Acta agriculturae Slovenica, 120/4, 1–11, Ljubljana 2024 doi:10.14720/aas.2024.120.4.18282 Original research article / izvirni znanstveni članek Evaluation of amino acid composition in different types of meat and plant-based burger patties Kaltrina BERISHA 1, 2, Zsuzsanna MEDNYÁNSZKY 2, Altin BERISHA 2, Tamara TARI 2, Hysen BYTYQI 3, 4 Received February 26, 2024; accepted October 04, 2024. Delo je prispelo 26. februarja 2024, sprejeto 04. oktobra 2024 1 University of Prishtina “Hasan Prishtina, Faculty of Agriculture and Veterinary, Department of Food Technology with Biotechnology, Prishtina, Republic of Kosova 2 Hungarian University of Agricultural and Life Sciences, Institute of Food Science and Technology, Department of Nutrition, Budapest, Hungary 3 University of Prishtina “Hasan Prishtina”, Faculty of Agriculture and Veterinary, Department of Animal Sciences, Prishtina, Republic of Kosova 4 Corresponding author, e-mail: hysen.bytyqi@uni-pr.edu Evaluation of amino acid composition in different types of meat and plant-based burger patties Abstract: The study examined the protein content and amino acid composition of various commercially available plant-based and meat-based burger patties. The aim of this study was to determine whether plant-based burger patties meet the requirement for essential amino acid content in the human diet. Amino acid profiles were determined using the Amino Acid Analyzer and FAO/WHO guidelines were con- sidered for essential amino acid requirements. In this study, the protein content and amino acid composition of various meat-based burger patties (ABB), including chicken, pork and beef, and plant-based burgers (PBB) were analysed. The results showed that among the plant-based samples, PBB 4 had the highest protein content (24.81 g / 100 g), which was almost equal to that of ABB 1 (26.48 g / 100 g). The most abundant amino acids detected were Glu, Asp, Leu, Lys, Arg, Ser, Pro and Gly, with samples PBB 6, PBB 3 and ABB 1 having the highest concentrations. PBB 1 stood out as a valuable pro- tein source with the highest content of essential amino acids (400.08 mg/g protein) among the plant-based burger patties. Some plant-based burger patties were deficient in essential amino acids, with PBB 3 and PBB 4 having the highest defi- ciency. The practical value of this study is that it helps people to make informed dietary choices. Key words: human nutrition, essential amino acids, pro- teins, meat substitutes, plant-based burger patties, nutrition value Določitev aminokislinske sestave mesnih in rastlinskih bur- gerjev Izvleček: V študiji smo preučevali vsebnost beljakovin in aminokislinsko sestavo različnih komercialno dostopnih mesnih in rastlinskih burgerjev. Namen študije je bil ugoto- viti, ali rastlinski burgerji izpolnjujejo zahteve po vsebnosti esencialnih aminokislin v humani prehrani. Aminokislinski profili so bili določeni z analizatorjem aminokislin, kemijski indeksi posameznih esencialnih aminokislin za odrasle pa izračunani na podlagi smernic FAO/WHO. V študiji smo analizirali vsebnost beljakovin in aminokislinsko sestavo različnih mesnih burgerjev (ABB) iz piščančjega, svinjskega in govejega mesa ter burgerjev iz sestavin rastlinskega izvo- ra (PBB). Rezultati so pokazali, da je imel med rastlinskimi vzorci PBB 4 najvišjo vsebnost beljakovin (24,81 g/100 g), ki je bila skoraj primerljiva z ABB 1 (26,48 g/100 g). Najbolj zastopane aminokisline v vzorcih so bile Glu, Asp, Leu, Lys, Arg, Ser, Pro in Gly, pri čemer so bile najvišje koncentracije le-teh izmerjene v vzorcih PBB 6, PBB 3 in ABB 1. PBB 1 se je pokazal kot dober vir beljakovin z najvišjo vsebnostjo esencialnih aminokislin (400,08 mg/g beljakovin) med ras- tlinskimi burgerji. Pri nekaterih rastlinskih burgerjih je bilo ugotovljeno pomanjkanje esencialnih aminokislin, največje pri vzorcih PBB 3 in PBB 4. Praktična vrednost te študije je v tem, da ljudem pomaga sprejemati premišljene prehranske odločitve. Ključne besede: humana prehrana, esencialne ami- nokisline, beljakovine, mesni nadomestki, rastlinski burgerji, prehranska vrednost Acta agriculturae Slovenica, 120/4 – 20242 K. BERISHA et al. 1 INTRODUCTION The traditional meat industry is reaching produc- tion limits and is associated with environmental prob- lems (e.g. impact on natural resources, greenhouse gas emissions, and use of land mass) (Godfray et al., 2018). Therefore, alternative protein sources, particularly plant-based meat analogues, are becoming increasingly popular as a potential solution to reduce the supply gap and environmental impact of conventional meat pro- duction. The popularity of plant-based meat substitutes valued for its ability to mimic traditional meat prod- ucts reflects a broader trend among consumers seeking sustainable and environmentally friendly food options. The study addresses the nutritional aspects of meat and meat substitutes, aligning with the current imperative to address the environmental consequences of dietary choices. This study contributes to informing people about the quality of alternative protein sources. A balanced diet is a key factor in human life and well-being. It consists of various foods from which hu- mans obtain the nutrients necessary for health and the performance of vital functions (Ahmad et al., 2018). As an important food group that contributes to a balanced and healthy diet, meat and its products are a rich source of nutrients (Pereira & Vicente, 