adding various copolymers, crosslinking agents, and plasticizers, although the effects of each additive have only been investigated at a basic level. This invention develops multifunctional biopolymer sheets / films / gels and studies their application effects in terms of the manufacturing process and resulting quality of wound dressings and leather substitutes, respectively.

[0017] Lignin is a remarkable organic polymer with diverse chemical components, making it highly suitable for burn and wound healing dressings. Its phenolic compounds possess natural antimicrobial and antioxidant properties, effectively preventing infection and reducing oxidative stress to promote healing. The major monolignols, coniferyl alcohol, cinapyr alcohol, and coumarill alcohol, play a crucial role in maintaining lignin's structural integrity and antimicrobial activity, supporting tissue repair. Methoxy groups enhance chemical reactivity, enabling the development of customized derivatives for improved moisturizing properties and controlled drug release. Furthermore, carboxylic acid groups increase hydrophilicity, leading to the formation of hydrogels that maintain a moist healing environment and enhance therapeutic agent binding capacity. In addition, lignin-carbohydrate complexes promote biocompatibility, while quinones introduce redox properties that help address infection and immunity, making lignin a versatile and highly effective material for advanced wound healing solutions.

[0018] On the one hand, alpha-lactalbumin has been reported to play an excellent role in promoting the regeneration of new skin on burns, leading to faster healing of the burn site, by stimulating the proliferation and adhesion of fibroblasts and promoting angiogenesis by increasing local serotonin concentration. Alpha-lactalbumin supports the supply of essential amino acids necessary for protein synthesis and tissue repair, especially at the highest levels of tryptophan. Furthermore, it modulates the immune response by minimizing excessive inflammation and controlling cytokine production, and helps to promote collagen synthesis for structural integrity. Also, when added to a burn dressing containing lignin, alpha-lactalbumin also helps to further improve the moist wound environment essential for cell migration and tissue formation, while the antioxidant properties of lignin relieve oxidative stress and further promote efficient skin regeneration.

[0019] In addition to the above uses for burn dressings, alpha-lactalbumin can be incorporated into multifunctional lignin biopolymers that constitute prototype leather substitute materials derived from the fibers of coconut husk and / or coirapis powder, regardless of the presence of additional leather reinforcing materials. The resulting leather substitute can provide excellent abrasion resistance from lignin and a soft, enveloping flexibility from alpha-lactalbumin if an ideal composition is provided.

[0020] More specifically, the present invention relates to a natural biocompatible multifunctional biopolymer comprising a) lignin extracted from coconut husk and / or coirapis powder obtained from the fruit of Cocos nucifera, a tropical plant belonging to the family Palmae, b) a copolymer, c) a crosslinking agent, d) a plasticizer, and e) at least one activator.

[0021] In other embodiments, the present disclosure relates to novel natural biocompatible multifunctional biopolymers in which at least one copolymer is selected from either natural polymers or water-soluble synthetic polymers. Natural polymers include hyaluronic acid and its derivatives, starch, modified starch, alginates, chitosan, chitin, natural gums, gelatin, collagen, casein, zein, gluten, soy protein isolate, whey protein isolate and other proteins, terrestrial plant extracts such as pectin, and combinations thereof. Water-soluble synthetic copolymers include cross-linked polyacrylic acid chains, polyvinylpyrrolidone, polyvinyl alcohol, modified cellulose ethers such as hydroxypropylmethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, carboxymethylcellulose, ion exchange resins, and combinations thereof. The effective amount of the copolymer ranges from about 10% to 90% by dry weight, preferably 30% to 70% by dry weight, depending on the use of the developed biocompatible multifunctional biopolymer.

[0022] In other embodiments, the present disclosure relates to novel natural biocompatible multifunctional biopolymers in which at least one cross-linking agent is selected from glutaraldehyde, formaldehyde, polyethylene glycol (PEG) and its derivatives, genipin, dextran, polyethyleneimine (PEI), ethylenediamine (EDA), and sugar alcohols / polyols such as erythritol, xylitol, maltitol, mannitol, lactitol, polyglycitol, isomalt, sorbitol, glycerol, and combinations thereof, which promote cross-linking chemical reactions. The effective amount of the cross-linking agent ranges from about 1% to 10% by dry weight, preferably 2% to 5% by dry weight, depending on the use of the developed biocompatible multifunctional biopolymer.

[0023] In other embodiments, the disclosure relates to novel naturally biocompatible multifunctional biopolymers in which at least one plasticizer is selected from sucrose ester derivatives, glycerin, propylene glycol, polyethylene glycol, magnesium stearate, animal fat, neatfoot oil, lanolin, mineral oil, silicone oil, jojoba oil, castor oil, coconut oil, whale oil, squalene, grapeseed oil, safflower oil, canola oil, almond oil, and combinations thereof. The effective amount of plasticizer is in the range of about 0.5% to 10% by dry weight, preferably 1% to 3% by dry weight, depending on the application of the developed biocompatible multifunctional biopolymer.

[0024] In other embodiments, the disclosure relates to novel naturally biocompatible multifunctional biopolymers in which the emulsifier is optionally selected from xanthan gum, lanolin, sodium alginate, lecithin, casein, whey protein, pectin, ammonium phosphatide, acacia, and oleylamine. The effective amount is in the range of approximately 0.5 to 5% (w / w), depending on the application of the developed biocompatible multifunctional biopolymer.

[0025] In other embodiments, the disclosure relates to novel naturally biocompatible multifunctional biopolymers in which the emulsifier is optionally selected from xanthan gum, lanolin, sodium alginate, lecithin, casein, whey protein, pectin, ammonium phosphatide, acacia, and oleylamine.

[0026] In other embodiments, the disclosure relates to a novel natural biocompatible multifunctional biopolymer used as a wound dressing or as part of a wound dressing on severe burns in patients to restore new tissue growth on burns and replace damaged tissue, wherein at least one activator may be loaded from turmeric extract (curcumin), honey, green tea extract, gotu kola extract (Centella asiatica), α-lactalbumin, whey protein isolate, comfrey extract, tea tree oil, lavender oil, chamomile extract, calendula extract, aloe vera gel, silver sulfadiazine, bacitracin, hyaluronic acid, collagen, vitamin E, allantoin, sphingosine- / phytosphingosine-1-phosphate, and deexpanthenol (vitamin B5), and combinations thereof. The effective amount of at least one activator ranges from about 0.01% to 50% by dry weight, preferably 1% to 20% by dry weight, depending on the application and the severity of the burn.

