OzlemAteş1 and Ebru Toksoy Oner2*
1 Department of Genetics and Bioengineering, Nisantasi University, Istanbul, Turkey
2 IBSB, Department of Bioengineering, Marmara University, Istanbul, Turkey
*Corresponding author e‐mail: ebru.toksoy@marmara.edu.tr
Biopolymers have received considerable research attention over recent decades since they have potential applications in various fields of the economy such as food, packaging and chemical industries, agriculture, and medicine due to their compatibility with human lifestyle as well as their biocompatibility, biodegradability, and environmental credentials. These naturally occurring polymers have many uses as adhesives, absorbents, lubricants, soil conditioners, cosmetics, drug delivery vehicles, textiles, high‐strength materials, implantable biomaterials, controlled drug carriers or scaffolds for tissue engineering (Guo and Murphy 2012; Ige et al. 2012; Kazak et al. 2010). Based on the production processes and origin of raw materials, these environmentally friendly biopolymers can be categorized as microbial biopolymers (polymers synthesized by micro‐organisms like bacterial exopolysaccharides), natural biopolymers (obtained from biosources like plant polysaccharides), and synthetic polymers (Poli et al. 2011; Spizzirri et al. 2015).
Bacterial polysaccharides, one of the main groups of microbially produced biopolymers and hydrocolloids, show great diversity and functions with unique and commercially relevant material properties. Therefore, there is increased interest in bacterial polysaccharides in industrial, biomedical, chemical engineering, and food science areas (Fahnestock et al. 2011; Nwodo et al. 2012; Rehm 2010).
Based on their biological functions, microbial polysaccharides can be subdivided into the exopolysaccharides (EPSs) (xanthan, levan, alginate, cellulose, etc.) and the intracellular storage polysaccharides (glycogen, starch) (Schmid and Sieber 2015). Bacterial EPSs such as dextran, xanthan, gellan, curdlan, pullulan, acetan, and levan are used commercially and are well‐known industrial microbial polysaccharides with numerous applications and a considerable market (Freitas et al. 2011; Kazak et al. 2010; Llamas et al. 2012; Ptaszek et al. 2015).
Most polysaccharides are hydrocolloids that are widely used in the food industry for their extensive range of functionalities and applications. They can be utilized as stabilizers, emulsifiers, thickeners, and gelling agents primarily in food products such as bread, sauces, syrup, ice cream, instant food, beverages and ketchup, and most of these hydrocolloids are classified as food additives (Ahmad et al. 2015; Pegg 2012). In 2008, the global hydrocolloid market, currently dominated by the algal and plant polysaccharides such as starch, galactomannans, pectin, carrageenan, and alginate, had a market value of US$4 million and it is expected to reach US$3.9 billion by 2012 and US$7 billion by 2019 (Patel and Prajapati 2013; Williams et al. 2002). However, production of these algal and plant polysaccharides is highly affected by climatological and geological conditions whereas microbial polysaccharides are produced by fermentation under controlled conditions with high and stable yields, which make them economically competitive with polysaccharides of algal and plant origin (Kaur et al. 2014).
In this chapter, after a brief description of microbial exopolysaccaharides and their use in the food industry, the bacterial EPSs xanthan, levan, gellan, and curdlan are discussed in more detail with a special focus on their uses as food additives.
In recent years, considerable attention has been given to the environmental and human compatible biopolymers which have been used as biotechnology products in numerous sectors such as food, chemical, agriculture, and health (Guo and Murphy 2012; Vroman and Tighzert 2009).
Polysaccharides are natural, non‐toxic, biodegradable polymers that cover the surface of most cells and play important roles in various biological mechanisms such as immune response, adhesion, infection, and signal transduction (Yasar Yildiz and Toksoy Oner 2014). Besides the interest in their applications in the health and bionanotechnology sectors, polysaccharides are also used as thickeners, bioadhesives, stabilizers, probiotics, and gelling agents in the food and cosmetic industries and as emulsifiers, biosorbents, and bioflocculants in the environmental sector (Toksoy Oner 2013).
The plant, algal, and microbial polysaccharides are adequate for the market but there is a demand for eco‐friendly technologies. Therefore, the use of bacterial polysaccharides for industrial applications is expected to increase sharply (Kumar et al. 2007). Since microbial sources allow high‐yield, sustainable and economical production of polysaccharides under controlled conditions to the specifications required at industrial levels, they are preferred to plants and algae (Kazak et al. 2010; Toksoy Oner 2013). Production by microbial sources is achieved within days to weeks whereas 3–6 months are required for production from plants. Moreover, geographical or seasonal variations and sustainable use of agricultural land are ever increasing concerns for production processes relying on plant resources. Production from microalgae is dependent on solar energy but there is no such requirement for microbial production, which also utilizes different organic resources as fermentation substrates (Donot et al. 2012; Toksoy Oner 2013)
The term exopolysaccharide was first used by Sutherland (1972) to define high molecular weight carbohydrate polymers produced by marine bacteria. EPSs are synthesized and secreted by various micro‐organisms (gram‐positive and gram‐negative bacteria, fungi, and some algae) into the extracellular environment as either soluble or insoluble polymers. Therefore their composition, functions, and chemical and physical properties that establish their primary conformation vary from one bacterial species to another. EPSs are composed mainly of carbohydrates (a wide range of sugar residues) and some non‐carbohydrate factors (such as acetate, pyruvate, succinate, and phosphate) (Llamas et al. 2012; Nicolaus et al. 2010; Staudt et al. 2012; Vu et al. 2009).
