Food Biotechnology
Elnaz Bayat asl; Marzieh Moosavi-Nasab; Mahbobe Fazaeli
Abstract
Introduction
Wheat germ is a valuable nutritional supplement and a by-product of the flour milling industry used for animal feed and oil extraction. Quinone compounds found in wheat germ have anti-cancer properties that are abundantly found in wheat germ. The aim of this study was to investigate the ...
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Introduction
Wheat germ is a valuable nutritional supplement and a by-product of the flour milling industry used for animal feed and oil extraction. Quinone compounds found in wheat germ have anti-cancer properties that are abundantly found in wheat germ. The aim of this study was to investigate the effect of fermentation conditions on the bioactive compounds in wheat germ with anti-cancer properties. For this purpose, the Saccharomyces cerevisiae 5022 and Lactobacillus plantarum strain 1058 were used for fermentation of wheat germ under different pH levels (4.5, 6, and 7.5) over different time (24, 48, and 72h). Response Surface Methodology was used to find the optimal fermentation conditions and to investigate the effects of above-mentioned conditions on DPPH radical scavenging activity, total phenolics and dimethoxy benzoquinone (DMBQ) content. Moreover, the amounts of bio-peptides and gamma aminobutric acid (GABA) were also determined under optimum conditions.
Materials and Methods
To accomplish the fermentation process, 10 g of wheat germ was suspended in 200 mL of sodium phosphate buffer solution. Bacterial and yeast cells were then separated from the culture medium by a centrifugation at 6,000×g for 5 min at room temperature. The harvested cells were then washed with sterile phosphate buffer multiple times, resuspended in water to achieve a cell population of 108 CFU/mL, and finally homogenized using a vortex unit. The yeast and bacterial cells were incubated at 28° C and 37° C, respectively, for 24, 48, and 72 h at pH levels of 4.5, 6.0, and 7.5. Upon the completion of each fermentation process, the obtained samples were lyophilized. Total phenolic content (TPC) was measured using the method adapted by Liu et al. (2017). Briefly, the Folin-Ciocalteu phenol reagent was diluted ten times using distilled water. Subsequently, 0.1 mL of the extract was mixed with 0.75 mL of the diluted reagent. After 10 min, 0.75 mL sodium carbonate solution (2% w/v) was added to the mixture and vortexed. The absorbance was measured at 765 nm by a spectrophotometer. The antioxidant activity of fermented wheat germs was assessed using the free radical scavenging activity of the samples evaluated through a DPPH radical assay. Briefly, 2 mL of wheat germ extract was diluted with 100 mL 90% methanol aqueous solution. The methanol extract was then mixed with 4 mL of DPPH stock solution. The tube was subsequently kept in the dark for 45 min. The absorbance of each sample was then read using a spectrophotometer at 517 nm (Adedoyin et al., 2013). Dimethoxy benzoquinone (DMBQ) content was measured by an HPLC system. Briefly, 10 g of lyophilized wheat germ sample was dissolved in 250 mL of distilled water and extracted three times by shaking with 200 mL of chloroform. The chloroform layers were collected, washed three times with distilled water, and exposed to sodium sulfate solution to induce drying of the sample. The filtrate was then evaporated using a vacuum evaporator at 30° C to achieve a stable dry material. The dried sample was thereafter dissolved in the mobile phase and injected into the HPLC column to determine the DMBQ content. The HPLC system was equipped with a C-18 column and a UV detector operating at 245 nm. The mobile phase consisted of 20% acetonitrile-80% water (v/v) mixture at a flow rate of 0.5 mL/min and a temperature of 25° C.
Results and Conclusion
The highest biological activity was found when fermentation proceeded by L. plantarum under pH 6 for 48 h. Under these optimal conditions, total phenol content (3.33 mg of GAE/g), free DPPH radical scavenging (86.49%), dimethoxy benzoquinone content (DMBQ) (0.56 mg/g), peptide content (607 μg/mL) and gamma aminobutyric acid (GABA) (19983.88 mg/kg) were significantly higher than those of raw non-fermented samples. During the fermentation process, increasing the pH levels led to enhancement of antioxidant activity, total phenolic and DMBQ contents up to 48 h followed by a decline. Also, the fermentation time had a positive effect in the amount of the antioxidant activity, while it allowed an increased followed by a decrease in the contents of total phenolic and DMBQ. These findings underscore the importance of fermentation conditions of wheat germ by L. plantarum and Saccharomyces cerevisiae and can potentially serve as a promising way for the development of valuable products with anti-cancer and antioxidant functions.