[0027] In other embodiments, the disclosure relates to a novel, naturally biocompatible, multifunctional biopolymer used as a wound dressing or as part of a wound dressing on severe burns in patients to restore new tissue growth on burns and replace damaged tissue, wherein at least one activator is epidermal growth factor (EGF), growth hormone / somatotropin (hGH / STH), vasoactive intestinal peptide (VIP), fibroblast growth factor family proteins, namely fibroblast growth factor-1 (FGF1), fibroblast growth factor-2 (FGF2), fibroblast growth Growth factor-7 / keratinocyte growth factor (FGF7; KGF), fibroblast growth factor-10 / keratinocyte growth factor-2 (FGF10; KGF2), fibroblast growth factor-19 (FGF19), fibroblast growth factor-21 (FGF21), fibroblast growth factor-23 (FGF23), etc., vascular endothelial growth factor (VEGF), thymosin beta-4 (TMS), stem cell factor / hematopoietic stem cell factor (SCF), platelet-derived growth factor (PDGF), prolactin (PRL), placental lactogen (hPL), nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), etc. Rear cell-derived neurotrophic factor (GDNF), ciliary neurotrophic factor (CNTF), interleukin-1 receptor antagonist (IL1Ra), interleukin family proteins, namely interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-13 (IL-13), interleukin-17 (IL-17), interleukin-18 (IL-18), adiponectin (ApN), TSLP (thymic interstitial lymphapoietin), and bone morphogenetic protein family proteins, namely BMP-2, BMP-4, BM P-7, etc., cluster differentiation 34 (CD34), desmoplakin (Dsp), erythropoietin (EPO), erythrocyte differentiation regulator 1 (ERDR1), fibronectin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), human chorionic gonadotropin (hCG), follistatin (FST), growth differentiation factor family proteins, namely growth differentiation factor 11 / osteogenesis imperfecta-11 (GDF11 / BMP-11), growth differentiation factor 15 (GDF15), etc., irisin, kisspeptin, klotho, matrixin (extracellular matrix-derived peptide),Human growth factors and / or cytokines, such as Scube3 (signal peptide, CUB and EGF-like domain-containing protein 3), osteopontin, oxytocin, pregnancy-associated plasma protein-A (PAPP-A), platelet factor 4 (PF4), reelin, stem cell antigen-1 (Sca-1), staniocalcin-1 (STC1), staniocalcin-2 (STC2), transforming growth factor (TGF) family proteins, i.e., TGF-alpha, TGF-beta-1, TGF-beta-2, TGF-beta-3, thyrotropin-releasing hormone (TRH), thyroid-stimulating hormone (TSH), heat shock protein family proteins, i.e., HSP90A and its derivatives, as well as combinations thereof, may be loaded. The effective amount of at least one activator is in the range of approximately 0.000001% to 1% by dry weight, depending on the application and the severity of the burn, preferably 0.00001% to 0.1%, and more preferably 0.00005% to 0.005%.

[0028] In other embodiments, the disclosure relates to a novel naturally biocompatible multifunctional biopolymer in which a combination of lignin and α-lactalbumin has been demonstrated to be effective in wound covering, wound healing, scar reduction, and promoting re-epithelialization in severe burns, and to restoring new tissue proliferation and replacing damaged burn tissue.

[0029] In other embodiments, the disclosure relates to a novel natural biocompatible multifunctional biopolymer used in the development of a leather substitute or a part of a leather substitute, wherein a prototype of the leather substitute, with or without the use of either α-lactalbumin or a leather reinforcing material obtained from coconut husk and / or koiapis powder fibers, exhibits the superior abrasion resistance and flexibility of natural leather. [Brief explanation of the drawing]

[0030] [Figure 1] a) shows the proportion of components in coconut husk, and b) shows the proportion of lignin in different plant materials. [Figure 2]The structure of the lignin component is shown. [Figure 3a] This shows the extraction of lignin from coconut husk. [Figure 3b] This shows the isolation of α-lactalbumin from whey protein. [Figure 4] This shows the zeta potential and particle size of lignin. [Figure 5] The ATR-IR graph of the extracted lignin is shown. [Figure 6] The X-ray diffraction (XRD) map of the extracted lignin is shown. [Figure 7] The differential scanning calorimetry (DSC) graph of extracted lignin is shown. [Figure 8] The polarized light microscope graph of the ferric chloride test is shown. [Figure 9] The lignin-based biopolymer films HLG (F1), XGPG (F2), and PXLG (F3) are shown. [Figure 10] This indicates the thickness of the film. [Figure 11] This demonstrates the folding durability of a lignin-based biopolymer film. [Figure 12] Graphs showing the texture and modulus of lignin-based biopolymer films - HLG(F1), XGPG(F2), and PXLG(F3) are shown. [Figure 13] The analysis results for the tensile strength and elastic modulus of lignin-based biopolymer films are shown. [Figure 14] This shows the degree of swelling of lignin-based biopolymer films. [Figure 15] This shows the water retention rate of lignin-based biopolymer films. [Figure 16] This shows the water vapor transmission rate of a lignin-based biopolymer film. [Figure 17a] This film shows the photodegradation of excess glutaraldehyde as CO2 at 2360 cm⁻¹. [Figure 17b] The IR spectrum of the HLG film is shown. [Figure 17c] The IR spectrum of the PXLG film is shown. [Figure 17d] The IR spectrum of the XGPG film is shown. [Figure 18] The XRD analysis graph of a lignin-based biopolymer film is shown. [Figure 19] This shows a DSC overlay of an α-lactalbumin-supported XGPG film. [Figure 20] This shows an SEM image of a lignin-based biopolymer XGPG(F2) film. [Figure 21] The histopathological findings of the tissue samples are shown (normal group - no injury was induced, kept for reference; negative control - injury was induced but no treatment was performed; blank film group - administered a blank lignin-based biopolymer film without α-lactalbumin; standard group - injury was induced and treated with commercially available silver sulfadiazine cream; treatment group - injury was induced and treated with a film containing α-lactalbumin). [Figure 22] This paper presents a prototype of an animal leather substitute made from lignin-based biopolymers. [Modes for carrying out the invention]

[0031] Those skilled in the art will recognize that this disclosure is subject to variations and modifications other than those specifically described. It should be understood that this disclosure includes all such variations and modifications. Furthermore, this disclosure includes all such steps, features, compositions, and compounds mentioned or indicated individually or collectively herein, as well as any and all combinations of any or more such steps or features.

[0032] Prior to further description of this disclosure, for convenience, the specific terms and examples used herein are set forth here. The terms used herein have meanings that are recognized and known to those skilled in the art, but for convenience and completeness, the specific terms and their meanings are set forth below.

[0033] The articles "a," "an," and "the" are used to refer to one or more (i.e., at least one) grammatical objects of the article.

[0034] The terms "comprise" and "comprising" are used in an inclusive and open sense, meaning that additional elements may be included. They are not intended to be interpreted as "consists of only."

[0035] Throughout this specification, unless otherwise required by context, the word “comprise,” and variations such as “comprises” and “comprising,” are understood to mean that they include the element or process, or group of elements or processes, described, but not that they exclude other elements or processes, or groups of elements or processes.

[0036] The term "including" is used to mean "including but not limited to." "Including" and "including but not limited to" are interchangeable.

[0037] In this specification, ratios, concentrations, quantities, and other numerical data may be presented in range format. It should be understood that such range formats are used merely for convenience and conciseness, and should be interpreted flexibly to include not only the numerical limits explicitly stated as range boundaries, but also all individual numerical values ​​or subranges encompassed within that range, as if each numerical value and subrange were explicitly stated.

