Source: UNIV OF CONNECTICUT submitted to NRP
ENGINEERING BIOPOLYMER BEADS FOR SIMULTANEOUS REMOVAL OF CHEMICAL AND BIOLOGICAL POLLUTANTS IN WASTEWATER FOR AGRICULTURAL IRRIGATION
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
Reporting Frequency
Annual
Accession No.
1025486
Grant No.
2021-67022-34415
Cumulative Award Amt.
$199,955.00
Proposal No.
2020-08763
Multistate No.
(N/A)
Project Start Date
Feb 15, 2021
Project End Date
Feb 14, 2024
Grant Year
2021
Program Code
[A1521]- Agricultural Engineering
Recipient Organization
UNIV OF CONNECTICUT
438 WHITNEY RD EXTENSION UNIT 1133
STORRS,CT 06269
Performing Department
Nutritional Sciences
Non Technical Summary
Our LONG TERM GOAL is to design inexpensive, multifunctional and reusable filtration devices via functionalization and engineering of ubiquitous biopolymers for high throughput applications in wastewater treatment. In this proposed study, we HYPOTHESIZE that multifunctional hydrogel based filtration device can be strategically designed by fundamental chemistry and engineering principles to simultaneously remove chemical and biological pollutants for efficient treatment of recycled wastewater. We will test the hypothesis by conducting 3 OBJECTIVES: 1) Prepare porous hydrogel beads and functionalize them with cationic quaternary ammonium and EDTA-likechelating groups; 2) Investigate the functionalized beads for biocidal activity against microorganisms and sorption kinetics against chemical pollutants commonly present in wastewater; and 3) Design a prototype filtration facility and validate the efficacy of as-developed hydrogel beads in a continuous flow system, mimicking the wastewater treatment in real scenario. The outcome of this study will lead to the invention of inexpensive biopolymer-based filtration device to control the quality and safety of recycled wastewater for irrigation. The findings from the proposed research will DIRECTLY CONTRIBUTE to U.S. agriculture by providing a revolutionary strategy to develop a low-cost, multifunctional and reusable filtration device that holds a high potential to fit the existing pipelines of wastewater treatment for high throughput applications, a step further towards SUSTAINABLE AGRICULTURE. This study addresses several emphasized areas of the Bioenergy, Natural Resources, and Environment Program, and the Program Area Priority, "Engineering for Agricultural Production Systems" focusing on gaining both fundamental and applied knowledge to enable engineering for natural resources.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
1330210200050%
1330210202050%
Goals / Objectives
Our long-term goal is to design inexpensive, efficient, multifunctional, and reusable filtration devices via functionalization and engineering of ubiquitous biopolymers for the high throughput applications in agricultural production. In this proposal, we will test the hypothesis that engineered filtration devices can be strategically designed by chemical modification of highly porous chitosan hydrogel beads with enhanced biocidal and adsorption properties for efficient treatment of wastewater. The hypothesis is based on fundamental chemistry and engineering principles coupled with our preliminary data, indicating that (1) chitosan hydrogel beads with nanosized pores can be easily prepared via coagulation for adsorption of dyes and metals, and (2) a powerful modification approach is feasible to endow porous beads with superior antimicrobial and adsorption properties simultaneously. Our hypothesis will be tested by conducting the following three objectives:OBJECTIVE 1: We will prepare hydrogel beads with various porosity and functionalize the porous beads with cationic quaternary ammonium and EDTA-like chelating groups.OBJECTIVE 2: We will investigate the engineered beads for their antimicrobial activity against microorganisms and sorption kinetics against chemical pollutants commonly present in wastewater.OBJECTIVE 3: We will design a prototype filtration system with continuous flow mimickingwastewater treatment and validate the efficacy of the engineered beads.
Project Methods
OBJECTIVE 1: Prepare and functionalize porous chitosan beadsa. Preparation and characterization of beads: To prepare beads, 50 mL of the chitosan solution will be extruded dropwise into a beaker containing coagulation medium (H2O: methanol: NaOH = 4:5:1, w/w/w), and the obtained beads will be solidified in coagulation medium for 2 h under gentle stirring.The preparation of porous beads will be systematically optimized by surface response methodology. The morphology of beads will be observed under SEM. The mechanical property (Young's modulus) will be studied by a texture analyzer to calculate the elasticity and stiffness of the beads prepared with different formulations. The porosity of beads will be calculated as total void volume (%) by comparing the weight of porous beads to solid control beads. The stability will be studied by mixing 1 g of beads with 50 mL of water at pH 5, 6.5 and 8, mimicking the wastewater conditions, for one week. Then, the beads will be collected and dried in a desiccator for 72 h to calculate mass loss. The beads with high porosity, high mechanical property and desirable stability will be selected for chemical modifications.b. Modification through quaternization of surface primary amines: Cationic quaternary ammonium will be added through quaternization of primary amines on porous chitosan beads. The beads (5 g) will be mixed with 20 g of 1,4-diiodobutane and methyl iodide(100:1 to 4:1, mol) in 50 mL of ethanol containing 20 g of potassium hydroxide. The reaction mixture will be refluxed for 2 days. The beads will be filtered and washed with ethanol and water to remove excess reactants. The quaternization