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Packaging accounts for more than 40% of all plastics produced every year and plastic accumulation in the environment has become a serious problem, thus the improvement of its circularity from origin to subsequent life cycles is now a priority. An effective and eco-friendly approach to solve such problem is the use of biodegradable materials, whose global production capacities are set to increase from around 1.1 million tonnes in 2022 to approximately 3.5 million tonnes in 2027. In particular, a new strategy for the management of plastics is based on the design of polymers that contain degrading enzymes in their formulation: the enzymes should be activated only at the end-of-life of the material. However, the harsh conditions used in most industrial plastic processes cause denaturation and loss of activity of enzymes, thus efficient protection strategies are necessary. In this way, thermally protected enzymes could be embedded in plastic formulation, obtaining novel “degradable on-demand” materials. In the present study, cutinase from Humicola Insolens, selected as a highly degrading polyester hydrolytic enzyme, is immobilized in Mg/Al layered double hydroxide structures. The immobilization efficiency results high and the enzyme thermal stability strongly improves after immobilization, since its half-life at 90°C increases by 6 times.

Thermally protected enzyme for degradable on-demand polymers Angela Romano, Antonella Rosato, Grazia Totaro, Giulio Zanaroli, Annamaria Celli, Laura Sisti Dipartimento di Ingegneria Civile, Chimica, Ambientale e dei Materiali, Università di Bologna, Via Terracini 28, 40131 Bologna, Italy. Plastic accumulation in the environment has become a serious problem, and adequate end-of-life strategies must be developed. The use of polymer-degrading enzymes can be an efficient method for the management of plastic waste: indeed, new “degradable on-demand” materials could be produced, by embedding them in plastic formulation. However, efficient protection strategies for the enzymes are necessary, since the high temperatures required for most industrial plastic processes cause their denaturation and loss of activity. A new, simple and successful strategy is here proposed. The study was carried out within the framework of the TERMINUS project, funded by the European Union under Horizon 2020 (Call: H2020-NMBP-ST-IND-2018, Grant Agreement: 814400), and it has been the topic of an oral presentation during the International conference “Bio-based polymers at the forefront of innovation in materials science”, held in Bertinoro (Italy), April 12-14, 2023., {"references":["Bioplastics market data. https://www.european-bioplastics.org/market/. Accessed on 03/10/2023.","A. Rosato, A. Romano, G. Totaro, A. Celli, F. Fava, G. Zanaroli, L. Sisti \"Enzymatic degradation of the most common aliphatic bio-polyesters and evaluation of the mechanisms involved: an extended study.\" Polymers 2022, 14, 1850.","EEA European Environment Agency. \"Plastics, the circular economy and Europe's environment-A priority for action. Report.\" 2021."]}

The use of degrading enzymes in polymer formulation is a very attractive strategy to manage the end-of-life of plastics. However, high temperatures cause the denaturation of enzymes and the loss of their catalytic activity; therefore, protection strategies are necessary. Once protected, the enzyme needs to be released in appropriate media to exert its catalytic activity. A successful protection strategy involves the use of layered double hydroxides: cutinase, selected as a highly degrading polyester hydrolytic enzyme, is thermally protected by immobilization in Mg/Al layered double hydroxide structures. Different triggering media are here evaluated in order to find the best releasing conditions of cutinase from LDH. In detail, phosphate and citrate–phosphate buffers, potassium carbonate, sodium chloride, and sodium sulfate solutions are studied. After the comparison of all media in terms of protein release and activity retained, phosphate buffer is selected as the best candidate for the release of cutinase from LDH, and the effect of pH and concentration is also evaluated. The amount of the enzyme released is determined with the Lowry method. Activity tests are performed via spectrophotometry.