Mohammad Taghi Golmakani; Marzieh Moosavi-Nasab; Malihe Keramat; Azin Azhand
Abstract
Introduction: Wheat germ is a by-product of wheat milling industry. It contains about 11% oil. Wheat germ oil is well known as a tocopherol rich food lipid. It also contains more than 55% polyunsaturated fatty acids, mainly linoleic and alpha-linolenic acid (Simopoulos 1999; Schwartz et al. 2008). Wheat ...
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Introduction: Wheat germ is a by-product of wheat milling industry. It contains about 11% oil. Wheat germ oil is well known as a tocopherol rich food lipid. It also contains more than 55% polyunsaturated fatty acids, mainly linoleic and alpha-linolenic acid (Simopoulos 1999; Schwartz et al. 2008). Wheat germ processing presents challenges due to its high content of bioactive compounds. Microwave-assisted extraction is a new extraction technology used for the extraction of bioactive compounds, which is based on combination of microwave and conventional solvent extraction. This technique which is used has many advantages such as short time, less solvent usage, and higher extraction yield (Hao et al. 2002).Common Kilka (Clupeonellacultriventris caspia) oil is considered as one of the most healthy and functional oils. It is highly rich in polyunsaturated ω-3 fatty acids such as eicosapentaenoic acid and docosahexaenoic acid. However, Kilka oil is highly vulnerable to oxidation due to its high content of poly unsaturated fatty acids. Oxidations of poly unsaturated fatty acids such as eicosapentaenoic acid and docosahexaenoic acid result in a number of oxidation products that have negative impacts on the flavor and odor of Kilka oil, and also can affect the amount of these fatty acids that are made available to the body (Lin and Lin 2004; Fazli et al. 2009; Pazhouhanmehr et al. 2015; Yu et al. 2002). In order to preserve polyunsaturated fatty acids of Kilka oil from oxidative degradation, the use of novel and effective antioxidants can offer methods to maintain the health of consumers.The objective of this study was to investigate the effect of microwave-assisted extraction method on extraction yield and some chemical characteristics of wheat germ oil in comparison with conventional Soxhlet method. Also, wheat germ oil was investigated as a natural antioxidant for improving oxidative stability of Kilka oil.
Materials and methods: Wheat germ used in this research was supplied from Sepidan Flour Mill (Shiraz, Iran). Crude Kilka oil with no added antioxidants was supplied by a local fishery factory (Rasht, Iran).Wheat germ samples were pretreated with microwave at 200 W for 5 min. Thereafter, the samples were extracted with Soxhlet method. Samples were analyzed at 2, 4, 6, 8, and 10 h of extraction process. Extraction yield, saponification value, acid value, iodine value, and fatty acid profile of wheat germ oil extracted with microwave-assisted method were compared with those extracted with conventional Soxhlet method. Fatty acid composition of wheat germ oil was determined according to the method described by Golmakani et al. (2012) with some modifications. Saponification, acid, and iodine values of wheat germ oil were determined by using the AOAC official methods (AOAC 2000). Wheat germ oil was added to Kilka oil at a concentration of 1000 ppm. For the control, Kilka oil without any added antioxidant was used. Peroxide, anisidine, and Totox values of wheat germ oil were measured during 15 days storage at 60 °C. Peroxide, anisidine, and Totox values of wheat germ oil were determined using the AOCS official methods (AOCS 2000). Induction period was considered as the number of days required for a sample to reach a PV of 20 meq O2/kg (Keramat et al. 2016).
Results and discussion: The microwave-assisted extraction method increased the extraction yield of wheat germ oil by 15-27%. Increase in extraction yield is due to cell membrane rupture by microwave which results in greater porosity, enabling the passage of oil from the cell membrane (Uquiche et al. 2008). The amounts of saturated fatty acids, monounsaturated fatty acids, and polyunsaturated fatty acids in samples extracted by microwave-assisted extraction method were similar to those extracted by conventional Soxhlet method. Acid value of samples extracted by microwave-assisted extraction method was slightly higher than those extracted by conventional Soxhlet method. This result is in agreement with the previous studies (Kiralan et al. 2014; Uquiche et al. 2008). The saponification value of wheat germ oil sample extracted by microwave-assisted extraction method was 9.65% higher than those extracted by conventional Soxhlet method. Thus, wheat germ oil sample extracted by microwave-assisted extraction method contained higher short chain fatty acids than those extracted by conventional Soxhlet method. The iodine value of wheat germ oil sample extracted by microwave-assisted extraction method was lower than those extracted by conventional Soxhlet method. Accordingly, microwave-assisted extraction method has a positive effect on the oxidative stability of wheat germ oil. Wheat germ oil significantly decreased the peroxide, anisidine, and Totox values of Kilka oil by 61.59%, 65.01%, and 61.97%, respectively, compared to the control. The induction period and protection factor of Kilka oil sample containing wheat germ oil (120.20 h and 1.42, respectively) was significantly higher than those of control sample (84.40 h and 1.00, respectively). The inhibitory effect of wheat germ oil against Kilka oil oxidation can be attributed to the presence of high amounts of biological active compounds. Based on the results of this study, microwave extracted wheat germ oil can be proposed as a natural antioxidant for improving oxidative stability of Kilka oil.
