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Materials Science, Biomaterials
Chronic ulcers are among the main causes of morbidity and mortality due to the high probability of infection and sepsis and therefore exert a significant impact on public health resources. Numerous types of dressings are used for the treatment of skin ulcers-each with different advantages and disadvantages. Bacterial cellulose (BC) has received enormous interest in the cosmetic, pharmaceutical, and medical fields due to its biological, physical, and mechanical characteristics, which enable the creation of polymer composites and blends with broad applications. In the medical field, BC was at first used in wound dressings, tissue regeneration, and artificial blood vessels. This material is suitable for treating various skin diseases due its considerable fluid retention and medication loading properties. BC membranes are used as a temporary dressing for skin treatments due to their excellent fit to the body, reduction in pain, and acceleration of epithelial regeneration. BC-based composites and blends have been evaluated and synthesized both in vitro and in vivo to create an ideal microenvironment for wound healing.
- bacterial cellulose
- polymer composites
- biomedical application
1. Introduction
Cellulose occupies the first position among the most abundant biomasses found in nature [69] and has two native forms: (1) pure cellulose, which is obtained directly from its natural state, such as BC and that produced by some algae species, and (2) complex cellulose, which contains impurities, such as lignin, pectin, and hemicellulose [70].
First reported by A. G. Brown in 1886 [71], Although bacterial cellulose (BC) consists of a translucent, gelatinous film composed of micro and nano fibrillary cellulose distributed in unsystematical directions.
This biopolymer is produced extracellularly, through an aerobic process, which can be of cell-free enzyme systems [72,73], by acetic-acid Gram-negative bacterial cultures of Aerobacter, Agrobacterium, Komagataeibacter, Pseudomonas, Achromobacter, Azobacter, Rhizobium, Salmonella, and Alcaligenes and Gram-positive bacterium Sarcina ventriculi. These aerobic and non-photosynthetic bacteria, or aerobes are generally found in alcoholic beverages, vinegar, fruits, and vegetables [74]. The most efficient BC producer belongs to the genus Komagataeibacter (previously called Gluconacetobacter) due to its greater production capacity and ability to grow in media with a diversity of carbon/nitrogen sources [75].
The use of a system free of cells is a possible cellulose production strategy, as it can improve the fiber strength and density. This process provides the carbon source uniquely for cellulosic production and for a long period, when compared with the production by microbial cell system [72,73]. The literature shows that the BC production in a cell-free system can operate even in anaerobic conditions. Ullah et al. [72,73] concluded that higher carbon source availability content (such as dextrose) in the production medium for longer fermentation time favours the microfibrillar synthesis by the cell-free system, in contrast to a bacterial cell system. This prolonged the synthesis of cellulose, resulting in larger pore diameter and a more compact cellulosic film fibrillar arrangement. The cell-free cellulose showed lower values for its crystallinity degree, a lower water release rate, higher tensile strength, a slightly lower elongation at break (strain), and higher thermal stability [72,73]. Such characteristics must be taken into consideration according to the intended application.
This unique nanofibrilated matrix is being extensively investigated for various applications. As it can be produced in several configurations, it is highly versatile polymer. The intrinsic characteristics of BC, namely its biodegradability, mechanical strength, biocompatibility, haemocompatibility, micro and nanoporosity, and its distinctive surface chemistry, show how this biopolymer satisfy the demands for a great many biomedical applications.
Inter and intramolecular covalent hydrogen bonds in the hydroxyl (–OH) group of the cellulose chain impede the solubility of cellulose in water. Such bonds play a substantial function as a ligand that maintains the polymeric chains of cellulose together, thereby conferring to the cellulose matrix’s a high tensile strength [67]. This makes a material adequate for tissue engineering applications, as the material is able to maintain its structure even if applied to the human body’s natural physiological conditions. BC’s great capacity of water absorption and strong mechanical properties are excellent attributes for biomedical applications [67].