Exopolysaccharides can be classified based on monomeric composition as homopolysaccharides, composed of a single type of monosaccharide, like dextran, levan or mutan, and heteropolysaccharides which have complex structures and are made up of various types of monosaccharides. They also vary in size from disaccharides to heptasaccharides like xanthans or gellans (Donot et al. 2012; Llamas et al. 2012; Nicolaus et al. 2010; Nwodo et al. 2012; Schmid and Sieber 2015).
Bacterial EPSs have long been accepted as significantly important biomaterials in industrial areas. EPSs have found broad application in the food, pharmaceutical, cosmetics, and petroleum industries in which emulsifying, viscosifying, suspending, and chelating agents are required since they have a variety of unique and complex chemical structures that offer beneficial bioactive functions and potential applications. The first bacterial polymer, dextran, was discovered by Louis Pasteur (1861) in the mid nineteenth century as a microbial product in wine.Numerous articles are published on new microbial EPSs each year but few EPSs have continued to exist industrially (Donot et al. 2012; Kumar et al. 2007; Liang and Wang 2015; Llamas et al. 2012; Ordax et al. 2010).
In more recent years, several novel bacterial EPSs have been isolated and identified, but only a few of them have achieved significant commercial value due to high production costs and poor properties (Freitas et al. 2011; Kumar et al. 2007; Llamas et al. 2012; Mata et al. 2006; Nicolaus et al. 2010). Some of the bacterial EPS with superior physical properties (xanthan produced by Xanthomonas campestris, gellan produced by Sphigomonas paucimobilis, dextran produced by Leuconostoc mesenteriodes and curdlan produced by Alcaligenes faecalis) can be used instead of plant (guar gum or pectin) or algae (carrageenan or alginate) polysaccharides in traditional applications (Freitas et al. 2011; Liang and Wang 2015; Nicolaus et al. 2010). Other bacterial EPSs with unique properties such as levan provide various new commercial opportunities due to their biological activities (Freitas et al. 2011).
Although there is a wide range of microbial EPSs (Table 7.1) that have industrially promising physicochemical properties, the global market is still dominated by plant and algal polysaccharides. For commercialization of new polymers in large markets, it is crucial to overcome economic problems by lowering their production costs. The main costs are the substrate and infrastructures required for production, including bioreactors and maintaining asepsis (Donot et al. 2012). Use of cheaper sources, optimization of fermentation conditions to improve product yield, development of higher yielding strains (e.g., by mutagenesis or genetic manipulation) and improvement of downstream processing are involved in approaches to decrease production costs. Moreover, there is increased interest in studies of metabolic pathways of EPS synthesis and regulation to optimize microbial EPS production (Sousa et al. 2011).
Table 7.1 Commercially available significant microbial polysaccharides and their characteristics.
| Microbial EPS | Monomeric units | Producer | Main properties | Industrial applications |
| Xanthan | Glucose, mannose and glucuronate | Xanthomonas spp. | Hydrocolloid, high viscosity yield at low shear rates even at low concentrations, stability over wide temperature, pH, and salt concentration ranges | Thickening, stabilizing agent, food additive |
| Levan | Fructose | Halomonas smyrnensis AAD6T, Zymomonas mobilis, Bacillus subtilis (natto) | Low viscosity, high water solubility, biological activity, antitumor activity, anti‐inflammatory, adhesive strength, film‐forming capacity | Emulsifier, stabilizer and thickener, encapsulating agent, food and feed additive, osmoregulator, and cryoprotector |
| Curdlan | Glucose | Agrobacterium spp., Rhizobium spp. and Cellulomonas spp. | Gel‐forming ability, water insolubility, edible and non‐toxic, biological activity | Thickening, stabilizing agent, food additive |
| Gellan | Glucose, rhamnose, glucuronic acid | Sphingomonas elodea, S. paucimobilis | Hydrocolloid, stability, gelling capacity, thermo‐reversible gels | Stabilizer, thickening agent, structuring and versatile gelling agent |
| Dextran | Glucose | Leuconostoc spp. and Streptococcus spp. | Non‐ionic, good stability, Newtonian fluid behavior | Blood plasma extender and chromatography media |
| Pullulan | Glucose | Aureobasidium pullulans | Water soluble, adhesive ability, forms fibers, films: transparent, printable, heat sealable | Thickening, stabilizing, texturizing, gelling agent |
| Alginate | Mannuronic acid and glucuronic acid | Pseudomonas spp. and Azotobacter spp. | Hydrocolloid, gellling and film‐forming capacity | Biomaterial, thickening, stabilizing, and gellifying agent |
| Cellulose | Glucose | Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and gram‐positive bacteria | High crystallinity, ınsolubility in most solvents, high tensile strength, moldability | Food (nata de coco), diaphragms of acoustic transducers and wound dressings |
EPS, exopolysaccharide.