[0038] In one embodiment of the present disclosure, a process for the extraction and purification of lignin from coconut husk and / or coiapis powder is provided, comprising: 1) the extraction of lignin from coconut husk using the Kraft pulp method; 2) 200 ml of 12.5% ​​w / v aq. NaOH aqueous solution was prepared by adding 10-80 g, more preferably 20-40 g, of husk powder, and the solution was held in an autoclave (Lequitron) at 130°C for 3 hours; 3) the NaOH solution containing the dissolved lignin was filtered through a muslin cloth using a vacuum filter, the residue / filter cake containing the deligninized lumps was stored separately, and the filtrate (black liquor) was collected and stored in a beaker. 4) The recovered black liquor was treated with concentrated sulfuric acid until the pH reached 2, and when the precipitated lignin became clearly visible, the solution was left to stand for 4 hours to allow it to precipitate (the lignin precipitate was at the bottom, while the acidic liquid formed a separate layer on top); 5) The liquid portion was carefully discarded, and the remaining solution was centrifuged at 10,000 RPM for 10 minutes, and the supernatant was discarded to obtain a residue in the form of lignin; 6) This lignin was further washed with distilled water and passed through a vacuum filter using filter paper (4.5 μm pore size, nylon 66); 7) The recovered lignin was dried overnight in a 60°C hot air oven, then ground into a fine powder using a mortar and pestle, and stored in a plastic bag (Figure 3a).

[0039] In one embodiment of the present disclosure, this specification exemplifies the development of novel naturally biocompatible multifunctional biopolymers comprising extracted lignin components, copolymers, crosslinking agents, and plasticizers from coconut husk and / or coiapis powder.

[0040] In one embodiment of this disclosure, this specification exemplifies the development of a novel natural biocompatible multifunctional biopolymer comprising extracted lignin components from coconut husk and / or coiapis powder, copolymers, crosslinking agents, and plasticizers, wherein the copolymers may be synthetic or natural. Examples of natural polymers include proteins such as hyaluronic acid and its derivatives, starch, modified starch, alginates, chitosan, chitin, natural gums, gelatin, collagen, casein, zein, gluten, soy protein isolates, and whey protein isolates, as well as land plant extracts such as pectin, and combinations thereof. Examples of water-soluble synthetic copolymers include crosslinked polyacrylate chains, polyvinylpyrrolidone, polyvinyl alcohol, modified cellulose ethers such as hydroxypropyl methylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, and carboxymethylcellulose, and ion exchange resins. The effective amount of copolymer ranges from about 10% to 90% by dry weight, preferably 30% to 70%, depending on the application of the developed biocompatible multifunctional biopolymer.

[0041] The term "copolymer" refers to a material that combines two or more polymers to form a network structure, where at least one polymer is synthesized or crosslinked in the immediate vicinity of the other polymer. These different polymers and / or copolymers physically intertwine with each other, forming a single intertwined network, resulting in improved mechanical and physical properties, ideal for a variety of applications such as bioadhesives and impact-resistant materials. The polymers and copolymers do not necessarily have to be covalently bonded to each other (i.e., they may be covalently bonded by, for example, a crosslinking agent), but they form a bonded structure that cannot be separated without breaking some chemical bond connecting them.

[0042] In one embodiment of the present disclosure, this specification exemplifies the development of a novel naturally biocompatible multifunctional biopolymer comprising extracted lignin components from coconut husk and / or coiapis powder, copolymers, crosslinking agents, and plasticizers, wherein the crosslinking agent is selected from glutaraldehyde, formaldehyde, polyethylene glycol (PEG) and its derivatives, genipin, dextran, polyethyleneimine (PEI), ethylenediamine (EDA), and sugar alcohols / polyols such as erythritol, xylitol, maltitol, mannitol, lactitol, polyglycitol, isomalt, sorbitol, glycerol, and combinations thereof, to facilitate the crosslinking chemical reaction. The effective amount of crosslinking agent is in the range of about 1% to 10% by dry weight, preferably 2% to 5%, depending on the application of the biocompatible multifunctional biopolymer developed.

[0043] In one embodiment of the present disclosure, this specification exemplifies the development of a novel natural biocompatible multifunctional biopolymer comprising extracted lignin components from coconut husk and / or coiapis powder, copolymers, crosslinking agents, and plasticizers, wherein the plasticizer may be selected from sucrose ester derivatives, glycerin, propylene glycol, polyethylene glycol, magnesium stearate, animal fat, neatfoot oil, lanolin, mineral oil, silicone oil, jojoba oil, castor oil, coconut oil, whale oil, squalene, grapeseed oil, safflower oil, canola oil, almond oil, etc., and combinations thereof. The effective amount of plasticizer is in the range of about 0.5% to 10% by dry weight, preferably 1% to 3%, depending on the application of the developed biocompatible multifunctional biopolymer.

[0044] In one embodiment of this disclosure, this specification exemplifies the development of a novel naturally biocompatible multifunctional biopolymer comprising extracted lignin components, copolymers, crosslinking agents, and plasticizers from coconut husk and / or coiapis powder, wherein the emulsifier is optionally selected from xanthan gum, lanolin, sodium alginate, lecithin, casein, whey protein, pectin, ammonium phosphatide, acacia, and oleylamine. The effective amount is in the range of about 0.5 to 5% (w / w), depending on the application of the developed biocompatible multifunctional biopolymer.

[0045] One embodiment of this disclosure provides the development of a novel, naturally biocompatible, multifunctional biopolymer. This biopolymer has been successfully studied as a wound dressing in severe burn accidents, restoring new growth on burns and replacing damaged tissue, and hereat least one activator may be loaded from turmeric extract (curcumin), honey, green tea extract, gotu kola extract (Centella asiatica), α-lactalbumin, whey protein isolate, comfrey extract, tea tree oil, lavender oil, chamomile extract, calendula extract, aloe vera gel, silver sulfadiazine, bacitracin, hyaluronic acid, collagen, vitamin E, allantoin, sphingosine- / phytosphingosine-1-phosphate, and deexpanthenol (vitamin B5). The effective amount of at least one activator ranges from about 0.01% to 50% by dry weight, preferably 1% to 20%, depending on the application and the severity of the burn.

[0046] One embodiment of this disclosure provides the development of a novel, naturally biocompatible, multifunctional biopolymer. This biopolymer has been successfully studied as a wound dressing in severe burn injuries, restoring new growth on burns and replacing damaged tissue, wherein at least one activator is epidermal growth factor (EGF), growth hormone / somatotropin (hGH / STH), vasoactive intestinal peptide (VIP), fibroblast growth factor family proteins, namely fibroblast growth factor-1 (FGF1), fibroblast growth factor-2 (FGF2), fibroblast growth factor-7 / keratinocyte growth factor (FGF7; KGF), fibroblast growth Factor-10 / Keratinocyte Growth Factor-2 (FGF10; KGF2), Fibroblast Growth Factor-19 (FGF19), Fibroblast Growth Factor-21 (FGF21), Fibroblast Growth Factor-23 (FGF23), etc., Vascular Endothelial Growth Factor (VEGF), Thymosin Beta-4 (TMS), Stem Cell Factor / Hematopoietic Stem Cell Factor (SCF), Platelet-Derived Growth Factor (PDGF), Prolactin (PRL), Placental Lactogen (hPL), Neurotrophic Factor (NGF), Brain-Derived Neurotrophic Factor (BDNF), Glial Cell-Derived Neurotrophic Factor (GDNF), Ciliary Cell Neurotrophic Factor Transtrophic factor (CNTF), interleukin-1 receptor antagonist (IL1Ra), interleukin family proteins, namely interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-13 (IL-13), interleukin-17 (IL-17), interleukin-18 (IL-18), adiponectin (ApN), TSLP (thymic interstitial lymphapoietin), bone morphogenetic protein family proteins, namely BMP-2, BMP-4, BMP-7, etc., clustered CD34, desmoplakin (Dsp), erythropoietin (EPO), erythrocyte differentiation regulator 1 (ERDR1), fibronectin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), human chorionic gonadotropin (hCG), follistatin (FST), growth factor family proteins, namely growth factor 11 / osteogenesis imperfecta-11 (GDF11 / BMP-11), growth factor 15 (GDF15), etc., irisin, kisspeptin, klotho, matrixin (extracellular matrix-derived peptide),Human growth factors and / or cytokines, such as Scube3 (signal peptide, CUB and EGF-like domain-containing protein 3), osteopontin, oxytocin, pregnancy-associated plasma protein-A (PAPP-A), platelet factor 4 (PF4), reelin, stem cell antigen-1 (Sca-1), staniocalcin-1 (STC1), staniocalcin-2 (STC2), transforming growth factor (TGF) family proteins, i.e., TGF-alpha, TGF-beta-1, TGF-beta-2, TGF-beta-3, thyrotropin-releasing hormone (TRH), thyroid-stimulating hormone (TSH), heat shock protein family proteins, i.e., HSP90A and its derivatives, as well as combinations thereof, may be loaded. The effective amount of at least one activator is in the range of approximately 0.000001% to 1% by dry weight, depending on the application and the severity of the burn, preferably 0.00001% to 0.1%, and more preferably 0.00005% to 0.005%.