2022). However, meat consumption is considered unfavorable by some due to its presumed impact on the environment, ethics, and certain religious traditions. Customer awareness of sus- tainability and environmentally-friendly food produc- tion practices is growing and there are trends towards adopting vegetarian or vegan diets or limiting meat consumption (Graça et al., 2019). An alternative could be plant-based meat ana- logues, which are plant-derived foods that have the sen- sory and chemical properties of traditional meat prod- ucts (Ismail et al., 2020). While not all meat analogues fall under the classification of ultra-processed foods, a significant number of modern meat analogues available in the food market today meet this criterion. Ultra-pro- cessed foods are food products that contain minimal or no whole foods and are instead produced with pro- cessed ingredients or substances that are obtained from whole foods. These processed ingredients can include protein isolates, oils, hydrogenated oils and fats, flours and starches, sugar variants, refined carbohydrates, and other added ingredients that increase the value of the product (Bohrer, 2019; Monteiro et al., 2019). Ultra- processed foods are associated with an increased risk of obesity, cardiovascular diseases, and metabolic disor- ders due to high quantity of added sugars, harmful fats, and low nutritional value (Monteiro et al., 2013). Some consumers prefer modern meat analogues because they fulfill their expectations by imitating meat in terms of appearance, quality, and taste. According to the forecast of the International Institute for Sustainable Development, the number of people on our planet will reach 9.9 billion by 2050 (Population Reference Bureau, 2020). Feeding such a mass population with meat could have a detrimental ef- fect on the environment. The benefits of reducing meat consumption are therefore manifold. Environmental studies have been conducted on protein-rich products, including plant-based meat analogues (soybeans, green peas, lupines, rice, etc.), animal proteins (milk, meat, in- sects, lab-produced proteins), and mycoproteins. Most studies have shown that plant-based meat analogues have less environmental impact than meat analogues that include a broader range of substitutes, including plant-based, fungi-based, and cultured meat products that appeal to a wider audience (Kyriakopoulou et al., 2019). Meat and its products are primary sources of protein and provide complete proteins containing all essential amino acids such as phenylalanine, valine, threonine, tryptophan, isoleucine, methionine, histi- dine, leucine and lysine. Complete proteins, which are found in animal foods such as eggs, milk, fish and meat, have the correct ratio of essential amino acids, making them sufficient sources of protein. Incomplete proteins lack some essential amino acids (Arentson-Lantz et al., 2021), but can be complemented by combining differ- ent incomplete or complete protein sources. Secondary proteins, commonly found in plants, may be deficient in some essential amino acids. Adequate dietary proteins are crucial at every stage of life and ensure the uptake of essential amino acids (Brestenský et al., 2019). For example, cereal proteins with low lysine content and legumes with a lack of sulphur-containing amino ac- ids (methionine and cysteine) pose a challenge for the utilisation of proteins in the human body. The amino acid composition and digestibility of proteins from dif- ferent plant sources are therefore of crucial importance. Studies by Day et al. (2022) and Foschia et al. (2017) on plant proteins as meat analogues emphasize the opti- mal mixture of certain grains and legumes to achieve a meat-like amino acid profile. This study aimed to detect amino acids in different burger patties (plant- and animal-based). In addition, this study investigated whether plant-based meat sub- stitutes (vegetable-based meats) meet the daily require- ments for the intake of proteins and essential amino ac- ids in the human diet when consuming these products. Acta agriculturae Slovenica, 120/4 – 2024 3 Evaluation of amino acid composition in different types of meat and plant-based burger patties 2 MATERIAL AND METHODS 2.1 BURGER SAMPLES Burger samples were collected from ten fast-food restaurants in Budapest, Hungary, during the study pe- riod of March–September 2023. A total of 50 burger samples of meat-based burgers (ABB) and plant-based burgers (PBB) were analyzed, with five samples for each type of burger. The patties of animal origin included chicken, pork, and beef, coded as ABB 1, ABB 2, ABB 3 and ABB 4. For the plant-based burgers, six different types were taken for analysis: PBB 1, PBB 2, PBB 3, PBB 4, PBB 5, and PBB 6 (Table 1). Samples were indepen- dently collected to ensure no cross-contamination and stored at −20 °C. The quality standards used for testing were based on the Crown Food Group – CFG (2023). 