Extremophilic micro‐organisms isolated from extreme environments, such as deep‐sea hydrothermal vents, Antarctic ecosystems, saline lakes, and geothermal springs, have also been studied as potential sources of highly valuable EPSs (Freitas et al. 2011). Since most commercial EPS‐producing strains have been pathogenic, more research has been focused on discovering and developing novel and functional EPSs produced by extremophilic strains. Several bacterial strains of thermophiles (heat loving), psychrophiles (cold adapted), halophiles (hypersaline bodies), acidophiles (acid resistance), and alkaliphiles (alkaline environment survivors) have been reported as EPS producers (Kazak et al. 2010, 2015; Nicolaus et al. 2010; Yasar Yildiz et al. 2014).
Microbial EPSs have rheological properties that match industrial demands and can be produced in large amounts and with high purity (Patel and Prajapati 2013). They can be used as viscosifying agents, stabilizers, emulsifiers, gelling agents, or water‐binding agents in foods. Xanthan and gellan, which are produced by Xanthomonas campestris and Pseudomonas elodea respectively, are the two main commercially available bacterial polysaccharides authorized for use as food additives in the United States and Europe (Morris 2006; Patel and Prajapati,2013). Dextran, curdlan, and cellulose, which are homopolysaccharides of glucose monomers, are neutral bacterial glucans (Nasab et al. 2010). Curdlan can be used as a gelling agent. The major market for pullulan is the food industry. Alginates produced by species of Pseudomonas and Azotobacter are widely used as thickening, stabilizing, and gelifying agents in the food, textile, paper, and pharmaceutical industries (Hay et al. 2014). Almost 30 species of lactic acid bacteria (LAB) are also known as polysaccharide producers but low production yields make it difficult to exploit them commercially. On the other hand, lactobacilli are GRAS (Generally Recognized as Safe) bacteria and their EPS could be utilized in foods (Badel et al. 2011). Besides the well‐known levan‐type and inulin‐type fructans commonly used in the food sector and synthesized from sucrose by fructosyltransferases, alternan is an example of an α‐glucan polymer often found in the food industry and produced by the alternansucrase enzyme of Leuconostoc mesenteroides (Ryan et al. 2015).
The market for hydrocolloids, which include many polysaccharides such as levan, pullulan, xanthan, alginate, dextran, cellulose, starch, agar, carrageenan, and pectin, was valued at more than US$4 million in 2008 and is estimated to reach US$7.911 billion by 2019. North America is the most important consumer and Asia‐Pacific is the fastest growing market for hydrocolloids. Archer Daniels Midland Company, B&V SRL, Ceamsa, Danisco A/S, FMC Corporation, Gelnex, Kerry Group PLC, Taiyo Kagaku Co. Ltd, Ashland Inc., Cargill Inc., CP Kelco, and Fiberstar Inc. are the major companies in the global market (www.marketsandmarkets.com/Market‐Reports/hydrocolloid‐market231.html). Xanthan gum, which is the second EPS to be approved by the FDA as a safe food ingredient, is the only significant bacterial EPS in this market, with 6% of the total market value.
The principal reason for the extensive use of hydrocolloids in the food industry is their ability to bind with water and modify the properties of other ingredients. The modification of rheological characteristics is helpful in enhancing the sensory properties of foods. Hence hydrocolloids are employed as extremely significant food additives (Li and Nie 2016). EPSs are alternatives to conventional food additives since there is a high consumer demand for products with low fat or sugar content and low levels of food additives, especially stabilizers and thickeners (Jolly 2002; Robitaille et al. 2009).
Since they improve the texture of fermented dairy products and the functional properties of foods (yoghurt, cheese, and bread) and also confer health benefits with their immunostimulatory, antitumoral or cholesterol lowering activity, EPSs are widely found in the dairy and food industries and in pharmaceuticals, where they are used directly as biological active agents, i.e., as a probiotic (Poli et al. 2011; Tieking and Gänzle 2005). EPS‐producing starters are used in dairy products since with EPS, the amount of added milk solids is reduced, yoghurt has better water‐binding capacity and its viscosity is improved, texture and mouth feel are enhanced and syneresis is avoided during fermentation or upon storage of fermented milk products such as dahi, yoghurt and cheeses, sour cream, and kefir. EPS cultures have also been shown to affect product rheology in European cultured dairy products (Patel and Prajapati 2013). Cheese fermented by EPS‐producing Streptococcus thermophillus and Lactobacillus delbrueckii subsp. bulgaricus culture showed significantly lower hardness, consistency, adhesiveness, chewiness and relaxation in texture profile analysis (Ahmed et al. 2005). Lactobacillus kefiri and L. kefiranofaciens synthesize Kefiran heteropolysaccharide which is found in the fermented dairy beverage kefir (Maeda et al. 2004).
