Biopolymers

Abstract

Significant progress has been made on biopolymers in recent years. Biopolymers are preferred to other materials because they have specific physical, chemical, biological, biomechanical, and degradation properties. Many natural or synthetic biopolymers can degrade hydrolytically or enzymatically and are used for many applications.

1. Introduction

Biopolymers are a large group of biomaterials, for which it is important that they are biocompatible and noncytotoxic and produce decomposition products that are themselves nontoxic. They should support cell adhesion and proliferation and be an active participant in the process of creating new tissues, ensuring that the dynamics of formation and resorption of these tissues take place efficiently, with a constant balance between the two processes. Biopolymers by their properties, although they include different types of hydrophilic and hydrophobic polymers (which include different types of complex polymers, such as grafted and block copolymers, hydrogels, different types of thermoplastic biodegradable polymers, and polypeptide polymers, which may be of natural origin or artificial, different functional properties, network kinetics, and biodegradation), mostly belong to the so-called family smart polymers, due to their ability showing significant changes in their structural properties when the environmental conditions in which they are changing.

Among the best-known polymers are polyethylene, polyvinyl chloride, polystyrene, polypropylene, polyvinyl alcohol, etc. Their basic properties are satisfactory strength, flexibility, ease of painting and modeling, and hardness. What is the basic specificity of smart polymers is their extreme response to slight changes in environmental conditions, such as changes in temperature, pH, water, and light. These changes cause changes in the structural characteristics of these polymers. Therefore, they have significant potential for different types of applications in the field of biotechnology and biomedicine. Knowledge of the chemistry of conformational changes in polymers, which are conditioned by changes in environmental conditions, offers the possibility of the synthesis of new smart polymers, whose changes can be controlled by changes in environmental factors. This is extremely important for biological systems, as it allows the efficient use of polymers for the delivery of drugs or other substances important for the control of metabolic mechanisms. Smart polymers can be designed in the form of hydrogels, patches, bags degradation of plastic, chewing gum, blood glucose detectors, and polymers for the targeted release of insulin.

Graft and block copolymers are a special group of polymers, which consist of two different polymers, which are linked together in a graft. There are many patents that offer different combinations of such polymers, which have different reactive groups. The products are a combination that has properties characteristic of both components involved in the construction of such a polymer, which gives a new dimension to the structures of smart polymers, thus creating conditions for increasingly diverse applications. The crosslinked hydrophobic and hydrophilic polymers form micelle-like structures in which the safe encapsulation and protection of the drug can be related to the delivery of the drug through an aqueous medium until it reaches the target location when degradation occurs by breaking the bonds between the two polymers. An example of such a polymer is a polyacrylic acid PAAc bioadhesive polymer. The polyacrylic acid adheres, swells rapidly, and degrades to pH = 7.4, resulting in the rapid release of the drug trapped in the matrix. The combination of polyacrylic acid with other polymers, which are less susceptible to changes in neutral pH, increases the retention time of the drug and slows its release, while showing an increase in its bioavailability and efficacy.

Hydrogels are networks of polymers that do not dissolve in water, but swell and collapse when the aquatic environment changes. They are used in biotechnology for phase separation because such polymers are suitable for recycling. Highly specialized hydrogels have also been developed for the delivery and release of drugs into specific tissues. Polyacrylic acid hydrogels have very pronounced bioadhesive properties and exceptional absorption.

Enzyme immobilization in hydrogels is a highly developed technique for introducing enzymes into biological systems. In such an enzyme application, the body’s response depends on the product of the enzyme reaction. The process of introducing enzymes, receptors, and antibodies takes place by establishing the connection of the given entities with the selected molecule, within the hydrogel. After the connection is established, a targeted chemical reaction takes place in the hydrogel, the product of which may be, for example, oxygen, due to the sensitivity of the system to the presence of redox enzymes or pH changes of hydrogels that are sensitive to pH changes. Thus, for example, combinations of glucose oxidase and insulin packaging are converted to pH-sensitive hydrogels. Namely, in the presence of glucose, the formation of gluconic acid by enzymes leads to the targeted release of insulin from the hydrogel. The two basic criteria for the efficient operation of this technology are stability enzymatic and rapid kinetics (rapid response to change activator and recovery after cessation of its action).