2.2 AMINO ACID DETERMINATION Protein-building amino acids were detected as de- scribed by Berisha et al. (2023). Briefly, 500 mg portions of burger patties were subjected to hydrolysis in a vessel contain- ing 10 mL of 6 mol L-1 hydro- chloric acid, while being exposed to a nitrogen atmosphere at a temperature of 110 °C for a duration of 24 hours in a controlled thermostat. The process of neutralization was carried out in a 25 ml volumetric flask by adding 10 ml of a 4 M L-1 NaOH solution to the hydrolyzed sample. The flask was then filled with buffer solution at a pH of 2.2. The samples that had been neutralized were passed through a mem- brane filter with a pore size of 0.25 µm. The determi- nation of amino acids was conducted using a AAA400 Automatic Amino Acid Analyzer (Ingos Ltd., Prague, Czech Republic), which was equipped with a cation- exchange column. The separation was carried out us- ing a stepwise gradient elution technique with lithium buffer systems. Following post-column derivatization using a ninhydrin reagent, colorimetric detection was performed at wavelengths of 570 nm and 440 nm spe- cifically for Pro. Three samples were produced in paral- lel for analysis (Berisha et al., 2023). 2.3 AMINO ACID SCORE CALCULATION The calculation of the essential amino acids scores or the so-called chemical score is done using the for- mula given by WHO (1991), equation 1, expressed ei- ther as a ratio of unity (recommended) or on a percent- age scale (Food and Agriculture Organization Expert Working Group, 2018). Reference protein amounts are shown in three Table 1: Ingredients for plant-based burger patties Burger type Composition PBB 1 Water, 18% pea protein, rapeseed oil, refined coconut fat, rice protein, natural flavour, dry yeast, butter, emulsifier: methylcellulose, less than 1% potato starch; table salt, potassium chloride, beetroot concentrate, apple extract, pomegranate concentrate, sunflower lecithin, food vinegar, lemon concentrate, vitamins and minerals (zinc sulfate, niacin [vitamin B3], pyridoxine [vitamin B6], cobalamin [B12]) pantothenic acid [vitamin B5]. PBB 2 Water, 20% pea protein, diced onion, sunflower oil, diced turnip, stabilizer: methylcellulose, pea fiber, potato starch, vinegar, maple syrup, spices, beet syrup, edible salt, smoked lemon concentrate. PBB 3 Water, 14% soybean meal, carrot, refined coconut fat, wheat flour, 4% wheat protein, rapeseed oil, acidity regulator: potassium lactate, potassium acetate; emulsifier: methylcellulose, yeast extract, flavouring, green pea starch, soy protein, table salt, spices, beetroot with colouring effect, mushroom powder, spice extracts, smoke flavour. PBB 4 Water, 11.8% soy protein, 9.9% wheat protein, carrot, refined coconut fat, rapeseed oil, emulsifier: methyl- cellulose, spices, yeast extract, flavouring, starch, natural flavour, acidity regulator: potassium lactate, acetate; table salt, vinegar, beet concentrate with colouring effect, bamboo fiber, spice extracts, smoke flavour, food acid: citric acid, antioxidant: ascorbic acid. PBB 5, PBB 6 ABB4, PBB5, PBB6 the proportions of the ingredients in the patty and their full list are secret. However, the ABB4 is known to be made from beef, the PBB5 is made primarily from potatoes, carrots, green peas and corn, while the PBB6 is made from soy and wheat from sustainable farms. Source: adjusted from food product label 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑆𝑆𝑆𝑆𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 = mg of amino acid in 1 g of the tested protein mg of amino acid in 1 g of the reference protein 𝑆𝑆𝑆𝑆𝑒𝑒𝑒𝑒 (1) 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑆𝑆𝑆𝑆𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 = mg of amino acid in 1 g of the tested p otein mg of amino acid in 1 g of the reference protein 𝑆𝑆𝑆𝑆𝑒𝑒𝑒𝑒 (1) 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴 𝑆𝑆𝑆𝑆𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝐴𝑆𝑆𝑆𝑆𝑆𝑆𝑆𝑆 = mg of amino acid in 1 g of the tested protein mg of amino acid in 1 g of the reference protein 𝑆𝑆𝑆𝑆𝑒𝑒𝑒𝑒 (1) (1) Acta agriculturae Slovenica, 120/4 – 20244 K. BERISHA et al. groups: ages 3–14 years, 15–18 years, and more than 18 years, approved by FAO (2013). In this research, the ratio of essential amino acids for adults (over 18 years) was evaluated. 2.4 ESSENTIAL AMINO ACID DEFICIENCY IN- DEX CALCULATION The essential amino acid deficiency index is determined by calculating the differences in the es- sential amino acid value compared to the reference protein (Table 2). their tryptophan content. The main components of the analyzed plant products can be classified as leg- umes, which are characterized by the limiting amino acid in the group of sulfur-containing amino acids (Met + Cys), so that the lack of tryptophan values does not pose a problem in the evaluation. This es- sential amino acid is rather limiting in maize. Table 3 shows the total protein content of the samples per 100 g of product and the standard devia- tion of the analyzed samples. The protein content of the ABB1 is 26.48 g / 100 g. The highest value of veg- gie patties is the PBB 3 (24.81 g / 100 g), approach- ing the protein content of the ABB 1 and surpass- Table 2: Procedure for calculating the essential amino acid (EAA) deficiency index Essential amino acids (EAA) Reference protein (mg/g) ABB4 (mg/g protein) EAA score Calculation 1 – EAA score Index of EAA deficiency Histidine 16 31.29 1.96 1–1.96 −0.96 Isoleucine 30 22.36 0.75 1–0.75 0.25 Leucin 61 79.80 1.31 1–1.31 −0.31 Lysine 48 82.06 1.71 1–1.71 −0.71 Methionine and Cysteine 23 18.66 0.81 1–0.81 0.19 Phenylalanine and Tyrosine 41 64.75 1.58 1–1.58 −0.58 Threonine 25 46.02 1.84 1–1.84 −0.84 Valin 40 37.78 0.94 1–0.94 0.06 If the EAA deficiency index is positive, this means that the amino acid is present in the sample in a lower proportion than the amino acid of the refer- ence protein. If it is negative, there is a surplus. The deficit is therefore indicated by positive values. The sum of these positive values gives the essential amino acid deficiency index. The calculation procedure is shown in Table 2 for the samples of ABB4 burgers; this procedure was used for all analyzed burger types. 