of surface primary amines will be characterized by Fourier-transform infrared (FT-IR), X-ray photoelectron (XPS) spectroscopies and solid-state 15N and 13C nuclear magnetic resonance. Degree of quaternization will be determined by potentiometry.c. Grafting chelating groups: Iminodiacetate anion is a tridentate ligand that complexes with transition metals ions with five membered chelate rings. It is one of the best ligands to strongly coordinate toxic metal ions, like Cu2+, Cr3+ and Cd2+. The residual alkyl iodide on the quaternized chitosan beads will further react with iminodiacetate anion. To do so, 0.5 g of beads and 1 g of iminodiacetic acid will be mixed in 10 mL of ethanol containing 2 g of potassium hydroxide. After refluxed for 2 days, the beads will be filtered and washed with ethanol. In subsequent experiments, by varying the ratio of 1,4-diiodobutane and methyl iodide, the amount of the iminodiacetate on the surface of chitosan beads is tunable. This allows us to optimize and balance the biocidal activity performance and the capacity to remove metal ions as demand.OBJECTIVE 2: Investigate adsorption and biocidal properties.a. Adsorption property: We will investigate the capability of prepared beads for the removal of chemical pollutants that are most commonly found in the wastewater, including heavy metal ions (Cu2+ & Cd2+), anionic and cationic dyes (Congo red & methylene blue), and organic matter (humic acid & fulvic acid), individually and more importantly in combination to mimic the complexity of wastewater. The organic matter in wastewater can decompose to generate organic acid, and thus most industrial and recycled wastewater is slightly acidic, i.e., pH 5-6.5; on the other hand, the acidic compounds may react with soils and minerals causing the reduction of H+ ion and thus the wastewater may become slightly alkaline, i.e. pH 8. Thus, the effect of pH will be studied by changing the initial pH of the pollutant solution to pH 5, 6.5 or 8. The batch adsorption studies will be carried out in a water bath shaker (30 °C) using glass vials containing either 0.2 g of wet functionalized or control beads, and 10 mL of contaminated aqueous phase containing individual or a combination of six pollutants (each at 100 mg/L). Equilibrium isotherm study will be carried out with a series of initial pollutant concentrations for 8 h. The non-linear forms of Langmuir and Freundlich isotherm models will be used to analyze the equilibrium adsorption isotherm data. The residual level of each pollutant in the aqueous phase will be analyzed using a UV/Vis spectroscopy (for dyes), inductively coupled plasma spectrometer (ICP-MS, for metal ions), and HPLC (for organic matter). The equilibrium adsorption capacity for each pollutant will be calculated based on mass balance equation Qe=V×(C0-Ce)/ W, where, Qe is the equilibrium adsorption capacity per gram dry weight of the adsorbent, mg/g; C0 is the initial pollutant concentration, mg/L; Ce is the final or equilibrium pollutant concentration, mg/L; V is the solution volume, L; and W is the beads weight, g. To test the reusability of the beads, desorption and regeneration experiments will be conducted. The beads (0.2 g) after saturated adsorption will be immersed in 0.1 M HCl solution for 24 h. The pollutant concentration in the aqueous phase will be analyzed to obtain recovery efficiency. If a recovery rate is > 95%, the beads will be considered as reusable in next adsorption.C.2.b. Antimicrobial experiments: To study the disinfection ability of beads, the two common bacterial species in wastewater will be tested, i.e., E. coli O157:H7 (Gram-negative) and S. aureus (Gram-positive). Bacteria will be grown in tryptic soy broth (TSB) and stored at −20 °C in 20% glycerol as stocks. Each type of bacteria will be transferred at least twice in TSB with an interval of 24 h prior to use. The beads (2 g) will be added into a beaker containing 100 mL of contaminated water sample inoculated with respective species or their combination. The contaminated water (100 mL) will be prepared by mixing with 100 µL of diluted culture ca. 108 CFU/mL in TSB. After incubation at room temperature for 0, 1, 2, 4, and 8 h under mild stirring, samples will be diluted with 0.1% peptone, and survivors will be enumerated after plating on TSA and incubating at 37 °C for 24 h. Since the antimicrobial activity is attributed to quaternary ammonium groups, their surface positive charges will not be affected by the metal/dye adsorption; and thus, we expect that the antimicrobial activity of beads will not be altered after regeneration.OBJECTIVE 3. Validate the efficacy of beads in a prototype filtration system.After optimizing the preparation and modifications of porous beads, we will design a lab-scale prototype to validate the efficacy of beads to control the quality of contaminated water in a filtration system with continuous flow. The contaminated water will contain a mix of chemical pollutants and bacteria, representing the complexity of chemical and biological species in a real scenario. Through a peristaltic pump, various flow rates (0.1, 0.2, and 0.5 mL/min) will be applied to pump wastewater into the filtration column filled with beads. The filtration device will be designed by filling wet beads into minicartridge columns (5-25 mL volume) with Luer-lock connections. One end will be attached to a sterile PVC pipe connecting a pump as the wastewater inlet, and the other end will be the outlet to collect treated water, which will be sampled to determine chemical and biological pollutants.Filtration capacity will be estimated by drawing a breakthrough capacity curve, which will help for the future design of a larger scale filtration unit in a future pilot study.