2. Bacterial Cellulose Synthesis and Production
BC is predominantly produced from C sources, mainly being glucose. However, other carbohydrate sources have also been reported, such as fructose [76], sucrose [77], galactose [78], corn steep liquor [77], and agro-industrial by products [79,80]. This biopolymer is produced via a progression of microbial enzymatic reactions, where the conversion of sugars into dextrose happens and then occurs their polymerization into cellulosic chains [81]. Molecular studies revealed the collaboration in the supramolecular assembly of cellulose fibrils of certain genetic operons in the cellulosic biosynthesis and extracellular transport. The structural and functional properties of BC, and its production yield have the possibility of genetic and metabolic modelling, through genome sequencing of the various BC-producing strains, this allows a functionalization for multipurpose applications [82].
The biosynthesis of BC serves various purposes for the microorganism, such as the protection of physiological, mechanical, and chemical stability, the enhancement of interactions and nutrient diffusion [83]. BC is produced by a biochemical process through oxidative fermentation in both non-synthetic and synthetic media, with the control of specific enzymes. Production starts in the microbial cells, cytoplasm, with the synthesis of β-1,4-glucan chains. These chains crystalize and form microfibrils that will later produce small pellicles that will then form membranes [70]. In a more detailed explanation, the biosynthesis pathway for BC initiates with the isomerization of a glucose sugar phosphorylated at the hydroxy group on carbon 6 molecule into the same molecule with a -OH group on carbon 1. Afterwards, it reacts with uridine-5′-triphosphate, forming uridine-50-diphosphate-alfa-D-glucose, that is later polymerized into 1,4 glucan chains in a linear conformation. The recently produced cellulose polymer chains are then secreted across the bacterial cell wall [84,85].
Production in a static culture result in a BC membrane, whereas an agitated culture results in the formation of suspended fibers in the form of irregular pellets [90]. Static BC production takes place at the air-liquid medium interface, whilst by agitated production, the pellets are formed submerged in the liquid fermentation medium. The usage or more complex techniques can result in cellulose with different morphologies, such as hollow spheres [91], aerogels [92] and even a water-in-oil emulsion [93]. BC’s resulting properties, micro and nanostructure, and morphology of BC are different. The production method of choice is conditional on the ultimate BC’s applications. However, the static culture method for production of BC still widely remains the selected approach.
In order to reduce BC production costs, there have been works focusing on the possibility and practicability of industrial, mainly agricultural-based residues and wastes being used as nutrient sources [76]. The BC from alternative mediums have similar physicochemical properties as those produced from the standard HS medium.
The unique synthesis process gives BC highly desirable physicochemical, structural, and biological properties, such as high purity, a high degree of polymerization, high hydrophilicity, high crystallinity (ranging from 60% to 90%) and even ex situ modifications into alternative formats [94,95,96]. Studies on the growth kinetics of microorganisms capable of producing BC membranes assist in the obtainment of specific thicknesses through changes to the production media or growth conditions, thereby facilitating the use of BC for the most diverse applications [97,98].
3. Bacterial Cellulose Applications
Unlike other types of synthetic membranes and even other types of biopolymers, the unique characteristics, properties and versatility make BC a biopolymer with considerable potential to be exploited by biotechnology in broad range of applications, such as medicine [67], foods [99], surgical and protection masks [100], cosmetics [5], electronics [101,102] and packaging [10,103], as well as for the separation of oily mixtures [7,104], among other fields.
BC can also be adapted to other applications through chemical modification or the functionalization of synthesized nanocellulose [104,105]. Such methods involve the modification of BC’s functional groups located on the surface of the membrane or pellets through different approaches, such as the addition of ionic charges by amidation, phosphorylation, acetylation, sulfonation, oxidation, carboxymethylation, etherification, silylation, etc. [106]. The benefits of the use of BC and its products for the treatment of wounds and infections and for patient recovery can revolutionize the biomedical field.
4. Bacterial Cellulose in Biomedicine
BC is a nontoxic, biocompatible, moldable, highly absorbent biopolymer [107]. Despite its promising chemical and physical properties, biomedical applications are limited because BC naturally lacks antimicrobial activity. However, studies have been conducted to overcome this problem with the incorporation of active molecules and materials with antimicrobial properties into the cellulosic matrix via several in situ and/or ex situ strategies for the development of novel polymeric materials [2,17,108,109,110].