[0047] In one embodiment of this disclosure, a process for isolating α-lactalbumin from whey protein (purchased from Suzhou Greenway Biotech Co. Ltd., China) is provided. This process includes extracting α-lactalbumin from the whey protein isolate (90%) and removing casein protein, which is responsible for the whiteness of milk, by centrifugation. Subsequently, NaCl was used as a washing and osmotic agent. Ca++ ions bound to the protein were released from α-lactalbumin by lowering the pH to 4.2 with the help of citric acid monohydrate (40 mM / L). As a result, the Ca++-bound α-lactalbumin was converted to a molten globule apo form without Ca++, which could be recovered through a vacuum filter. Upon restoration of the pH balance, the apo form returned to its original form in the presence of Ca++. By this method, α-lactalbumin was extracted effectively and efficiently (Figure 3b).

[0048] In one embodiment of this disclosure, the development of a novel, naturally biocompatible, multifunctional biopolymer has been successfully studied for wound dressing in severe burn accidents to rejuvenate new tissue in place of damaged burn tissue. Here, α-lactalbumin, a tryptophan-rich protein, has been demonstrated in various studies to possess healing properties, but the use of whey-extracted α-lactalbumin as a wound dressing has not yet been investigated. In this study, the combination of lignin and α-lactalbumin was demonstrated to be effective in promoting wound healing in Swiss albino mice. Furthermore, the combination of lignin and α-lactalbumin was found to reduce scarring and accelerate re-epithelialization at a faster rate than commercially available silver sulfadiazine cream. Notably, animals treated with the α-lactalbumin-containing film showed greater stimulation of hair follicles and faster hair growth than control animals. These results suggest that the combination of lignin and α-lactalbumin is promising as an effective wound dressing that promotes wound healing and reduces scarring.

[0049] In one embodiment of the present disclosure, the development of a novel natural biocompatible multifunctional biopolymer used herein for further development of leather substitute prototypes is provided, wherein the production of the prototype involves complete adhesion and sealing of both the leather substitute material and the leather reinforcement material. This was achieved by placing a heavy object on the material for 48 hours at 25°C and finishing the coating with a wax emulsion, softener, and wetting agent. As a result, a completely sealed prototype was obtained, which could then be cut to specifications. Here, the leather reinforcement material can be obtained from coconut husk and / or coir apice powder fibers uniformly adhered to a cellulosic aqueous adhesive present in the leather substitute material, foam padding using EVA (ethylene vinyl acetate), mesh fabrics such as polyester or nylon, glass fibers, polyethylene or polycarbonate, rubber, and combinations thereof. [Examples]

[0050] Next, examples of the present disclosure will be described. These examples are not intended to limit or restrict the scope of the present invention. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art in which this disclosure pertains. Similar or equivalent methods and materials may be used in carrying out the methods and compositions disclosed herein, but exemplary methods, devices and materials are described herein. Since such methods and conditions may vary, it should be understood that this disclosure is not limited to the specific methods and experimental conditions described herein.

[0051] <Example 1> (Extraction and characterization of lignin from coconut husk and / or coiapis powder) (material) Coconut husk and / or coi apis powder was purchased from various vendors in the local market of Shirpur, Maharashtra, India; sodium hydroxide, sulfuric acid, glutaraldehyde, other organic solvents, and starch were sourced from Rankem, Mumbai, India; xanthan gum, sorbitol, and glycerol were obtained from Loba Chemie Pvt. Ltd. in Mumbai; HPC (hydroxypropyl cellulose) and PVA Hot (polyvinyl alcohol) were purchased from Research Lab Pvt. Ltd. in Mumbai. The coconut fibers were removed by hand, the husk was ground in a kitchen grinder, passed through 20 and 40 sieves (IP; The Indian Pharmacopoeia), the powder was dried in an oven overnight to remove excess moisture, and then stored in a sealed plastic bag at room temperature (25°C).

[0052] (Lignin extraction method from coconut husk powder) The Kraft pulp method was used to extract lignin from coconut husk powder. To extract the lignin, two sample coconut husk powder solutions were prepared, each containing 10-80g, more preferably 20-40g, of dried coconut husk powder in 200ml of 12.5% ​​w / v aq. NaOH. One solution was kept in an autoclave (Lequitron) at 130°C for 3 hours, and the other solution was kept in a microwave oven at 140°C for 1 hour (Figure 3a).

[0053] (Evaluation of lignin properties) Extract yield: The lignin yield percentage was calculated as the weight of extracted lignin divided by the weight of coconut husk before lignin extraction. The coconut husk powder was dried in a 60°C hot air oven for 12 hours to remove excess moisture. The dried and pre-weighed coconut husk powder (W2) was treated with alkali, and lignin was extracted using a muslin filter. The lignin in the filtered black liquor was then precipitated by adding concentrated sulfuric acid dropwise to adjust the pH to 2. The precipitated lignin was separated and collected by either centrifugation or filtration through Whatman Paper (grade 42, pore size 2.5 μm), dried in a 60°C hot air oven for 8 hours, washed twice with water to remove impurities, and dried again in a 60°C hot air oven for 12 hours. The dried lignin was then ground in a mortar and pestle and weighed again (W1).

[0054]

number

[0055] Approximately 8.4 g of lignin (W2) was obtained from an initial weight of 30 g of coconut husk powder (W1), resulting in a lignin yield of 28% from coconut husk powder. Since the theoretical yield in various studies is approximately 34%, the percentage yield by Kraft extraction was approximately 82.35%. This yield may be improved in future embodiments by improving the heating method and optimizing the pH during extraction. Further pulverization of the coconut husk powder should also improve the yield.

[0056] (Particle size and zeta potential) The particle size and surface charge of the lignin prepared in this manner were measured, and the effect of pH on the resulting lignin particles was evaluated. In this experiment, 10 mg of lignin was dissolved in 100 ml of neutral (pH 7) and alkaline (pH 12) buffer solutions, and the solutions were transferred to a Zeta Sizer (Malvern) cuvette to 3 / 4 of its volume using a syringe. The particle size and zeta potential were measured.

