NOVEL ENHANCED SOYBEAN PROTEIN CONTENT AND ANTIOXIDANT ENZYMES WITH CHITOSAN-ALGINATE THEREOF

Abstract

ABSTRACT The invention enhances soybean protein content and antioxidant enzyme activity through the application of natural biopolymers, specifically chitosan and alginate. These biopolymers, known for their biocompatibility and biodegradability, were utilized to treat soybean seeds at critical growth stages, including post-germination, mid-growth, and pre-flowering. The study demonstrated that combined treatments significantly increased the protein content of soybeans by at least 14%, along with notable improvements in the levels of essential storage proteins, glycinin and ß-conglycinin. Furthermore, the treatments boosted the activity of key enzymes involved in nitrogen assimilation and enhanced antioxidant defense mechanisms, improving resilience to environmental stress. This sustainable approach not only elevates the nutritional value of soybeans but also promotes better growth and yield, offering a valuable strategy for improving crop quality in environmentally friendly agricultural practices. The findings underscore the potential of chitosan and alginate as effective agents for optimizing soybean production. Dated this …….Day of September, 2024 Dr. Monica Gulati Registrar Lovely Professional University

Specification

Description:The following specification particularly describes the invention and the manner it is to be performed.

TECHNICAL FIELD
[001] The technical field of the invention encompasses agricultural biotechnology, specifically the enhancement of crop quality and nutritional value through the application of natural biopolymers. This includes the use of chitosan and alginate as growth promoters and enhancers of protein content in soybean plants (Glycine max L.). The invention focuses on optimizing the expression of key storage proteins, such as glycinin and ß-conglycinin, while also improving nitrogen assimilation and antioxidant enzyme activity. The methods involve seed soaking and foliar spraying during critical growth stages, aiming to provide a sustainable approach to increasing protein levels and resilience against environmental stressors in soybean cultivation. This innovation contributes to advancing sustainable agricultural practices and improving the nutritional profile of plant-based protein
BACKGROUND
[002] The global demand for plant-based protein sources, especially soybeans, is on the rise due to their nutritional value and versatility in food products. As a major source of protein and essential amino acids, soybeans are crucial in both human diets and livestock feed. However, environmental factors, such as nutrient deficiencies and climate stress, often hinder their growth and protein synthesis, necessitating effective agricultural practices to enhance their yield and quality.
[003] Chitosan, a biopolymer derived from chitin, has been extensively studied for its role in promoting plant growth. Its properties include improving nutrient uptake, enhancing chlorophyll levels, and stimulating the production of defense-related enzymes. These effects contribute to increased resilience against biotic and abiotic stress, making chitosan a valuable tool in sustainable agriculture. Research has shown that chitosan can enhance protein synthesis, potentially leading to higher protein content in crops, including soybeans.
[004] Alginate, extracted from brown seaweed, also presents significant advantages in agricultural applications. It functions as a natural soil conditioner, helping retain moisture and nutrients, thus promoting better root development and overall plant health. Additionally, alginate can encapsulate bioactive compounds, facilitating their release to plants and improving nutrient availability. This synergistic effect can be particularly beneficial for crops that require enhanced nutrient uptake for optimal growth.
[005] US4964894A, highlight the potential of chitosan as a plant growth regulator, but often focus on its effects on growth characteristics rather than on enhancing protein content specifically in soybeans. Similarly, WO2024003795A1 discusses biopolymer formulations for agricultural applications but does not delve into the combined effects of chitosan and alginate on soybean protein enhancement. This indicates a research gap that the current invention aims to address.
[006] Recent studies indicate that the use of biopolymers like chitosan and alginate can lead to significant improvements in nitrogen metabolism within plants. Enhanced activity of enzymes such as glutamine synthetase and nitrate reductase is crucial for efficient nitrogen assimilation, directly influencing protein synthesis in soybeans. By optimizing these enzymatic pathways, the treatments can increase the levels of essential storage proteins like glycinin and ß-conglycinin, which are vital for the nutritional profile of soybeans.
[007] The combined application of chitosan and alginate in soybean cultivation during critical growth stages presents a novel approach to enhancing seed quality. The strategic timing of seed soaking and foliar spraying can maximize the beneficial effects of these biopolymers, leading to improved protein content and yield. This innovative treatment method offers an environmentally sustainable alternative to synthetic fertilizers, aligning with current trends toward organic and sustainable agriculture.
[008] The antioxidant defense mechanisms in plants are vital for combating oxidative stress, which can be exacerbated by environmental factors. Chitosan and alginate treatments have been shown to boost the activity of key antioxidant enzymes, such as superoxide dismutase (SOD) and catalase (CAT). This enhancement not only improves stress resilience in soybeans but also contributes to overall plant health and productivity, making the crops more robust against varying climatic conditions.