Smart polymers are not yet suitable for drug delivery, although their properties are suitable for bioseparation. The time and cost involved in protein purification can be reduced with the significant use of smart proteins, which undergo rapid reversible changes in response to changes in average properties. Conjugate systems have long been used in physical and chemical separations and immunoassays. Microscopic changes in the structure of the polymer are manifested by the formation of a precipitate, which can be used to help separate the trapped proteins from the solution. These systems work in such a way that if a protein or other molecule is separated from the mixture forms a biconjugate with the polymer, then it precipitates with the polymer when the properties of the medium change. The precipitate is removed from the medium, separating the desired conjugate component from the rest of the mixture. The removal of these components from the conjugate depends on the speed of recovery of the polymer and its return to baseline. Another possibility of controlling biological reactions using smart polymers is the preparation of recombinant proteins, with characteristic binding sites embedded in ligand-like proteins or binding sites in cells. This technique is used to control both ligands and cell binding activities, which are responsible for various stimuli of activity, such as temperature or light [1, 2]. Future applications of smart polymers will be related to the development of polymers that will be able to learn and correct their own behavior over time. In the near future, smart toilets will use such polymers for analysis, for example, urine, and provides assistance in identifying health problems. Many creative approaches to targeted drug delivery systems and their self-regulation based on their unique cellular environment are already the subject of intense research.

Intelligent natural polymers and their response to external stimulation.

Polymers found in living systems (proteins, carbohydrates, and nucleic acids) play a significant physiological role in biological systems. Smart polymers have become especially important after studying their chemistry and the conditions that serve as activators of their conformational changes. The new polymeric materials are formulated to “feel” specific changes in biological systems by adjusting to respond to them in a predictable way, allowing them to be used as a useful tool for the administration of drugs and the control of various metabolic mechanisms. The nonlinear response of smart polymers makes them unique and efficient. Significant changes in the structure and properties of smart polymers can be caused by a small change in stimuli. Once a change occurs, there are no changes, which means that it is predictable whether or not there will be a response. It is important to note that the change in the conformation of the polymer is homogeneous for the system as a whole. Smart polymers change their conformational, adhesive, or retention properties when present in water, with small changes in pH, ionic strength, temperature, or other change activators. Another factor that defines the response efficiency of smart polymers is their inherent nature. The response of molecules to stimulus-induced change occurs simultaneously in a number of individual monomeric units. Although the change, which is related to only one of the monomer molecules, is almost insignificant when it comes to the whole set of such molecules (because the total change is equal to the sum of these identical or similar minor changes and theirs), corresponding responses (hundreds or thousands) such a change has a significant value, conditioning the appearance of a very significant force of stimulation of the appropriate biological responses (processes).

2. Classification and basic applications of polymers

Today, the most common application of polymers in biomedicine is for the targeted delivery of drugs. With over-the-counter pharmaceuticals, the basic problem for scientists is to find an effective way to deliver drugs to a certain place in the body without first degrading it to very acidic stomach acid. Preventing side effects on healthy bones and tissues is also a significant issue. It is advisable to monitor the release of medicines until the delivery system is in the desired location [3].

The release of the drug is controlled by chemical or physiological activation. Intelligent linear or matrix polymers are in different varieties and may have different properties depending on the type of functional groups or side chains present in them. These groups may be sensitive to pH, temperature, ionic power, electric or magnetic field, and light. Some polymers crosslink with non-covalent bonds and can cleave and reform depending on external conditions. Dendrimers are known as typical particle-free polymers, which are most commonly used for drug delivery. Lactic acid polymers are used in the traditional encapsulation of medicines. The release of drugs from smart polymer matrices is achieved by chemical or physiological reactions, such as hydrolysis, in which the bonds are broken and the drug is released when the matrix is ​​divided into biodegradable components. Artificially obtained polymers, such as polyanhydrides, polyesters, polyacrylic acids, poly (methyl methacrylates), and polyurethanes, are most often used for this purpose. Low-molecular-weight hydrophilic amorphous polymers containing heteroatoms (other atoms relative to carbon) have been shown to degrade most rapidly. By controlling the rate of degradation, today’s scientists are trying to control the rate of drug delivery [3, 4, 5]. Another very important field of application of polymers is tissue engineering, where many polymers of natural and artificial origin can be part of such composite structures that allow better adhesion and attachment of suitable cells (stem cells), their differentiation according to a certain phenotype, and finally, of them proliferation to new tissue [3, 4, 5].

