MINISTRY OF EDUCATION AND TRAINING HO CHI MINH CITY UNIVERSITY OF TECHNOLOGY AND EDUCATION FACULTY FOR HIGH QUALITY TRAINING GRADUATION PROJECT FOOD TECHNOLOGY PREPARATION OF FERMENTED SOYBEAN MEAL BY SOLID-STATE FERMENTATION LECTURER: ASSOC. TRINH KHANH SON STUDENT: TRAN KIEU HUONG LUONG THI NGOC SANG SKL 0 0 9 1 2 4 Ho Chi Minh City, August, 2022 HO CHI MINH CITY UNIVERSITY OF TECHNOLOGY AND EDUCATION FACULTY FOR HIGH QUALITY TRAINING GRADUATION PROJECT Thesis code: 2022-18116020 PREPARATION OF FERMENTED SOYBEAN MEAL BY SOLID-STATE FERMENTATION TRAN KIEU HUONG Student ID: 18116020 LUONG THI NGOC SANG Student ID: 18116034 Major: FOOD TECHNOLOGY Supervisor: ASSOC. TRINH KHANH SON Ho Chi Minh City, August 8th, 2022 i HO CHI MINH CITY UNIVERSITY OF TECHNOLOGY AND EDUCATION FACULTY FOR HIGH QUALITY TRAINING GRADUATION PROJECT Thesis code: 2022-18116020 PREPARATION OF FERMENTED SOYBEAN MEAL BY SOLID-STATE FERMENTATION TRAN KIEU HUONG Student ID: 18116020 LUONG THI NGOC SANG Student ID: 18116034 Major: FOOD TECHNOLOGY Supervisor: ASSOC. TRINH KHANH SON Ho Chi Minh City, August 8th, 2022 i ii ACKNOWLEDGEMENTS First of all, we would like to express our sincere and deep thanks to the teachers in the Department of Food Technology – Faculty of High-Quality Education – University of Technology and Education of Ho Chi Minh City for their dedication to conveying their knowledge during the past four years, creating all conditions for facilities and equipment to help us complete the thesis in the best way.
In particular, we would like to thank Assoc. Trinh Khanh Son who was very dedicated to helping, guiding, and imparting knowledge and experience to help us complete the graduation thesis on time. Finally, we would like to thank our family and friends by our side for supporting us, giving us time to research the topic and wholeheartedly supporting us mentally during the course of the thesis. We have tried to research and research to best complete the graduation thesis, but we are still limited in knowledge and experience, so there are bound to be shortcomings.
We hope to receive the contributions of teachers and friends to improve the topic. Thank you very much! iii iv v vi vii viii ix x xi xii xiii xiv xv xvi TABLE OF CONTENTS ACKNOWLEDGEMENTS. Error! Bookmark not defined. Error! Bookmark not defined.
TABLE OF CONTENTS. xvii LIST OF FIGURES. xix LIST OF TABLES. xx LIST OF ABBREVIATIONS.
Overview of Soybean Meal. Introduction of Soybean Meal. Properties of Soybean Meal. The role of B.
Overview of Fermented Soybean meal. The reason for creating the topic. MATERIALS AND METHODS. Materials, chemicals and equipment.
Materials and Chemicals. Investigation of the growth ability of microbial strains. Evaluation of proteolytic activity. Solid-state fermentation (SSF) in soybean meal.
Determination of protein content. Determination of total carbohydrate by Phenol-Sulfuric Acid method. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Gel Permeation Chromatography (GPC) analysis.
Selection of microorganisms with intense protease activity. Chemical composition of soybean meal (SBM). Enzyme-degrading protein concentration. Protein molecular weight.
24 CHAPTER 4: CONCLUSION AND RECOMMENDATIONS. 31 xviii LIST OF FIGURES Figure 2. Solid-state fermentation process SBM. Procedure for measuring soluble protein according to the Lowry method.
Growth curve of strain LM 1……… …………………………………………………………………………………. Growth curve of strain LM 2. Growth curve of strain LM 3. Growth curve of strain LM 4.
