An Overview of Ingredients Used for Plant-Based Meat Analogue Production and Their Influence on Structural and Textural Properties of the Final Product

Climate change and the sustainability of the production process are some of the most pressing problems in food production. The impact of food production and type of diet on climate change is measured by the carbon footprint, and, according to the research available so far, the change in the eating habits of consumers from the predominant Western type of diet to a Mediterranean type, vegetarian or vegan diet also contributes to the carbon footprint reduction by minimizing greenhouse gas emissions and reduction in water consumption required for production [1,2]. The earlier mentioned is one of the reasons why consumers are continuously turning to healthier alternatives to the Western diet. Accordingly, the demand for vegetarian and vegan food products is growing, as evidenced by the fact that the size of the vegan food market was estimated at USD 19.7 billion dollars in 2020, and is expected to reach a market worth of USD 36.3 billion by 2030 [3]. In accordance with the increased demands for vegan food, the technology for the production of such food is also being developed, which includes novel drying processes, protein extraction, protein purification, microencapsulation, 3D printing and more [4,5,6]. One of the new technologies whose application is increasingly common in the production of vegan food is additive technology, or 3D printing technology. Additive technologies are characterized by advantages that include the possibility of creating complex shapes, the possibility of regulating texture and color, the protection of thermolabile ingredients and the possibility of creating personalized products [7]. Additive technology involves construction of solid geometries in multiple layers, which are then connected by chemical reactions or phase transition processes [8]. Some of the most commonly used 3D printing technologies are extrusion printing, ink jet printing, bio printing and printing by deposition and bonding powder (binder jetting) [7,9]. Briefly, the extrusion process uses semi-solid pastes, which are then squeezed out of syringes with different nozzle diameters and geometries to form a specific shape, previously defined using a 3D modeling software (CAD software). An ink jet requires liquid materials with a significantly lower viscosity in comparison to pastes used for extrusion, which are then sprayed or dripped to a desired form [9]. Binder jetting is a specific process where powders are used as raw materials to form final products, while bioprinting is usually connected to printing forms comprised of living tissues. According to literature data, in the food industry, 3D printing is applied to cereal products, confectionery, pasta, cheese, fruit products, surimi, etc. [8,10,11,12,13].

In the production of plant-based meat analogues, besides 3D printing, other technologies (e.g., shear cell, high- or low-moisture extrusion and others) can also be applied to produce a product that resembles meat, but is completely plant-based. The most common type of technology used is extrusion printing, where it is of importance to adjust and optimize the composition of the printing mixture so that, first of all, printing is possible, and then so that the said product has satisfactory nutritional, sensorial and textural properties, which are of primary importance to the end consumer. Plant-based meat analogues are often characterized as mixed gel systems produced with the texturization of various plant-based ingredients (e.g., cereals, legumes, algae, fungi) [14]. The most common ingredients in plant-based meat alternatives include soy, legumes (e.g., chickpeas, lentils, peas), grain proteins (gluten from rice, oats, barley or wheat), colorings (cumin, carotene, lycopene, beetroot juice) and flavorings (spices, herbs, yeast extract, paprika, sugar, mushrooms). In addition to the above, to ensure the texture and properties similar to meat, oils (sunflower, coconut, palm, etc.) are used, as well as various “structural” ingredients responsible for the connection and stability of the shape (carrageenan, starch, cellulose, etc.). Furthermore, the mentioned mixtures can be enriched with various vitamins and other bioactive ingredients [15].

This review provides an insight into the most common ingredients used for production of plant-based products, with a special focus on their structure and the ability of the ingredients to affect and regulate the texture, structure and nutritional properties of the final product. It is divided in sections, which focus on specific groups of ingredients: proteins and fats, ingredients responsible for maintaining acceptable texture of the products; spices, colorings and functional ingredients, which can act as stabilizers; and enrichment agents.

Proteins are a group of macronutrients, which are considered to play a major role in building muscles and tissues and are crucial in maintaining proper functions of the human body. Humans mostly ingest proteins of an animal origin, but the ever-growing shift towards healthier alternatives has put the plant-based protein sources in perspective. Figure 1 depicts some of the plant-based protein sources.

As seen in Figure 1, based on their origin, plant-based protein sources can be divided into several groups: (1) legumes (soy, pea, chickpea, lentils, beans, peanuts); (2) cereals and pseudocereals (wheat, corn, oats, quinoa, amaranth, buckwheat); (3) oil seeds (hemp, sunflower, rapeseed, sesame); and (4) algae. While some of the mentioned groups (e.g., legumes and cereals) contain proteins that are able to affect the structure of meat analogues and can therefore be used to produce meat analogues of a desired texture and mouthfeel [16], others are primarily used as good sources of amino acids and bioactive peptides [17,18]. Furthermore, besides proteins, seeds can also act as sources of fats necessary to ensure texture, flavor, mouthfeel and juiciness, but also as sources of fat-soluble vitamins and fatty acids [15].