[0057] The purpose of this study was to investigate the effect of pH on the size change of lignin particles due to differences in the pH range of the solvent. It was found that the particle size of lignin decreased significantly as the pH increased from 5 to 13. This is thought to be due to the free dissolution of lignin in alkaline aqueous solutions (≥pH 12), and its isoelectric point is estimated to be pH 2. At neutral pH, the particle size of lignin was 323 nm, but it decreased to 205 nm when the pH exceeded 9. At neutral pH, lignin showed a value of -13.5 mV (anion). This falls within the stable range of -30 mV to +30 mV (Figure 4).

[0058] (ATR-IR (Attenuated Total Reflectance-Infrared) spectroscopy of lignin) Prior to analysis, the lignin powder was dried in a hot air oven for 8 hours to remove moisture and reduce noise in the results below. The lignin powder was ground in a mortar and pestle and pressed under an ATR-IR sampler. The chemical composition of the lignin was analyzed using a Brookfield ATR-IR Spectrophotometer at 4000 cm⁻¹.-1 from 400 cm -1 and analyzed within the range of

[0059] The provided FTIR (Fourier Transform Infrared) spectrum shows the transmittance (%) of the lignin sample over a wavenumber range (cm -1 ). This spectrum shows important peaks indicating the presence of functional groups in lignin. Specifically, there is a broad peak corresponding to O-H stretching around 3400 cm 1 , and a sharp peak indicating C-H stretching around 2930 cm -1 . In addition, characteristic peaks related to aromatic C=C stretching around 1600 cm -1 and 1510 cm -1 , a peak corresponding to C-O stretching (aryl ether) around 1260 cm -1 , and peaks related to the C-O stretching of the guaiacyl unit and alcohol and ether around 1120 cm -1 and 1030 cm -1 appear respectively (Figure 5).

[0060] (X-ray Diffraction (XRD)) To measure the properties of lignin powder and the purity regarding the crystalline state versus the amorphous state, an X-ray diffractometer was used. The sample was pulverized with a mortar and pestle, transferred to a cuvette, and then analyzed using a Bruker D2 Phaser X-ray diffractometer. The voltage and current used were 40 kV and 30 mA respectively, and the X-ray diffraction pattern was obtained in the 2θ range of 5° - 50° at a step of 0.02° and a scan speed of 5° / min.

[0061] The extracted lignin was 86% amorphous and 14% crystalline, and no sharp peaks were observed. All peaks remained in a relatively short range of 200–500 counts. Lignin generally exhibited a broad and diffuse scattering pattern. The amorphous nature of lignin arose from its complex and irregular polymer structure. Phenolic monolignols formed a three-dimensional network lacking the regular repeating units and long-range order characteristic of crystalline structures. The slight crystallineity was inherited from the lignin precipitation process and attributed to sulfur impurities remaining after the washing process (Figure 6).

[0062] (Differential scanning calorimetry) To evaluate the thermal properties of lignin, thermal phenomena during the curing process were monitored using a differential scanning calorimeter (TA Instruments, DSC Discovery 250). 2.4 mg of lignin powder was placed in an aluminum open pan and thermal analysis was performed in the temperature range of 30°C to 325°C. DSC curves were recorded under an N2 (50 mL / min) atmosphere at a heating rate of 10°C / min.

[0063] The initial gradient at 40°C was due to instrument calibration. Lignin showed a clear, sharp endothermic peak, likely due to the initial desorption of water from the lignin in the presence of moisture. Drying lignin powder at high temperatures may reduce moisture content, but this could affect the appearance and solubility of the lignin. The absence of any secondary peaks indicates that lignin was successfully extracted and separated from cellulose, hemicellulose, and xylan, as it decreased well in this range. Lignin showed no phase changes in the 300°C range (Figure 7).

[0064] (Ultraviolet-visible spectrophotometer) Lignin was identified and its linearity determined using wavelength-versus-absorption spectroscopy. Using a Shimadzu 1900 UV-Visible spectrophotometer, 10 mg of lignin was dissolved in 10 ml of deionized water using a sonicator to obtain a 1000 ppm stock solution, which was then diluted to 100 ppm. λ maxThe values ​​and linearity range were measured.

[0065] In ultraviolet-visible light spectroscopy, lignin is found at 272 nm (λ). max ) showed a peak typical of Kraft lignin, derived from the aromatic ring / unconjugated phenol group. Lignin has typical absorption peaks at both 230 nm and 275 nm, which are due to electron transitions from the unconjugated phenol structure in the aromatic. Linearity was confirmed in distilled water in the concentration range of 20 μg / ml to 100 μg / ml (R 2 (=0.999).

[0066] (Other tests) The ferric chloride test for phenol was performed by dropping a freshly prepared ferric chloride solution into a 1% w / v lignin aqueous solution, and observing the visible color change of the resulting solution. The shape, uniformity, and particle size were observed using a polarizing microscope (PLM). Liquid solutions of lignin in deionized water were observed under PLM at 40x and 100x magnification.

[0067] At a neutral pH, undissolved particles appeared in the range of 10–37 μm, with very few particles around 70 μm. Zeta Sizer analysis revealed that most dissolved lignin particles were in the range of 200–300 nm at a neutral pH, and 200 nm at pH 8. The particle shapes ranged from uniformly circular to rhomboid. When the extracted lignin was dissolved in distilled water, the resulting solution turned dark green, indicating a positive ferric chloride test (Figure 8).

[0068] <Example 2> (Applications of lignin extracted from coconut husk and / or coiapis powder in pharmaceuticals, nutritional supplements, and cosmetics) (Biopolymer formulation) In this invention, the following approaches were taken to produce three different biopolymers using different methods.

[0069] Glutaraldehyde was used as a crosslinking agent at a concentration of 2.5%, and HPC and lignin were mixed in a 3:2 ratio to obtain a fluid solution. Sulfuric acid was added dropwise during mixing. The resulting solution was named HLG biopolymer (F1). A thick, viscous solution named XGPG biopolymer (F2) was obtained by mixing 10% polyethylene glycol, xanthan gum, and lignin in a 1:1 ratio. The mixture was stirred at 2000 rpm and 85-90°C (high-speed homogenizer, IKA T25 Digital Ultra Turrax, Mumbai, India). Polyvinyl alcohol, xanthan gum, and lignin were mixed in a 1:1:1 ratio to obtain a thick solution, named PXLG biopolymer (F3); this mixture was stirred with glycerol (1.5%) as a plasticizer at 2000 rpm and 85-90°C (Figure 9).

[0070] Subsequently, α-lactalbumin (ALA), extracted from whey protein as described in the embodiment, was added to each of the three biopolymers, and the solutions were mixed. Finally, the solutions were cast into a silica mold (15 × 15 cm) and dried at 30°C for 48 hours. These biopolymers have the potential to revolutionize several fields, including medicine, food, packaging, and leather substitutes. Further research is needed to explore these properties and their potential applications in other fields.

[0071] <Example 3> (Characterization of lignin-based biopolymers) (thickness) The width or thickness of the film was measured by randomly selecting five samples of similar size from each film, then carefully clamping the lower jaw of the Mitutoyo Digimatic Caliper to fit the film, and taking three measurements for each sample. Particular care was taken to avoid applying excessive pressure when fitting the caliper jaws, and the average value was calculated after various measurements.