In bread production, EPSs produced by lactobacilli are used, since they favorably influence bread properties such as water absorption, softness, gluten content of dough and also improve the structure build‐up, increase the specific volume of a loaf and prolong shelf‐life (Tieking and Gänzle 2005).
EPSs like carrageenans, guar gum, and xanthan gum are used as ingredients and additives in flavored milks to provide sweetness, increase viscosity and enhance perception of texture and mouth feel. Some ice cream formulations also include fructans that influence texture and mouth feel and can allow the reduction of sugar and fat since they contain less energy (Early 2012).
In recent years, there has been a significant increase in the functional food market, including products formulated to maintain a “healthy” gut microbiota, sich as probiotics and prebiotics. EPSs that are related to the interaction of probiotics with the gut ecosystem are used in probiotics to improve bacterial persistence and colonization of a given niche (Salazar et al. 2016). Fructose‐oligosaccharides (FOS) have interesting properties for food applications as they have a low sweetness compared to sucrose, are essentially calorie free, and non‐carcinogenic. Food applications of inulin and FOS are based mainly on their prebiotic properties. Remarkably, fructose‐based EPSs can be fermented by gut microflora which leads to improvement of intestinal flora and increases mineral absorption (Patel and Prajapati 2013).
To encapsulate probiotic cells, various polysaccharides are employed to improve probiotic survival during processing and storage in food products or in gastrointestinal transit (Tripathi and Giri 2014). Gums and polysaccharides generally act as viscosifiers or stabilizers that are added to food products to avoid sedimentation of particles or creaming of emulsion droplets by increasing the viscosity of the product (Alting and van de Velde 2012).
Due to their flocculating ability, EPSs clarify liquids by precipitating colloidal suspensions. Therefore they can be employed as fining agents in the alcoholic beverage industry to remove hazes (haze stabilization) and to help filtration by increasing particle size (Buglass and Caven‐Quantrill 2012).
Microbial polysaccharides such as gellan, pullulan, xanthan, bacterial cellulose, curdlan, and levan have been applied in the development of edible and/or biodegradable membranes and edible coatings, to serve as barriers in food packaging. (Espitia et al. 2014; Freitas et al. 2014).
Commercially available gum‐based food thickeners have been widely used as additives in cold beverages and also they are used to obtain the optimal swallow response because of ease of preparation, convenience, reasonable cost, and the suspending ability of the thickened fluids (Cho and Yoo 2015).
Polysaccharides have been also used as fat replacers to improve technological characteristics or to enhance meat protein functionality (Villamonte et al. 2015).
Among several novel microbial EPSs, xanthan, levan, gellan, and curdlan are seen as the most promising polysaccharides for various industrial sectors (Donot et al. 2012).
Xanthan is the most commercially important microbial EPS with a rapidly growing demand (Frese et al. 2014; Vorhölter et al. 2008). Xanthan is a heteropolysaccharide composed of repetitive pentasaccharide units consisting of two glucose, two mannose, and one glucuronic acid residue with a backbone chain consisting of (1 → 4)‐β‐D‐glucan cellulose (Khan et al. 2007). Xanthan is a natural product produced by the plant pathogen proteobacterium Xanthomonas campestris pv. campestris (Xcc). Xanthan is a high molecular weight and anionic polysaccharide that can hinder the gel formation of myofibrillar proteins and enhance surface hydrophobicity under high pressure (Khan et al. 2007; Villamonte et al. 2015). It can be recovered easily from culture broth by precipitation using ethanol or isopropanol (Schatschneider et al. 2013).
Xanthan shows high viscosity at low concentrations in solution and strong pseudoplasticity, and it is stable over a wide range of pH, temperature, and ionic strengths (Patel and Prajapati 2013). It shows weak gel‐like properties at high polymer concentration (Milani and Maleki 2012). Since xanthan is resistant to pH variations, and stable in both alkaline and acidic conditions, it is used in a wide range of products. Due to its superior properties and rheological characteristics, xanthan is employed as a thickening or stabilizing agent in a wide range of industrial areas such as food, cosmetics, and oil drilling industries (Chivero et al. 2015; Schatschneider et al. 2013). It has been described as a “benchmark” product based on its significance in food and non‐food applications which include dairy products, drinks, confectionery, dressing, bakery products, syrups and pet foods, as well as the oil, pharmaceutical, cosmetic, paper, paint, and textile industries (Cho and Yoo 2015; Patel and Prajapati 2013). Xanthan is also employed in non‐food applications such as stabilizing cattle feed supplements, calf milk substitutes, agricultural herbicides, fungicides, pesticides, and fertilizers, and to impart thixotropy to toothpaste preparations (Morris 2006).
Xanthan was discovered in the 1950s by Allene Rosalind Jeanes at the United States Department of Agriculture, and it is the second microbial EPS approved as a food additive in 1969 by the US Food and Drug Administration. It was approved in Europe in 1982 under the E number E415. In 1988 the ADI value (acceptable daily intake) of xanthan was changed to “not specified” which confirmed it as a safe food additive (Khan et al. 2007; Petri 2015). Xanthan is commercially produced by several companies such as Monsanto/Kelco, SKW Biosystems, Cargill, Rhodia, and Archer Daniels Midland (Born et al. 2005). China has been one of the largest xanthan producers since 2005 (Petri 2015).
