Plant-based meat substitutes and their
nutritional composition
A study on iron content, zinc content, calcium content and protein quality in meatballs and plant-based substitutes and how they contribute to the goal of reaching recommended daily intakes
Degree Project
Author: Linnea Thyrén Supervisor: Maria Bergström Examiner: Cornelia Witthöft Semester: VT20
Abstract
The purpose of this thesis was to study similarities and differences between a meat product and its plant-based substitutes in terms of how they enable people to reach the recommended daily intakes (RDI). The meat product included in the study were meatballs and its substitutes were three different plant-based alternatives. One was mainly based on soy, one on pea protein and one consisted of several different vegetables. The parameters studied were iron, zinc and calcium content as well as protein quality. By analyzing the four different products with flame atomic absorption spectroscopy (FAAS), the mineral content was calculated and protein quality was determined using amino acid analyzis carried out by the laboratory analysis company ALS. The method used to determine protein quality was DIAAS (Digestible Indispensable Amino Acid Score). Example meals and example days were created to visualize any differences and similarities when the products were put in a wider perspective. The results showed that there were differences between the products when they were compared individually, but that the differences were negligible when the products were included in an example meal or example day. This indicates that the products fulfill comparable dietary requirements and that the plant-based products were good substitutes for the meat product. The soy-plant-based product was the best plant-based alternative when it comes to iron content, calcium content and protein quality. However, the soy product and the remaining substitutes reached recommended intakes for the same parameters when included in example days, which shows that the differences between the products are only present when analyzed individually.
Sammanfattning
Syftet med denna studien var att undersöka likheter och skillnader mellan en köttprodukt och dess växtbaserade substitut sett till möjliggörandet att uppnå rekommenderade dagliga intag (RDI) av kalcium, järn, zink och essentiella aminosyror. Köttprodukten som inkluderades i arbetet var köttbullar och dess substitut var tre olika växtbaserade ersättningar. En var främst baserad på soja, en på ärtprotein och en bestod av flertalet olika grönsaker. De parametrar som studerades var järn-, zink-, och kalciuminnehåll samt proteinkvalitet. Genom att analysera de olika produkterna med atomabsorptionsspektroskopi med flamma (FAAS) beräknades mineralinnehållen och proteinkvalitet bestämdes med hjälp av aminosyraanalyser från analysföretaget ALS. Metoden som användes för att bestämma proteinkvalitet var DIAAS (Digestible Indispensable Amino Acid Score). Exempelmåltider och exempel på heldagsmenyer togs fram för att visualisera eventuella skillnader och likheter då produkterna sattes i ett större perspektiv. Resultaten visade att det fanns skillnader mellan produkterna då de jämfördes individuellt, men att skillnaderna var försumbara då produkterna inkluderades i en exempelmåltid eller heldagsmeny. Det indikerar att produkterna fyller en liknande funktion och att de växtbaserade substituten var goda ersättare för köttprodukten. Produkten baserad på soja var det substitutet med bäst näringsinnehåll sett till järn, kalcium och proteinkvalitet.
Key words
Acknowledgements
I would like to express my gratitude to my supervisor Maria Bergström at Linnaeus University for her support and guidance throughout this thesis project.
Table of content
1 Introduction 1
1.1 Meatballs and vegan alternatives 1
1.1.1 Limitations with meat products 1
1.1.2 Description of the products studied in the thesis 2
1.2 Descriptions of the minerals studied 3
1.2.1 Iron 3
1.2.2 Calcium 4
1.2.3 Zinc 5
1.2.4 Determination of mineral concentrations 5
1.2.5 Limitations mineral analysis 6
1.3 Proteins and how they are analyzed 7
1.3.1 Meat protein 8
1.3.2 Plant protein 8
1.3.3 HPLC with fluorescence detection for amino acid profile 9
1.3.4 Evaluation of protein quality 11
1.4 The principle for example meals and example days 12
1.4.1 Example meals 12
1.4.2 Example days 13
1.5 Dietary recommendations 13
2 Aim 14
3 Method and materials 14
3.1 Quantitative determination of iron, zinc and calcium with FAAS 14 3.2 Determination of protein quality, example meals and example days 15
4 Results 16
4.1 Analysis of mineral content 16
4.2 Analysis of protein quality 17
4.3 Analysis of example meals and example days 17
5 Discussion 18 5.1 Protein quality 18 5.2 Zinc 19 5.3 Calcium 20 5.4 Iron 21 5.5 Example meals 22 5.6 Example days 23
5.7 Social relevance, ethics, and impact on the environment 23
6 Conclusions 24
7 References 25
1 Introduction
1.1 Meatballs and vegan alternatives
Meatballs began to be consumed in Sweden in the early 1700s, in the form of boiled meatballs. But since the 1800s, the most common way to cook them is by frying them in a frying pan or in the oven. The food has since then had a clear role in Swedish home cooking. A traditional Swedish way of serving meatballs is with potatoes, lingonberry jam, cream sauce, and green peas [1]. Recently, both new and old companies have started producing meat-free alternatives, where the idea is that they should be able to replace the meat product. The ingredients vary and there are several different protein sources used, of which some common ones are pea and soy protein. Retail sales-data has showed that the total retail market for the plant-based food sector increased their sales with 20% in 2018, a number that continues to increase [2].
1.1.1 Limitations with meat products
The main ingredients in meatballs are ground beef and minced pork [1], which also is the reason why the product is not an optimal choice in all aspects. The livestock industry causes large greenhouse gas emissions, contributes to waste of resources, uses large amounts of fossil fuels [3, 4], can increase the risk for certain diseases [4-6] and is called into question in terms of ethics [4]. Waste of resources and fossil fuels mainly comes from the feed production. Large amounts of food are used in feed production that humans could have consumed themselves. The use of landmass had been made more efficient if the cultivated crops had gone to human food instead of to animal feed. The use of landmass also becomes ineffective as alternatives that have less impact on the environment could have been cultivated. Large quantities of fossil fuels are used in the production, processing and transport of feed [3]. To provide a growing world population with sustainable food systems, changes must be made. The global food industry has many issues, but the fundamental problem lies in the fact that too many people consume a diet that can contribute to deficiency diseases, obesity, and non-communicable diseases. Changes need to be made to make food production and consumption of food sustainable. A prospective cohort analysis has shown that by replacing protein from animal sources with protein from plants sources, overall mortality is substantially reduced [5]. Plant sources also have lower environmental effects per serving than food from animal sources, see Figure 1. Meat from ruminants, such as beef, has the highest environmental effect. Seen to greenhouse gases, the emissions from ruminant meat are 1,200 g CO2-eq/serving whilst
vegetable products such as fruits, vegetables, cereals and roots have an environmental effect of 10-50 g CO2-eq/serving. The energy use for ruminant meat is more than 1,500 kJ/serving and
Figure 1. The effects of greenhouse gases, land use, energy use, acidification potential and eutrophication potential per serving of food produced [5].With permission from Elsevier.
The issues mentioned above are some reasons why many vegan alternatives have been developed lately. The meatless alternatives in this project are plant-based.
1.1.2 Description of the products studied in the thesis
The products selected for analysis are four different products from the Swedish food market. Three of the products are plant-based and one includes meat. They are different in terms of ingredients, nutritional values and to some extent appearance and texture. However, the common factor is the area of application, for which the products are seen as replaceable with each other. There are two different types of meat-free alternatives: the analogue meat product that mimics the original product and the alternative product that can differ in taste and texture but can still be used in the same way as the original meat product. Even if the products vary in taste or texture, they can be used for the same purpose and in dishes composed the same way. The following products are renamed and irrelevant ingredients, seen to this project, have been removed from the ingredients list.
The MeatB is a meatball of the traditional Swedish kind. The product contains 14 g of protein per 100 g.