[009] This invention addresses a critical need in agricultural biotechnology by integrating chitosan and alginate treatments to optimize soybean protein content and overall quality. By filling the existing research gaps, it offers a sustainable and effective method to enhance crop yield and nutritional value while promoting resilience against environmental stressors. This innovation holds the potential to significantly impact both agricultural practices and food industry standards.
SUMMARY
[010] The invention focuses on enhancing soybean protein content through the combined application of chitosan and alginate, two natural biopolymers known for their positive effects on plant growth and health. By treating soybean seeds and plants during critical growth stages, the invention aims to optimize the expression of key storage proteins, specifically glycinin and ß-conglycinin, which are essential for both nutritional quality and industrial applications. This method leverages the unique properties of chitosan to improve nutrient uptake and stimulate defense-related enzymes, while alginate enhances nutrient absorption and retention, promoting overall plant vitality.
[011] The application process involves seed soaking in chitosan and alginate solutions, followed by foliar sprays at key developmental stages. These treatments not only increase protein content by enhancing nitrogen utilization efficiency but also improve the overall antioxidant defense system of the plants. The resulting increase in enzymatic activities, including those of superoxide dismutase, catalase, and peroxidase, enhances the plants' resilience to environmental stressors, such as drought and disease, thereby ensuring better growth outcomes.
[012] Experimental results demonstrate that the combined treatment significantly boosts soybean protein levels, with increases in glycinin and ß-conglycinin content, along with improvements in nitrogen metabolism-related enzymes. The findings reveal a 14% rise in overall protein content and notable enhancements in seed yield, size, and germination rates. This indicates that the combined application of these biopolymers not only elevates nutritional value but also contributes to greater agricultural productivity.
[013] In addressing existing research gaps, this invention provides a novel and sustainable approach to soybean cultivation that minimizes reliance on synthetic fertilizers and chemicals. By integrating natural biopolymers, the method aligns with contemporary agricultural practices focused on sustainability and environmental stewardship. The innovative use of chitosan and alginate reflects a strategic advancement in biotechnology that can significantly impact crop quality and yield.
[014] Ultimately, this invention offers significant benefits for both farmers and the food industry, aiming to produce soybeans with higher nutritional value and enhanced resilience against environmental challenges. By optimizing protein content and promoting plant health, the invention contributes to the development of more robust agricultural practices, paving the way for future research and applications in sustainable crop management.
BRIEF DESCRIPTION OF THE DRAWINGS
[015] The foregoing detailed description of embodiments is better understood when read in conjunction with the appended drawings. For the purpose of illustrating of the present subject matter, an example of the construction of the present subject matter is provided as figures; however, the invention is not limited to the specific method disclosed in the document and the figures.
[016] The present subject matter is described in detail with reference to the accompanying figures. In the figures, the left-most digit(s) of a reference number identifies the figure in which the reference number first appears. The same numbers are used throughout the drawings to refer to various features of the present subject matter.
[017] Figure 1provides the Comparison of Protein Content and Enzyme Activities in Soybean Seeds Between Control and Chitosan + Alginate Groups
[018] The given figures depict an embodiment of the present disclosure for illustration and better understanding only.
DETAILED DESCRIPTION
[019] Some of the embodiments of this disclosure, illustrating all its features, will now be discussed in detail. The words "comprising," "having," "containing," and "including," and other forms thereof, are intended to be equivalent in meaning and be open-ended in that an item or items following any one of these words is not meant to be an exhaustive listing of such item or items or meant to be limited to only the listed item or items. It must also be noted that as used herein and in the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise.
[020] The invention focuses on enhancing soybean protein content by employing chitosan and alginate as treatments. These natural biopolymers were chosen for their biocompatibility and ability to positively influence plant growth and metabolism. Chitosan is derived from chitin, while alginate comes from brown seaweed, making them suitable for sustainable agricultural practices. The invention aims to upregulate the expression of essential storage proteins, glycinin and ß-conglycinin, crucial for improving the nutritional profile of soybeans.