Sareh Boostani; Marzieh Moosavi-Nasab; Mahmoud Aminlari; Mehrdad Niakosari; Gholam Reza Mesbahi
Abstract
Introduction: Proteins play a fundamental role in biological systems and are often sensitive against organic solvents, heat and other damaging factors. Proteins are the basic component of food formulations and enhancement the functional characteristics and stability of the proteins has always been the ...
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Introduction: Proteins play a fundamental role in biological systems and are often sensitive against organic solvents, heat and other damaging factors. Proteins are the basic component of food formulations and enhancement the functional characteristics and stability of the proteins has always been the main goal of food industry engineers. One of the natural ways used for protein modifications is Maillard reaction. Maillard reaction as a result of covalent binding between the available amino groups of the proteins and carbonyl containing moiety of the polysaccharides, causes a loss in free amino group content of the mixture. Protein- polysaccharide hybrids, as a result of dry heating of two biopolymers mixture under controlled reaction conditions, cause the emergence of conjugates with novel functionalities. Much research has shown that conjugation can increase thermal stability and functional characteristics of food proteins and also reduce the allergenicity of suspected proteins. Although many studies have been conducted on the effects of conjugation on functional properties of proteins, the impacts of conjugation on proteins behavior after food processing have been less investigated. So, in this paper the influences of Maillard reaction on functional properties of soy proteins have been investigated. In addition, characteristics of conjugated proteins after pasteurization treatment was also studied. Materials and methods: Construction of protein- polysaccharide conjugates was done in several steps. First, soy proteins and dextran were mixed with phosphate buffer (0.1 M, pH: 8.5) and 1 to 4 ratio of protein to polysaccharide. After mixing and incubating at ambient temperature for some hours, solutions were frozen at –80 ℃ and freeze dried. Then the lyophilized powder was incubated at different times, at 60℃, under the 79 percent relative humidity in presence of saturated KBr. For each treatment a non conjugated sample was prepared in the exact same condition. Conjugation of proteins to polysaccharides was monitored by SDS-PAGE electrophoresis, browning intensity measurement and UV absorbance analysis. SDS-PAGE was conducted according to Laemmli procedure using a discontinuous buffer system. A vertical gel electrophoresis unit was used with 3% acrylamide stacking gel and 10% acrylamide running gel. Evaluation of the color changes as an indicator of grafting intensity was investigated by monitoring absorption at a wavelength of 420 nm. To investigate the UV absorption of conjugated proteins, the samples were diluted with SDS (Sodium dodecyl sulfate) solution and the absorption was read by a UV-visible spectrophotometer at 294 nm. The impact of modification on characteristics of soy proteins was monitored by examining the functional properties changes of protein samples. In the last stage soy drinks were prepared from conjugated and non conjugated proteins then the prepared beverages were subjected to thermal processing conditions and the influences of Maillard conjugation on the stability of soy drinks was monitored over time. Results and Discussion: SDS-PAGE electrophoresis profile showed that proteins-polysaccharide conjugates were formed. As a result of conjugation, the protein-dextran covalent binding occurs, leads to the formation of higher molecular weight components, resulting in its accumulation on the top of the separating gel. When heating time increased a wider and higher molecular weight bands appeared near the top of the running gel however they were not observed in the native soy pattern. Covalent linkage between amino group of proteins and carbonyl group of polysaccharides causes a color changes from light yellow to brown, browning intensity results showed that the even during early incubation time, a significant absorption was observed at 420 nm compared to the control samples. UV absorption results showed similar trend of changes in browning intensity measurement. Increasing UV absorption is due to the intermediate Maillard reaction products (MRP). Increasing UV absorption with increasing heating time indicates the fact that Maillard reaction products (MRP) formation are more favorable in the long incubation time. Data of UV absorption are a good evidence for SDS-PAGE and browning intensity results. Functional properties results indicated that grafted proteins had better functional properties. The storage stability of soy drinks prepared from conjugated proteins was significantly higher than the samples prepared from non conjugated proteins. Stability of beverages after thermal processing and during storage is one of the most important features of protein drinks and many efforts have been made to develop mentioned characteristics. Stability of soy drinks produced from the conjugated proteins was significantly higher than those prepared from non conjugated soy proteins. Functional characterization of proteins is dependent on several factors, the majority of soy drink composed of proteins that could be denaturated by heating applied during thermal processing, as the results showed conjugation with dextran caused an increase in denaturation temperature of soy proteins which enhance the resistance of proteins during thermal processing treatment. In addition, the solubility and emulsifying properties of soy proteins increased with conjugation which can be a good reason for improvement the relation between protein and surrounding water molecules and therefore increases the protein storage stability. It can be concluded that Maillard reaction could be applied as a means to prepare soy proteins–dextran conjugates with better functional properties and more stable during processing and storage.