Main ingredients: Beef, pork, onion, breadcrumbs, egg.
The GreenB is a vegetable-based ball, which is a vegan substitute to meatballs. The vegetable pieces are large and visible in the product. It contains 7.7 g of protein per 100 g.
Main ingredients: Chickpeas, green peas, carrot, red peppers, corn, kale, pea protein, onion,
rapeseed oil.
The PeaB is an analogue meatball with pea protein as its main protein source. It is a vegan alternative to a meatball and is supposed to look, taste and feel similar to it. The product contains 11 g protein per 100 g.
Main ingredients: Pea protein, rapeseed oil, potatoes, onion, oat bran.
The SoyB is an analogue meatball with soy protein as its main protein source. It is a vegan alternative to a meatball and is, like PeaB, supposed to look, taste and have the same texture as a meatball. The product contains 15 g protein per 100 g.
1.2 Descriptions of the minerals studied 1.2.1 Iron
Iron is a micronutrient found in for example liver, meat, egg, legumes, and wholegrain. The nutrient is a part of hemoglobin, which transports oxygen in the blood, and myoglobin which transports oxygen in muscles. Iron is also an important cofactor in enzymes. Iron deficiency (anemia) is the most common disease globally that occurs due to a lack of micronutrients [7]. It can, for example, be obtained as a result of bleeding, non-varied diets, diseases, intolerances or iron-deficient diets [8]. Iron in the diet is found in two biochemical forms: haem iron and non-haem iron. Haem iron is a complex of iron and porphyrin found in animal products (see example in Figure 2). Porphyrin is an aromatic compound which occurs naturally and consists of four modified pyrrole rings. Non-haem iron consists of inorganic ferrous (Fe2+) or ferric iron
(Fe3+) bound to salts, peptides or organic acids and is the predominant dietary form. It can be
found in both vegetable and animal products. However, it has a low bioavailability in contrast to haem iron whose bioavailability is high. In a diet containing both iron sources, about 50% of the iron absorbed by the body is haem iron even though the dietary amount is low. Non-haem iron bioavailability is low because of iron’s interaction with other dietary factors. Most dietary iron is also in its oxidized form, (Fe3+), whilst it has to be in its reduced, soluble form (Fe2+), to
be absorbed. Because of these reasons the bioavailability of iron varies greatly between different foods and is for example high in meat products but low in vegetables, eggs and legumes, mainly due to phytates, which is a molecule found in plants that is used in energy storage. The molecule consists of inositol and phosphate and leads to the formation of an oxidized complex together with iron (see Figure 2) [9]. Phytates can be broken down by food processing and it is rare that they cause problems for people who eat a balanced diet. It is also hard to predict mineral bioavailability in products only by using the phytate content [10]. Ascorbic acid (Vitamin C) is the most significant enhancer of iron bioavailability due to its capacity to reduce and chelate iron [9]. Ascorbic acid facilitates the iron absorption with chelated iron at low pH, which makes the molecule soluble in the duodenum [11].
Figure 2. The structure to the left is an example of Fe-porphyrin subunit in heme [12]. The structure to the right
1.2.1.1 Dietary recommendations for iron
Gender, age, country, pregnancy, lactation, and iron bioavailability are factors that affect the recommended daily intake (RDI). WHO has developed guidelines, and females of childbearing age are recommended an intake of 20 mg/day and other adults 7.5 mg/day [9]. The recommendations from NNR (Nordic Nutrition Recommendations) are 15 mg/day for females in childbearing age and 9 mg/day for other adults [7]. Organic pan-fried eggs are an example of a food rich in iron, and they contain 2.1 mg of iron per 100 g. One egg (one portion) is 51 g and contains 1.1 mg iron [14].
1.2.1.2 Iron and food processing
Food processing can change iron bioavailability. Cooking reduces the levels of haem iron as high temperatures break down the porphyrin ring. In this way, haem iron is partially converted to non-haem iron. Processing can also affect bioavailability by the degradation of enhancers, such av Vitamin C. Fortification of foods with other macronutrients may also change iron bioavailability. Single meal studies have demonstrated a decrease in iron absorption when co-ingested with either calcium or zinc. Although, the overall effects of these interaction in a freely choosen diet is unclear [9]. Pork contains lower levels of iron than beef and higher iron content is found in whole pieces of meat such as roast beef [14].
1.2.2 Calcium
Calcium is a micronutrient found in most food, but the highest amounts are found in milk products, leaf vegetables and nuts. The human body consists of about 1-2% calcium and 99% of all calcium in the body is found in bones and teeth. Anyone who consumes milk products usually has no problem getting enough calcium. Vegans, on the other hand, need to find different ways to include enough calcium in their diets [15]. In the diet, calcium occurs in many forms. Some examples are calcium-phosphate complexes, milk protein (casein) and calcium phytate salt complexes. Meat protein and sodium can significantly alter calcium status, as they contribute to increased levels of calcium in the urine. It is unclear how protein affects calcium status. Sodium and calcium are proposed to compete for kidney reabsorption by usage of the same transporters. In populations that eat less meat protein and sodium, it may also be expected that they need lower levels of calcium in the diet [9].
1.2.2.1 Dietary recommendations for calcium
Calcium requirements depends on life stage. The WHO recommend adult females and males an intake of 750 mg/day. The recommended intake for adolescents is 1000 mg/day. Elderly men, pregnant woman and postmenopausal woman are recommended 800 mg/day [9]. The recommendations from NNR are 800 mg/day for adults [16]. Northern European countries are presumably the only countries with an intake that is high enough, which is due to a high consumption of milk. Countries in Africa and South America have the lowest calcium intakes: between 400 and 700 mg/day. Also, many countries in Asia have a very low intake [17]. Consequences of deficiency are for example tremors, rickets, and convulsions in newborn babies [9]. Milk is an example of a food rich in calcium, and it contains 122 mg of calcium per 100 g. One glass is equivalent to 44 g of milk, and thus contain 54 mg calcium [18].
1.2.2.2 Calcium and food processing
1.2.3 Zinc
Zinc is a mineral whose main function is as cofactor to about 100 enzymes in the body. The enzymes affect the metabolism of protein, carbohydrates, fats, nucleic acids and certain vitamins. Zinc is also needed for the immune system. Meat, dairy products, whole grains and intestinal foods are good sources of zinc. The uptake of zinc is facilitated by meat proteins [19]. Phytate can, as with calcium and iron, form complexes with zinc which inhibit absorption [13].
1.2.3.1 Dietary recommendations for zinc
WHO has a recommended daily intake of 2-3 mg/day [9]. The recommendations from NNR are 7 mg/day for females and 9 mg/day for men [19]. It is estimated that zinc deficiency affects one third of the world’s population, but severe zinc deficiency is rare [20]. Consequences of deficiency are for example night blindness, growth retardation and immune deficiency [9]. Cheese (17% fat) is an example of a food rich in zinc, and it contains 3.8 mg of zinc per 100 g. Two slices can be seen as a portion and is 30 g, which means that it contains 1.1 mg zinc [21]. 1.2.4 Determination of mineral concentrations
The total iron, calcium and zinc content in food can be determined by atomic absorption spectrometer with flame atomization (FAAS). Before FAAS can be used, the sample needs to be prepared. Sample preparation can be done in the form of dry ashing. A muffle furnace is common to use and it helps the sample to dry, volatile materials to evaporate and organic materials to be destroyed [22]. The remaining ash then needs to be in a solution to enable analysis with FAAS. The solution is made of diluted nitric acid, which counteracts precipitation by transferring the metals into nitrate salts, which are highly soluble. The nitric acid also increases the efficiency of the atomization process, which is described in the following paragraph [23].