[0072] The film thickness should not be so thick as to be bothersome to the patient, while being deep enough to effectively protect and seal the wound. For this purpose, the average film thickness was measured and calculated three times. The XGPG film was the thickest at 0.24±0.01 mm, PXLG was intermediate at 0.22±0.01 mm, and HLG was the thinnest at 0.21±0.01 mm (Figure 10).

[0073] (Foldable) To measure the foldability of the film, five samples of each film were cut to 3 x 3 cm, and the process of pressing down the center with clippers, folding by hand, and then unfolding was repeated until a visible crack appeared in the film. The measurements were recorded and the average value was calculated.

[0074] All wound dressings designed for wound covering must have good fold durability, as they may be folded repeatedly depending on the location of the wound. Therefore, we evaluated the foldability of the films. Among all the films, HLG showed the best fold durability, exhibiting no cracks even after 500 folds, while PXLG showed slight bending and cracking after approximately 250 and 290 folds, respectively. Film XGPG also showed considerable durability, but cracking occurred after 190 folds. The following graph was created using the average values ​​(Figure 11).

[0075] (Texture analysis and elastic modulus) Each film was cut to a size of 3 x 1 cm and analyzed using a CT3 texture analyzer (Brookfield Engineering Lab) under conditions of 30°C and 1 bar. Using the data obtained in this study, stress data, deformation at the fracture point, and elastic modulus were determined.

[0076] To establish the physical properties of the film, the elastic modulus was determined using the following formula, based on the force applied to a film of known length.

[0077]

number

[0078] PXLG exhibited the highest tensile strength, followed by XGPG and HLG. The maximum loads before fracture for PXLG, XGPG, and HLG were 339g, 270g, and 191g, respectively. Film PXLG also showed the largest deformation at the fracture point at 4.97mm, followed by HLG at 2.75mm and XGPG at 2.65mm (Figure 12).

[0079] For wound dressings, resistance to shape changes is crucial. Shrinkage and stretching can expose the wound to the external environment, potentially leading to infection. It also relates to the level of physical protection the dressing should provide.

[0080] The load was converted from grams to Newtons (1 gram-force is equal to 0.0098 N), and the moduli of elasticity of PXLG, HLG, and XGPG were determined as follows using the aforementioned formula (Figure 13).

[0081] Of all the films, PXLG had the highest total saturation (TS) due to the addition of PVA and glycerol-crosslinked xanthan gum (GCX). Since the stabilization of GCX and PVA would have been enhanced in the absence of moisture, further dehydration would also have increased its TS. This is because the presence of moisture weakens the bond between the hydroxyl groups of glycerol and the hydrophilic functional groups of xanthan gum. The XGPG film had a lower TS than PXLG due to the lower degree of crosslinking by PEG compared to glycerol in the presence of PVA. Glycerol was immobilized, while PEG maintained its mobility within the matrix (Domjan, Bajdik et al. 2009).

[0082] Film XGPG is significantly stiffer than both Film PXLG and Film HLG, while Film PXLG and HLG have similar stiffness, with Film HLG being slightly stiffer than Film PXLG. Film XGPG is the stiffest, but Film PXLG is the strongest in terms of tensile strength. Film HLG has moderate stiffness but the lowest tensile strength. This correlation suggests that stiffness (modulus of elasticity) and tensile strength may be related, but do not necessarily increase proportionally. A material is not necessarily strong just because it is stiff, and vice versa.

[0083] (Degree of swelling) The degree of swelling was determined by weighing three 1 x 1 cm samples from each film (W1), immersing them in a beaker containing 15 ml of pH 7.2 phosphate buffer for 5 hours, and weighing each sample every hour (W2). The degree of swelling was calculated using the following formula.

[0084]

number

[0085] Swelling degree is a measure of not only the degree of crosslinking but also the ability to absorb water without dissolving itself. Similar to the inventors' theory that PEG acts as an external crosslinking agent, the uncrosslinked fraction played the main role in swelling, as evidenced by XGPG's highest swelling index of 430.56% after 5 hours, significantly lower than PXLG's 324.44%. This is due to the glycerol-crosslinked-xanthanum functional group, which creates a more stable GCX in an anhydrous environment. HLG showed the least swelling at 14.28%, which was due to extensive crosslinking by glutaraldehyde between lignin and hydroxypropylcellulose (Figure 14).

[0086] (water retention) The dry weight of a 2x2 cm film was measured (W1) and stored in a beaker containing 50 ml of water for 5 hours. The film was then transferred to a centrifuge tube and centrifuged at 500 rpm for 5 minutes using a mini centrifuge (BioEra). The supernatant was discarded, and the film residue was weighed again as wet weight (W2) according to the method of Wang et al. (2019).

[0087] The film's water retention value (WRT) was calculated using the following formula.

number

[0088] HLG showed a hydrophobic tendency and had the lowest water retention capacity, while the other two films, due to the presence of xanthan gum in the film, showed three times the water retention capacity of HLG (Figure 15).

[0089] (Water vapor transmission rate) Three samples of each film were cut to a size of 1 x 1 cm and attached to the neck of a vial with Teflon tape, so that the opening of the 5 mm radius vial containing 10 ml of deionized water could be completely sealed without the film coming into contact with the water. The vial was left open to allow free water vapor to pass through. The vial was weighed initially (W1) and again after 12 hours (W2). The water vapor transmission rate for each film was calculated using the following formula.

[0090]

number

[0091] Films used as wound dressings must be able to create a moist wound environment to help promote cell migration and wound healing. An ideal coverage range is approximately 2000 g / m². 2 The results were as follows: Among all the films, HLG had the lowest vapor transmission rate, followed by PXLG. While these films all had low vapor transmission rates due to their high crosslinkability, the vapor transmission rate of XGPG, which had PEG acting as an external crosslinking agent, was almost twice that of the other films (Figure 16).

[0092] (ATR-IR) Place the sample under the ATR-IR sample holder, press it slightly, then press it down to 4000 cm². -1 From 400cm -1 The scan was performed within the specified range. The chemical composition of the polymer film was analyzed by ATR-IR spectroscopy using a Brookfield ATR-IR Spectrophotometer.

[0093] While PEG 400 acted like an external crosslinking agent, glutaraldehyde exhibited the best chemical crosslinking by introducing new bonds in the form of hemiacetals and esters. On the other hand, glycerol is known to impart flexibility with excellent tensile strength to the film at low concentrations (1-3% w / v), while sorbitol tends to rapidly increase flexibility at high concentrations (8-10% w / v). These results support our theory that excess and unreacted glutaraldehyde can be easily removed by photodegradation (Figure 17a).

[0094] HLG has a range of 3500-3200 cm² due to the bonded OH group. -1 A clear and broad peak is observed, along with 2924 cm², which is attributed to the CH stretching vibration of methyl (CH3) present in unsaturated aromatic systems. -1 It shows the peak at 1462cm. -1 The peak is due to CH bending of the methyl group. 1043 cm in the hemiacetal and ester band region. -1 The new peak was attributed to COC of the acetal or hemiacetal group and was considered to be the result of chemical crosslinking (Figure 17b).