The use of polymer-degrading enzymes in polymer formulation is a very attractive strategy for the life-cycle of plastics. The design of a material able to degrade on-demand could help reach the ambitious targets pursued by recent European policies. Only 32% of the resulting waste from the 50 million tonnes of plastics consumed each year in Europe is actually recycled. Since packaging accounts for more than 40% of all plastics produced every year, the improvement of its circularity from origin to subsequent life cycles is now a priority. The present research suggests a solution to improve the recyclability of plastics via a life cycle approach employing thermally stable enzymes as innovative materials providing a new potential for plastic and its end life. More specifically, cutinase, selected as a highly degrading polyester hydrolytic enzyme, was thermally protected by immobilization in Mg/Al layered double hydroxide structures. The cutinase immobilization efficiency was found to be high, as well as its release ability in an appropriate medium. The thermal stability of cutinase was strongly improved after immobilization, as highlighted by a 6-times increase of its half-life at 90 °C, compared to the free enzyme, and by a high activity retention (>60%) after short exposure to temperatures up to 200 °C. Moreover, it was demonstrated that a film of poly(butylene succinate-co-adipate) formulated with 5 wt% of immobilized cutinase, completely degraded within 24 h.

The constant increase of the plastic production over the world has become a serious problem, since most conventional plastic materials come from fossil resources and are not biodegradable. This causes significant plastics accumulation in the environment, whose end-life must be managed. An effective and eco-friendly approach to solve such problem is the use of biodegradable materials. Biopolymers such as poly(butylene succinate) (PBS), poly(butylene succinate-co-adipate) (PBSA), poly(caprolactone) (PCL), poly(lactic acid) (PLA) and poly(propylene carbonate) (PPC) are among the most promising biodegradable commercial polyesters thanks to their high susceptibility to hydrolytic enzymes and to many microorganisms naturally occurring in the environment [1-2]. The current study is an investigation of the degradation ability of several hydrolytic enzymes belonging to different subclasses (i.e. lipase, esterase, proteinase, etc.) against some common aliphatic commercial polyesters. A deeper elucidation on the degradative ability and the mechanism of hydrolytic biodegradation can be useful for the management of biodegradable plastic wastes, the bioremediation of plastic-polluted environments, as well as the design of innovative biodegradable plastic materials. The end-of-life of biopolymers indeed, should also be addressed in view of a circular economy concept. The enzyme screening was first carried out by investigating the capacity of fully degrading the target polymers in 24 h, then weight loss measurements of selected polyesters and target enzymes were performed. Solid residues after enzyme degradation were characterized by proton nuclear magnetic resonance (1H NMR), gel permeation chromatography (GPC), infrared spectroscopy (FT-IR), differential scanning calorimetry (DSC) and thermogravimetry (TGA). Liquid fractions were studied via GPC, 1H NMR and high-performance liquid chromatography (HPLC). This, in order to understand molecular and chemical modifications induced at the surface and/or in the bulk of polymer materials over time. The results showed that the enzymatic degradation occurred homogenously from the surface through an erosion mechanism and involved both the amorphous and the crystalline regions. Cleaving action mode for each enzyme (endo- and/or exo-type) is also proposed.

The production and use of plastics have been growing in the past decades. Most of these plastics are fossil fuel-based and non-degradable, thus they accumulate in the environment causing severe problems. The use of plastic-degrading enzymes in the treatment of plastic waste, thus the use of biodegradable polymers has gained growing attention in different fields, including packaging, agriculture and medicine. Aliphatic polyesters are the most promising biodegradable plastics because of their high susceptibility to the attack of hydrolytic enzymes and of many microorganisms naturally occurring in the environment. Among them, poly(ε-caprolactone) (PCL) is a semicrystalline aliphatic polyester, widely used in biomedical and packaging applications, because of its biocompatibility and biodegradability. This study investigates the degradation mechanism of two commercial PCL with different molecular weights (150*103 and 215*103 g/mol) by a commercial lipase from Candida sp. (CALB). The degradation mechanism is studied through weight loss measurements of polyester films combined to other analyses (i.e., GPC, 1H NMR and DSC) performed on the residual solids after incubation with the enzyme, in order to understand molecular and chemical modifications induced by lipase. Moreover, a cleaving action mode for the enzyme (endo- and/or exo-type) on both PCL was also proposed based on the identification and quantification, via GPC, 1H NMR and HPLC-RID analyses, of residual oligomers and monomers released as degradation products in the liquid fraction. Results showed that lipase CALB degrade the polymers homogeneously from the surface through an erosion mechanism, with an exo-type cleaving action mode, and the degradation rate is slower with a higher molecular weight.