FAAS is based on ions atomizing as they pass through a gas flame. A high temperature in the flame allows electrons in it to connect with the ions to form atoms, and the flame is in that way reducing. The gas composition affects how effective the reducing flame is. The instrument also has a lamp, which is specific for the metal being analyzed (a hollow cathode lamp). The light from the lamp goes through the flame, which allows the metal atoms to absorb some of the light. The wavelengths of the absorbed light are characteristic for the metal, and it is produced by the hollow cathode lamp since it contains the same metal as being measured. A spectrometer measures how much light that is absorbed, and thus the content of metal atoms in the solution can be calculated [24]. FAAS is widely used because it is well established and validated, and it has a comparatively low cost and is time efficient [25]. Inductively coupled plasma mass spectrometry (ICP-MS) is another technique that can be used. It can measure several metals at the same time as well as lower levels. However, the instrument is more expensive [26].
Figure 3. The steps to obtain analyte (M) atoms from a sample containing the salt (MX). Excited state is marked
with (*) [25].
The most common flame to use when analyzing iron, zinc and calcium is based on a mixture of air and acetylene. The fuel and oxidant are mixed in a chamber before combustion in the burner. In the premix chamber, the large droplets of nebulized sample are allowed to condense and drain whilst the finer droplets can be mixed with the air and acetylene. The mixture is then transferred to the burner head. Combustion with a premix burner is preferable because of the less noisy signals produced. The premixture also makes the flame stable and allows a temperature from 2100-2400°C. The flame consists of the zones: the outer zone, the interconal zone, and the inner zone. The interconal zone is the hottest part and it is where the most analytical measurements are made. For each element, a hollow cathode lamp containing the specific metal being analyzed has to be used to visualize the absorption. By exciting the metal in the lamp, light is obtained with exactly the right wavelength that can be absorbed by the metal atoms in the flame [25].
1.2.4.1 Calculation of mineral concentrations
The concentration of iron, zinc or calcium in a solution can be calculated according to Eq. (1). The equation is obtained from the standard curve for the specific mineral by analyzing solutions with a known concentration of the mineral. Y is the absorbance, k is the response factor, x is the concentration in ppm, and m is the intercept on the y-axis. In order to obtain the final result, expressed as unit mg mineral/100g wet weight, the amount of sample, the volume of the solution, and dilution factor are considered (cf. Appendix 1 for an example.
𝑌 = 𝑘 ∙ 𝑥 + 𝑚 (1) 1.2.5 Limitations mineral analysis
products. Therefore, it was included as a parameter to investigate how the selected products counteract deficiency. The selected minerals were also advantageous to study since they could be analyzed in the same sample that had undergone a single sample preparation, which means that one method could be used for analysis of all minerals and the time needed was considered appropriate for the size of the project. Other minerals, such as iodine, selenium, potassium and magnesium, were excluded as this work highlights differences and similarities between meat products and plant-based substitutes. The intakes of the unanalyzed minerals are not perceived as diet-specific as iron, zinc, and calcium, since and the minerals excluded in this thesis can be consumed in several different types of diets [9]. Still, they and other nutritional parameters would have been interesting to include to get an overall picture.
1.3 Proteins and how they are analyzed
Protein consists of 20 different amino acids and out of these there are nine that an adult body cannot make (Table I). These are called the essential amino acids and it is vital to absorb them from food. Amino acids are bound together by peptide bonds to create peptides or proteins (a chain of more than fifty amino acids). An important role of proteins is to provide building material for synthesis of muscle and other tissues. But proteins are also involved in a variety other biological processes, such as in transport of nutrients, cell signaling and energy transformations. The proteins in food contribute with flavor and texture, but it can also have negative effects. For example, allergens are proteins that can be very harmful to some people and certain food enzymes might accelerate degradation processes. But enzymes can also be useful and may have a technical function during processing or storage [27].
Table I. The estimated daily requirements for the essential amino acids in adult humans [33].
Amino acid mg per 70 kg body weight
Histidine 700 Isoleucine 1400 Leucine 2730 Lysine 2100 Methionine + Cysteine 1050 Phenylalanine + Tyrosine 1750 Threonine 1050 Tryptophan 280 Valine 1820
Protein quality was chosen to be studied to reveal differences between amino acid intake in different types of products. Amino acid composition was analyzed by ALS as it was not possible to do it at the university during the project timeframe.
1.3.1 Meat protein
Meat has been consumed by humans as a protein source all throughout the history and protein from meats are nutritionally more complete compared to plant sources. Animal protein sources, such as meat, fish, dairy, poultry, and eggs, contain all the essential amino acids in a good mixture. That means that the amounts of the different amino acids are matching with the needs of the human body. The protein that is consumed through meat comes from the muscle of the animal, in which 60% are myofibrillar proteins [27]. Globally, pork is the most common type of meat that is eaten, followed by poultry and beef [34].
1.3.2 Plant protein
Avoiding meat protein goes far back in time. The first known group to abstain from meat were the Pythagoreans, followers of the Greek philosopher Pythagoras, in the 6th century B.C. Also,
Buddhist leaders have long advocated a vegetarian diet, which was based on the fact that the animals would avoid suffering. The first Buddhist leader to be a vegetarian was the Indian Emperor Ashoka (304 B.C.) [35]. Even till this day, many Buddhists eat a vegetarian or vegan diet [36]. Today, there is a great interest in plant-based food among many different populations and it continues to increase [15]. Common plant sources of protein used in meat substitutes are different types of lentils, soy products, beans, and peas [15]. Their nutritional composition is influenced by, for example, growth conditions, processing and refinement [27]. Methionine is the most common limiting amino acid in legumes and vegetables [31].
The following sources are the main plant sources of protein in the vegan products analyzed in this thesis and examples of amino acid compositions can be seen in Table II:
• Chickpeas are a type of legume. They are rich in protein, fiber, several key vitamins and minerals [39]. Cooked chickpeas contain approximately 9 g of protein per 100 g. The product is a good substitute for animal-based protein and is used in for example meat substitutes, dairy-free yoghurt and plant-based beverages [40]. It is not a listed allergen [41].
• The majority of pea protein used in food is extracted from yellow peas. Yellow peas are a type of legume with a protein that has a neutral taste, and is therefore used widely. The protein quality is not as high as meat protein as pea protein have lower contents of the essential amino acids, but peas also contain significant quantities of minerals, vitamins, fiber and carbohydrates. Pea protein can for example be used in dairy alternatives, meat substitutes and meal-replacement shakes. It is not a listed as an allergen [42]. Peas contain approximately 8 g protein per 100 g [43]. An industrial pea protein concentrate contains approximately 71% protein [44].
Table II. Amino acid composition in plant sources proteins according to literature examples (mg/100 g) [45].
Protein His Ile Leu Lys SAA AAA Thr Val
Soy, 92% 2500 4120 6620 5450 1960 7360 3680 4120
Chickpea,
dried 560 920 1540 1410 550 1170 790 920
Pea, dried 480 930 1480 1620 450 1450 860 1000
1.3.3 HPLC with fluorescence detection for amino acid profile
For amino acid profile analysis, free amino acids can be determined by high performance liquid chromatography (HPLC) with fluorescens light detection (FLD). Strongly acidic conditions are used to hydrolyze the proteins and obtain free amino acids. HPLC can then be used and is a form of separation of different molecular species from a mixture. The sample consists of analytes and matrix, of which it is the analytes that are the molecules of interest. In a chromatographic separation, the sample is injected into a flowing mobile phase which passes a stationary phase. The stationary phase is found in a column which is packed with small, porous particles between 1-5 µm in diameter. The particles are usually made of silica and the stationary phase is found on the particles. The stationary phase helps separating analytes with different characteristics that interact with the stationary phase. The liquid mobile phase moves through the column with the help of a pump. The delay, due to the interaction with the stationary phase in the column, causes different types of molecules to leave it at separate times, which enable detection of specific analytes (Figure 4) [46].