BC’s fibrillar structure with its nanoscale and high porosity is an appropriate macromolecular support for the absorption and adsorption of active substances, pharmaceuticals, and drugs. As it has a neutral electrostatic charge, this facilitates the incorporation of bioactive compounds with both negative and positive charges [15], facilitating their incorporation onto the polymeric matrix. Therefore, they are ideal for the development of innovative specific controlled release systems, especially in biomedical engineering, including wound dressing and transdermal drug delivery systems [107]. In addition, when used as a membrane, it can contribute to an increase in cellular adhesion as well as the proliferation, migration, and differentiation of cells, thereby accelerating the re-epithelialization, which results in a faster wound healing process [15,111]. As previously stated, its performance as a biomaterial has attracted attention for use in drug delivery systems [15,111,112,113], wound dressings [114], tissue scaffolds [115], and implants [116].
Due to its unique characteristics previously mentioned, BC used in skin wound treatment has great potential. Studies offer promising results regarding the modification of BC to give it antimicrobial properties. The different modifications include the incorporation of antibiotics (such as Bacitracin and Amoxicillin) [15], silver nanoparticles [117,118], copper nanoparticles [119,120], silver chloride nanoparticles [121], silver montmorillonite nanoparticles [122], impregnation of montmorillonite nanocomposites [123], benzalkonium chloride solution [124] the use of immobilized lysozyme in bacterial cellulose nanofibers [125], the incorporation of propolis extract [5] and the creation of a BC–chitosan composite [123]. BC can also be applied as scaffolds for seeding of cells. The literature has shown several types of cells that can grow in the presence of BC [47,126,127,128,129].
One of the proliferation phases that occur following a wound consists mainly of the migration of fibroblasts from different sources, which results in the development of new connective tissue and microscopic blood vessels (granulation tissue). Fibroblasts are responsible for repairing damaged tissue by providing a new extracellular matrix, which is followed by the closure of the wound [130]. Studies have investigated the combination of cellulose and other biomaterials to enhance the proliferation, infiltration, and adhesion of fibroblasts [46,120,131].
As mentioned above, BC can be used for the development of innovative materials. Specially designed materials whether of a synthetic or natural origin that regulate the environmental conditions of a wound are at the forefront of regenerative medicine [111].
The fabrication of BC-based biomaterials includes biological, chemical, or physical methods to enhance the properties of these materials for application in a specific field [132]. A considerable number of studies have been carried out to improve the BC properties to enable its use in biomedicine by incorporating other polymers [30], nanoparticles [109], active molecules, or extracts [5].
To gain a better understanding of the biocompatibility potential of BC, Pértile et al. [39] surgically performed the subcutaneous implantation of BC in rats to provide greater contact with biological tissues in comparison to wound dressings. The results demonstrated that the BC structure exerted a positive influence on cell invasion and the behavior of the implant over time. The macroscopic examination revealed that the BC implants maintained their shape, but internal fissures lined with migratory stem cells were evident in the histological analysis. The authors found an absence of clinical signs of inflammation at the incision sites. The cellular response evolved progressively to chronicity, with a reduction in inflammatory cells around the implants and a predominance of macrophages over neutrophils. After three months, macrophages, fibroblasts and endothelial cells were predominantly found on the implants. All animals implanted with cellulose nanofibers survived and presented development throughout the period studied. As expected, the BC implants in this experiment did not provoke an uncommon response of body systems. Thus, it was clear that BC did not cause a foreign-body reaction beyond the formation of a thin fibrous layer [39].
BC has also been used in biomedical tissue engineering and bone grafting. Such applications are possible due to the composition of bones, which consists of a inorganic mineral phase (also called (also called inorganic bone phase, bone salt, or bone apatite, which consists mainly of calcium hydroxyapatite) and an organic phase (composed mostly of collagen and non-collagen proteins) [133]. Thus, BC can be used as a matrix for the obtainment of calcium carbonate crystals, which are believed to improve biocompatibility. Studies conducted by Stoica-Guzun et al. [133] revealed that BC nanofibrils can reproduce the characteristics of collagen nanofibrils for the deposition of calcium and phosphorus through biomineralization.