Progress 02/15/21 to 02/14/24

Outputs
Target Audience:One PhD student worked on this project during its lifetime and received comprehensive training on technical skills and fundamental knowledge. One undergraduate student was also recruited to assist the PhD student during the second and third year of this project. In particular, the undergraduate student engaged in research activities and learned a wide range of skills during the past two years. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Two Ph.D. students and one undergraduate student received comprehensive training from this project during the last year, and interdisciplinary collaboration between food science and chemistry programs have led to conference presentations and journal article publications by the trained students. How have the results been disseminated to communities of interest?During the lifetime of this project, the research findings were presented to multiple conferences, including international, national, and regional events. We have disseminated the research findings to a broader audience worldwide. One of the publications is published as an open access source to ensure reaching the broader impact worldwide. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Chitosan(CS)-based double-network antibacterial hydrogels were developed, and the hydrogels exhibitedoutstanding mechanical and antibacterial properties. The composite hydrogels had excellent antibacterial efficacy against Gram-positive (Listeria monocytogenes) and Gram-negative (Escherichia coli) bacteria, mainly attributed to the presence of quaternary ammonium groups in the modifiedpolymers. The major objective of this project has been achieved. The filtration prototype system was tested in the lab for potential future scale-up applications.

Publications

  • Type: Journal Articles Status: Published Year Published: 2024 Citation: Zhu, H., Yang, T., Chen, S., Wang, X., He, J., & Luo, Y. (2023). Construction and characterization of chitosan/poly (acrylamide-[2-(methacryloyloxy) ethyl] trimethylammonium chloride) double-network hydrogel with enhanced antibacterial activity. Advanced Composites and Hybrid Materials, 6(6), 192.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2023 Citation: Characterization of P(MTAC-AAm)/Chitosan Composite Hydrogels: Rheology, Texture, Antibacterial Activity and Cytotoxicity. Annual Meeting of International Associate of Food Protection, Toronto, Canada, 2023, July 16-19.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2023 Citation: Composite hydrogel for wastewater treatment to improve food safety. Graduate Research Forum, University of Connecticut, 2023, April.


Progress 02/15/22 to 02/14/23

Outputs
Target Audience:The target audiences rearched this reporting period included students, researchers, scientists, as well as professionals from the food and agricultural industry. We presented our data to a few international conferences where the professionals from both academia and industry were gathered to learn about our research findings. One research paper and one review article have been published during this reporting period to disseminate our findings to a broader audience for a greater impact. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Two PhD students received significant training for heavy metals removal and microbiological research. In particular, one PhD student from Nutritional Sciences and one PhD student from Chemistry (CoPI's lab) are closely collaborating to achieve the reserach goals in this project. One undergraduate was also recruited in the academic semesters to assist the two graduate students to conduct experiments in both departments. How have the results been disseminated to communities of interest?Yes, we have published a total of two journal articles, one research paper and a review paper. In the research paper, we reported our major research findings onthe efficacy of dyes removal. In the review paper, we comprehensively introduced and summarized the mechanisms (mode of action) of chitosan-based hydrogels on removing chemical dyes and adsorbing heavy metals. The review paper is published in an open-access journal, which we envision will be widely disseminated among the scientific community. What do you plan to do during the next reporting period to accomplish the goals?During the next reporting period, we will primarily focus on two major improvements: (1) modification of chitosan to prepare high functional hydrogels that can simultaneously remove chemical dyes and adsorb heave metal ions, as well as kill pathogenic microorganisms. (2) We also plan to further investigate the antimicrobial activity at the molecular level.