Sareh Boostani; Mahmoud Aminlari; Marzieh Moosavi-Nasab; Mehrdad Niakosari; Gholam Reza Mesbahi
Abstract
Introduction : Soybean is an excellent plant protein source with diverse applications in food systems. Despite numerous commercial applications and rich nutritional properties of soybeans, soy proteins are sensitive to heat and other damaging agents during food processing which can limit their applications ...
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Introduction : Soybean is an excellent plant protein source with diverse applications in food systems. Despite numerous commercial applications and rich nutritional properties of soybeans, soy proteins are sensitive to heat and other damaging agents during food processing which can limit their applications in food industries. Maillard reaction includes a series of chemical reactions between the free carbonyl groups of carbohydrates and the un-protonated amino groups of proteins under mild experimental conditions. This is one of the most desirable approaches for applying in food systems, because of the safety of the procedure and the independency of adding extra chemicals. Natural occurring Maillard reaction can be a relatively safe and inexpensive method in order to improve functionalities of food proteins. The production of conjugates haspositive influences on food proteins functionality such as solubility, water holding capacity, emulsion activity and stability, foaming properties, thermal stability, whipping ability, textural and gelation properties and also reduce allergenicity of proteins. Due to the positive characteristics and reasonable price of both soy proteins and maltodextrin in food industries, the aim of current study was to enhance the heat stability and functional properties of soy proteins through glycosylation with maltodextrin. In addition, assessment of changes in protein properties as a function of incubation time were evaluatedMaterials and methods: Preparation of purified soy proteins (Acid precipitated soy proteins) was done by a multistage process of washing, centrifugation, dialysis and freeze drying. The resulting powder contained pure soy globulins. Conjugation of acid precipitated soy proteins with maltodextrin was performed according to the method described as follows: protein-polysaccharide at a weight ratio of 1: 3 were dissolved in 0.01 M phosphate buffer, at pH values of 8, and were incubated at ambient temperature for 1 hr. Solutions were frozen at –80°C and then freeze dried. Lyophilized powders were incubated at 60 °C for 1, 3, 5 and 7 days, under 79% of relative humidity provided by saturated KBr. For each treatment an un-conjugated (control) sample was prepared under the same conditions. The formation of protein-polysaccharide conjugates was confirmed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and gel filtration chromatography (Sephadex G-100 chromatographic system was used to separate the conjugated proteins from the un conjugated samples). Determination of protein denaturation temperature changes were carried out using METTLER TA Q100-DSC thermal analyser.The emulsifying properties of the samples including emulsifying activity and emulsion stability were assessed according to the procedure established by Pearce and Kinsella. Protein solubility was measured by calculating the amount of nitrogen in the supernatant and total nitrogen content of the samples and reported as percentage of protein solubility at pH 3, 5, 7 and 9. Foaming properties of the samples including foaming capacity and foaming stabilitywere determined using calibrated measuring cylinderDiscussion & Results: When the heating duration is increased, wider and heavier molecular weight bands emerge near the top of the running gel of SDS-PAGE, and yet these were not observed in the control. As a result of conjugation, the protein-carbohydrate covalent binding occurs, producing heavier molecular weight species, and thus leading to its accumulation on top of the separating gel. Compared with un-modified soy proteins, the conjugated soy proteins eluted in the void volume of G-100 gel permeation chromatography column, suggesting increase in the size and molecular weight of soy proteins due to the covalent attachment of maltodextrin. According to differential scanning calorimetry (DSC) analysis, thermal stability of soy proteins was remarkably increased by conjugation with maltodextrin and maximum denaturation temperature was observed for the mixture incubated for 7 days. The improved thermal stability is manifested in increase in denaturation temperature of globular proteins, hence conjugation leads to significant improvement of soy proteins stability. Increase in thermal stability is the result of inclusion of the hydrophilic carbohydrate moiety to the surface of the proteins. Compared to control sample, the solubility, foaming characteristics and emulsifying properties were significantly improved by increasing incubation