Figure 4. The separation process in a chromatography. Black and white stars indicate different types of molecules
[46].
Fluorescence detectors are based on emission of light by a molecule after absorbing an initial radiation (excitation light). A molecule can go from a ground state, which is a lower energetic state, to an excited state. The change in state is caused when the molecule absorbs energy from a light source, such as a laser. When the molecule bounces back to the initial state and emits a photon, the fluorescence radiation has a lower frequency than excitation radiation (Figure 5). The excited state of a molecule lasts for about 10-8 seconds, if not disturbed by collisions. The
quantitation of the fluorescence species is determined by measuring the fluorescence intensity using fluorescence light detectors. They generate an electrical signal of intensity which is linear with the concentration of a specific molecule [46].
Figure 5. The change in electronic transitions during fluorescence which enable fluorescence detection [46].
The analysis of amino acids by this method can for example be performed at the analysis company ALS. They have the reporting limit 0.02 g/100 g. The measurement uncertainty for each amino acid can be seen in Appendix 2 [49].
1.3.4 Evaluation of protein quality
The Food and Agricultural Organization (FAO) developed the method named Digestible Indispensable Amino Acid Score (DIAAS) in 2013 to evaluate protein quality. The method replaces Protein Digestible Amino Acid Score (PDCAAS) from 1991. A protein’s nutritional value is based on its composition and digestibility. DIAAS is preferred to be used as it reflects the true nutritional value of dietary protein for humans better than PDCAAS. It is also more accurate in assessment of protein quality in a mixture of different dietary protein sources. PDCAAS and DIAAS are very much alike, but differ in the way that PDCAAS includes the true fecal protein digestibility and DIAAS includes the true ileal protein digestibility [32]. The protein digestibility includes digestion in the small intestine and stomach, absorption of peptides, absorption of free amino acids, metabolism, in situ protein synthesis, and efflux back to the intestinal lumen and to the portal blood. As mentioned above, DIAAS uses the value of the true ileal protein digestibility [27]. It is determined at the end of the small intestine. The true ileal digestibility amino acids in specific protein sources has been compiled in Table III. The true ileal digestibility is preferably determined in humans. Growing pigs can be used if it is not possible to determine in humans. Growing rats can be used if it is not possible to determine in pigs [32].
Table III. True ileal digestibility for amino acids in different protein sources. XP indicates that the determination was made in pigs, XH in human and XR in rat [52-54]. The protein sources contain more than 4,4 g protein per 100g of food.
Protein source Ileal digestibility in amino acids (%)
His Ile Leu Lys Met Phe Thr Val
BeefP 95 94 99 99 99 99 96 98 PorkP 93 91 91 93 95 89 86 89 ChickpeaR 87 82 88 90 94 90 80 83 PeaH 81 77 77 81 76 76 74 74 Pea concentrateR 99 99 98 99 98 99 97 97 SoyH 97 97 97 98 96 97 97 97
To determine DIAAS, there are recommended scoring patterns for amino acids. The scoring patterns for young children can be seen in Table IV. The pattern is recommended by FAO to use in protein quality calculations for all foods and population groups, except for infant formulas [32].
Table IV. Reference scoring patterns for protein requirements with the unit mg/g protein. SAA are the Sulphur
Amino Acids and AAA are the Aromatic Amino Acids [32].
Age group His Ile Leu Lys SAA AAA Thr Val
- Child (6 months to 3 years) - 20 - 32 - 66 - 57 - 27 - 52 - 31 - 43
DIAAS is defined in equation (2). The “Digestible IAA reference ratio”, expressed in equation (2), is for the amino acid with the lowest calculated quantity.
Protein quality is determined according to Table V. The score “No protein quality claim” is interpreted as suboptimal, whilst “Good protein quality” is considered good but does not provide an optimal supply of essential amino acids. The score “High protein quality” is the optimal alternative [32].
Table V. How the DIAAS Score is used to determine protein quality [32].
DIAAS Score Quality
DIAAS < 75% No protein quality claim
DIAAS ³ 75 % - < 100 % Good protein quality
DIAAS ³ 100 % High protein quality
1.4 The principle for example meals and example days
An example meal or day can be included in a study to highlight how the product is consumed. By including other foods and thus creating a meal, not only the individual product is studied but also how nutritional quality from different products together contribute to reaching the recommended daily intake. In this way, it is made visible which aspects of the main products that can be problematic even when the product is a part of a meal or a day’s intake [9].
1.4.1 Example meals
An example meal includes the product being analyzed and other foods with which it is usually combined. For example, a meatball and its meat-free alternatives can be matched with potatoes, lingonberry jam, cream sauce, and green peas. This makes the meal a mixed meal with both plant-based and animal-based products. The common ingredients’ content in terms of iron, zinc, calcium, and essential amino acids is presented in Table VI.
Table VI. Iron, zinc, calcium and essential amino acid content in mg per 100 g product in boiled potatoes, lingonberry jam, cream sauce and green peas [14, 45, 55].
Food Iron Zinc Calcium His Ile Leu Lys Met Cys Phe Tyr Thr Val
- Boiled potatoes - 0.4 0.2 - 4.0 - 8 - 58 - 84 - 90 - 21 - 13 - 63 - 29 - 53 - 93 - - Lingonberry sauce - 0.2 - 0.1 - 9.9 - 0.66^ - 0.20^ - 0.30^ - 0.30^ - 0.09^ - -^ - 0.38^ - 1.25^ - 0.76^ - 1.00^ - Cream sauce - 0.0 - 0.2 - 47.7 - 61* - 120* - 205* - 180* - 54* - 17* - 110* - 135* - 99* - 150* - Green peas 1.3 - 0.7 - 21.7 - 110 - 195 - 320 - 310 - 82 - 32 - 200 - 115 - 205 - 230 -
* means that the value is for cream (not cream sauce). ^ means the value is for lingonberry (not lingonberry jam)
A meal, such as lunch or dinner, should preferably contain 25-35 % of the daily energy intake [56]. In a standard meal, 120 g of potatoes, 128 g of meatball/substitute, 35 g of lingonberry sauce, 70 g of sauce, and 40 g of green peas is included and it is these quantities that are used in compiling the example meal in this thesis [57]. The amount of a certain parameter in an example meal is calculated according to Eq. (3). QM is the quantity (mg) of iron, zinc, calcium
or an essential amino acid in a meal. QP is the amount (mg) of the specific parameter in 100cg
is in green peas. The amount of certain minerals in example meals or example days can also be calculated with the program Dietist Net.
𝑄! = 𝑄"∙ 1.2 + 𝑄#∙ 1.28 + 𝑄$∙ 0.35 + 𝑄%∙ 0.70 + 𝑄& ∙ 0.40 (3)
1.4.2 Example days
Two example days can be included to visualize different types of diets and how the contribute to reach the recommended intakes. Examples of diets to include are one mixed diet with animal-based products and one plant-animal-based diet (see Table VII). Unless otherwise stated, water was the drink consumed with the meals.
Table VII. The products included in the different meals in the example days.