Besides water, proteins, flavoring, coloring and binding agents, meat alternatives contain between zero and fifteen percent of fats [120]. Combination of the above-mentioned ingredients is responsible for sensorial and textural characteristics of meat alternatives.

Lipid ingredients, such as vegetable oils and fats, improve juiciness, tenderness, mouthfeel and flavor of meat analogues [9,77]. Different types of lipid ingredients (fats/oils) are used during preparation of plant-based meat analogues, with the most common oils being canola, coconut, corn, sunflower and sesame oil, and cocoa butter [78]. Although meat analogues traditionally have low lipid content, their second generation consists of saturated-fat-rich semi-solid constituents (e.g., coconut oil and cocoa butter) [25,70,121,122]. Currently available emulsion-type meat analogues on the market have up to 25% fat content. However, these products are subjected to criticism due to a high amount of saturated fat [78]. Lipids are bioactive components (functional ingredients), and from a health point of view, their inclusion in plant-based meat analogues is of great importance [123]. According to the World Health Organization, dietary fat intake should account for 15% to 30% of total diet energy with cholesterol intake limited to 300 mg/day [124]. Furthermore, nutritionists and dieticians have also recommended limitation of saturated fat and trans-fat intake [125,126,127]. The fatty acid composition of fats or oils derived from plants differs significantly: different vegetable oils contain different quantities of specific fatty acids such as palmitic acid, stearic acid, oleic acid, caprylic acid, capric acid, lauryc acid, myristic acid and linoleic acid [128]. Besides that, plant-based oils/fats include variations in saturated, monounsaturated, polyunsaturated and trans fatty acids, as well as their solid or liquid state [129].

Since abundant consummation of fats is related to a negative impact on human health, researchers are focused on the reduction in actual fat content in meat products [130]. One of the solutions is to introduce fat substitutes, consisting of water and functional ingredients [131,132,133,134]. Fat substitutes could also be used for the preparation of low-fat meat analogues, where textural properties need substantial improvement [25]. Protein particles (from milk, eggs, soy protein isolate), modified (synthetic) lipids and carbohydrates (starch, fibers, cellulose) are already used in the food industry as fat substitutes [129,135,136]. Another example is combination of konjac gels with ingredients such as starch, carrageenan and gellan gum, which can also be utilized for the formulation of low-fat plant-based meat analogues [137]. It is worth noting that fat substitutes have no effect on the sensory properties of meat analogues [25]. However, significant attention should be paid regarding the effect of the oils/fats during preparation of meat analogues with the extrusion process. Oil or fat addition increases juiciness and affects textural properties, such as fiber formation, of the final product [138]. Lipids decrease viscosity of the mixtures used for extrusion and can lubricate the extruder, resulting in a decrease in shearing and friction, and lowering the mechanical energy required to run the extrusion process. This will consequently change the reaction behavior of the proteins, leading to a less fibrous structure. Parameters such as fat/protein interactions, structure formation mechanisms, oil stabilization and component interactions differ for every protein–oil combination and it is difficult to predict the degree of interference of vegetable oils/fats on meat analogue production. Lipids influence the mechanical properties of proteins, as well as the organoleptic perception of the final product [139]. Interactions of fat particles with proteins affect the water-binding capacity, lubricating properties and protein gel strength [140]. Characteristic of the application of oils in plant-based meat products is that the oil needs to be added in the form of droplets. Incorporation depends on the product type and oil type used. If it is very difficult to stabilize incorporating fats, then pre-emulsified oils are used. The advantages of oil pre-emulsion technology include improvement of fat binding ability into a protein matrix and improved oxidative stability of lipids. Additionally, pre-emulsified oils are easier to disperse into water-based systems [141]. Microencapsulation of oils (fish oil emulsions) will facilitate their incorporation in meat analogues, inhibit lipid oxidation, mask undesirable flavors and improve bioavailability of n-3 PUFAs. The amount of oil in this form varies between 1% and 30% while low-molecular-weight compounds are often used as an emulsifier [123]. Structured oils (oleogels) are solid materials, made from fats, which have previously been structurally modified by oleogelators into a three-dimensional thermoreversible gel network. Polymers are often used as structurants, due to the fact that many of them are food-grade and inexpensive. Physical characteristics of edible oleogels enable their use as fat replacers: palm oils, oleins and stearins in combination with additives are mainly used for preparation of meat analogues because they provide desirable textural characteristics. Monoglycerides, lecithin, phytosterols and vegetable waxes are among the most researched structural agents for edible oils. Their combination with other ingredients plays a vital role in the final preparation of plant-based meat analogues [142,143]. An overview of the fats used in plant-based meat production, along with their main textural and structural properties, is summarized in Table 6.