Studies have also evaluated the use of BC for cardiovascular applications, with BC used in the production of blood vessels in in vivo tests. These “synthetic veins” were developed by Klemm et al. [1] and used to repair the carotid artery in a rat. After one month, the BC/carotid artery complex was found to be enfolded with connective tissue, demonstrating BC’s potential as replacement of blood vessels. BC have also been reported as an advanced biosensing [134,135] and diagnostic materials [43].
Considering the properties of BC and possible modifications and functionalization to obtain novel biomedical functions, this biopolymer has considerable potential in the treatment of wounds. Indeed, BC demonstrates superiority over conventional dressings due to its easy attachment to the skin without restricting the movements of the patient. Thus, microbial cellulose has been increasingly used for wound dressings and tissue engineering applications [136,137].
4.1. Functionalized Bacterial Cellulose for Biomedical Applications
According to Lima-Júnior et al. [138], wound dressings need to be effective, offer practical applicability and be easy to produce, obtain and market. Thus, the materials that compose dressings need to be inexpensive, enable easy storage and prolonged stability, must not have antigenicity, should have good flexibility and stretch resistance, offer good adherence to the wound site, as well as good adaptation around the wound while facilitating joint movements. Such materials should also be applied in a single surgical session, offer easy handling, attenuate pain, accompany body growth and maintain body temperature. BC is compatible with such characteristics, which suggests the considerable potential of this biopolymer for applicability as a biomaterial in medical applications [138]. The characteristics that are essential for optimal wound dressing is thermal insulation, biocompatibility, cost efficacy, mechanical stability, non-toxicity, to maintain a moist wound environment, infection prevention, and adequate gaseous exchange [138]. All the aforementioned characteristics are inherited to BC.
During the wound treatment process, proper moisture is required to enable a shorter recovery time. The high water-holding capacity enables microbial cellulose to conserve the ideal moisture of the skin’s wound and even ulcers site. Due to the nanofibril network, these membranes can have additives incorporated into the matrix and create a physical protection that impedes the infiltration of microorganisms, thereby diminishing the risk of infection while also reducing both pain and healing time [139].
As specified by Czaja et al. [140], BC’s biocompatibility for bandages and dressings is related to its distinct structure, which serves as an adequate environment for wound healing. This nanofibrillar structure is able to aid in eliminating the discomfort symptoms by increasing the adsorption of exudate from the wound and isolating the skin’s nerve endings. Excessive exudate results in the separation of tissue layers, which hinders the healing and tissue regeneration process. Thus, the adequate absorption of exudate is an important aspect in the development of modern dressings [140]. In comparison to conventional dressings, such as moist and ointment-impregnated gauze, BC enables a faster skin healing process. Moreover, BC has shown good cytological and histological compatibilities [141], diminishing the risk of infection and sepsis.
A balance is needed between the absorption and adsorption of these fluids and the release of moisture, as the dehydration of the wound surface hinders the successful recovery of the tissues. Because of its high water-holding capacity (WHC) and water/moisture release rate (WRR), BC is a material with great potential for wound dressing applications. Moreover, its membranes structures can be adapted to diverse situations in this type of application [142]. Studies have demonstrated that BC-based linings diminish the pain of the wound, quickens the re-epithelialization, and lessens would infection rates and scars [19,69,118,131].
The main characteristics of currently available wound treatment materials are good absorption and permeability. However, these materials often cause trauma and harm upon removal from the wound site. Comparing the properties of BC to conventional materials used in the treatment of wounds, membranes produced by microbiological fermentation can be used directly after the fermentative process following rinsing with running water. BC membranes can also be processed in different forms suitable for various wound dressing applications, as previously mentioned [136].
Another important characteristic of wound dressings is the capacity to remain structurally intact between placement and removal, especially when placed close to a joint, as the movement of the body can lead to the exposure of the wound. The tensile strength of BC membranes is an important factor and depends on both the culture conditions and treatment. Tensile strength can reach 260 MPa, with stretching up to 32% prior to breaking. Such mechanical properties of strength and flexibility demonstrate that BC is adequate for a variety of dressings in different types of treatment and sites [136,142,143].