Aqueous dispersions of xanthan gum are thixotropic in either hot or cold water which means that products which are shaken and thinned during distribution will thicken when stored at the point of sale (Ahmad et al. 2015; Early 2012; Morris 2006). The thixotropy of xanthan samples allows manipulation and control of processes such as spreading, pumping, pouring, and spraying and has led to the development of a number of dry mix formulations such as sauces, gravies, and desserts. Moreover, xanthan can be heated or refrigerated without losing its desirable textural characteristics (Khan et al. 2007; Morris 2006).
The colloidal stability of emulsions and the shelf‐life of edible emulsions of oil in water increase in the presence of xanthan. These superior properties and its shear‐thinning behavior make xanthan gum an excellent emulsion stabilizer in salad dressings and sauces (Khan et al. 2007; Petri 2015).
In beverages, xanthan gum is effective for suspending fruit pulp and also enhances the taste of the drink. Therefore it can also be used in low‐calorie drinks (Cho and Yoo 2015; Khan et al. 2007).
Xanthan gum in whipped cream to a level of 0.1% increases the consistency of the cream (Zhao et al. 2009). Simsek (2009) reported that the application of various xanthan concentrations influenced the syruping formation of refrigerated dough formulation. Texture and apparent viscosity of ice cream are very important since they determine the quality of the product (Dogan et al. 2013). Use of xanthan in ice cream has several advantages as it provides viscosity to the liquids during processing, reduces the freezing point and enhances both the taste and texture of the product (Dogan et al. 2013; Maletto 2005). Halim et al. (2014) observed that ice cream containing xanthan gum showed the best effect on overrun and the quality of ice cream was increased in terms of pH value and total plate (colony) count. Buttered syrups and chocolate toppings containing xanthan have excellent consistency and flow properties and because of their high viscosity at rest, appear thick and appetizing on products such as pancakes, ice cream, and cooked meats (Palaniraj and Jayaraman 2011).
Xanthan improves processing and storage and enhances elasticity of batters and doughs, and it can be used as a gluten replacement in the development of gluten‐free breads (Khan et al. 2007; Morris 2006). For instance, the specific volume of wheat breads was significantly increased by xanthan at a dosage of 0.1% (w/w flour), whereas the crumb hardness of fresh breads could not be reduced by the addition of xanthan at all tested dosages (Guarda et al. 2004).
When xanthan is mixed with starch, the swelling of the starch effectively concentrates xanthan, leading to enhanced rheology of the bulk phase containing the enlarged swollen granules (Morris 2006; Petri 2015).The use of innovative packaging methods is possible due to the ability of xanthan to control pouring, pumping, spreading, and splashing. The mixture of xanthan and galactomannans, carob or tara forms transparent thermo‐reversible gels (Morris 2006). Gellan‐xanthan gum mixtures have been used for immobilization of bifidobacteria in beads to increase their tolerance of high acid environments (Sun and Griffiths 2000).
Levan is a homopolysaccharide of β(2‐6)‐linked fructose residues which is synthesized outside the cell from sucrose by the extracellular enzyme levansucrase (EC 2.4.1.10) by several bacteria, including Acetobacter, Aerobacter, Azotobacter, Bacillus, Corynebacterium, Erwinia, Gluconobacter, Mycobacterium, Pseudomonas, Streptococcus, and Zymomonas (Donot et al. 2012; Freitas et al. 2011; Nakapong et al. 2013). In addition, halophilic bacterium Halomonas smyrnensis AAD6T has also been reported as the first high‐level levan producer extremophile (Poli et al. 2009).
As a homopolysaccharide with many distinguished properties such as high solubility in oil and water, strong adhesivity, good biocompatibility and film‐forming ability, levan has great potential as a novel functional biopolymer in foods, feeds, cosmetics, pharmaceutical and chemical industries (Kang et al. 2009; Kazak et al. 2010). In fact, a recent literature analysis on microbial EPSs named levan together with xanthan, curdlan, and pullulan as the most promising polysaccharides for various industrial sectors (Donot et al. 2012). Levan has many potential uses as an emulsifier, stabilizer and thickener, encapsulating agent, osmoregulator and cryoprotector in addition to its uses in medicine as plasma substitute, prolongator of drug activity, radioprotector, antitumor and antihyperlipidemic agent (Freitas et al. 2011; Kang et al. 2009; Sezer et al. 2011).
Levan is used as an industrial gum, a sweetener, a formulation aid, emulsifier, a carrier for flavor and fragrances, a surface‐finishing agent, stabilizer and thickener, encapsulating agent, osmoregulator, and cryoprotectant (Esawy et al. 2013; Shih et al. 2010). It is also used as a food or feed additive with prebiotic and hypocholesterolemic effects (Esawy et al. 2013; Shih et al. 2010). In addition, it is used in edible food coatings (Shit and Shah 2014). A combination of fermented red ginseng and levan significantly inhibited body weight gain in HFD‐induced obese mice and prevented insulin and leptin resistance (Oh et al. 2014).