In addition to all of the above, a comprehensive understanding and analysis of lipid sources and their functional roles in plant-based meat analogues, as well as their molecular interactions with other components, are of great importance for the development of the plant-based meat analogue industry [150].

For commonly used proteins, which were previously discussed, it is necessary to achieve the desired functionality and acceptability by users [151]; so, addition of stabilizers and structural ingredients is necessary [152]. An exceptional challenge in the preparation of plant alternatives to meat is the extremely large number of ingredients, which is usually over 20 [153], as it can be seen in listed ingredients for vegan/vegetarian bratwurst (Table 7).

Additives that are added in plant-based meat analogues should assure plant-protein functionality through foaming capacity and stability, emulsification, rheology, oil and water holding capacity, gelling and solubility [154]. To ensure functionality and mimic meat, preservatives, stabilizers and dyes, some of which will not be found in meat products, such as lecithin, methylcellulose and titanium dioxide [78,155], are often added to plant-based meat analogues. All added ingredients should assure attributes in accordance with consumer expectations [156].

Meat analogues almost always contain carbohydrates as structural ingredients (as can be seen from Table 4), and they can be in the following forms: starch/flour and gums or binding ingredients [155]. Mattice and Marangoni [153] state that the function of the carbohydrate additives (in the form of starch or flour) is to improve the texture and consistency of the product while the gums/binders will stabilize and shape the product (e.g., methylcellulose, gum acacia, xanthan gum, carrageenan, etc.) [15,156]. There is also an example of using konjac glucomannan in the development of the fermented soybean patty [148], where the produced patty analogue had higher firmness and cohesiveness, and lower stickiness.

The addition of these ingredients enhances the interaction of other components (protein, fat, water) during the preparation process [31,157] and thus the higher carbohydrate content in plant-based meat analogues. At high temperatures, methylcellulose forms a gel network and can create thermo-reversible gel. Addition of a biopolymer to proteins enhances the stability of the mixture needed to form a 3D structure [36]. As presented by Shabazi et al. [36], some of the most frequently used biopolymeric surfactants include the following: (i) ethyl (hydroxyethyl) cellulose stabilizes oil-in-water emulsion through an associate thickening mechanism; (ii) octenyl succinic anhydride attaches to the hydrophilic backbones; (iii) acetylated starch forms a densely packed interface layer on the surface of the oil droplet; and (iv) hydrophobically modified inulin. Recently, there have been significant discussions about the effect of added gums and binders in meat analogues on consumer health [15,151,158]. The focus of recent research is methylcellulose (modified cellulose dietary fiber), which is a very effective binder [159] used also in food packaging and creates a viscous solution (during the digestion), consisting of pseudo-crystalline sequences of trimethyl glucose units [160]. There is a growing trend for using different replacements for methylcellulose as the most commonly used binding ingredient in the plant-based meat analogues. For example, binding performance of methylcellulose hydrogel and a new binder made of pea protein and sugar beet pectin was evaluated on burger meat analogues [161] and bacon-type meat analogues [162], with a conclusion that the homogeneity of the produced analogues was positively affected by the binder addition.

The gums used in the production of meat analogues have not shown any impact on health; however, when purchasing, end users opt for products whose labels indicate environmentally friendly production (such as Eco-Score, Table 4) as well as less processing (NOVA score, Table 4). Precisely because of the above mentioned, efforts are being made to minimize the use of gums as well [156,157,163], which are currently an indispensable ingredient in vegetable meat analogues. Based on literature data, a 2.5% gum arabica/soy protein concentrate can be efficiently used to improve sensory properties of mushroom-based sausage analogues [164]. Study of the effect of different concentrations of a carrageenan, casein, xanthan gum and soy protein concentrate on quality characteristics of mushroom-based sausage analogues showed significant improvement in textural properties of samples prepared with carrageenan [165]. As mentioned before, in plant-based meat analogues, soy and pea proteins are typically the most commonly utilized components. However, those proteins provide poor characteristic juiciness and fibrousness of actual meat. Wheat gluten is also frequently added to the formulations in order to overcome this restriction [166]. Under hydrated conditions, gluten makes plant-based meat mixtures viscous and significantly strengthens plant-based meat mixture adhesion [167]. Besides gluten, there is also research focused on rheological and textural properties of soy-protein-isolate-based meat analogues prepared with guar and xanthan gum, carrageenan, hydroxypropyl and cross-linked tapioca starch as binding agents [168], as well as on organoleptic quality of cowpea protein meat analogues with cocoa pod husk extracts as binding agents [169]. Generally, it can be concluded that, based on the added amount, some ingredients can behave as binders (ingredients with high protein levels), while some can behave as fillers or extenders (ingredients with low or no protein levels) [25].