Depending on the usage of the BC’s polymeric matrix, it is interesting to modify its porosity, as appropriate porosity such characteristic of the biomaterial needs to be similar to the replaced tissue that is going to take place. Nicoara et al. [111], modified BC through in situ and ex situ and obtained a BC/Hydroxyapatite (HAp) composite with the incorporation of silver nanoparticles (AgNPs) with 10–70 nm size. All obtained materials demonstrated a homogenous porous structure and high-water absorption capacity, <5% degradation rates in artificial human blood plasma, and good antimicrobial action due to the AgNPs. The obtained composite’s prepared via in situ showed a wider porosity distribution and better homogeneity [107]. Other innovative approaches to BC’s modification have been widely reported in literature [139], including 3D-bioprinting. Studies show bioprintable BC as a medical material to be utilized in different tissues and scaffolds [44,61,98,114,128].
Traditional bandages, such as gauze dressings, are convenient for drug delivery. However, such dressings adhere easily to the exudate, causing secondary wounds and even infection [144]. Infection prevention should be of low cost to assist in wound healing and to be easily removed [114]. Modern dressings have been produced with the objective of reducing inflammatory and immunological diseases while also preventing dehydration and enhancing the healing process [132]. Several hydrogel polymers that can be easily manufactured have been used for wound dressings, such as collagen [108], alginate [145], cellulose [146], and composites [112].
BC is an inherit occurring material with nanoporosity, and this highly desired property has attracted scientists as shown by the increasing numbers of annual publishing that appear in ‘Google Scholar’ involving the descriptors ‘bacterial cellulose’ and ‘medicine’ and ‘bacterial cellulose’ and ‘wound dressing’ or their combinations as the search words.
Due to BC’s polar molecules and porous geometry, the cellulosic matrix has been widely processed with other materials and polymers resulting in blends and composites for targeted applications. The coalescence of additives onto its polymeric chains has led to unique features, such as transparency [147] bactericidal activity [10], and enhanced biocompatibility [148,149].
Literature shows other very interesting manuscripts on BC modifications. Some of which are: In situ and ex situ obtention of BC/Hydroxyapatite (HAp) composite incorporated with AgNPs [111]; Printable BC/polycaprolactone (PCL) composite loaded with antibiotics [113]; In situ composite of transparent antimicrobial AgNPs/BC films [150]; BC whiskers and poly (2-hydroxyethyl methacrylate) (pHEMA) hydrogel incorporated with AgNPs via ex situ [151]; BC/gelatin (Gel) membrane guided with electrofield (EF) stimulation [152]; BC/Gel/selenium nanoparticles (SeNPs) in situ nanocomposite hydrogel synthesis [153]; BC impregnated via ex situ with antibacterial bioactive extracts [154]; Addition of semi-dissolving microneedles and TEMPO-oxidized BC nanofibers [155]; Nanopolymer blend of BC and polyacrylamide mesh [156]; BC polymeric blend with low molecular weight deacetylated chitin biopolymer [157]; BC membrane reticulated with citric acid and additivated with inorganic catalysts [158], and curcumin-loaded BC nanocomposite prepared by ex situ method [159].
BC’s porous network exerts a positive response human body’s cell. That is, dressing materials with adequate moisture degree are able to accelerate the wound healing procedure and to protect from eventual microbial contamination. Thus, the ability to manage BC’s porosity can be utilized to modify its WHC and WRR. Both parameters are essential in determining the applicability of BC as wound dressing material [162,163,164].
According to Dahman (2009) [165], the hydrophilic polarity, high number of free fibrils and high surface area are responsible for BC’s high WHC, reaching 100–200 times its dry weight. This characteristic makes microbial cellulose a successful material for burn and scalds treatments, assisting the skin’s thermoregulation of surface moisture content [166].
Another important point is the purification of BC-based dressings. Efficient purification of BC’s raw material must be performed to guarantee the complete removal of residues from the culture medium and bacterial cells that can cause contamination during the use of the product. Generally, the thermal purification of the membrane with a basic pH solution, such as NaOH solution between 0.1 and 1M at 60–100 °C for 1–3 h, followed by pH neutralization with the aid of organic acids, or rinsing in running water until achieving the desired pH [140].