3 RESULTS 3.1 PROTEIN CONTENT ON MEAT PATTY AND PLANT-BASED MEAT ANALOGUES The total protein content of the samples was cal- culated using the results of the amino acid analysis (Table 3). Tryptophan is not included in the amino acid values obtained by chromatography, as its indole group is degraded by the acidic sample preparation, so that this amino acid cannot be detected by this method. Due to the variety of ingredients in burger patties, there is no information in the literature on Table 3: Total protein content of samples per 100 g of product Type of burger patty Protin Content (g/100 g) ± standard deviation (n = 3) PBB 1 16.96 ± 0.54 PBB 2 15.76 ± 0.42 PBB 3 24.81 ± 2.23 PBB 4 12.90 ± 2.81 PBB 5 4.34 ± 0.44 PBB 6 24.72 ± 2.15 ABB 1 26.48 ± 2.24 ABB 2 22.74 ± 0.72 ABB 3 18.26 ± 0.22 ABB 4 19.14 ± 0.18 ing all three meat samples in terms of protein. This burger contains 14 % soy flour, 4 % wheat protein, and mushrooms also contribute to the development of favorable protein content, so the product can be considered a good substitute for meat in this regard. The lowest protein, only 4.34 g / 100 g, is found in the PBB 5. Acta agriculturae Slovenica, 120/4 – 2024 5 Evaluation of amino acid composition in different types of meat and plant-based burger patties 3.2 PROTEIN-BUILDING AMINO ACIDS The results for the composition of protein-forming amino acids in plant-based burger patties are presented in Table 4. The main amino acids detected in the ana- lyzed samples include Glu, Asp, Leu, Lys, Arg, Ser, Pro, and Gly which account for more than 50 % of the total amount of amino acids detected. By comparing the total amount of amino acids detected in plant-based burger patties, it was found that the PBB 6 (247.21 mg/g) had the highest amount of the amino acids, followed by PBB 3 (248.1 mg/g). The lowest amount of amino acids was detected in PBB 5 (43.42 mg/g). Table 6 shows that PBB 1 with 400.08 mg/g protein has the highest total content of essential amino acids among the plant-based samples. PBB 2 and PBB 6 con- tain 381.57 mg/g and 351.96 mg/g, respectively, while PBB 3 has the lowest total essential amino acid con- tent at 304.01 mg/g protein. PBB 2, containing highest proportion of pea protein, has the highest content of leucine and isoleucine among the plant-based variants and is therefore a good source of these essential amino acids. PBB 3 has the lowest proportion of branched- chain amino acids, which make up around a third of the essential amino acids in muscle tissue. PBB 3, PBB 4, PBB 5 and PBB 6, which contain wheat protein, have Table 4: Amino acid composition of plant-based burger patties (mg/g) Amino acids PBB 1 PBB 2 PBB 3 PBB 4 PBB 5 PBB 6 Glutamic acid 38.36 35.98 86.72 42.17 10.34 67.59 Aspartic acid 19.46 18.53 18.53 9.85 5.52 24.33 Leucine 15.24 14.26 14.33 10.16 2.84 21.02 Lysine 13.1 11.75 10.49 5.04 2.29 13.94 Arginine 12.61 14.15 10.64 6.85 2.39 17.4 Serine 10.09 9.43 13.6 6.83 2.37 13.11 Proline 8.3 5.37 23.33 13.33 2.59 15.19 Glycine 8.24 7.38 10.86 5.4 2.12 12.49 Phenylalanine 7.52 7.39 9.99 5.47 1.59 10.95 Threonine 7.48 6.65 8.14 5.18 2.14 9.78 Valina 6.91 6.04 7.64 4.3 1.83 9.33 Alanine 6.87 6.63 9.25 4.24 2.39 9.97 Tyrosine 5.73 5.41 11.6 3.95 2.43 8.15 Histidine 4.26 3.95 7.24 2.82 1.64 6.35 Isoleucine 4.24 3.99 3.87 2.68 0.7 6.21 Methionine 1.17 0.69 1.84 0.74 0.23 1.4 Sum mg/g 169.6 157.6 248.1 129 43.42 247.21 Table 5 shows the results for the amino acid com- position of meat-based burger patties (ABB). Amino acids such as Glu, Gly, Asp, Lys, Leu, Arg, Ala, Thr, and Ser were detected in the highest amounts in these burg- ers. ABB 1 (264.82 mg/g), and ABB 2 (227.4 mg/g) had the highest amount of amino acids, followed by beef burgers (191.41 mg/g), while the lowest amount of ami- no acids was detected in pork burgers (182.64 mg/g). The total essential amino acid content is presented in Tables 6 and 7. All types of analyzed burgers con- tained all essential amino acids. Essential amino ac- ids are present in plant products in amounts between 304.01 to 400.08 mg/g of protein. a lower lysine content than the other two plant-based meat analogues. This is because lysine is the limiting amino acid in whole grains. According to the product label (Table 1), the soy flour content in the PBB 3 patty was higher (14 %) than in the PBB 4 patty (11.8 %). This is due to the fact that soy supplements the missing lysine content in wheat flour, which could explain the higher lysine content (42.35 mg/g and 39.29 mg/g re- spectively). The methionine content was relatively low in all plant-based products, as they contained a high proportion of pulses, which are low in sulphur-contain- ing amino acids such as methionine and cysteine. The PBB 3 patties contained the highest level of sulphur- Acta agriculturae Slovenica, 120/4 – 20246 K. BERISHA et al. containing amino acids (7.52 mg/g) and the lowest level in PBB 2 (4.36 mg/g). PBB 2 has the highest content of legume protein (20 % pea protein) and some other vegetables (turnips and onions), which explains the low value of the limiting amino acid. Table 7 shows the content of essential amino ac- ids in meat-based burgers. The ABB 1, made from raw animal material, contains 382.72 mg/g of protein, while the ABB 2, ABB 3, and ABB 4 contain higher amounts, ranging from 409.36 to 477.70 mg/g of protein. In general, plant-based products do not fall signifi- cantly short of the total amino acid content of a tradi- tional animal-based burger. 