Due to the relevant economic and environmental aspects, the growing interest in natural polymers is becoming more important in natural polymers due to their biodegradability, low toxicity, low cost, low availability, and renewable costs. Moreover, they offer a wide range of benefits for tissue engineering applications, such as biological signaling, cell adhesion, degradation, and responsible cell remodeling. However, their inadequate physical properties, such as solubility, rapid degradation, and possible loss of biological properties during casting, often limit their use as scaffolding materials. In addition, the risk of immune rejection and transmission of diseases require further purification. Physical or chemical modification of such polymers is an effective way to improve the stability of the material and its physical characteristics [6].

3. Polymers of natural origin

Natural polymers used in tissue engineering are very diverse in origin and composition. Such polymers include proteins, collagen, silk fibroin, polysaccharides, chitosan, and its derivatives, hyaluronic acid, alginates, starch, cellulose, dextrans, and polyesters of microbial origin.

3.1 Proteins

There have been numerous studies on proteins that make up the extracellular matrix (ECM) of natural tissues, such as scaffolding in tissue engineering. The use of biomacromolecules, such as collagen and fibronectin, which transmit biological information and provide the necessary physicochemical functions, is attempting to mimic ECM in the development of targeted tissues. A group of collagen-based biomaterials has been investigated with a particular interest in their application in bone regeneration [7].

Despite attempts to replace natural fibrous protein materials with artificial polymeric materials, natural protein fibers, such as silk, wool, and leather fibers, have not yet been successfully reproduced as artificial materials. Bone tissue and cartilage engineering based on the use of silk fibroin in the construction of scaffolds based on biomaterials is a very current topic of modern research [7, 8, 9].

3.2 Collagen

Collagen is a biological protein with a high content of glycine (almost 33%) and other amino acids (almost 20%). It is rich in the ECM of many tissues (skin, bones, cartilage, blood vessels, and teeth) where it provides the basic structure and mechanical support of these systems. Type I collagen is found in bones, tendons, cartilage, and corneal fibers, type II in hyaline cartilage and the nucleus pulposus (the gelatinous substance in the central part of the spine), type III in the intestines and uterine walls, type IV in the endothelium tissues and epithelial membrane, and type V in the cornea, placenta, bones, and heart valves. In the structure of collagen fibers, glycine is the most abundant, which is in place of every third residue in the polypeptide chain of collagen, forming (Gly-XY) a motif repeatedly adapted to the helical structure oriented to the left (α chain) with a length of about 1400 amino acids and three residues per reason. The most common collagen sequences are composed of glycine (gly), proline (pro), and hydroxyproline (hyp) (Gly-Pro-Hyp). Three α chains are wrapped around each other, twisted to the right, and densely wrapped in a triple helix (Figure 1) [10, 11].

Figure 1.

Collagen triple helix [http://www.rcsb.org/].

Collagen fibers are stabilized by specific covalent crosslinkers between collagen molecules. Biodegradability, low antigenicity, and cell binding properties make collagen a valuable material for use in tissue engineering. Collagen sponge accelerates the growth and growth of cells and tissues and improves bone formation by promoting the differentiation of osteoblasts.