Aliphatic polyesters are the most promising biodegradable plastics because of their high susceptibility to the attack of hydrolytic enzymes and of many microorganisms naturally occurring in the environment. Poly(butylene succinate-co-adipate) (PBSA) is a biobased semicrystalline copolymer of poly(butylene succinate) (PBS), with adipic acid as co-monomer. It is highly biodegradable because of its lower crystallinity and higher flexibility of polymer chains compared to PBS. In the present study, thebiodegradability mechanism of PBSA was evaluated using cutinase from Humicola insolens, which is a commercial hydrolytic enzyme presenting both lipase and esterase features. The polyesterdegradation is based on weight loss measurementscombined to other analyses (i.e., GPC, 1H NMR, and DSC) performed on the residual solids of PBSAfilms, in order to understand the molecular and chemical modifications induced at the surface and/or in the bulk of polymer materials over time. The results showed that the enzymatic degradation mechanism of cutinase proceedby degrading the polymer from the surface of the film. An exo-type cleaving action mode was proposed based on the identification of the oligomers and monomers released as degradation products.

Currently, multilayer packaging is widely used for the preservation and distribution of food, beverages, pharmaceuticals and other consumable products. However, if packaging materials are excellent in preserving, protecting and extending the useful life of the packaged goods, on the other hand their recycling is very demanding because of multimaterial layers, which cannot be simply separated and reused. Actually the majority of such packaging is either incinerated or bound for landfills. The European project H2020 TERMINUS [1] aims at designing a smart multilayer packaging able to self-delaminate at the end of its life, thus enabling the recycling of the different layers. The delamination strategy involves the use of protected enzymes within the multilayer formulation. After a specific trigger, the enzyme is released and degrades the adhesive anchoring the layers, causing debonding, thus layers can be separated and recycled. In detail, in the present study some enzymes with high degrading activity against some polymeric adhesives were protected, in order to increase their thermal stability and allow their use during the processing of the multilayer. With the successful implementation of TERMINUS project, the expected results will lead to a 15% of improved economy efficiency, 80% reduction of the landfilling rates for multilayer packaging as well as 50% decrease of overall landfilling rates of plastic, along with a 65% decrease in carbon dioxide emissions. [1] TERMINUS has received funding from the European Union’s Horizon 2020 research and innovation programme, under grant agreement number 814400.

One of the main limitations to the bioremediation of oil-contaminated marine sediments is the low hydrocarbons bioavailability. Aim of this PhD research project is to identify an environmental friendly approach to increase hydrocarbons bioavailability and biodegradation in oil-contaminated marine sediments. Several surfactants/pollutant mobilizing agents have been selected and applied to slurry microcosms: two microbial surfactants (sophorolipids and rhamnolipids), two types of cyclodextrins (hydroxypropyl- and randomly methylated- β-cyclodextrins; HPB-CD and RAMEB-CD), two commercial soy lecithin products (de-oiled and raw) and bile acids. Sophorolipids, cyclodextrins and to less extent, soy lecithins stimulated n-alkanes anaerobic degradation of actual oil-contaminated marine sediment from Gela (Sicily). Molecular analysis of 16S rRNA gene suggested that an Acidobacteria was probably responsible for the anaerobic biodegradation. Under aerobic condition, the same surfactants had inhibitor effects on n-alkanes degradation in Gela sediment, while HPB-CD and de-oiled soy lecithin were able to increase the degradation in crude oil-contaminated sediment from Ravenna. Increase of n-alkane bioavailability in oil-contaminated Gela sediment occurred in the presence of the two cyclodextrins and raw soy lecithin, both immediately after oil contamination as well as after the complete adsorption of hydrocarbons to sediment. Investigations of surfactant releasing formulations for the deployment of these agents to the sediments showed that HPB-CD could be efficiently encapsulated in agar hydrogels, while sophorolipids in polybutylene succinate (PBS) microspheres. The release rate of encapsulated HPB-CD was higher than encapsulated sophorolipids when formulations were incubated in marine water and similar in oil-contaminated sand slurries. Both encapsulated surfactants remarkably reduced adsorption of freshly spiked hydrocarbons to sand; conversely, only agar-encapsulated HPB-CD were able to desorb n-alkanes in weathered contaminated sand and increase their bioavailability and biodegradation similarly to free cyclodextrins. Therefore, encapsulation of HPB-CD in agar capsules might be the most promising solution for the enhancement of the bioremediation in marine sediment.