3.3 ESSENTIAL AMINO ACIDS SCORE AND LIM- ITING AMINO ACIDS The amount of each essential amino acid in mg/g protein in each sample was divided by the amount of mg/g protein of each of the essential amino acids of the FAO/WHO reference protein to give the score for each essential amino acid (EAAS- essential amino acid score). The closer this score is to 1 for each essential amino acid, the more the food corresponds to the FAO/ WHO reference protein composition. If the essential amino acid has a ratio less than l then it is counted as the limiting amino acid. Table 5: Amino acid composition of meat-based burger patties (mg/g) Amino acids ABB 1 ABB 2 ABB 3 ABB 4 Glutamic acid 53.23 46.65 36.59 34.47 Glycine 26.30 9.11 7.71 11.62 Aspartic acid 24.09 20.68 16.14 18.72 Lysine 21.71 22.07 17.42 15.91 Leucine 21.16 22.85 18.43 16.26 Arginine 19.54 17.68 14.27 12.46 Alanine 17.47 11.23 8.59 10.68 Threonine 12.20 12.16 9.30 9.26 Serine 12.03 10.16 7.71 8.08 Proline 10.80 6.38 4.35 9.49 Valine 10.00 10.97 8.20 10.77 Phenylalanine 9.23 10.00 8.04 6.45 Histidine 8.27 4.44 7.83 6.75 Tyrosine 7.92 8.28 6.60 5.68 Isoleucine 5.93 7.62 6.04 7.84 Methionine 4.93 7.14 5.42 6.98 Sum mg/g 264.82 227.40 182.64 191.41 Table 6: Essential amino acid composition of plant-based samples (mg/g protein) Essential amino acids PBB 1 PBB 2 PBB 3 PBB 4 PBB 5 PBB 6 Histidine 25.15 25.05 29.00 22.12 37.23 25.68 Isoleucine 25.03 25.30 15.75 20.60 16.46 25.10 Leucine 89.94 90.49 58.89 79.07 66.40 84.96 Lysine 89.94 74.61 42.35 39.29 53.08 56.27 Methionine 6.90 4.36 7.52 6.02 5.40 5.50 Phenylalanine and Tyrosine 78.16 81.23 87.04 73.06 92.65 77.27 Threonine 44.16 42.21 32.67 40.40 49.09 39.51 Valine 40.80 38.32 30.79 33.68 42.17 37.67 Sum 400.08 381.57 304.01 314.24 362.48 351.96 Acta agriculturae Slovenica, 120/4 – 2024 7 Evaluation of amino acid composition in different types of meat and plant-based burger patties The results for the essential amino acid score are shown in Table 8 for plant-based burger patties and in Table 9 for animal-based burger patties, in which the limiting amino acids are marked. The amino acids methionine and cysteine, which are normally found in larger quantities in meat, are the limiting amino acids in all plant-based burger patties, which contain significant amounts of legumes. The low- est amount is found in PBB 2, which contains 20 % pea protein. The ABB 1 patty contains three essential amino acids (isoleucine, methionine and valine) with a value of less than 1 compared to the FAO/WHO reference proteins. It can be assumed that a vegetable protein- based additive was added to the minced meat as an en- hancer, binder or fat substitute, which can be measured as protein together with the meat content of the sample, resulting in values for essential amino acids below 1. Among meat-free products, PBB 3 has the highest protein content per 100 g, but qualitatively it is consid- ered the most unfavorable source of protein, as it has Table 7: Essential amino acid composition of animal-based samples (mg/g protein) Essential amino acids ABB 1 ABB 2 ABB 3 ABB 4 Histidine 31.29 19.67 42.86 35.29 Isoleucine 22.36 33.46 33.03 40.96 Leucine 79.80 100.44 100.87 84.95 Lysine 82.06 97.04 95.37 83.11 Methionine 18.66 31.37 29.66 36.51 Phenylalanine and Tyrosine 64.75 80.35 80.13 63.33 Threonine 46.02 53.40 50.90 48.38 Valine 37.78 48.19 44.90 56.40 Sum 382.72 463.92 477.72 431.05 Table 8: Essential amino acids score and limiting amino acids of plant-based burger samples Essential amino acids PBB 1 PBB 2 PBB 3 PBB 4 PBB 5 PBB 6 Histidine 1.57 1.57 1.81 1.38 2.33 1.60 Isoleucine 0.83 0.84 0.52 0.69 0.55 0.84 Leucine 1.47 1.48 0.97 1.30 1.09 1.39 Lysine 1.87 1.55 0.88 0.82 1.11 1.17 Methionine and Cysteine 0.30* 0.19* 0.33* 0.26* 0.23* 0.24* Phenylalanine and Tyrosine 1.91 1.98 2.12 1.78 2.26 1.88 Threonine 1.77 1.69 1.31 1.62 1.96 1.58 Valine 1.02 0.96 0.77 0.84 1.05 0.94 Table 9: Essential amino acids score and limiting amino acids of meat-based burger samples Essential amino acids ABB 1 ABB 2 ABB 3 ABB 4 Histidine 1.96 1.23 2.68 2.21 Isoleucine 0.75* 1.12 1.10 1.37 Leucine 1.31 1.65 1.65 1.39 Lysine 1.71 2.02 1.99 1.73 Methionine and Cysteine 0.81 1.36 1.29 1.59 Phenylalanine and Tyrosine 1.58 1.96 1.95 1.54 Threonine 1.84 2.14 2.04 1.94 Valine 0.94 1.20 1.12 1.41 Acta agriculturae Slovenica, 120/4 – 20248 K. BERISHA et al. low levels of essential amino acids (isoleucine, leucine, lysine, methionine, valine), compared to the reference protein. PBB 1 and PBB 5 are deficient only in methio- nine and isoleucine. The amount of histidine, phenyla- lanine, tyrosine, as well as threonine, is above the mini- mum values determined by FAO/WHO experts. 