Studies involving the seeding of mesenchymal stem cells (MSCs) in collagen gel, when implanted in osteochondral defects in rabbits, lead to bone and hyaline cartilage formation, although the mechanical properties of regenerated tissue are significantly lower than in normal tissues. It is important to note that, after the implantation process, no degeneration of the surrounding tissues was observed in the first 24 weeks. The main disadvantage of using collagen as a biomaterial for tissue repair is its high rate of degradation, which leads to the rapid loss of mechanical properties of scaffolds based on it. Many attempts have been made to overcome this problem by further processing to ensure that the collagen remains insoluble during a certain critical period, by adding suitable mineral crystals or a combination of collagen with other natural materials, such as glycosaminoglycans (GAGs) or synthetic polymers, derived from methacrylate, or by applying specific methods of collagen crosslinking [10, 11, 12]. Highly porous hydroxyapatite/collagen composite constructions seeded with chondrocytes showed an increase in the stability of the foam composite in the culture medium upon ECM deposition. A matrix made by crosslinking collagen fibers with modified hydroxyapatite, implanted in the cranial defects of mice, showed good biocompatibility and improved osteoconductivity in relation to the collagen material itself. The improvement of mechanical properties can be achieved by crosslinking the collagen chains by amino groups of lysine and hydroxylysine using glutaraldehyde or other agents, such as 1-ethyl-3-(3-dimethyl aminopropyl), carbodiimide (1-ethyl-3-(3)-dimethyl aminopropyl) carbodiimide, and hexamethylene diisocyanate. However, these treatments are not sufficiently cytocompatible due to the potential toxicity of any of the network agents used.

Other strategies for modifying collagen fibers are related to the chemical modification of the side chains of the collagen fiber to form double bonds, which allow subsequent crosslinking with chemical or photoinitiated free radicals or crosslinking with the enzyme lysyl oxidase. Takayama and Mizumachi have shown that lactoferrin adsorbs type I collagen, thus accelerating the calcification of collagen in vitro and the differentiation of human osteoblastic cells. Biocompatible scaffolds derived from type I collagen electrospun and hydroxyapatite nanoparticles show a significant increase in tensile strength and modulus. The mechanical properties of the scaffold are further improved by crosslinking with the vapor phase glutaraldehyde.

A new method for obtaining a biostyclic-nanofibrocollagen nanocomposite, in the form of a membrane or macroporous scaffold, has recently been published. The nanocomposite matrix exhibits “in vitro,” bioactivity induced by the rapid formation of bone-like mineral apatite on its surface when incubated in body fluid (SBF). Osteoblastic cells show a favored growth on nanocomposite scaffolds, and their alkaline phosphatase activity is significantly higher than in pure collagen scaffolds [13].

3.3 Silk fiber

Silk is a natural polymer that has been used clinically for centuries. Naturally extruded by insects or larvae, the silk took the form of a fiber consisting of a protein fiber called fibroin and a sticky coating made of sericin protein. Today, fibroin fibers are increasingly used as biomaterials for new biomedical applications, especially, in the field of tissue engineering, due to their biocompatibility, slow degradability, and exceptional mechanical properties, as well as better ability to control molecular structure and morphology through various forms of processing and surface modification.

Silk fiber in various forms (films, fibers, nets, membranes, sponges, etc.) supports the adhesion, proliferation, and differentiation of stem cells “in vitro” and promotes the repair of tissues “in vivo.” 3D fibrous silk scaffolds have a special application in skeletal tissue engineering’s, such as bones, ligaments, and cartilage, as well as in connective tissue engineering (skin tissues). The dominant source of silk-based materials to date is fibroin produced from Bombyx mori silkworm caterpillars. Silk fibroin from spiders and modified transgenic species is also a very important source. Such a silk fibroin shows the possibility of modifying certain fibroin sequences from natural silk and thus indirectly new possibilities for the production of materials inspired by silk fibroin, for various medical applications.

4. Structure and properties of silk

Silk belongs to a group of natural polymers that produce a wide variety of insects and spiders. In nature, silk has different structures and functions that are evolutionarily oriented toward the needs of the animal species that produce it in a given environment. The different functions of silk are classified as a series of tissue constructions, from tissue in the form of a victim trap (spider weaving), through safety rope (pull rope) to reproductive form (cocoon shape). Silk offers an excellent combination of lightweight (1.3 g/cm3), high tensile strength (over 4.8 GPa, as the strongest fiber known in nature), and outstanding hardness and elasticity (with a tightening length, before breaking the fiber by more than 35%). The tensile strength of silk yarn is comparable to most synthetic Kevlar-49 fibers, while the elasticity of silk fibers is 4–7 times higher than Kevlar-49 fibers, and the energy required to break the fibers is 3–4 times bigger. In addition to these remarkable mechanical properties, silk has thermal stability of up to about 250°C, which allows it to be processed over a wide temperature range [14].