Example day with animal
ingredients
Breakfast Oatmeal porridge, whole grains 200g
Apple 125g Milk, 1.5% fat 200g Coffee 150g Lunch Meatball/substitute 128g Boiled potato 120g Cream sauce 70g Lingonberry jam 35g Green peas 40g
Snack Bread, levain 100g
Butter, 80% fat 10g
Cheese, 26% fat 30g
Red paprika 30g
Dinner Homemade lasagna 250g
Example day with plant-based
ingredients
Breakfast Oatmeal porridge, whole grains 200g
Apple 125g Oat drink 200g Coffee 150g Lunch Meatball/substitute 128g Boiled potato 120g Oat cream 70g Lingonberry jam 35g Green peas 40g
Snack Bread, levain 100g
Oat spread 30g
Red paprika 30g
Dinner Bolognese on soy 120g
Pasta 150g
1.5 Dietary recommendations
2 Aim
The aim of this degree project is to study nutritional similarities and differences between products that are replaceable with each other. This is carried out with the goal to strengthen or disprove the general beliefs that exist in society about different types of plant-based and animal-based products. To analyze this, plant-animal-based and animal-animal-based products are compared with each other in terms of protein quality, zinc content, iron content and calcium content. The comparison is made between the products themselves, between the products as part of an example meal, and as part of example days. The ambition is to, with the help of the different analyzes, create a discussion on how the different products contribute to the goal of reaching daily recommended intakes and to see if the plant-based products are good substitutes to the meat product.
3 Method and materials
The following methods are the standard methods used to study iron, zinc, and calcium content in food and for the determination of protein quality.
3.1 Quantitative determination of iron, zinc and calcium with FAAS
Two samples of each product were included to obtain more reliable results. The pre-treatment included cutting the samples and then drying them in foil molds. One sample of a product was one ball (from 10.81 to 16.41 g) and the exact weight of each product was noted. The drying was done for 7 hours in a heating cabinet which had the temperature 80 ° C.
The standard solutions were made from 1000 ppm iron, zinc, and calcium solutions with 0.1 mol/l nitric acid. The standard solutions used to analyze iron were of the concentrations 0 ppm, 1 ppm, 2 ppm, and 4 ppm. The standard solutions used to analyze zinc were of the concentrations 0 ppm, 0.2 ppm, 0.4 ppm, 0.6 ppm and 0.8 ppm. The standard solutions used to analyze calcium were of the concentrations 0 ppm, 1 ppm, 2 ppm and 4 ppm. Lanthanum solutions (5%) were included in the calcium standards.
The FAAS used was PerkinElmer AAnalyst 400. The wavelength 248.33 nm was used for iron, 213.86 nm was used for zinc and 422.67 nm was used for calcium. The iron, zinc and calcium concentrations in the liquid samples were calculated according to Eq. (1). The weighed amount and dilution factor were used to calculate the concentration in the original sample (wet weight). 3.2 Determination of protein quality, example meals and example days
To determine the amino acid composition, samples of the four products were sent to the analysis company ALS in Danderyd, Sweden. The samples were sent in a cooler bag containing freezer packs and arrived at ALS 24 h after they were sent. Only one sample of each product was analyzed as a result of the automated manufacturing process assumed to produce uniform results. The method used by ALS to analyze amino acid profile was HPLC with fluorescence detector. When the result with amino acid composition was obtained, DIAAS could be calculated according to Eq. (2), see Appendix 3.
4 Results
4.1 Analysis of mineral content
SoyB, GreenB and PeaB had similar iron mean concentrations (0.9-1.1 mg/100 g). The MeatB had the lowest iron content with a concentration of 0.6 mg/100 g. The product with the highest zinc concentration was the MeatB (1.9 mg/100 g). GreenB and PeaB had once again similar concentrations, approximately 1 mg/100 g. The SoyB had the lowest zinc content with a concentration of 0.5 mg/100 g. The product with the highest calcium concentration was SoyB (73.6 mg/100 g). GreenB had a calcium concentration of 37.9 mg/100 g, whilst PeaB had a value of 15.7 mg/100 g. The MeatB had the lowest calcium content with a concentration of 9.9 mg/100 g. See Table VIII for the concentrations obtained in each sample.
Table VIII. The iron, zinc and calcium concentrations for each sample with the mean and standard derivation for
each product.
Sample Iron conc.
4.2 Analysis of protein quality
The MeatB was the only product with high protein quality, whilst the other products had good quality (see Table IX). The GreenB has a DIAAS of 76%, PeaB had 75%, SoyB had 91% and MeatB 101%. The limiting amino acid for MeatB was leucin and the sulfur-containing amino acids (cysteine and methionine) were limiting for GreenB, PeaB and SoyB.
Table IX. The limiting amino acid, DIAAS and protein quality for the products analyzed.
Product Limiting aa DIAAS (%) Quality
MeatB Leu 101 High
GreenB SAA 76 Good
PeaB SAA 75 Good
SoyB SAA 91 Good
4.3 Analysis of example meals and example days
All the varieties of meals had contents of essential amino acids that was higher than the recommended intake for the meal, whilst none of the meals reached the recommended intake of iron or calcium (Table X). The meals with MeatB, GreenB and PeaB reached the recommended intake for zinc and the meal with SoyB had the lowest zinc content (1.4 mg). However, the meal in which SoyB was included had the highest intake of iron (2.4 mg) and calcium (144.6 mg). The meal with MeatB had the lowest intakes of these minerals, with 1.8 mg of iron and 62.9 mg of calcium.
Table X. Intake of iron, zinc, calcium and the essential amino acids (mg) in an example meal consisting of
potatoes, meatballs or substitute, lingonberries, cream sauce and green peas. The intakes of iron, calcium and zinc are to compare with 25% of NNR’s RDI (mg), whilst the RDI for the amino acids are from WHO.
Meal
including Iron Zinc Calcium His Ile Leu Lys SAA AAA Thr Val
MeatB 1.8 3.2 62.9 673 934 1440 1589 617 1444 916 1081 GreenB 2.1 2.0 98.9 276 624 990 959 338 1084 616 741 PeaB 2.2 1.9 70.4 336 734 1150 1119 367 1284 676 840 SoyB 2.4 1.4 144.6 516 1064 1600 1479 557 1764 996 1111 --- Recommended intake (meal) 3.8 1.8 200.0 175 350 683 525 263 438 210 455
As seen in Table XI, in which different versions of an whole day diet with mixed food are presented, all the examples reached the recommended daily intake for zinc and calcium. None of the example days reached the RDI for iron, but the day with SoyB had the highest content with 6.1 mg. The day with MeatB had the lowest iron intake (5.5 mg).
Table XI. Intake of iron, zinc, calcium (mg) in an example day with mixed food (both plant and animal-based). The intakes are to compare with the recommended daily intakes made by NNR (mg).
Day including Iron Zinc Calcium
In the example days with a plant-based diet (Table XII), none of the recommended daily intakes were reached.
Table XII. Intake of iron, zinc, calcium (mg) in an example day with plant-based food. The intakes are to compare with the recommended daily intakes made by NNR (mg).
Day including Iron Zinc Calcium
MeatB 7.6 6.5 422.0 GreenB 8.0 5.3 457.9 PeaB 8.0 5.3 429.4 SoyB 8.2 4.7 503.6 Recommended intake (day) - 15.0 - 7.0 - 800.0
5 Discussion
5.1 Protein qualityMeatB was the only one of the four analyzed products that had a high protein quality [32]. DIAAS was over 100%, which means that it provides an optimal supply of essential amino acids [32]. This makes it a good source of complete protein that the body can use efficiently. MeatB does not need to be mixed with different protein sources to get a good amount of the essential amino acids, but contributes itself with sufficient amounts, as expected. Leucine was the limiting amino acid, but it is not relevant since the supply of essential amino acids was sufficient.