Impacts
What was accomplished under these goals? We have revised our hydrogel formulation and included carboxymethyl cellulose in order to improve the textural and mechanical properties as well as further enhance the adsorption capability. A benign chemical crosslinker, epichlorohydrin, was introduced to strengthen the polymeric network. With the new formulation, polyethylene glycol (PEG) as a pore-forming agent, has played an important role in further enhancing the adsorption capability of hydrogelsfor anionic (Congo red, CR) and cationic (methylene blue, MB) dyes removal from aqueous solutions. We also systematically studied the adsorption mechanism using mathematical models under various conditions. We shed light on different models for different dyes by as-prepared hydrogels, i.e.,the pseudo-second-order for CR and pseudo-first-order kinetics model for MB. This will lay the scientific foundation for future design of hydrogels targeting different industrial dyes. The hydrogels also revealed strong antimicrobial activities against both Gram-negative and positive bacteria, and thestudies on their antimicrobial mode of action are stillundergoing.

Publications

  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Zhu, H., Chen, S., Duan, H., He, J., & Luo, Y. (2023). Removal of anionic and cationic dyes using porous chitosan/carboxymethyl cellulose-PEG hydrogels: Optimization, adsorption kinetics, isotherm and thermodynamics studies. International Journal of Biological Macromolecules, 231, 123213.
  • Type: Journal Articles Status: Accepted Year Published: 2023 Citation: Zhu, H., Chen, S., & Luo, Y. (2023). Adsorption mechanisms of hydrogels for heavy metal and organic dyes removal: A short review. Journal of Agriculture and Food Research, 100552.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Zhu H, Luo Y. Facile preparation of carboxymethyl cellulose-chitosan composite hydrogel using PEG as pore-forming agent for enhanced anionic and cationic dyes removal. Annual Meeting of Society of Food Engineering, Raleigh, North Carolina, 9/18-21, 2022.


Progress 02/15/21 to 02/14/22

Outputs
Target Audience:The target audiences researched this reporting period included students, researchers, scientists, as well as professionals from the food and agricultural industry. One book chapter has been completed and is currently revision for production. Changes/Problems:We did not have any major problems in fabricating the porous chitosan hydrogels during the first year, and we will continue with chemical modification to functionalize the hydrogels with potent antimicrobial activities. What opportunities for training and professional development has the project provided?One PhD student was recruited last year to focus on this project. Comprehensive trainings have been provided to the student to conduct the experiment with the state-of-the-art facilities, such as scanning electron microscopy,Fourier Transform Infrared (FTIR) spectroscopy, UV-Vis spectroscopy, as well as the trainings on comprehensive statistical and analytical software was accomplished. One undergraduate student was also recruited as a research assistant to work with the Ph.D. student together on this project, and similar trainings were also provided to the undergraduate student. How have the results been disseminated to communities of interest?A book chapter was written and is currently under revision for publication and production. We are also working on a research paper and will submit the paper to a peer-reviewed journal later this year for publication. A poster presentation was accepted to the Annual Conference of the Society of Food Engineering in October 2022. What do you plan to do during the next reporting period to accomplish the goals?We are now working on the evaluation of chemical modificationto functionalizeas-prepared hydrogels with strong antimicrobial activities. We also anticipate completing the antimicrobial experiments during the next reporting period.

Impacts
What was accomplished under these goals? In the past year, we have completed the preparation of chitosan (CS)/carboxymethyl cellulose (CMC) hydrogelsby using a chemical cross-linking method, with polyethylene (PEG) as a pore-forming agent. Hydrogels made with two biopolymers at various mass ratios were optimized for their swelling properties under both acidic and alkaline conditions where interestingly, the CS2/CMC2 hydrogel (2% (w/v) CS in the acetic acid/water; 2% (w/v) CMC in the water) had the highest swelling ratio of 315% and 265%, respectively. With the optimal PEGconcentration, the highest adsorption rate ofCS2/CMC2-PEG hydrogels were achieved at above 97% for Congo Red, and 63% for Methylene Blue. The surface and cross-section of CS2/CMC2-PEG hydrogels observed under scanning electron microscopy (SEM) appeared more porous before adsorption and more compact with disappearance of porosity after adsorption. The results of Fourier Transform Infrared (FTIR) showed the CS2/CMC2-PEG hydrogel peak was narrowed and enhanced and shifted to a lower frequency of 3292 cm-1, suggesting the hydrogen bonds between CS and CMC were reduced, which could also mean that -OH from CS and CMC participated in the reaction. What's more, the PEG could be washed entirely, reflected by no obvious difference observed between the spectra of CS2/CMC2 and CS2/CMC2-PEG hydrogels. Besides, the CS2/CMC2-PEG hydrogels exhibited stable adsorption capacity after five adsorption/desorption cycles.

Publications