The zone of clearance on YNB agar plates by strain LM 1 at 37˚C for 24h, 48h, 72h (from left to right). The zone of clearance on YNB agar plates by strain LM 2 at 37˚C for 24h, 48h, 72h (from left to right). The zone of clearance on YNB agar plates by strain LM 3 at 37˚C for 24h, 48h, 72h (from left to right). The zone of clearance on YNB agar plates by strain LM 4 at 37˚C for 24h, 48h, 72h (from left to right).
Results of identification of microorganisms of strain LM3. Protein molecular weight based on SDS-PAGE electrophoresis results of soybean meal samples sterilized before LM 3T fermentation under condition 1. Protein molecular weight based on SDS-PAGE electrophoresis results of unsterilized soybean meal samples before LM 3K fermentation under condition 3. Gel Permeation Chromatography (GPC) results determined the molecular weight distribution of FSBM under condition 1.25 xix LIST OF TABLES Table 1.
Chemical composition, amino acid profile, and quality parameters of soybean meal protein and its net energy value 1. Standard curve construction table of Lowry method. Diameter size (mm) hydrolysis of 4 microbial strains on YNB agar plate at 37˚C…16 Table 3. Chemical composition of SBM and FSBM.
Results of total protein and water-soluble protein content of SBM samples sterilized at condition 1 (Appendix 4). Results of total protein and water- soluble protein content of unsterilized SBM samples at condition 3 (Appendix 4). Results of total protein and water-soluble protein content of SBM samples sterilized at condition 2 (Appendix 4). Results of total protein and water-soluble protein content of marketed FSBM samples (Appendix 4).22 xx LIST OF ABBREVIATIONS Abbreviations Explanation SBM Soybean meal SB Soybean FSBM Fermented soybean meal SSF Solid-state fermentation B.
subtilis Bacillus subtilis ANFs Antinutritional factors TI Trypsin inhibitor activity xxi ABSTRACT Soybean meal is a by-product of oil extraction. It contains many nutrients suitable for animal feed production. In addition to being a high-protein product, SBM (soybean meal) also contains significant amounts of anti-nutritional factors, and those elements need to be removed to increase the absorption of SBM. Therefore, SBM fermentation was performed from mold or bacterial strains (predominantly Bacillus sp, Aspergillus oryzae, and Lactobacillus subtilis, respectively) to eliminate compound anti-nutrients and improve the nutritional value of SBMs and also the degradation of allergen SB during fermentation by proteolytic microorganisms.
In the study in this article, four bacterial strains were screened to select the strain with the most substantial protease ability and then put into solid state fermentation within five days (0h, 24h, 48h, 72h, and 96h) and survey on the change of chemical composition of SBM during fermentation (total protein content, soluble protein content. The results showed that FSBM (fermented soybean meal) significantly increased in total protein (from 46.30%), water- soluble protein (from 8.23%), nitrogen-soluble index (from 17.53%); while reducing the content of protein soluble in KOH (from 86.6 to 75%), trypsin inhibitor activity (from 2. Overview of Soybean Meal 1. Introduction of Soybean Meal Soybean meal (SBM) is one of the essential protein sources used in the poultry and livestock industry, accounting for about two-thirds of the total protein feed production worldwide.
The protein value in SBM is higher than that of other plant protein sources and is considered a benchmark against which other protein sources are compared [1]. Soybean meal is a by-product of oil extraction. Its crude protein content often classifies SBM for marketing purposes. Soybeans, from the hulled seeds, the high-protein varieties have been harvested and contain 47-49% protein and 3% crude fiber, while another soybean meal contains less than 47% protein and more than 6% fiber, including the shell or part of the cover [2].
Soybean meal (or okara) is the fibrous part of beans, the insoluble part of soybeans left after soybeans have been pureed and filtered to produce soy milk and tofu. Generally, they are white or yellowish and are rich in nutrients, including insoluble and soluble fiber, protein, and other minerals (such as calcium, potassium, and niacin). Most of the isoflavones, B vitamins, and fat- soluble nutritional elements (including linoleic acid, soy lecithin, phytosterols, linolenic acid, vitamin D, and tocopherol) present in soybeans are still present in the SBM [3]. Several anti-nutritional factors are present in the SBM, such as saponins, trypsin inhibitors, and hemagglutinin.