3.4 ESSENTIAL AMINO ACID DEFICIENCY IN- DEX The greatest deficiencies in essential amino acids were found in PBB 3 (1.53) and PBB 4 (1.39) (Table 10). PBB 5, which probably contains mainly potatoes, was not the worst product due to the added egg content, but still showed a large deficit of 1.22. PBB 2 and PBB 6 have almost identical values of 1.01 and 0.98, respec- tively. Although PBB 1 is deficient in only 2 essential amino acids, their sum amounts to 0.87. Since the to- tal ratio of essential amino acids in most meat-based burgers is above 1 (Table 9), their essential amino acid deficiency index is 0. However, ABB 1 has a deficiency of 0.50, which may be due to the addition of vegetable protein-based additives. The ratios of the essential ami- no acids threonine (2.14), lysine (2.02), phenylalanine and tyrosine (1.96), leucine (1.65) in ABB 2, histidine (2.68), threonine (2.04), lysine (1.9), phenylalanine and tyrosine (1.95), leucine (1.65) in ABB 3, histidine (2.21), threonine (1.94), lysine (1.73), methionine (1.59), phe- nylalanine and tyrosine (1.54) in ABB 4 are much higher than those of the reference protein. Most of the plant- based samples contain less methionine, isoleucine and valine. In PBB 4, the amount of lysine is also below the recommended value, while in PBB 3 the leucine content is suboptimal compared to the reference protein. 4 DISCUSSION The amino acid composition of meat-based burg- er patties and plant-based burger patties can be very different, primarily depending on the protein source. Meat-based burger patties typically provide more com- plete and balanced protein nutrition, including essen- tial vitamins and minerals naturally present in meat, while plant-based meat analogues vary based on the in- gredients used in their formulation, as in the study con- ducted by Day et al. (2022). Meat and its products are thought to have been part of the human diet from 2–6 million years ago and increased over time as a result of increases in income and population number. However, the trend towards the use of plant-based meat substi- tutes in the human diet is increasing (Godfray et al., 2018; Filin et al., 2023). When comparing meat-based burger patties and plant-based burger patties, consumers should consider their dietary goals, preferences, and ethical concerns. If someone relies heavily on meat analogues, they may need to supplement their diet to ensure they are get- ting all the essential amino acids. It is also important to know the specific amino acid composition of the two types of burger patties and how it meets nutritional needs and individual health status. A study conducted by Bryant et al. (2019) examined the consumer accept- ance of plant-based and clean (cultured) meat products in the United States, China, and India. The findings in- dicate that urban, well-educated, and high-income con- sumers in India and China demonstrate a higher pro- pensity to purchase clean meat and plant-based meat compared to consumers in the USA. The research con- ducted by Ismail et al. (2020) also highlights a signifi- cant bias towards urban, educated, and wealthy groups in China and India, as opposed to the overall popula- tion. The study revealed that disgust plays a crucial role in determining the adoption of plant-based and clean meat, a distinctive observation limited to the United States. In China, there is a notable deviation from the generally observed demographic pattern in the West- ern countries regarding the acceptance of clean meat, particularly in relation to gender. The attitudinal fac- tors influencing the acceptance of both plant-based and clean meat in China include perceptions of healthiness, nutritional value, excitement, goodness, and necessity. Given the recommended dietary allowance (RDA) for protein intake in adults is 0.8 grams per kilogram of body weight per day (Nishimura et al., 2023) and in- corporating these plant-based options can help meet protein needs without the associated health risks of ex- cessive red meat consumption. High intake of red meat, particularly processed varieties, has been linked to in- Table 10: Index of the deficiency of essential amino acids of the samples compared to the reference protein Type of burger Index of the deficiency of essential amino acids PBB 1 0.87 PBB 2 1.01 PBB 3 1.53 PBB 4 1.39 PBB 5 1.22 PBB 6 0.98 ABB 1 0.50 ABB 2 0.00 ABB 3 0.00 ABB 4 0.00 Acta agriculturae Slovenica, 120/4 – 2024 9 Evaluation of amino acid composition in different types of meat and plant-based burger patties creased risks of chronic diseases such as cardiovascular disease, type 2 diabetes, and certain cancers due to its saturated fat, cholesterol content, and potential carcin- ogens formed during cooking (Fogelholm et al., 2015; Larsson and Orsin., 2014; Zheng et al., 2019). The analysis of various burger patties used in pop- ular fast-food offerings reveals that these meat substi- tutes can be made from a variety of plant-based ingre- dients, such as soy, peas, beans, or mushrooms, and may serve as satisfactory protein substitute in the human diet, as they contain all essential amino acids. However, it should be noted that while meat contains a balanced profile of essential amino acids, plant-based burger pat- ties (i.e. legume-based meat analogues) are often defi- cient in amino acids, as methionine, lysine, and cysteine. PBB 1 stands out as a valuable protein source among these options due to its high content of essential amino acids. A review paper by Kyriakopoulou et al. (2021) in functionality of ingredients in plant-based meat ana- logues showed that the main ingredients in commer- cial meat analogues are soy, pea, and gluten, which are widely available, and by-products of established food production lines. This provides additional opportunities to enhance functionality and optimize resource utilization. For ex- ample, pea protein and rice protein can be combined