[0095] The film PXLG shows overlap between the functional group spectra of lignin and xanthan gum, and the decrease in intensity or disappearance of peaks associated with unbonded OH stretching vibrations indicates a decrease in free hydroxyl groups, likely because the material is chemically modified, reducing its tendency to interact with or absorb water. 2950 cm -1 The splitting of the OH peak is due to the protons in the conjugated system. In a water-free environment, the degree of crosslinking is so extensive that it obscures the crosslinking caused by other compounds, clearly indicating the extent of xanthan gum binding by glycerol (Figure 17c).

[0096] The film XGPG spectrum showed significant changes. This corresponds to the OH stretching vibration at 3200-3600 cm². -1The broad peaks in the vicinity showed a significant decrease in intensity, indicating the consumption of hydroxyl groups during bridging along other intrinsic peaks of lignin, such as new peaks in the carbonyl and CO stretching regions, as well as changes in aromatic ring vibrations, suggesting the formation of new chemical bonds and structural changes in lignin. 2865 cm -1 The sharp peaks were due to -CH3 stretching, similar to those reported by Marcos et al. (2017). Compared to PXLG, lignin showed its characteristic peaks with weaker intensity; these were either involved in crosslinking or present in smaller amounts than the intrinsic lignin. This supports the idea that PEG acts as an external crosslinking agent, improving film stability by assisting in the formation of the polymer network (Figure 17d).

[0097] (XRD) The samples were cut into small squares, placed on XRD cuvettes, and analyzed using a Bruker D2 Phaser X-ray diffractometer. The voltage and current used were 40kV and 30mA, respectively, and the XRD patterns were obtained in a 2° range from 5° to 50° with a 0.02° step and a scan rate of 5° / min.

[0098] XRD analysis in the 2° range from 5° to 50° showed that several HPC peaks were above lignin, indicating that the film was amorphous. However, it remained amorphous overall. The film lacked crystalline peaks, indicating the absence of crystalline impurities that may have been generated during the lignin extraction process (Figure 18).

[0099] (DSC) Alpha-lactalbumin was supported on XGPG film and analyzed using a differential scanning calorimeter (TA Instruments, DSC Discovery 250) to understand the thermal properties during the healing process. 2.3 mg of biopolymer film samples were placed in an aluminum open pan and analyzed at temperatures ranging from 30°C to 325°C. DSC curves were recorded under an N2 (50 mL / min) atmosphere at a heating rate of 10°C / min.

[0100] DSC studies revealed that the film can protect alpha-lactalbumin from thermal degradation. The initial endothermic peak of α-lactalbumin was in the 70-80°C range, which was presumed to be due to protein denaturation. However, within the film, this slope was slanted, indicating the protective properties of the film against alpha-lactalbumin. Furthermore, the film showed no peaks other than the 125°C peak due to the xanthan gum present in the film. The exothermic peak of the film in the 90-100°C range may be due to the glass transition of the blend (Figure 19).

[0101] (SEM) The film matrix was analyzed using a scanning electron microscope (JEOL JSM-6390 LA). SEM was used to clearly understand the surface morphology and structure of the film. SEM images of the polymer film surface were acquired with a point resolution of 0.23 nm by operating the SEM at 200 Kv using a LaB6 electron gun.

[0102] SEM images showing the surface of the XGPG film show undissolved lignin particles in areas (a) and (b) of the first image, but their occurrence is minimal, indicating that most of the lignin is bound to the xanthan gum film matrix (Figure 20).

[0103] <Example 4> (In vivo clinical evaluation using Swiss albino mice) This study demonstrated that a combination of lignin and α-lactalbumin promotes wound healing in Swiss albino mice. Furthermore, the combination of lignin and α-lactalbumin was found to reduce scarring and accelerate re-epithelialization at a faster rate than commercially available silver sulfadiazine cream. In particular, animals treated with α-lactalbumin-containing films showed significant stimulation of hair follicles, and hair regrowth occurred earlier than in other animals. Here, α-lactalbumin (ALA) is a protein commonly found in the milk of almost all mammalian species and plays an important role in regulating lactose production. In its multimer form (Eugenia Lucena, Alvarez et al. 2006, 2007), α-lactalbumin strongly binds to calcium and zinc ions, suggesting potential bactericidal and antitumor effects (Sharma, Hanson et al. 2015). This affinity has made ALA (Eugenia Lucena, Alvarez et al. 2006, Eugenialucena, Alvarez et al. 2007), which possesses a wide range of activities including bactericidal, antitumor (Krunic, Rakin et al. 2018), and wound healing ability (Guo, Liu et al. 2020), the target of numerous studies aimed at extracting and isolating it. Of all proteins, ALA has the highest percentage of tryptophan content (Layman, Lφnnerdal et al. 2018). Various studies and hypotheses have been proposed to support the potential mechanisms of wound healing by exogenous tryptophan application, including the reduction of cellular stress levels through inhibition of indoleamine 2,3-dioxygenase (IDO) and tumor necrosis factor-α (TNF-α) (Kiank, Zeden et al. 2010, Bandeira, Bortolot et al. 2015). Topical application of 1 μM tryptophan has been previously demonstrated to reduce pain and increase re-epithelialization in patients with lower extremity ulcers, showing promise as a natural component of wound healing (Barouti, Mainetti et al. 2015).These findings suggest that the combination of lignin and α-lactalbumin is a promising and effective wound dressing that promotes wound healing and inhibits scar formation. Histopathological analysis of wound tissue samples revealed clear differences between the groups. In the normal group, the structure of the epidermis, dermis, and subcutaneous layers was well preserved, including intact sebaceous glands and hair follicles. In contrast, the negative control group showed significant changes, including hyperplasia, edema, and increased epidermal thickness. In addition, there was mild infiltration of inflammatory cells, accompanied by epidermal thickening and hyperkeratosis.

[0104] In the blank film group, typical epidermal and dermal structures were maintained, showing normal structure, but hair follicles were noticeably absent. Both the standard group and the treatment group showed similar skin structures in the epidermal and dermal layers. However, the treatment group had a significantly higher number of hair follicles compared to the standard group, suggesting that α-lactalbumin may stimulate hair follicle growth (Figure 21).

[0105] (Normal group - no injury induced, kept for reference; Negative control - injury induced but no treatment; Blank film group - administered a blank lignin-based biopolymer film without alpha-lactalbumin; Standard group - injury induced and treated with commercially available silver sulfadiazine cream; Treatment group - injury induced and treated with a film containing alpha-lactalbumin).

[0106] <Example 5> (Application to the leather industry) Leather is an extremely versatile material used in a wide range of applications, including interior furniture and decoration, clothing, footwear, bags, handbags, accessories, and automotive applications. Traditionally, leather was made from animal hides. However, synthetic alternatives have emerged, including natural or synthetic fibers coated with synthetic polymers. Despite its popularity, the production of synthetic leather presents several environmental challenges due to the use of non-biodegradable plastics. These include not only difficulty in processing but also a considerable time for decomposition. Most synthetic leathers contain a polyester knit base coated with polyurethane and polyvinyl chloride. Furthermore, while synthetic leather can be cheaper than natural leather, its quality and durability are typically inferior.