GreenB had a good protein quality [32] with a DIAAS of 76%. Also, PeaB had a good protein quality, but with a DIAAS of 75%. Their limiting amino acid was SAA (cysteine and methionine). It was expected that SAA would be limiting as methionine is the most common limiting amino acid in vegetables [31]. The fact that the protein quality is good (not high) means, according to FAO [32], that the product is decent from an amino acid perspective, but that it does not contribute to an optimal supply of essential amino acids. For GreenB to have a high protein quality, not only increased content of SAA would be required, but also leucine (Table IX) as the leucine level was 95% of what is needed for an optimal supply of essential amino acids. For PeaB to have a high protein quality, only an increased content of SAA would be required. A way to increase the content of leucine, methionine and cysteine in the products is to use dried pea or chickpea as it contains high levels of amino acids (see Table II). For example, dried pea or chickpea could have been included in the form of a powder or flour which could possibly also contribute to the thickening of the product. In this way, a proportion of the thickener could be replaced and a higher protein quality could be achieved. SoyB had good protein quality with a DIAAS of 91%. The limiting amino acid was once again SAA and it was expected as methionine [31] is the most common limiting amino acid in legumes [31]. For SoyB to have a high protein quality, only an increased content of SAA would be required. To increase the content of methionine and cysteine in the product, dried soy beans or higher amounts of soy protein could have been included.
is difficult to obtain sufficient amounts. Although all the essential amino acids are present in the products, the challenge seems to be to obtain them in high enough quantities. All plant-based alternatives would benefit from having a higher protein content, as it allows for a higher protein quality to be achieved. Also, by varying protein sources, a better amino acid composition could be obtained. For example, protein from peas and cereals could be mixed so that a good amino acid composition is achieved by the complementary properties of the foods as cereals and peas are rich in various amino acids [28]. Cereals can in this way contribute with methionine and cystine, while peas can contribute with leucine [45]. Of the plant-based products, PeaB and SoyB do contain cereals, but as an optimal protein quality was not reached the quantities were not sufficient. However, a change in protein content or protein source might affect the cost, taste and texture of a product.
Limitations with DIAAS are that data for all protein sources are not yet available. The method is relatively new [32] and all the values that can be needed for protein quality calculations are not available. This means that there is a risk that the results obtained differ to some extent from reality. When DIAAS was calculated for MeatB, GreenB, PeaB and SoyB, the digestibility was used for the main protein sources of the products. This means that the digestibility of protein sources of which there was less in the products was not included, which could possibly affect the result. It is still preferable to use DIAAS compared to PDCAAS as DIAAS uses true ileal protein digestibility, which is considered more accurate. DIAAS also allows one to calculate protein quality for a mixture of different protein sources and to include the bioavailability of amino acids.
5.2 Zinc
MeatB had the highest content of zinc; 1.93 mg/100 g. This is a consequence of meat products being good sources of zinc [19], and the result was therefore expected. However, the zinc content is lower than in for example 100g of cheese (3.8 mg/100 g) [21], which is seen as a product rich in zinc. But it is important to remember that 100g of cheese is not consumed at once like MeatB is, and that MeatB contributes to a higher intake of zinc in a portion perspective. The large intake of MeatB therefore makes it a good source of zinc. WHO has a recommended daily intake of 2-3 mg/day [9], which means that the consumption of MeatB would help achieve a large part of RDI. However, NNR has a recommendation of 7 mg/day for women and 9 mg/day for men [19]. An intake of 1.93mg/100 g contributes to a big part of RDI also when NNR's RDI is followed.
or several different sources. It is also conceivable that for zinc content it does matter what type of soy product that is included.
The fact that the zinc content is lower in GreenB, PeaB and SoyB than in MeatB can become problematic as the body's zinc uptake is stimulated by meat proteins [19]. This means that lower levels than the ones originally found in GreenB, PeaB and SoyB can be used by the body. In the plant-based products, the zinc content that the body can use may also be lower than the actual content due to phytates [13]. However, it does not appear that this is a widely-spread problem at the moment as zinc deficiency is uncommon, especially in the Nordic countries [20]. But it may be an aspect that needs to be kept in mind and studied further as a plant-based diet becomes more common. Still, can be assumed that phytates do not affect zinc intake significantly when a balanced diet is consumed and to some extent due to the food processing. Even if the phytate content in the products had been known, it would have been difficult to predict how much it would affect the body's absorption of the mineral.
5.3 Calcium
The calcium content in MeatB was 9.85 mg/100 g. MeatB had the lowest amounts of iron and calcium of the four products. The fact that its calcium content was lower than the other products can be explained by MeatB mainly containing of beef and pork, ingredients who are known to not be very rich in calcium [15]. An example of a product that is rich in calcium is milk [18] in which one glass contains more than five times as much as a portion of MeatB. The daily recommended intake of calcium is 800 mg/day according to NNR [16] and 750 mg/day according to WHO [9]. MeatB contributes with a small amount of calcium in comparison with how high the intake should be in a day, and it is required that the product is accompanied with other products rich in calcium during the day.
that phytates do not affect calcium intake significantly when a balanced diet is consumed and to some extent due to the food processing [9, 58].
5.4 Iron
The iron content of MeatB was 0.64 mg/100 g. An example of a product that is rich in iron is egg [14] which contains higher amounts than the ones in MeatB, both seen to portions and mg/100 g. The daily recommended intake of iron for adults is 9 mg/day according to NNR [7] and 7.5 mg/day according to WHO [9], but is higher for females of child bearing age. MeatB contributes with a relatively low amount of iron in comparison with how high the intake should be in a day, and it is required that the product is accompanied with other products during the day that are rich in iron. The fact that MeatB had the lowest content of iron was unexpected as meat products are considered to be rich in iron [7]. However, the low content may depend on the types of meat used and how it was processed. The types of meat found in the product are beef and pork in the form of mince. Pork contains lower levels of iron than beef [14 ]and the fact that mince is used can also be a factor. Higher iron content is found in whole pieces of beef such as roast beef [14]. Meat containing more blood may also contain higher amounts of iron [14]. The general perception that meat contains a lot of iron may be the result of some meat products having higher amounts, but that is not true for all meat products – which this thesis shows. That meat is a good source of haem-iron [9] can also contribute to the perception that all meat products are good sources of iron generally.
The iron content in GreenB and PeaB was 0.88 mg/100 g and 0.93 mg/100 g, respectively. The content of iron in GreenB can be explained by the fact that it contains legumes such as peas and chickpeas, which are seen as relatively rich in iron [7]. The content of iron in PeaB can be explained by the pea protein [7]. Compared to a portion of eggs [14], both GreenB and PeaB contain less iron, but the difference is not as great as for MeatB. GreenB and PeaB contributes with a relatively low amount of iron in comparison with the RDI [7, 9], and it is required that the products are accompanied with other products during the day that are rich in iron. SoyB had the highest iron content of the four products; 1.11 mg/100 g. The content of iron in SoyB can be explained by it containing soy protein [7], which is rich in iron. A portion of SoyB contained the same amount of iron as a portion of organic eggs did [14], but when seen to mg/100 g, the SoyB contained less. That means that it is a good source of iron but it is not necessarily rich in iron and that it needs to be supplemented with other iron-rich foods to reach the RDI.
and to some extent due to the food processing. The products rich in Vitamin C can also increase the absorption of iron, as the vitamin enhances it [9].
5.5 Example meals
In all example meals (Table X), the amounts of essential amino acids reached the recommended intake for a meal and the recommended intake for zinc was met by MeatB, GreenB and PeaB. The fact that all meals reached the recommended intake of essential amino acids [33] was unexpected as MeatB had a high protein quality, but the plant-based products only a good protein quality. The differences in protein quality for the products becomes very small when they are combined with other food in a meal, the plant-based products are shown to be good contributors with essential amino acids as the recommended intakes are reached. However, neither the recommended iron nor calcium intake [7, 16] was reached for any of the meals. But by drinking a glass of milk, or fortified plant-based milk, with the meal or including a salad with spinach, the meal could become complete also with regard to iron and calcium. The example meals visualize that the meal composition is of importance. If one would have looked only at the products themselves, the recommended intakes for several parameters would have been difficult to reach. But when the products are put into the context of a complete diet, it becomes clear that all the products studied play a good part in reaching recommended intakes for especially the essential amino acids and to a certain level zinc.