Flavor is one of the essential sensory components of food products [170]. It is important to comprehend the processes involved in the synthesis of flavor molecules in animal flesh to add a meaty flavor to plant-based equivalents [171]. For instance, the flavors of raw and cooked meats change significantly. Raw meat has no aroma and just tastes of blood, metal and salt, while fragrance and flavor compounds in cooked and roasted meat become more complex as processing temperatures rise [172,173]. According to Li and Li [171], flavor compounds in cooked meat are produced with (i) the Maillard reaction between amino acids and reducing sugars; (ii) oxidation of fatty acids; and (iii) thermal degradation of thiamine. The primary ingredients of plant-based meat substitutes are soy protein and wheat gluten, neither of which has the essential intermediate compounds required to give meat-like flavor [155,174]. As a result, to replicate the tastes and scents of meat, several flavorings that resemble meat are added to meat substitutes [175]. To mimic the flavors of processed meat, a wide variety of spices and herbs are added, including those that are also used in the meat processing industry [120]. As previously described by Wang et al. [42], compounds used to improve taste of the plant-based meat analogues can be grouped as follows: (i) natural spices and herbs used to prevent lipid oxidation, (ii) Maillard reaction precursors including reducing sugars, amino acids and thiamine to generate meat-like flavor, (iii) hydrolyzed vegetable proteins used to improve meat-like flavor, (iv) yeast extract contributing to the roasted, meat-like aroma and (v) vegetable oils contributing to mouthfeel of the plant-based meat product. When aromatizing meat analogues, the production process also has to be considered, because analogues that are flavored during extrusion undergo a series of physicochemical changes in the premix [176], and in the case of plant analogues that are produced at a high temperature and/or pressure, the volatile components can significantly change [177]. All of the above greatly change the taste perception, which must remain stable during shelf life [26].

Dried onions, dried garlic, curry powder, black pepper, garlic, chili, paprika and ginger are among the most widely used spice mixtures [120]. According to Vlaic et al. [178], the most widely used aromatic plants in plant-based meat production are parsley, dill, basil, oregano, sage, coriander, rosemary, marjoram, tarragon, bay, thyme and mint. The mentioned herbs contain bioactive molecules that can have a beneficial effect on human health. According to Zioga et al. [179], the addition of herbs and spices masks beany off-flavors, and organic acids, which can be present in those herbs and spices, can also contribute to shelf life. Even yet, a lot of plant-based substitutes still have an aftertaste [180]. As described by Amyoony et al. [181], plant-based meat analogues are characterized by beany and chalky flavors, bitter tastes and “off-flavors” overlapping with unwanted tastes, odors and sensations including astringency that may be brought on by intrinsic components or as a result of storage and processing. The same authors also stated that aftertaste intensity of the plant-based meat analogues was negatively correlated to the overall acceptance of the products based on the sensory evaluation. In addition, vegetable oils, which can influence taste, are combined with the protein components to simulate mouthfeel and juiciness. This applies in particular to coconut oil, which can be mixed with liquid oils such as sunflower and rapeseed oil [182].

The distinct beany smell, which is believed to be associated with the byproducts or derivatives of secondary lipid oxidation, including hexanal and methanethiol, along with the naturally occurring bitter-astringent tastes resulting from saponins and isoflavones, may pose a challenge for the use of soy protein as a building block for meat substitutes [56,183].

Yuan et al. [184] analyzed the influence of four selected spices (black pepper, red pepper, onion and garlic) on the flavor of 50 commercial meat analogues. According to their findings, incorporating those spices into the extrusion process decreased the production of certain volatile chemicals linked to heat-related treatments. So, in addition to improving flavor profiles of extrudates, adding spices can delay the production of some volatile chemicals with bad flavors, such as alcohols, ketones and aldehydes. Similarly, Yuan et al. [185] used a five-spice powder mixture in the production of a mushroom-based meat sausage analogue. They analyzed the effect of the water content, type of mushroom and amount of added powder on physicochemical and structural properties and the aroma profile of prepared meat analogues. They successfully produced a plant-based meat analogue whose aroma was close to that of beef and concluded that, besides the spice mixture used in the formulation, the aroma profile is governed by volatile flavor substances, which are mostly a result of the complexities of lipid oxidation and bacterial metabolism.