time. The protein solubility of conjugate remarkably increased at all pH’s compared with the un-conjugated proteins. Covalent links between hydrophilic maltodextrin and soy proteins could enhance the reaction tendency between proteins and water molecules under unfavorable conditions. Improvements in the emulsifying properties of the conjugated samples can be explained by the fact that there is a combination among the emulsifying activity of proteins and the stabilizing impacts of polysaccharides per molecule. Foaming capacity of proteins can be affected by the solubility of proteins. Furthermore, maltodextrin is a hydrophilic carbohydrate which can improve the stability of soy proteins foams by acting as a thickener, thus increasing the strength of bubbles. It should also be considered that functionality of proteins are frequently influenced by protein solubility, and this could also serve the understanding of why improvements occur in functional properties of conjugated proteins, compared to un-conjugated ones. The results indicate that physiochemical and functional properties of soy proteins were modified and improved by conjugation with maltodextrin.
Sareh Boostani; Mahmoud Aminlari; Marzieh Moosavi-Nasab; Mehrdad Niakosari; Gholam Reza Mesbahi
Abstract
Introduction: Engineering of food proteins with improved functional properties and higher resistance to heat is the main goal of food scientists. Food technologists are always seeking for innovative, simple and effective methods to manipulate proteins, hence natural modification has had the most attention ...
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Introduction: Engineering of food proteins with improved functional properties and higher resistance to heat is the main goal of food scientists. Food technologists are always seeking for innovative, simple and effective methods to manipulate proteins, hence natural modification has had the most attention in last decades. One of the natural ways used for protein modifications isMaillard grafting reaction. Maillard reaction as a consequence of covalentbindingbetween the available amino groups of the proteins and carbonyl containing moiety of the polysaccharides, causes a loss in free amino group content of the mixture that can be measured through different methods. Protein-polysaccharide hybrids, as a result of dry heating of two biopolymers mixture under controlled reaction conditions, cause the emergence of conjugates with novel functionalities.Selecting appropriate reaction conditions has a significant impact on the properties of the final conjugates. Among the factors affecting the degree of conjugation, temperature, pH and incubation time have considerable roles in determining the degree of conjugation. There are various methods by which conjugation degree can be assessed and factors such as accuracy, cost, accessibility and etc. must be considered when selecting a measuring method. Therefoe, in this study the effect of different Maillard reaction conditions on the conjugation degree of soy proteins-dextran heated mixtures have been investigated. In addition, the glycation degree was compared and reported by both OPA assay and UV absorbance methods.Materials and methods: Soy proteins–dextran conjugates were prepared as described later. First, soy proteins and dextran were mixed with phosphate buffer (0.1 M, pH: 8.5 and 7) and 1 to 4 ratio of protein to polysaccharide. After mixing and incubating at ambient temperature for some hours, solutions were frozen at –80 ℃ and freeze dried. Then, the lyophilized powder was incubated at 40 ℃ for 0, 4, 8, 24 hours and 2, 4, 6, 8, 12 days, at 60 ℃ for 0, 1, 2, 3, 4, 6, 8 days and at 80 ℃ for 0, 1, 2, 4, 8, 16, 24, 48 hours, under the 79 percent relative humidity in presence of saturated KBr. For each treatment a non conjugated sample was prepared in the exact same condition. Conjugation of proteins to polysaccharides was monitored by two methods (OPA assay and UV absorbance). Determining degree of glycation by OPA method was done as follows, in this procedure the level of available amino groups was estimated using the ortho- phthaldialdehyde (OPA) reagent, the absorbance was measured at 340 nm and degree of glycation was measured using a formula. To investigate the UV absorption of conjugated proteins, the samples were diluted using distilled water and the absorption was read using a UV-visible spectrophotometer.Results & Discussion: Covalent linkage between amino group of proteins and carbonyl group of polysaccharides causes depletion in the amount of available amino groups. The extent of soy proteins-dextran conjugation under various Maillard reaction conditions was evaluated by the reduction of available amino groups of proteins, the more reduction in amount of amino groups, the more conjugation between protein and polysaccharide. OPA results showed that, in the samples heated at 40 °C and 80 °C (at both pH 7 and 8.5), the amount