to create a complete amino acid profile, making the prod- uct more nutritionally comparable to animal protein. Similarly, combining soy protein with wheat gluten can improve the texture and binding properties of meat substitutes, as noted by Asgar et al. (2010). Additionally, the incorporation of algae or seaweed extracts can en- hance the nutritional value and provide unique flavours and textures (Abdel-Moatamed et al., 2024; Schuler et al., 2020). These combinations not only enhance func- tionality but also optimize resource utilization, open- ing new possibilities for sustainable and nutritious food products. The shift towards individual dietary patterns such as vegetarian or vegan diets is now being driven by a variety of factors, including health concerns about sat- urated fats in animal products, environmental impact, greenhouse gas emissions, climate change and ethical concerns about animal husbandry and slaughter prac- tices. However, consumers may need to pay more atten- tion to their overall diet to ensure they are getting all the essential amino acids. In addition, a combination of plant and animal proteins can be explored to produce healthy burger patties that meet essential amino acid requirements while reducing saturated fat content from animal products. The evaluation of amino acid com- position highlights the importance of considering the overall quality of the protein and the completeness of the nutritional content when choosing between meat- based and plant-based burger patties. It is crucial to educate consumers about the differences in amino acid composition and the nutritional impact of choosing between meat and meat substitutes to help them make dietary choices in line with their needs and preferences. Ongoing research to improve the amino acid composi- tion of meat substitutes and fortify them with essential nutrients could bring products closer to meat products and have a siginificant impact on the food industry. 5 CONCLUSIONS This study provides a comprehensive analysis of the protein content and amino acid composition of a wide variety of burger patties, including both plant- based and meat-based options. The results of the amino acid composition studies showed a wide range of ami- no acids in the products, with the major amino acids Glu, Asp, Leu, Lys, Arg, Ser, Pro, and Gly accounting for more than half of all amino acids found. The analysis of the protein composition revealed that plant-based sub- stitutes (e.g. PBB 3) can have competitive protein levels comparable to those of traditional meat products. Cer- tain differences in the composition of essential amino acids were found, especially in the plant-based patties with lower amounts of methionine and lysine. Plant- based patties with a high proportion of legumes had limiting values for cysteine and methionine, confirming their status as limiting amino acids. However, it is also important to consider the bioavailability of proteins. Factors that affect the bioavailability of plant-based proteins include the presence of antinutritive factors such as phytates, which can inhibit mineral absorption, and the protein matrix itself, which can affect digestion and absorption. Although the amino acid profiles and protein content of plant-based burgers are promising, further bioavailability studies are needed to fully un- derstand their nutritional impact. The results of the study point to the potential of mixing animal and plant protein sources to address the lack of essential amino acids in plant-based burger patties. This approach could reduce meat consumption while ensuring adequate nu- trient intake. This study aims to provide information to make better dietary decisions. 6 REFERENCES Abdel-Moatamed, B. R., El-Fakhrany, A. E. M., Elneairy, N. A., Shaban, M. M., & Roby, M. H. (2024). The Impact of Chlo- rella vulgaris Fortification on the Nutritional Composition Acta agriculturae Slovenica, 120/4 – 202410 K. BERISHA et al. and Quality Characteristics of Beef Burgers. Foods, 13(12), 1945. https://doi.org/10.3390/foods13121945 Ahmad, R., Imran. A., & Muhammad, B. (2018). Nutrition- al composition of meat. In Meat Science and Nutrition. London: IntechOpen. https://doi.org/10.5772/intecho- pen.77045 Arentson-Lantz, E., Von Ruff, Z., Harvey, M., Wacher, A., & Paddon-Jones, D. (2021). A moderate serving of a low- er-quality, incomplete protein does not stimulate skel- etal muscle protein synthesis. Current Developments in Nutrition, 5(2), 487–487. https://doi.org/10.1093/cdn/ nzab041_002 Asgar, M. A., Fazilah, A., Huda, N., Bhat, R., & Karim, A. A. (2010). Nonmeat protein alternatives as meat extenders and meat analogues. Comprehensive reviews in food science and food safety, 9(5), 513–529. https://doi.org/10.1111/ j.1541-4337.2010.00124.x Berisha, K., Gashi, A., Mednyánszky, Z., Bytyqi, H., & Simon Sarkadi, L. (2023). Nutritional characterisation of home- made beef sausage based on amino acid, biogenic amines, and fatty acid composition. Acta Alimentaria, 52(3), 439–448. https://doi.org/10.1556/066.2023.00071 Bohrer, B.M. (2019). An investigation of the formulation and nutritional composition of modern meat analo- gueue products. Food Science and Human Wellness, 8(4), 320–329. https://doi.org/10.1016/j.fshw.2019.11.006 Brestenský, M., Nitrayová, S., Patráš, P., & Nitray, J. (2018). Di- etary requirements for proteins and amino acids in hu- man nutrition. Current Nutrition & Food Science, 