[0107] A prototype of a leather substitute was prepared by using 2.5% glutaraldehyde as a crosslinking agent in a lower ratio (3:2) of HPC to lignin, and adding sulfuric acid dropwise, resulting in a fluid solution named HLG. A 10% concentration of polyethylene glycol was used in a 1:1 ratio of xanthan gum and lignin, and stirred at high temperature and high RPM to obtain a thick, viscous solution named XGPG. A thick solution was obtained by adding 1.5% glycerol as a plasticizer to polyvinyl alcohol, xanthan gum, and lignin in a 1:1:1 ratio, and stirring at high temperature and 2000 RPM, and named PXLG. All solutions were then cast into silica molds. Leather reinforcement materials obtained from coconut husk and / or coir apice powder fibers were uniformly attached to a cellulose-based aqueous adhesive present in the leather substitute.

[0108] The prototype manufacturing involved the complete adhesion and sealing of both the leather substitute and leather reinforcement materials. This was achieved by placing heavy objects on the materials for 48 hours at standard room temperature, followed by a finishing coating of wax emulsion, softener, and wetting agent. The result was a perfectly sealed prototype that could be cut according to specifications.

[0109] The final product (shown in Figure 22-XGPG below) exhibits excellent abrasion resistance and flexibility, particularly as a leather substitute. Furthermore, it is unaffected by weather changes and ultraviolet radiation.

[0110] The modulus of elasticity was determined using the force acting on a prototype finished leather of known length. The results are shown in Table 1.

[0111] [Table 1]

Claims

1. Naturally biocompatible multifunctional biopolymers including the following: a) Lignin extracted from coconut husk and / or coiapis powder obtained from the fruit of Cocos nucifera, a tropical plant belonging to the family Palmae; b) Copolymer; c) Crosslinking agents; d) Plasticizers; and e) At least one activator.

2. The natural biocompatible multifunctional biopolymer according to claim 1, wherein at least one copolymer is selected from either a natural polymer or a water-soluble synthetic polymer, the natural polymer being proteins such as hyaluronic acid and its derivatives, starch, modified starch, alginate, chitosan, chitin, natural gum, gelatin, collagen, casein, zein, gluten, soy protein isolate, and whey protein isolate, as well as land plant extracts such as pectin, and combinations thereof, and the water-soluble synthetic copolymer being a natural biocompatible multifunctional biopolymer comprising crosslinked polyacrylic acid chains, polyvinylpyrrolidone, polyvinyl alcohol, modified cellulose ethers such as hydroxypropyl methylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, and carboxymethylcellulose, and ion exchange resins, and combinations thereof.

3. The natural biocompatible multifunctional biopolymer according to claim 1, wherein at least one crosslinking agent is selected from glutaraldehyde, formaldehyde, polyethylene glycol (PEG) and its derivatives, genipin, dextran, polyethyleneimine (PEI), ethylenediamine (EDA), and sugar alcohols / polyols such as erythritol, xylitol, maltitol, mannitol, lactitol, polyglycitol, isomalt, sorbitol, glycerol, and combinations thereof, to promote the crosslinking chemical reaction.

4. The natural biocompatible multifunctional biopolymer according to claim 1, wherein at least one plasticizer is selected from sucrose ester derivatives, glycerin, propylene glycol, polyethylene glycol, magnesium stearate, animal fat, neatfoot oil, lanolin, mineral oil, silicone oil, jojoba oil, castor oil, coconut oil, whale oil, squalene, grapeseed oil, safflower oil, canola oil, almond oil, and combinations thereof.

5. The natural biocompatible multifunctional biopolymer according to claim 1, wherein the emulsifier is optionally selected from xanthan gum, lanolin, sodium alginate, lecithin, casein, whey protein, pectin, ammonium phosphatide, acacia, and oleylamine.

6. A natural biocompatible multifunctional biopolymer according to claim 1, used as a wound dressing or as part of a wound dressing on severe burns in a patient, to restore new tissue growth on the burn and replace damaged tissue, wherein at least one activator may be loaded from turmeric extract (curcumin), honey, green tea extract, gotu kola extract (Centella asiatica), α-lactalbumin, whey protein isolate, comfrey extract, tea tree oil, lavender oil, chamomile extract, calendula extract, aloe vera gel, silver sulfadiazine, bacitracin, hyaluronic acid, collagen, vitamin E, allantoin, sphingosine- / phytosphingosine-1-phosphate, and deexpanthenol (vitamin B5), and combinations thereof.

7. Used as a wound dressing or as part of a wound dressing on severe burns in patients, it restores new tissue growth on the burn and replaces damaged tissue, wherein at least one activator is epidermal growth factor (EGF), growth hormone / somatotropin (hGH / STH), vasoactive intestinal peptide (VIP), fibroblast growth factor family proteins, namely fibroblast growth factor-1 (FGF1), fibroblast growth factor-2 (FGF2), fibroblast growth factor-7 / keratinocyte growth factor (FGF7; KGF), fibroblast growth factor-10 / keratinocyte growth factor FGF-2 (FGF10; KGF2), fibroblast growth factor-19 (FGF19), fibroblast growth factor-21 (FGF21), fibroblast growth factor-23 (FGF23), etc., as well as vascular endothelial growth factor (VEGF), thymosin beta-4 (TMS), stem cell factor / hematopoietic stem cell factor (SCF), platelet-derived growth factor (PDGF), prolactin (PRL), placental lactogen (hPL), nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF), ciliary neurotrophic factor (CNTF), interleukin-1 receptor antagonist Nist (IL1Ra), interleukin family proteins, namely interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-13 (IL-13), interleukin-17 (IL-17), interleukin-18 (IL-18), etc., adiponectin (ApN), TSLP (thymic interstitial lymphapoietin), bone morphogenetic protein family proteins, namely BMP-2, BMP-4, BMP-7, etc., cluster differentiation 34 (CD34), desmoplakin (Dsp), erythropoietin (EPO), erythrocyte differentiation Regulatory factor 1 (ERDR1), fibronectin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), human chorionic gonadotropin (hCG), follistatin (FST), growth and differentiation factor family proteins, namely growth and differentiation factor 11 / osteogenesis imperfecta-11 (GDF11 / BMP-11), growth and differentiation factor 15 (GDF15), etc., irisin, kisspeptin, klotho, matrixin (extracellular matrix-derived peptide), Scube3 (signal peptide, CUB and EGF-like domain-containing protein 3), osteopontin, oxytocin,A naturally biocompatible, multifunctional biopolymer according to claim 1, which may be loaded with human growth factors and / or cytokines such as pregnancy-related plasma protein-A (PAPP-A), platelet factor 4 (PF4), reelin, stem cell antigen-1 (Sca-1), staniocalcin-1 (STC1), staniocalcin-2 (STC2), transforming growth factor (TGF) family proteins, i.e., TGF-alpha, TGF-beta-1, TGF-beta-2, TGF-beta-3, thyrotropin-releasing hormone (TRH), thyroid-stimulating hormone (TSH), heat shock protein family proteins, i.e., HSP90A and its derivatives, and combinations thereof.

8. The combination of lignin and α-lactalbumin has been shown to be effective in wound covering, wound healing, scar reduction, and promoting re-epithelialization in severe burns, as well as restoring new tissue growth and replacing damaged burn tissue, according to claims 1, 6, and 7, a naturally biocompatible multifunctional biopolymer.

9. A natural biocompatible multifunctional biopolymer according to claim 1, used in the development of a leather substitute or a part of a leather substitute, wherein a prototype of the leather substitute, with or without the use of either α-lactalbumin or a leather reinforcing material obtained from coconut husk and / or koiapis powder fibers, exhibits the excellent abrasion resistance and flexibility of natural leather.

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