The most commonly studied silk is derived from the larvae of Bombyx mori and the silk thread of the spider Nephila clavipes. Structurally, fibroin silks of these species are characterized by natural block copolymers, consisting of hydrophobic blocks with very regular repeated sequences, consisting of short chains of amino acids, such as glycine and alanine, and hydrophilic blocks with more complex sequences. Consisting of long chains of amino acids, such as charged amino acids (Figures 2 and 3) [15, 16].

Figure 2.

The primary structure of silk fibroin [wikipedia.org].

Figure 3.

a) Negatively charged disintegrated Anaphe moloneyi silk, dyed with sodium phospholphramate, with fibrous plaques, magnification 120.000x b) and c) fibers twisted by 900, fiber width, magnification 350,000x; d) a film poured from a fibroin solution of Bombyx silkworm obtained by treatment with cupriethylenediamine negatively charged, stained with sodium phospholphramate, magnification 120,000 x; e) fibers obtained from a sequentially charged polypeptide (alanine-glycerol-alanine-glycerol-serine) polypeptide, stained with sodium phospholphramate. Magnification 120.000x [J.Cell biol, 33, (1967), 289].

Hydrophobic blocks tend to form β-plates or crystals bound by hydrogen bonds and hydrophobic interactions, forming the basis of the tensile strength of silk fibroin. These arranged hydrophobic blocks are responsible for the high elasticity and hardness of the silk fibers [17].

The method of processing fibroin solutions into fibers by various bodies is still insufficiently studied and still remains in the field of intensive research. The process of obtaining fibroin fibers involves spinning a highly concentrated aqueous solution of silk fibroin into a non-Newtonian liquid crystalline state, in which the silk fibroin is lubricated and stabilized with water and forms micelle-like structures by separating the phases due to the process, and internal hydrophilic and hydrophobic block structures. The process is also supported by the water content and location. During the natural process of producing fibroin fiber, the concentration of the solution of silk fibroin in the glands gradually increases to form mycelium, which further aggregates to form globules or gel-like structures. At this stage, the fibrous silk protein is organized into a metastable state that maintains enough water to avoid premature conversion to the structure of the β-plate. The tensile stress during the spinning of the fibers (movements of the head in a silk caterpillar or pushing of a spider’s legs) determines the final assembly of the β-plate into crystal blocks [18].

In the final stage of spinning a silkworm, hydrophilic proteins, such as sericin, form a composite matrix with a silk fibroin core. The resulting silk fibers are insoluble in many solvents, such as water, ethanol, dilute acids and bases, hexafluoroisopropanol (hexafluoroisopropanol (HFIF)), calcium nitrate, or LiBr solution. The crystalline region of silk fibroin contains repetitive sequences rich in alanine or alanine glycine. These repetitive sequences are used as a basis for the genetic engineering of silk fibroin-like polymers in host systems, such as Escherichia coli, yeast, mammalian cells, and plants. Similar to natural fibroin fibers, most recombinant fibroin polymers have low water solubility due to their hydrophobicity [17, 19].

Strategies used to self-organize recombinant polymers similar to silk fibroin to increase solubility include, but are not limited to the following: i) the inclusion of molecular activators (triggers) of reactions, such as methionine reduction/oxidation reactions to control β-plate formation or phosphorylation/phosphorylation kinase action, ii) construction of chimeric polymer fibers of fibroin-like silk to incorporate α helical structures, and iii) inclusion of GVGVP (glycine-valine-glycine-valine-proline) domain characteristic of elastin to reduce crystallinity. The latter approach, based on copolymers of fibroin silk and elastin, whose hydrogels are exposed to physiological conditions, makes this combination of copolymers attractive for use in injectable systems for the controlled administration of therapeutic agents.

5. Conclusions

This review detailed the classification, types, and some important applications of biomaterials used in different applications in our life. From the biomaterial’s unique properties, these materials are perfect candidates for different bio-related applications. Biopolymers have attractive properties, particularly biodegradability, biocompatibility, selective permeability, and modifiable physical-mechanical properties. These properties find targeted applications in a variety of fields, particularly in the field of pharmacy and medicine. The use of natural biopolymers makes it possible to manufacture sustained-release forms that should avoid post-consumer peaks (diminish side and undesirable effects) and spread the effectiveness of this molecule over time (decrease the number of daily doses).