Research has been reported on the potential use of levan by Halomonas smyrnensis as a bioflocculating agent (Sam et al. 2011), its suitability for peptide and protein‐based drug nanocarrier systems (Sezer et al. 2011). With this microbial system, productivity levels were improved by use of cheap sucrose substitutes like molasses (Kucukasik et al. 2011) as well as other cheap biomass resources (Toksoy Oner 2013) as fermentation substrate. Moreover, levan and aldehyde‐activated levan were successfully deposited by matrix‐assisted pulsed laser evaporation (MAPLE) resulting in uniform, homogeneous, nanostructured, biocompatible, thin films (Sima et al. 2011, 2012) and furthermore, the feasibility of phosphanate‐modified levan as adhesive mulitilayer film was demonstrated (Costa et al. 2013).
The antioxidant potential of this levan polysaccharide in high glucose conditions in pancreatic INS‐1E cells by demonstrating a correlation between reduction in oxidative stress and apoptosis with its treatment has been reported (Kazak et al. 2014). In vitro anticancer activity of linear levan and its oxidized forms containing increasing amounts of aldehydes was also investigated and anticancer activity was found to depend on the dose as well as on the cell type and this effect became more apparent with increasing degree of oxidation (Kazak Sarilmiser and Toksoy Oner 2014). Ternary blend films of chitosan, PEO, and levan were also prepared to evaluate their morphological, thermomechanical, surface, and biological properties (Sennaroglu Bostan et al. 2014). Moreover, the generic metabolic model of H. smyrnensis AAD6T was reconstructed to elucidate the relationship between levan biosynthesis and other metabolic processes (Ates et al. 2013). Recently, its whole genome sequence was announced (Sogutcu et al. 2012).
Fructan type polysaccharides that ferment in intestinal flora can enhance intestinal flora and mineral absorption. There are several reports of the prebiotic effect of levan‐type fructans. Oligofructans obtained from hydrolization of levan from Zymomonas mobilis in a microwave oven beneficially affect the host by selective stimulation of probiotic bacteria in the colon (de Paula et al. 2008). The levan from L. sanfranciscensis LTH 2590 also exhibits prebiotic effects as demonstrated in vitro by different experimental approaches (Bello et al. 2001). The possibility of acting as prebiotic substrate has been demonstrated successfully for a fructan‐type EPS produced by one strain of L. sanfranciscensis by Korakli et al. (2003). The bifidogenic effect was also reported for the levan‐type EPS produced by another strain of the same species (Patel and Prajapati 2013). Recently, prebiotic activities of three different kinds of fructan, levan from H. smyrnensis AAD6T, inulin and Agave tequilana fructans, were compared and investigated for the effective formulation of symbiotics with antibiotic activity (Arrizon et al. 2014).
Levan can act as a prebiotic, cholesterol and triacylglycerol‐lowering agent in the food industry (Nakapong et al. 2013). Commercial food‐grade levan is produced by a number of companies, including RealBiotech Co. (www.realbio.com), Chungnam, Korea, and Advance Co. Ltd (www.advance.jp), Japan. Levan production and levan derivatives have also been developed by Montana Biotech and the Sugar Processing Research Institute (Kang et al. 2009).Various forms of levan like chewing gum and powder can be found as “functional food” at markets in Korea. Levan was initially industrialized as a food additive to support functional soluble fiber (Kang et al. 2009). Low molecular weight levan was found to show in vitro inhibitory effects against pathogenic bacteria and to have potential as a sweetener in food (Byun et al. 2014).
During 1‐week storage of fresh wheat breads, an increased volume, clear softening effect and retarded staling were observed as a result of fructans of several acetic acid bacterial strains (Hermann et al. 2015). Dough rheology and bread texture were enhanced by levan from the sourdough isolate Lactobacillus sanfranciscensis (Brandt et al. 2003) and improved antistaling properties of bread were observed with levan addition (Kaditzky et al. 2008).
Levan is accepted as a non‐toxic substance and has been verified as safe by many scientific studies (Kang et al. 2009). LD50 and NOEL (no observed effective level) values for levan are known to be 7.5 ± 0.5 g/kg bw and 1.5 g/kg/day, respectively (Kang et al. 2009). The Department of Food Chemistry, Ministry of Health, performed general toxicology tests and mutagenicity tests on levan from B. subtilis and verified its safety. Levan from Z. mobilis was approved as a food by the Korea Food and Drug Administration (KFDA) in 2001, and was listed in the Korea Health and Functional Food Code. A levan with the trade name FRUCTAN is approved as a food and food additive and marketed in many countries such as the USA, EU, Japan, Australia, and New Zealand (Kang et al. 2009).
Many applications for levan, including as a food additive, have been patented. Levan from S. salivarius was reported as a food additive with a hypocholesterolemic effect (Kazuoki et al. 1996). A GRAS‐grade levan producer, Lactobacillus reuteri, can be used as a probiotic (van Hijum et al. 2004). The levan synthesized by L. sanfranciscensis can be used in human or pet food products (Vincent et al. 2005).