15(7), 638–645. https://doi.org/10.2174/15734013146661805071 23506 Bryant, C., Szejda, K., Parekh, N., Deshpande, V., & Tse, B. (2019). A survey of consumer perceptions of plant-based and clean meat in the USA, India, and China. Fron- tiers in Sustainable Food Systems, 11, article number 11. https://doi.org/10.3389/fsufs.2019.00011 Crown Food Group (CFG). (2023). Food safety documentation. Food safety and quality policy. Retrieved from https://www. crownnational.co.za/_downloads/food-safety-policy. pdf?version=13#:~:text=Crown%20Food%20Group%20 (CFG)%20is,are%20safe%20for%20human%20consump- tion Day, L., Cakebread, J.A., & Loveday, S.M. (2022). Food pro- teins from animals and plants: Differences in the nutri- tional and functional properties. Trends in Food Science & Technology, 119, 428–442. https://doi.org/10.1016/j. tifs.2021.12.020 Deutz, N. E., Bauer, J. M., Barazzoni, R., Biolo, G., Boirie, Y., Bosy-Westphal, A., ... Caldern, C. P. (2014). Protein in- take and exercise for optimal muscle function with aging: Recommendations from the ESPEN Expert Group. Clini- cal Nutrition, 33(6), 929–936. https://doi.org/10.1016/j. clnu.2014.04.007 Filin, S., Bal-Prylypko, L., Nikolaenko, M., Holembovska, N., & Kushnir, Yu. (2023). Development of technol- ogy for plant-based minced semi-finished products. Animal Science and Food Technology, 14(2), 100–112. https://doi.org/10.31548/animal.2.2023.100 Fogelholm, M., Kanerva, N., & Männistö, S. (2015). Asso- ciation between red and processed meat consumption and chronic diseases: the confounding role of other di- etary factors. European journal of clinical nutrition, 69(9), 1060–1065. https://doi.org/10.1038/ejcn.2015.63 Food and Agriculture Organization Expert Working Group. (2018). Protein quality assessment in follow-up formula for young children and ready to use therapeutic foods. Rome: Food and Agriculture Organization of the United Na- tions. Retrieved from https://www.fao.org/3/ca2487en/ CA2487EN.pdf Foschia, M., Horstmann, S. W., Arendt, E. K., & Zannini, E. (2017). Legumes as functional ingredients in gluten-free bakery and pasta products. Annual Review of Food Sci- ence and Technology, 8, 75–96. https://doi.org/10.1146/ annurev-food-030216-030045 Godfray, C., Aveyard, P., Garnett, T., Hall, W. J., Key, J. T., Lorimer, J., ... & Jebb, A.S. (2018). Meat consump- tion, health, and the environment. Science, 361(6399). https://doi.org/10.1126/science.aam5324 Graça, J., Godinho, C.A., & Truninger, M. (2019). Reducing meat consumption and following plant-based diets: Cur- rent evidence and future directions to inform integrat- ed transitions. Trends in Food Science & Technology, 91, 380–390. https://doi.org/10.1016/j.tifs.2019.07.046 Ismail, I., Hwang, Y. H., & Joo, S. T. (2020). Meat analogue as future food: A review. Journal of Animal Science and Technology, 62(2), 111–120. https://doi.org/10.5187/ jast.2020.62.2.111 Kyriakopoulou, K., Dekkers, B., & van der Goot, A. J. (2019). Plant-based meat analogueues. In C.M. Galanakis (Ed.), Sustainable meat production and processing (pp.103– 126). London: Academic Press is an imprint of Elsevier. https://doi.org/10.1016/C2017-0-02230-9 Kyriakopoulou, K., Keppler, J. K., & van der Goot, A. J. (2021). Functionality of ingredients and additives in plant-based meat analogueues. Foods, 10(3), article number 600. https://doi.org/10.3390/foods10030600 Larsson, S. C., & Orsini, N. (2014). Red meat and processed meat consumption and all-cause mortality: a meta-anal- ysis. American journal of epidemiology, 179(3), 282–289. https://doi.org/10.1093/aje/kwt261 Monteiro, C. A., Cannon, G., Levy, R. B., Moubarac, J. C., Louzada, M. L. C., Rauber, F., ... Jaime, P. C. (2019). Ul- tra-processed foods: What they are and how to iden- tify them. Public Health Nutrition, 22(5), 936–941. https://doi.org/10.1017/S1368980018003762 Monteiro, C. A., Moubarac, J. C., Cannon, G., Ng, S. W., & Popkin, B. (2013). Ultra-processed products are becom- ing dominant in the global food system. Obesity Reviews, 14(S2), 21–28. https://doi.org/10.1111/obr.12107 Nishimura, Y., Højfeldt, G., Breen, L., Tetens, I., & Holm, L. (2023). Dietary protein requirements and recommenda- tions for healthy older adults: a critical narrative review of the scientific evidence. Nutrition research reviews, 36(1), 69–85. https://doi.org/10.1017/S0954422421000329 Pereira, P. C., & Vicente, F. (2022). Meat nutritive value and human health. In New aspects of meat quality (second edi- tion) (pp. 561–577). https://doi.org/10.1016/B978-0-323- 85879-3.00024-6 Acta agriculturae Slovenica, 120/4 – 2024 11 Evaluation of amino acid composition in different types of meat and plant-based burger patties Population Reference Bureau. (2020). 2020 world popula- tion data sheet. Retrieved from https://interactives.prb. org/2020-wpds/ Schüler, L., Greque de Morais, E., Trovão, M., Machado, A., Carvalho, B., Carneiro, M., ... Varela, J. (2020). Isolation and characterization of novel Chlorella vulgaris mutants with low chlorophyll and improved protein contents for food applications. Frontiers in Bioengineering and Biotech- nology, 8, 469. https://doi.org/10.3389/fbioe.2020.00469 Zheng, Y., Li, Y., Satija, A., Pan, A., Sotos-Prieto, M., Rimm, E., ... Hu, F. B. (2019). Association of changes in red meat consumption with total and cause specific mortality among US women and men: two prospective cohort stud- ies. BMJ, 365. https://doi.org/10.1136/bmj.l2110