Table X uses NNR’s recommendations for iron, calcium and zinc [7, 16, 19] to calculate the recommended intake of a meal, but if the recommendations from WHO [9] would have been used all the products would have met the recommendations for zinc. It is positive that the recommended intake for zinc is reached from a global perspective. Still, the intake of calcium and iron is not optimal for the example meal. The key lies in varying the diet and in consuming several different sources of nutrients over a meal, day, and week. In this way, the analyzed products can form a good basis that allows all RDIs to be reached. The strength of the example meal is that it provides a clear picture of which RDIs can be difficult to reach even when the specific products are part of a meal. In the analysis, it would have been interesting to compare the content of the minerals and essential amino acids with the minimum requirements to see if the content may be sufficient from that point of view.
Enriching (fortifying) the products with the minerals that are insufficient in relation to the recommended intake is an alternative, and calling a product rich in calcium or iron would make it competitive. However, fortification of a product is not welcomed by everyone and consumers might appreciate a “natural” product over a fortified. The priority for the products is therefore that they should be replaceable with each other without fortification. The plant-based products are good substitutes for MeatB and their equal contents of nutrients underlines why they do not need to be enriched.
5.6 Example days
For the example days that included animal-based food, the recommended daily intake for zinc and calcium were reached by all products. Unfortunately, none of the them reached RDI for iron. In the example days where the common meals were plant-based, RDI was not reached for any of the minerals. It again shows how similar the four products are and that they are good substitutes for each other. Seen over an entire day, none of the products stand out in terms of enabling or preventing RDI from being reached more than the others. This again indicates that MeatB and the plant-based alternatives play a similar function in the role of contributing with iron, zinc and calcium. But what matters is the diet in general and that the example days with plant-based meals did not reach the recommended intake shows the good nutritional values of animal foods, even if the products studied contributes equally.
The example days showed that although the products can be good substitutes for each other, there is much more that matters than just a single product. It is the composition of the day’s diet that determines if RDIs are reached. It also shows that it is more important in some types of diets than others to think about what you eat. The example day with a plant-based diet shows that it can be a challenge to reach RDI in terms of iron, zinc and calcium and one may need to have a bit of understanding of how to reach RDI for all nutrients if a completely plant-based diet is consumed. Even if the plant-based products analyzed are good substitutes to each other and to the meat product, it is showed by the example days that they are only as good as the products they are accompanied with. However, it is important to remember that dietary advice refers to intake over a longer period, which is at least one week.
5.7 Social relevance, ethics, and impact on the environment
6 Conclusions
This thesis shows that the differences between MeatB and the meat substitutes analyzed are small, making the plant-based alternatives good substitutes. There are clear differences when the products are compared to each other, but when placed in an example meal or example day these become less distinct. When the products are analyzed individually, MeatB has the highest protein quality and zinc content, while the plant-based alternatives have higher calcium and iron content. Although MeatB was the only product that had a high protein quality, the example meals including GreenB, PeaB and SoyB also reached the recommended intakes for essential amino acids. The products did have varying levels of different nutrients, but still they reached the recommended intakes for the same nutrients in the example days and to a certain extent in the example meals. This thesis shows that the animal product and the plant-based products play the same role when placed in a bigger perspective. The plant-based products are preferred over MeatB from an environmental and animal rights perspective. To study differences between MeatB and the plant-based products in terms of how much of the mineral contents the body can use, further studies are needed.
7 References
1. Nationalencyklopedin, Köttbullar.
2. Plant Based Food Association, Retail Sales Data 2018. 2019.
3. Gerber, P.J., et al., Tackling climate change through livestock - A global assessment of emissions and
mitigation opportunities. 2013, Food and Agriculture Organization of the United Nations (FAO): Rome.
4. Naturskyddsföreningen, Kött, klimat och miljö - dina val påverkar! Available from:
https://www.naturskyddsforeningen.se/kott-klimat-miljo (Access date: 2020-04-06)
5. Willett, W., e. al., Food in the Anthropocene: the EAT-Lancet Commission on healthy diets from
sustainable food systems. 2019: The Lancet. p. 447-492.
6. Livsmedelsverket, Matvanor och sjukdom. 2019; Available from:
https://www.livsmedelsverket.se/matvanor-halsa--miljo/sjukdomar-allergier-och-halsa/matvanor-sjukdom (Access date: 2020-04-06)
7. Livsmedelsverket, Järn. 2020; Available from:
https://www.livsmedelsverket.se/livsmedel-och-innehall/naringsamne/salt-och-mineraler1/jarn (Access date: 2020-04-06)
8. Vårdguiden, Brist på järn. 2019; Available from:
https://www.1177.se/sjukdomar--besvar/hjarta-och-blodkarl/blodsjukdomar/blodbrist-pa-grund-av-for-lite-jarn/ (Access date: 2020-04-08)
9. De la Guardia, M. Handbook of mineral elements in food. 2015, Wiley Blackwell: England
10. Lopez, H.W., et al., Minerals and phytic acid interactions: is it a real problem for human nutrition? 2002, International Journal of Food Science & Technology.
11. Lynch, S., Cook, J., Interaction of Vitamin C and iron. 1980, Ann N Y Acad Sci. 355: p. 32-44.
12. Wikipedia, Heme, in Structure of Fe-porphyrin subunit of heme B. 2020; Available from:
https://en.wikipedia.org/wiki/Heme (Access date: 2020-05-19)
13. FAO/IZiNCG, Global food composition database for phytate version 1.0 - PhyFoodComp1.0. 2018:
Italy.
14. Livsmedelsverket, Livsmedelsdatabasen. 2020; Available from: http://www7.slv.se/SokNaringsinnehall
(Access date: 2020-04-22)
15. Livsmedelsverket, Vegetarisk mat. 2020; Available from:
https://www.livsmedelsverket.se/matvanor-halsa--miljo/kostrad-och-matvanor/vegetarisk-mat-for-vuxna (Access date: 2020-04-10)
16. Livsmedelsverket, Kalcium. 2020; Available from:
https://www.livsmedelsverket.se/livsmedel-och-innehall/naringsamne/salt-och-mineraler1/kalcium (Access date: 2020-04-07)
17. Balk, E., et al., Global dietary calcium intake among adults: a systematic review. 2017, Osteoporosis
International. p. 3315-3324.
18. USDA, Milk, reduced fat, fluid, 2% milkfat, without added vitamin A and vitamin D. 2019; Available
from: https://fdc.nal.usda.gov/fdc-app.html#/food-details/172205/nutrients (Access date: 2020-05-09)
19. Livsmedelsverket, Zink. 2019; Available from:
https://www.livsmedelsverket.se/livsmedel-och-innehall/naringsamne/salt-och-mineraler1/zink (Access date: 2020-04-10)
20. WHO, The world health report. 2002.
21. USDA, Cheese, edam. 2019; Available from:
https://fdc.nal.usda.gov/fdc-app.html#/food-details/173419/nutrients (Access date: 2020-05-20)
22. Miller-Ihli, N., Techniques and Instrumentation in Analytical Chemistry, in Dry ashing. 1992.
23. Luque de Castro, M. and J. Luque García, Techniques and Instrumentation in Analytical Chemistry, in
Nitric acid. 2002.
24. Niedzielski, P., et al., Determination of Iron Species in Samples of Iron-Fortified Food. 2014. 7: p. 2023-2032.
25. Sanz-Medel, A., Atomic absorption spectrometry: an intoduction. 2014, Momentum Press, LLC: New
York.
26. Wilschefski, S.C., Baxter, M.R., Inductively Coupled Plasma Mass Spectrometry: Introduction to
Analytical Aspects. 2019: Clin Biochem Rev. p. 115-133.
