COFUND-BLUEBIO-BIOSHELL“Recycling crustaceans shell

wastes for developing biodegradable wastewater cleaning composites – BIOSHELL”

Project Partners and Institutions:

Project Director:

CS I. Dr. Eng. RADU Anita-Laura, Advanced Polymer Materials and Polymer Recycling Group, Polymers Department,

Coordinator:

National Research & Development Institute for Chemistry and Petrochemistry-ICECHIM, Bucharest, Romania (www.icechim.ro)

Project Responsible:

CS III. Dr. biochim. Andreea Gabriela OLARU, Director of the R&D Dept. and Quality Dept., S.C. EDAS-EXIM S.R.L.

Prof. Dr. Artur Jose Monteiro VALENTE, University of Coimbra, Faculty of Sciences and Technology – Department of Chemistry,

Dr. Alexandre Cabral CRAVEIRO, Brinova Bioquimica Lda.,

Prof.Dr.eng. Lisa PARUCH, Professor at Environment and Natural Resources Division,

Partners:·        

  • S.C. EDAS-EXIM.SRL, Bucharest, Romania, http://www.edas.ro·        
  • University of Coimbra, Faculty of Science and Technology, Coimbra,
  • Portugal, http://www.uc.pt·  Brinova Bioquimica Lda., Évora, Portugalia, http://www.brinova.eu·        
  • Norwegian Institute of Bioeconomy Research –NIBIO, Division of Environment and Natural Resources As, Norway, http://www.nibio.no

National Funding Agencies

Unitatea executiva pentru finantarea invatamantului superior, a cercetarii, dezvoltarii  si inovarii (UEFISCDI): https://www.uefiscdi.ro/

UEFISCDI and CE: 250 000 €

Co-financing amount from EDAS-EXIM. SRL 30.000 €

Fundação para a Ciência e a Tecnologia (FCT): http://www.fct.pt/

FTC and CE: 99.431 €

Co-financing amount from University of Coimbra: 88.122 €

Co-financing amount from BRINOVA Lda: 84.000 €

Duration of project: 36 months (start date: 10 April 2020)

Abstract

Wastes from agriculture and fishery cause harmful effects on the environment and implicitly on humans. But, many of these wastes can berecycled. One of the current global issues refers to minimizing waste production, effective wastewater treatment, biosafe food production,and reducing hazards from the exposure to pathogens. Most of the threatening microorganisms especially emerging pathogens (EPs) derivefrom wastewater. Moreover, antibiotics residues present in wastewater lead bacterial pathogens to develop antibiotic resistance genes(ARGs). In addition, heavy metals are among the most harmful non-microbial pollutants due to their toxicity to humans. BIOSHELL aims atsynergistically solving economic, environmental and health problems. The project focuses on utilizing the wastes from sea foodpreparation such as crustacean carcasses in the development of innovative and efficient inorganic-organic functionalized hydrogelnanocomposites, suitable to facilitate the sustainable wastewater purification technologies about heavy metals retention, antibioticselimination, EPs and ARGs removal.

Objective

Functional biopolymer-based hydrogels starting from valorized crustacean’s shell wasteswill be developed both for the metal and antibiotics retention in waters as well as for anti-bacterial treatment. These competitive materialswill be ion imprinted polymers (IIPs) or molecularly imprinted polymers (MIPs). They will benefit from new synthesis methodologies appliedfor chelating the chitosan nanocomposites and for the chemical grafting of the bactericidal hybrid surfaces. The development of newapproaches for the valorization of crustacean wastes, by the new functionalized biohydrogels, will improve the on-site wastewater treatmentin EU. The regeneration of new bio-based agents is also targeted.

Estimated Results

Consortium Results:

(i) 6 scientific ISI papers;

(ii) 2 patent applications;

(iii) 6 communications at prestigious Symposia ;

(iv) 3 organized workshops;

(v) 1 participation to Invention Salons;

(vi) project website.

STAGE I. Studies for obtaining raw chitosan and chitosan-based materials

PERIOD: 10.04.2020-31.12.2020

OBJECTIVES

Act 1.1 – Obtaining chitosan starting from commercial chitin and obtaining crude chitosan starting from chitin extracted from preconditioned waste of crustaceans. Results: Experimental model for obtaining chitosan.

Act 1.2– Studies on obtaining MIP pearls based on chitosan (via commercial chitin) for the treatment of antibiotics. Results: Concept for the MIP pearls.

Act 1.3 – Studies on the synthesis of antibacterial hydrogels based on chitosan (via commercial chitin) and quaternary ammonium salts. Results: Concept for the antibacterial hydrogel.

Act 1.4 – Characterization of raw materials, intermediates and final materials. Results: 3 Characterization reports.

Act 1.5 – Communicating the existing results of the project partners by organizing a kick-off meeting at the headquarters of the Project Coordinator. Results: Report of dissemination, communication and travel.