To ensure biosafety in this and other fields of application, Nascimento et al. [167], conducted a study to determine whether gamma irradiation could be used for a simple, effective sterilization of the BC membranes. Gamma irradiation is often used as a sterilization method for medical products and equipment. However, due to its high penetration power, it was necessary to assess its reactions on the physicochemical and structural properties of the membranes. The researchers used cobalt-60 as the irradiation source. The results demonstrated that gamma irradiation (at 25 kGy) did not cause any relevant alterations to the polymeric properties of membranes and therefore constitutes an effective sterilization method for this material.
Active principles that inhibit microbial growth represent another crucial characteristic for dressing used in the treatment of wounds, chronic ulcers, and burns. The antimicrobial function can be added through a modification of the structure and impregnation with antimicrobial agents, such as biopolymers [168], cationic antiseptics [169], antimicrobial peptides [170], antibiotics [171], natural active compounds [5], or inorganic nanocomposites with bactericidal properties [172].
Some works that study BC for transdermal drug delivery include cellulose membranes embedded with anti-inflammatory drugs [173], reducing agent compounds [174], nanoparticles [175], etc., aiming for similar human skin permeation rates to the already commercialized patches.
4.2. Bacterial Cellulose in Medicine
According to the American Academy of Alternative Routes of Drug Administration [176], drugs can be administered to the human body through different anatomic routes. The chosen used material conditions the most adequate route of administration, and that is essential to ensuring the success of the therapeutic process.
Studies on polymers of a natural and artificial origin have demonstrated different drug delivery systems (DDS) ensuring minimal or no side effects. Nanotechnology used jointly with nanomembranes is a novel, promising strategy for DDS [177]. According to the authors cited, the nanocarriers’ properties responsible for enhancing the efficiency of DDS include biocompatibility, biosafety, encapsulation capacity, molecular polarity, bioavailability, and therapeutic efficiency (controlled distribution and release, cellular absorption, excretion, pharmacokinetics, toxicity and depuration).
The development of biomaterials that enable the controlled release of medications is of considerable interest, as the administration of medications in pure form has undesirable effects, such as the rapid degradation of substances in the organism, distribution to non-target tissues and organs and a possible significant reduction in the effective concentration in the target tissue. Moreover, the combination of these factors can lead to systemic toxicity [176]. The controlled administration of medications enables the extension of treatment with the use of controlled concentrations that reduce the risk of irritation and enable the use of substances with a short biological half-life [176].
Different materials can be used as the base, vehicle or matrix for the development of controlled DDS. BC is a viable option for this purpose and has been used as a topical agent for the encapsulation and delivery of different types of active compounds, including insoluble drugs [176,178]. Recent advances in the use of BC for controlled DDS include oral, ocular, intratumor, topical and transdermal delivery. Besides its use as a transporting system, BC can also be employed to encapsulate drug excipients, such as thickeners, emulsifiers, stabilizers and surfactants [179].
The drug release process is controlled by diffusion, which depends directly on pH and can result in different responses that can be adapted through the use of physical treatment or chemical modification [180]. BC can modify the drug release process through water retention, enhanced adhesion, or the formulation of a film [179].
4.3. Bacterial Cellulose for Drug Delivery
Hydrogels are materials with 3D network formed by cross-linked hydrophilic polymers. These materials have great uptake/holding and release capabilities for water and other polar fluids [180,181,182,183,184]. Amidst the several polymeric materials, nanocellulose-based hydrogels have received attention [184].
At the end of the 20th Century, BC was used for the first time as non-permanent skin replacement and under commercialization named BioFill®, currently known as Dermafill™. The product is made of partially dried BC membranes for the treatment of damaged skin by thermal burns, abrasions, lacerations and ulcers. The performance of the dressing was better than conventional dressing with regards to pain relief and the acceleration of the healing process [35].
This entry is adapted from the peer-reviewed paper 10.3390/molecules27175580
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