of free amino groups slightly reduced compared to 60 °C heated samples. The disappearance of available amino groups at 60°C was faster than other temperatures and formation of conjugates in this temperature was more successful. A stepwise reduction in free amino group content observed with increasing incubation time. When soy proteins were incubated at pH 8.5 for 8 days at 60 °C, a considerable decrease in available amino group contents occurred. UV absorption results showed similar trend of changes in OPA method. Increasing UV absorption is due to the intermediate Maillard reaction products (MRP). Increasing incubation time, temperature and pH cause a significant increase in UV absorbance. Increasing UV absorption with increasing heating time indicates the fact that Maillard reaction products (MRP) formation is more favorable at alkaline pH. Data of UV absorption are a proper evidence for OPA assay results. Conclusion: As a conclusion, both OPA assay and UV absorption methods are cost effective, accurate and simple methods which can represent valuable information concerning the Maillard conjugation.
Marzieh Moosavi-Nasab; Nazanin Darabzadeh
Abstract
Lysine is nutritionally essential for humans and animals. It cannot be synthesized biologically in body. Children and growing animals have a high requirement of lysine, since it is needed for bone formation. Therefore, it must be added to food and feed supplement to improve the protein quality. In addition, ...
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Lysine is nutritionally essential for humans and animals. It cannot be synthesized biologically in body. Children and growing animals have a high requirement of lysine, since it is needed for bone formation. Therefore, it must be added to food and feed supplement to improve the protein quality. In addition, lysine has some pharmaceutical applications in the formulation of diets with balanced amino acid composition. Chemical, enzymatic and biotechnological processes have been used to synthesize lysine. However, biotechnological methods (fermentation) seem to be the most economical and practicable means of producing lysine. In the present study, production of lysine by Brevibacterium linens PTCC 1603 from whey and molasses medium, in the purpose of using agricultural by-products as low-cost sources for production of value-added products, was investigated. Whey medium was prepared by hydrolyzing of lactose with HCl and molasses medium was prepared by addition of some chemicals to it. The production of lysine was examined qualitatively and quantitatively using ascending thin layer chromatography (TLC) and spectrophotometry. Results showed that lysine concentration in both media was increased significantly by the time. The highest concentration of lysine produced from whey and molasses media were 11.8 and 10.5 g.L-1, respectively. Increasing lysine concentration resulted in pH increment in molasses medium, while the pH of whey medium was decreased. After 6 days of fermentation, the amount of lysine in whey medium was significantly higher than in molasses medium.
Keywords: Lysine, Fermentation, Brevibacterium linens, Whey, Molasses
Marzieh Moosavi-Nasab; Afsaneh Dehghan
Abstract
Some wastes especially wastes of food industries need to be treated in order to recover some value-added products. One of these valuable products is biological pesticides. Today, the most successful biological pesticides are produced by Gram-positive, rod-shape bacteria, the genus Bacillus that produces ...
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Some wastes especially wastes of food industries need to be treated in order to recover some value-added products. One of these valuable products is biological pesticides. Today, the most successful biological pesticides are produced by Gram-positive, rod-shape bacteria, the genus Bacillus that produces crystalline endotoxin proteins during spore formation. These toxins will be activated when they enter the insect intestine under alkaline condition and will damage the cell membrane and make holes in the membrane which result in lysing of epithelial cells. In this research, the waste of starch producing industry was inoculated with Bacillus thuringiensis var. kurstaki and then the crystalline protein with MW of 65 KDa was produced during fermentation. The larvaes of Galleria melonella were treated to this protein and the rate of mortality was assessed after 24 and 48 h, and its effect was compared with that of control. Researches showed that the production of delta endotoxin has direct correlation with sporulation. The most toxicity of these bacterial proteins occurs at the highest number of spores. Thus, the number of total bacteria and their spores were enumerated using PCA during fermentation that was 12.5×108 and 48×107 cfu/ml, respectively.
Key words: Microbial pesticide, Starch producing industry wastewater, Endotoxin, Bacillus thuringiensis var. kurstaki