SUMMARY

The main objective of the research activity was to perform studies of obtaining crude chitosan and chitosan-based materials, starting from commercial chitin, and tracking the effects on the final properties. In this respect, the following secondary objectives have been established:

OS1. Obtaining chitosan from commercial chitin and obtaining crude chitosan from chitin extracted from preconditioned shellfish waste. Physico-chemical

characterization of raw materials, intermediates and final materials.

OS2. Studies on obtaining MIP beads based on chitosan (via commercial chitin) for antibiotic retention. Physico-chemical characterization of raw materials, intermediates and final materials.

OS3. Studies regarding the synthesis of antibacterial hydrogels based on chitosan (via commercial chitin) and quaternary ammonium salts. Physico-chemical and

bacteriological  characterization of raw materials, intermediates and final materials.

OS1. In order to obtain chitosan from commercial chitin, two synthesis methods were used in which the percentage of NaOH was varied. To obtain the chitosan, the deacetylation process involved the homogenization of chitin with NaOH in a 250 mL round-bottomed flask. The suspension was refluxed under stirring and the obtained chitosan was washed until the pH of the filtered product was neutral. Laboratory synthesized chitosan samples were compared with commercial chitosan (Ccom) of medium molecular weight, in which case some similarities were found in both FTIR and TGA analysis. In fact, the results were compared with a sample of commercial chitin that was used as raw material in the synthesis of chitosan.

However, the Micro-CT images and the degrees of deacetylation obtained from the NMR analysis for the synthesized chitosan were different compared to the commercial chitosan sample. The chitosan synthesized in the laboratory indicated a degree of deacetylation of over 86% compared to the commercial one of 75% and the

degree of compaction of the synthesized chitosan was lower, the proof being the Micro-CT analysis which indicated towards a material with a more increased intrinsic porosity than that of the commercial reference.

OS2. The study on obtaining MIP pearls based on chitosan allowed to obtain information regarding the production of hybrid pearls or granules with applications in the retention of antibiotics from wastewaters. In this regard, molecularly imprinted cryogels based on chitosan and biocellulose were obtained for penicillin retention. Thus, molecularly imprinted and non-imprinted cryogels based on chitosan were obtained (4 series of NIP/MIP pairs).

The study of swelling degrees showed that all cryogels adsorb a lot of water, but also the fact that some of them fragment due to low mechanical strength (gels being physically crosslinked with ammonium acid carbonate). UV-Vis analysis allowed to highlight the washing/extraction degree of penicillin for the obtained cryogels. In fact, it also proved that the MIP sample re-adsorbed 5.24 more specifically penicillin than the reference cryogel NIP.

Thermal analysis also showed that imprinting took place for all MIP series and highlighted differences in stability between the samples obtained with commercial chitosan vs the one synthesized in the laboratory. In the next stage, the method of obtaining cryogels will be optimized in order to

obtain more stable materials from a mechanical point of view.

OS3. Obtaining new polymer networks with interpenetrated structure consisted in the polymerization of a monomer, generator of quaternized ammonium groups, in the presence of chitosan (commercial or synthesized). Hydrogels based on commercial/synthesized chitosan were characterized in terms of swelling degree. The results showed that the type of chitosan influenced the degree of swelling for each hydrogel. For the hydrogels obtained using a smaller amount of crosslinker, differences were observed between the values of maximum swelling degrees. This showed that the use of a higher concentration of crosslinker led to much more rigid polymer networks which prevented the absorption of a larger amount of water.

Regarding the commercial/synthesized chitosan-based hydrogels, FTIR spectra showed the characteristic bands for chitosan. TGA/DTG analysis confirmed the presence of chitosan in the final structure of polymer networks.

Bactericidal tests reflected the potential of the synthesized materials, especially the chitosan-based hydrogel synthesized in the laboratory, to destroy both coliforms and clostridia in proportions of 83% and 69%, respectively.

DISSEMINATION

During stage 1/2020, a project kick-off meeting was organized with the physical participation of the project partners on 03.03.2020.

The Project Director participated in the BLUEBIO projects kick-off meeting (Online kick-off of the BlueBio ERA-NET COFUND 9 June 2020) and in a networking event (Connectivity among Blue Bioeconomy Cofund projects 20 November 2020).

An ISI indexed article was published and one was submitted for publication as follows:

1.Teodor Sandu, Maria Luiza Jecu, Iuliana Raut, Mariana Calin, Elvira Alexandrescu, Tanta Verona Iordache, Marinela Victoria Dumitru, Andrei Sarbu, Hybrid Beads Bearing Immobilized Bacteria As Advanced Means For The Removal Of Acid Blue 93 Dye, Materiale Plastice (Mater. Plast.),Year 2020, Volume 57, Issue 3.


2.Ana-Mihaela Gavrilă, Simona Nedelcu-Flor, Andrei Sârbu, Teodor Sandu, Andreea Gabriela Olaru, Gheorghe Hubca, Dan Donescu, Tanța-Verona Iordache, Synthesis And Properties Of Organosilica Particles With Quaternary Ammonium Bearings As Bacteriostatic Interfaces, Trimis spre publicare U.P.B. Sci. Bull., Series.