SBM fermentation can aid in digestion and absorption of nutrients and improve nutritional value. It clears the smell of beans and increases the amount of edible fiber, fatty acids, sugars, free amino acids, vitamin B2, vitamin B12, and flavoproteins. SBM is a functional food to prevent diabetes, hyperlipidemia, and obesity. However, due to its high protein, soluble and insoluble fiber content, and high perishability, SBM is usually used immediately or cooked and frozen for storage.
The most attractive point of SBM that can be mentioned is the low price and low calories. More recently, okara has been used as an ingredient in cookies, cakes, and other confectionery, as well as in traditional side dishes. Their low-calorie composition makes them an ideal diet food [4]. SBM is the most popular soybean product in animal diets, with other products being used to varying degrees.
These products include full-fat soybeans, soy protein concentrate (SPC), soybean oil soy protein isolate (SPI), and soybean hulls. Each of these products has unique nutritional properties that make them suitable for inclusion in the diets of certain animals. In particular, in the composition of SBM, amino acids (AA) have supplemented the AA components of different grains with the practical use of AA in soybean meal, also explaining why in the food industry. For pet food, soybean meal is more and more widely used [5].
Properties of Soybean Meal The SBM's size and particle size distribution were determined using the standard method provided by ASAE (1993). The medium range (0.595 - 2,380 mm) consists of 70% soybean residues; 30% fine grain (<0. Analysis of normal and log-normal particle size distributions showed similar results for the mass-average particle diameters, 0. Humidity has a significant effect on both actual density and bulk density.
As the humidity increases, the solid and bulk density both decreased. Particle size significantly affects bulk density but does not affect absolute density. Fine particles have a higher bulk density than coarse particles. The particle shape and characteristic dimensional properties such as length, width, thickness, arithmetic mean diameter, and geometric mean diameter increase linearly with increasing moisture content.
The sphericity decreases linearly as the moisture content increases. The grain mass and surface area increase linearly as moisture content increases. Actual density, bulk density, and porosity decrease linearly with the increase in humidity [6]. Regarding the nutritional value of soybean meal, the chemical composition and nutritional value of soybean meal can vary depending on the processing process [7] and storage conditions, as well as the origin of the beans [8] and storage conditions.
Chemical composition of soybean meal, it also varies depending on the latitude of the growing area, the number of hours of light, weather, and environmental conditions during the harvest season [8] [9]. Accordingly, soybean meals from the US, Brazil, and Argentina (the top exporters worldwide) may have higher nutritional differences than expected. Chemical composition, amino acid profile, and quality parameters of soybean meal protein and its net energy value 1 [8][10]. Specific analysis, % 1 Soybean meal (Argentina) Ashes 6.5y Ethereal Extract (hydr.0y Amino acids profile, CP % Lysine 6.37x CP quality indicators UA3, mg N/g 0.2y TIA6, mg/g MS 2.8y Estimated analysis EN7, kcal/kg 2.051y 1 All results are given for a dry matter content of 88%; 2 Neutral detergent fibre; 3 Urease activity; 4 Protein Dispersibility Index; 5 Protein solubility in KOH; 6 Trypsin inhibitory activity; 7 Net energy is estimated by Noblet et al.
Bacillus sp Bacillus sp is a large and heterogeneous assembly of diverse aerobic or anaerobic, rod-shaped, spore-forming bacteria widely dispersed throughout the environment. pumilus have been identified as 'original members' of the genus Bacillus [12]. In 1943, Fukumoto isolated B. amyloliquefaciens for the first time, a bacterium that produces liquefied amylase from soil [13].
amyloliquefaciens was combined with the closely related B. subtilis species into the “B. subtilis species complex”, based on phylogenetic and magnetic evidence [14]. This classification was based on the highly conserved nature of the protein-coding sequences in the B.
subtilis species complex [15]. For many years, these closely related species were difficult to classify using classical taxonomic parameters such as morphology, physiology, guanine-cytosine content, and phylogenetic analysis 16S rRNA gene sequencing. Bacillus velezensis Some Bacillus plays a progressively important role in many fields such as agriculture and the fermentation industry.