Gellan is a high molecular weight, linear, anionic heteropolysaccharide that consists of tetrasaccharide chemical repeat units in which β‐(1 → 4)‐linked glucose, glucuronic acid, glucose, and rhamnose in α‐(1 → 3) linkage are bonded together (Ahmad et al. 2015; Wüstenberg 2015). Gellan, which is marketed as a broad‐spectrum gelling agent, is highly viscous and has shear‐thinning behavior in dilute aqueous solutions (Morris 2006). It is soluble in water, forming a viscous solution, and is insoluble in ethanol. The functionality of gellan and type of gellan gel are varied based on the degree of acylation and the ions present (Khan et al. 2007; Pegg 2012). An interesting feature of low acyl gels is their excellent flavor release properties (Morris 2006). Research on gellan gum is intensive since it is an appropriate model for the study of thermo‐reversible sol–gel transition (Prajapati et al. 2013).
Gellan is extracellularly secreted by Sphingmonas elodea (originally called Pseudomonas elodea) by aerobic submerged fermentation in batch culture (Ryan et al. 2015). Kelco isolated the bacterium from water plant (elodea) tissue as part of a screening program intended to identify interesting new bacterial polysaccharides (Khan et al. 2007). Gellan was patented and produced commercially by Kelco. Gellan was first used in food in Japan in 1988 and the FDA has approved it for use in the United States and in Europe it is classified as E 148. It is sold commercially under the trade name Gelrit (Ahmad et al. 2015; Kang et al. 2014; Morris 2006). Specifications for gellan such as molecular mass, solubility, and functional uses were prepared at the 46th Joint Expert Committee on Food Additives (JECFA) in 1996 (Prajapati et al. 2013). It was also approved as a safe food ingredient and additive in various countries such as Australia, Canada, United States, Mexico, Chile, Japan, South Korea, and Philippines (Milani and Maleki 2012). Gellan gum products are available with the names Gelrite, Kelcogel, and Gel‐Gro (Prajapati et al. 2013).
Toxicological analysis has been performed and the acute oral toxicity (LD50) value for rats was found to be >5 g/kg. Eye and skin irritation studies suggest that gellan is not an irritant. No significant effect on plasma biochemistry, hematological indices, urinalysis parameters, blood glucose, plasma insulin concentrations, or breath hydrogen concentration was observed for 23‐day human dietary studies (Anderson et al. 1988).
Transparent and stable hydrogels of gellan gum have a wide range of applications in tissue engineering and drug delivery. Gellan, which is a non‐toxic, biodegradable and biocompatible polymer, forms hydrogels with mechanical properties similar to normal human under definite conditions. Therefore it can be used as a versatile biomaterial (Kang et al. 2014).
Gellan can be employed as a thickener, binder, film former, and stabilizer in a wide range of food applications such as confectionery products, jams and jellies, fabricated foods, water‐based gels, pie fillings and puddings, icings and frostings, and dairy products such as ice cream, yoghurt, milkshakes, and gelled milks. It can be used as a hydrocolloid coating for cheese. Gellan can replace agar in Asian foods: Japanese foods such as hard bean jelly, mitsumame jelly cubes, soft red bean jelly, and tokoroten noodles, in the preparation of other Japanese products including karasumi‐like food and laver, and in the manufacture of tofu curd (Ahmad et al. 2015; Morris 2006; Prajapati et al. 2013; Wüstenberg 2015). Interactions with other polysaccharides and between the two gellan types are an advantage for using gellan gum for food product development since this allows the production of a wide range of textures and provides structure with superior properties (Danalache et al. 2015).
Gellan gum can be used in the pet food industry, often in combination with other hydrocolloids, to produce both meat chunks and gelled pet foods. The moisture barrier properties of edible gellan films have been investigated (Morris 2006).
Gellan gum is used as a stabilizer in water‐based gels such as desserts, drinking jellies, and mango bars (Danalache et al. 2015; Wüstenberg 2015). In bakery products, it improves bake‐stability of fillings with low to high soluble solids content. It is also used as a replacement for gelatin in cultured dairy products like yoghurt and sour cream in vegan, Kosher or Halal nutrition (Wüstenberg 2015). Fruit purees/juices along with gelling agents such as gellan can be used for development of novel food products with a variety of textures (Danalache et al. 2015). Gellan reduces clouding and pulp settling in beverages. It is employed in systems with low milk amounts, low‐quality milk protein, or heat‐damaged proteins that are found in spray‐dried milk powders. It also provides a short texture to gelled confectionery, improves gelatin gummy candies, and reduces stickiness. Low‐calorie (sugar‐free) jams, fruit preparations for yoghurt, sauces, no‐fat salad dressings with herbs, and films and adhesion systems (Wüstenberg 2015) and oil reduction in fried foods (Khan et al. 2007) are other applications.
Curdlan is a high molecular weight polysaccharide of glucose residues with the general formula (C6H10O5)n. It is a a linear β‐(1 → 3)‐glucan that is extracellularly produced under nitrogen‐limited conditions by the non‐pathogenic bacteria Agrobacterium biobar and mutants of Alcaligenes faecalis var. myxogenes, a group of micro‐organisms occurring in the soil (Wu et al. 2015). Curdlan is insoluble in water, alcohols and the majority of organic solvents and is soluble in aqueous alkaline solutions and it shows gel strength between agar and gelatin (Khan et al. 2007; Morris 2006).