27. Ustunol, Z., Applied Food Protein Chemistry. 2015, John Wiley & Sons.
28. Livsmedelsverket, Protein. 2020; Available from:
https://www.livsmedelsverket.se/livsmedel-och-innehall/naringsamne/protein (Access date: 2020-04-20)
29. Hoffman, J., Falvo, M., Protein - Which is best? 2004: J Sports Sci Med. 3(3): p. 118-130.
30. FAO, Protein quality assessment in follow-up formula for young children and ready to use therapeutic
foods. 2018: Rome.
31. ASN, Protein Complementation. 2011; Available from: https://nutrition.org/protein-complementation/
32. FAO, Dietary protein quality evaluation in human nutrition - Report of an FAO Expert Consultation. 2011: New Zealand.
33. WHO, Protein and amino acid requirements in human nutrition: report of a joint FAO/WHO/UNU
expert consultation. 2002: Geneva, Switzerland.
34. Our world in data, Meat production by animal. 2018. Available from:
https://ourworldindata.org/meat-production (Access date: 2020-04-19)
35. Hultin, G., The History of Vegetarian Diets: Explore the Progression of Plant-Based Eating. 2019,
Food & Nutrition. Available from: https://foodandnutrition.org/from-the-magazine/the-history-of-vegetarian-diets-explore-the-progression-of-plant-based-eating/ (Access date: 2020-05-13)
36. Wilson, C., Plant-based Populations. 2013, Food & Nutrition. Available from:
https://foodandnutrition.org/july-august-2013/plant-based-populations/ (Access date: 2020-05-13)
37. USDA, Soybeans, mature seeds, raw. 2019; Available from:
https://fdc.nal.usda.gov/fdc-app.html#/food-details/174270/nutrients (Access date: 2020-05-01)
38. USDA, Soy protein concentrate, produced by alcohol extraction. 2019; Available from:
https://fdc.nal.usda.gov/fdc-app.html#/food-details/172447/nutrients (Access date: 2020-05-01)
39. Ware, M., What are the benefits of chickpeas? 2019: Medical News Today.
40. Wallace, T., Murray, R., Zelman, K., The Nutritional Value and Health Benefits of Chickpeas and
Hummus. 2016: Nutrients. 8(12): p. 766.
41. Cabanillas, B., Jappe, U., Novak, N., Allergy to Peanut, Soybean, and Other Legumes: Recent Advances
in Allergen Characterization, Stability to Processing and IgE Cross-Reactivity. 2018, Molecular Nutrition & Food Research. 62(1).
42. Noswoethy, M., e. al., Effect of Processing on the in Vitro and in Vivo Protein Quality of Yellow and
Green Split Peas (Visum sativum). 2017, Journal of agricultural and food chemistry. 65(35): p. 7790-7796.
43. USDA, Green or yellow split peas, dry, cooked, fat not added in cooking. 2020; Available from:
https://fdc.nal.usda.gov/fdc-app.html#/food-details/784275/nutrients (Access date: 2020-05-03)
44. Buffo, R., Jung, H., Innovations in Food Packaging. 2005, Academic Press. p. 277-300.
45. Livsmedelsverket, Aminosyror i livsmedel. 2020; Available from:
https://www.livsmedelsverket.se/globalassets/livsmedel-
innehall/naringsamnen/livsmedelsdatabas/aminosyrorper-100g.pdf (Access date: 2020-05-04)
46. Moldoveanu, S., Essentials in modern HPLC separations. 2012, Elsevier: Amsterdam.
47. Nielsen, S.S., Food Analysis. 2012, Springer-Verlag New York Inc.
48. Perucho, J., et al., Optimal excitation and emission wavelengths to analyze amino acids and optimize
neurotransmitters quantification using precolumn OPA-derivatization by HPLC. 2015: Amino Acids.
47(5): p. 963-973.
49. ALS, Orderbekräftelse. 2020: Danderyd.
50. ALS. Om ALS. 2018; Available from: https://www.alsglobal.se. (Access date: 2020-04-07)
51. European Commission, Commission Regulation (EC) No 152/2009 of 27 January 2009 laying down the
methods of sampling and analysis for the official control of feed, European Union. 2009. 52. Gilani, S., et al., The assessment of amino acid digestibility in foods for humans and including a
collation of published ideal amino acid digestibility data fort human foods. 2012.
53. Hodgkinson, S., et al., Cooking Conditions Affect the True Ileal Digestible Amino Acid Content and
Digestible Indispensable Amino Acid Score (DIAAS) of Bovine Meat ad Determined in Pigs. 2018, The Journal of Nutrition. 148(10): p. 1564-1569.
54. Bailey, H., Berg, E., Stein, H., Pork Products Have Digestible Indispensable Amino Acid Scores
(DIAAS) That Are Greater Than 100 When Determined in Pigs, but Processing Does Not Always Increase DIAAS. 2020: The Journal of Nutrition. 150(3): p. 475-482.
55. Lee, J., Finn, C., Lingonberry (Vaccinium vitis-idaea L.) grown in the Pacific Northwest of North
America: Anthocyanin and free amino acid composition. 2012, Journal of Functional Foods. 4(1) p. 213-218.
56. Johansson, U., Näring och hälsa. 2014, Studentlitteratur: Lund.
57. Livsmedelsinfo, Omvandla vikter och volymer.
58. Nordiska ministerrådet, Nordiska näringsrekommendationer 2012 - Rekommendationer om näring och
8 Appendices
Appendix 1. Calculation minerals
Figure 4. The standard curve for iron with the equation of a straight line and R2 value. Absorption as a function
of concentration (ppm).
Figure 5. The standard curve for zinc with the equation of a straight line and R2 value. Absorption as a function
of concentration (ppm).
Figure 6. The standard curve for calcium with the equation of a straight line and R2 value. Absorption as a
function of concentration (ppm). y = 0.1033x + 0.0104 R² = 0.9935 -0,1 0 0,1 0,2 0,3 0,4 0,5 0 1 2 3 4 5 Ab so rb an ce Concentration (ppm)
Standard curve for iron
y = 0.8075x + 0.0114 R² = 0.9969 -0,2 0 0,2 0,4 0,6 0,8 0 0,2 0,4 0,6 0,8 1 Ab so rb an ce Concentration (ppm)
Standard curve for zinc
y = 0.0971x + 0.0258 R² = 0.9995 0 0,1 0,2 0,3 0,4 0,5 0 1 2 3 4 5 Ab so rb an ce Concentration (ppm)
Example on how to calculate the concentration of a mineral, in this case iron in MeatB 1.
The equation of a straight line from the iron standard curve. 𝑦 = 0.1033𝑥 + 0.0104
Include the sample absorbance in the equation and determine x. 0.239 = 0.1033𝑥 + 0.0104
𝑥 ≈ 2.21 𝑝𝑝𝑚
Convert to appropriate unit. 2.21 𝑝𝑝𝑚 = 2.21𝑚𝑔𝐿 2.21𝑚𝑔
𝐿 = 2.21 µ𝑔/𝑚𝐿
Include correct amount of sample. 2.21 µ 𝑔
𝑚𝐿∙ 50 𝑚𝐿 = 110.5 µ𝑔/50𝑚𝐿 → 110.5 µ𝑔/17.15𝑔
Convert to the right unit. 110.5 µ𝑔 17.15 𝑔 ≈ 6.44 µ𝑔/𝑔 6.44 µ𝑔 𝑔 ∙ 100𝑔 = 644 µ𝑔 100𝑔 644 µ𝑔/100𝑔 1000 ≈ 0.64 𝑚𝑔/100𝑔
Appendix 2. Measurement uncertainty amino acids
Table XIII. Measurement uncertainty for different amino acids.Amino acids Measurement uncertainty