Similar to other constituents of plant-based meat products, spices also have the ability to affect structural properties of the mixtures, and, once again, the complexity of their interactions with other components (proteins, fats, stabilizers) is still an area where much research is needed.

Color is one of the most important factors. Also, hydrated alginate and maltodextrin are coloring agents that reduce color migration during production, which governs consumers’ product acceptance [186]. Generally, raw fresh meat has a red color, which turns to brown during cooking and it is a challenge to obtain plant-based meat to mimic and resemble traditional meat. Gluten and soy, as some of the most widely used plant-based proteins, are originally yellow or beige in nature, which raises the need to improve their color to be more similar to that of meat, by using coloring ingredients or precursor substances [187].

Consequently, natural pigments have been developed [188,189] but they cannot be used directly from renewable sources because the incorporation of raw materials as coloring agents has many limitations (e.g., natural pigments are unstable at a high pressure and high temperature, chemically degrade when subjected to oxygen and lose their acceptance and functionality during storage) [188]. Nevertheless, natural colorants have remarkable antioxidant properties, suggesting that they could be used as nitrite replacers in meat products, as well as flavor and textural property enhancers [182,188]. The simulation of the color of cooked/roasted meat is achieved by using caramel colors (annatto or malt; carotene, cumin, turmin) [190] and beet root extracts [191], but also by adding reducing sugars (e.g., dextrose, maltose, lactose, xylose, galactose, mannose and arabinose). Reducing sugars can react with amine groups in proteins (Maillard reaction) [192,193]. In addition, color additives can also be used to impart a reddish-brown color. One of the color additives described is soy leghemoglobin, which was approved by the Food and Drug Administration in 2019 [194].

In the production of plant-based meat products, the thermal stability and pH sensitivity are of great importance. The degradation of thermally unstable agents can lead to an unacceptable color appearance [187]. To ensure stability, juices rich in polyphenols and ascorbic acid (apple extracts or citrus fruit extracts) are often added to plant-based meat products [15,195]. Furthermore, juices can also act as antimicrobial and preservative agents [195]. Other examples of coloring agents include carotene, cumin and turmin, which are considered heat-stable and preferred by consumers [196]. The red meat color can also be mimicked by leghemoglobin, lycopene, annatto and beet juice extracts, while chicken meat color can be mimicked by titanium dioxide [15]. As mentioned earlier, coloring agents can be combined with proteins before the structuring treatment or mixed with the semi-structured plant-based materials during structuring [187]. Also, hydrated alginate and maltodextrin are coloring agents that reduce color migration during the production [187].

Ryu et al. [186] investigated the use of different natural pigments in plant-based meat to imitate the color of meat after cooking and the results confirmed that natural pigments can be used as alternative colorants in plant-based meat. Also, Bakhsh et al. [197] applied natural colorants such as lactoferrin and red yeast rice to plant-based patties and their results showed that the use of natural colorants had a negligible effect on the chemical composition and textural attributes, and the colors of the products were lighter and had a brownish tinge in comparison to the control. A similar investigation was carried out by Akramzadeh et al. [198], where the authors analyzed three different natural color agents (red yeast rice, lycopene and paprika oleoresin) used as ingredients in non-meat sausages. Respectively, the products provided superior sensory acceptance.

Plant-based meat analogue production is on the rise. It is mainly driven by an ever-rising awareness of climate change and the need for the food industry to reduce CO₂ emissions. Plant-based food holds the promise toward a cleaner and healthier future, from one side due to the reduction in the environmental impact, and, on the other side, because plant-based foods are known to possess many beneficial effects. However, the biggest problem that still remains to be solved is the difference in texture, mouthfeel and taste of the plant-based meat analogues in comparison to real meat. This review paper has shown that there is an abundance of literature available on the topic of texture and structure of plant-based meat analogues, but somehow, a majority of literature data come to the same conclusion: the interactions of particular components of plant-based meat analogues are extremely complex and highly dependent on the processing conditions during production and, subsequently, the processing (cooking, baking, etc.) conditions at home, after the product has been purchased by the consumer. Therefore, the need for optimization of the composition of the mixtures, as well as the processing conditions, has to be highly stressed. In general, the development of plant-based meat analogues is a complex work that requires the expertise of many different specialists—from nutritionists who can define optimal nutritive content and food technologists who can optimize the structural properties of the analogue to engineers who can adapt the manufacturing process to specific needs.