The term “curdlan” has been derived from “curdle” – describing its gelling behavior at high temperatures with different characteristics. A high‐set, thermo‐irreversible, strong elastic gel is formed upon heating its aqueous suspensions above 80 °C followed by cooling. Alternatively, a low‐set, thermo‐reversible gel can be obtained by heating it to 55 °C and subsequently cooling to ambient temperatures. The latter type of gel can be formed without heating, by the neutralization of its solutions with an alkali (Khan et al. 2007; Wu et al. 2015; Zhang and Edgar 2014).
Curdlan is widely used as a biothickening and gelling agent in foods such as tofu, noodles, and jellies since it has remarkable rheological properties among natural and synthetic polymers (Khan et al. 2007; Morris 2006). There are numerous applications of curdlan as a gelling agent in the food, construction, and pharmaceutical industries. Gelatin and agar are replaced with curdlan in the production of jellies, desserts, and confectionery. It can also be used as a thickener and binder in foods such as salad dressings, desserts, and pasta. Edible coatings of curdlan can be used to prolong shelf‐life (Hu et al. 2015; Khan et al. 2007). Curdlan as a dietary fiber also provides a nutritional benefit and has the potential to be produced at a higher quality for related foods (Wu et al. 2015). Curdlan and microbial transglutaminase would improve the gel strength, water‐holding capacity, and whiteness of heated fish meat gel (Hu et al. 2015).
Curdlan was first discovered in 1964 and research was started for production of curdlan for both food and general industrial applications by Takeda Chemical Industries Ltd in 1968. Although development work ceased in 1973, industrial‐scale production was restarted in 1988 (Morris 2006). Its usage for food was approved in Korea, Taiwan, and Japan in 1989 (Hu et al. 2015).
After xanthan and gellan, curdlan was approved as the third micro‐organism‐fermented hydrocolloid food additive by the US FDA in December 1996 (Wüstenberg 2015). Curdlan has been used in a wide range of food applications in Japan where bacterial polysaccharides are considered as natural products (Morris 2006). Toxic effects of curdlan have been extensively studied and it has been estimated as a safe product (Hu et al. 2015; Khan et al. 2007).
Curdlan is tasteless, odorless, colorless and has a high water‐holding capacity that makes it an ideal textural agent in various food applications such as meat, dairy, sauces, baking, and nutraceutical products (Hu et al. 2015; Khan et al. 2007). Curdlan can be directly added to regular food systems at low levels (less than 1%) or used as a building block of the food structure at high levels to develop new food products. In Japan, it is employed mostly in the meat‐processing industry for improvement of texture by enhancement of the water‐binding capacity. Elasticity and strength of noodles are also enhanced by curdlan (Khan et al. 2007).
Rheological and thermal behaviors of curdlan have led to its application for the integrity of oil‐moisture barriers in batter and coating systems, imitation seafood delicacies, vegetarian meat analogues, and tofu products having freeze/thaw and retort stability (Khan et al. 2007; Zhang and Edgar 2014). Curdlan gels have been used to develop calorie‐reduced items, since there are no digestive enzymes for curdlan in the upper alimentary tract, and curdlan can be used for fat replacement (McIntosh et al. 2005; Zhang and Edgar 2014).
Non‐food applications of curdlan have also been studied and antitumor, antioxidative, and immunomodulating activities have been discovered due to curdlan’s unique helical structure, gel‐forming capacity, and potent pharmacological properties. Several studies have been employed to improve the solubility of curdlan to increase its applications, develop nanostructures for encapsulation and release of biological active reagents, provide curdlan derivatives with antitumor, anti‐HIV, and anticoagulation activity, improve wound healing activities and inhibit microbes, fungi, and viruses (Li et al. 2014; McIntosh et al. 2005; Zhang and Edgar 2014).
Exopolysaccharides produced by a diverse group of microbial systems are rapidly emerging as new and industrially important biomaterials. Microbially produced EPSs show great diversity and functions with unique and commercially relevant material properties. EPSs are mainly associated with high‐value applications in various fields such as food, feed, packaging, chemicals, textiles, cosmetics and pharmaceutical industry, agriculture and medicine, and they have received considerable research attention over recent decades due to their biocompatibility, biodegradability, and both environmental and human compatibility.
Exopolysaccharides are widely used to enhance the quality, texture, mouth feel and flavor of food as thickeners, emulsifiers, stabilizers, texturizers, and gelling agents. Xanthan, levan, gellan, and curdlan, which are currently used as additives in the food industry, are well known industrial microbial polysaccharides with numerous applications and a considerable market. There is a growing body of research associated with these high‐value microbial EPSs for development of low‐cost production processes, improvement of properties, and optimization of performance as food additives. These natural polysaccharides will become increasingly essential since they are natural non‐toxic alternatives to synthetic food additives with their potentially harmful characteristics.