[021] The methodology involved a controlled greenhouse study at Lovely Professional University (LPU), where soybean seeds were planted in sterilized soil pots. Chitosan was prepared as a 0.5% solution by dissolving it in 1% acetic acid, while alginate was made as a 2% solution in distilled water. Both solutions were stirred for 24 hours to ensure complete dissolution. The seeds underwent surface sterilization using 0.1% mercuric chloride before being rinsed and soaked in the biopolymer solutions for 24 hours prior to sowing.
[022] In one embodiment it is provided that, theexperimental design included four treatment groups: a control group with no treatment, a group treated with chitosan, a group treated with alginate, and a group receiving the combined treatment of chitosan and alginate. This setup allowed for comparison of the effects of each treatment on soybean growth and protein synthesis. Treatments were administered at three critical growth stages: post-germination (Day 15), mid-growth (Day 30), and pre-flowering (Day 45) to maximize their impact.
[023] The results demonstrated that the combined treatment of chitosan and alginate significantly increased soybean protein content by 14%, from 35.0% in the control group to 40.0% in the treated group. The levels of glycinin and ß-conglycinin also saw notable improvements, with increases of 25% and 24%, respectively. These enhancements indicate the effectiveness of the treatments in optimizing the nutritional value of soybeans.
[024] In one embodiment it is provided, that the Further analysis revealed that the activity of key enzymes involved in nitrogen assimilation, such as glutamine synthetase and glutamate synthase, increased by 28% and 30%, respectively, in the treated group. This upregulation facilitates more efficient nitrogen utilization, leading to enhanced protein synthesis. Additionally, nitrate reductase and phenylalanine ammonia-lyase activities also improved significantly, further contributing to the overall metabolic efficiency of the plants.
[025] The study also assessed the antioxidant defense mechanisms of the treated soybeans. The activity of antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), increased substantially, indicating improved resilience to oxidative stress. This enhancement is critical for maintaining plant health under adverse environmental conditions, making the plants more robust against stresses such as drought and disease.
[026] Seed yield measurements showed that the combined treatment resulted in a 16% increase in yield per plant, rising from 50.0 g in the control group to 58.0 g in the treated group. The size and weight of the seeds also improved, with an increase in average seed weight from 0.20 g to 0.23 g. This demonstrates that the treatments not only enhance protein content but also contribute to better overall productivity.
[027] In one embodiment it is provided, that the germination rate of seeds from the treated group was higher at 97%, compared to 90% in the control group, reflecting improved seed viability and vigor. Additionally, the stress tolerance of the treated plants rated higher on a scale from 1 to 5, moving from 3 (moderate tolerance) to 5 (high tolerance), highlighting the effectiveness of the biopolymer treatments in improving plant resilience.
[028] The advantages of this invention include a significant boost in soybean protein content and yield through sustainable practices that utilize natural biopolymers. By reducing reliance on synthetic fertilizers and chemicals, the method aligns with modern agricultural trends focused on environmental sustainability. This not only benefits farmers but also contributes to healthier food sources for consumers.
[029] The integration of chitosan and alginate treatments offers a flexible application method that can be tailored to various growth stages, allowing for targeted interventions at critical times. This adaptability can enhance the effectiveness of the treatments and improve the overall health and yield of soybean crops.
[030] Discussions around the invention emphasize the importance of further research to explore the underlying mechanisms by which chitosan and alginate enhance protein synthesis and stress resilience. Investigating the specific signaling pathways activated by these biopolymers could provide insights for optimizing their application in other crops and agricultural contexts.
[031] This invention presents a promising approach to improving soybean protein content and yield through the synergistic effects of chitosan and alginate. The demonstrated benefits in protein levels, enzyme activity, and stress resilience position this method as a valuable strategy in sustainable agriculture, paving the way for future innovations aimed at enhancing crop quality and productivity.
[032] Referring to Figure 1, presents a comparative analysis of soybean seeds treated with chitosan and alginate versus a control group, illustrating the differences in various biochemical parameters. It features a bar graph displaying protein content, glycinin and ß-conglycinin levels, and the activities of key enzymes such as glutamine synthetase, glutamate synthase, nitrate reductase, and phenylalanine ammonia-lyase, all measured in units per milligram of protein. Additionally, antioxidant enzyme activities, including superoxide dismutase, catalase, and peroxidase, are showcased, indicating the enhanced antioxidant capacity of the treated seeds. The bars for the treated group consistently show higher values across all parameters, demonstrating the significant improvements achieved through the combined application of chitosan and alginate. The visual representation effectively highlights the beneficial impact of these treatments on the nutritional and metabolic profile of soybeans, underscoring their potential to enhance both protein synthesis and plant resilience.
Dated this …….Day of September, 2024

Dr. Monica Gulati
Registrar
Lovely Professional University
, Claims:We claim:
1. A method for enhancing soybean protein content and antioxidant enzyme activity comprising the steps of:
A. Treating soybean seeds with a combination of chitosan and alginate solutions at specific growth stages, including post-germination, mid-growth, and pre-flowering, to increase the expression of glycinin and ß-conglycinin proteins.
2. The method of claim 1, wherein the chitosan solution is prepared by dissolving chitosan in 1% acetic acid to achieve a concentration of 0.5% (w/v), ensuring optimal solubility and bioactivity for plant uptake.
3. The method of claim 1, wherein the alginate solution is prepared by dissolving sodium alginate in distilled water to create a 2% (w/v) solution, facilitating effective nutrient encapsulation and delivery to the soybean plants.
4. The method of claim 1, wherein the soybean seeds are soaked in the chitosan and alginate solutions for a duration of 24 hours prior to sowing, allowing for enhanced absorption of the biopolymers and maximizing their effects on seed germination and early growth.
5. The method of claim 1, wherein the foliar spray applications of the chitosan and alginate solutions are conducted at three specific growth intervals: on Day 15 post-germination, Day 30 during mid-growth, and Day 45 prior to the pre-flowering stage, to optimize the timing of nutrient delivery.
6. The method of claim 1, wherein the combined treatment results in a significant increase in soybean protein content, specifically achieving an enhancement of at least 14% compared to untreated control seeds, thereby improving the nutritional profile of the harvested soybeans.
7. The method of claim 1, wherein the application of chitosan and alginate significantly enhances the activities of antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), thereby improving the plant's capacity to withstand oxidative stress and environmental challenges.
8. The method of claim 1, wherein the treatment with chitosan and alginate leads to an increase in nitrogen utilization efficiency in soybean plants, promoting enhanced growth rates and yield outcomes by improving nitrogen assimilation processes essential for protein synthesis.
Dated this …….Day of September, 2024
Dr. Monica Gulati
Registrar
Lovely Professional University

Documents