The aim of this study was to develop a packed bed reactor (PBR) process for the on-site aerobic cometabolic bioremediation of a groundwater contaminated by trichloroethylene (TCE) and 1,1,2,2-tetrachloroethane (TeCA). With regard to the growth substrate selection, the comparison of the CAH degradation performances obtained with 5 candidate substrates led to the selection of butane and to the development from the site’s indigenous biomass of a suspended-cell consortium capable to degrade TCE (TCE first-order constant = 96 L gprotein-1 d-1 at 30 °C and 4.3 L gprotein-1 d-1 at 15 °C) with a 90% mineralization of the organic Cl. Based on PCR-DGGE analysis of the 16S rRNA genes followed by band excision and sequencing, the microbial consortium enriched was mainly composed of Bacteroidetes and Alpha- and Beta-Proteobacteria that were distantly related to known CAH-cometabolizing bacteria. The low TeCA degradation rate by the developed consortium suggested the introduction of a chemical pre-treatment based on the TeCA to TCE conversion via -elimination, a very fast reaction at alkaline pH. The choice of the best-performing carrier was made by means of a 2-level procedure: the 1st level consisted of batch tests, operated both at 30 and 15 °C, whereas the 2nd level consisted of continuous-flow tests, operated at 30 °C. A porous ceramic carrier named Biomax resulted the best-performing one. On the basis of the kinetic parameters relative to TCE aerobic cometabolic biodegradation by the selected consortium, a 31-L packed-bed reactor, consisting of 14 columns connected in series, packed with Biomax and immerged in a temperature-controlled bath, was designed and set-up. The preliminary data, obtained by operating the plant with a 4-day residence time, indicate the attainment of a 0.1 mg L-1 d-1 TCE biodegradation rate.

A procedure for the design of a packed bed reactor (PBR) process for the on-site aerobic cometabolic bioremediation of a groundwater contaminated by trichloroethylene (TCE) and 1,1,2,2-tetrachloroethane (TeCA) was developed. The comparison of the CAH degradation performances obtained with five substrates led to the selection of butane and to the development from the site’s indigenous biomass of a consortium capable to degrade TCE, with a 90% mineralization of the organic Cl. A porous ceramic carrier (Biomax) resulted the best option for the PBR process. On the basis of the kinetic analysis, a 31-L PBR, consisting of 14 columns connected in series, was designed.

Chlorinated aliphatic hydrocarbons (CAHs) are widespread groundwater contaminants. Aerobic cometabolism, that requires the supply of a suitable growth substrate, represents an interesting option for the remediation of CAH-contaminated aquifers, thanks to its capability to lead to the complete mineralization of a very wide range of CAHs. The aim of this study was to develop and validate a procedure relative to the lab-scale tests required to obtain the essential information for the design of a process of CAH aerobic cometabolism in a packed bed reactor (PBR). To validate the procedure, the latter was applied to the development of a PBR process for the on-site aerobic cometabolism of an aquifer contaminated by trichloroethylene (TCE) and 1,1,2,2-tetrachloroethane (TeCA). The specific goals of this study, corresponding to the main steps of the above-mentioned procedure, were: (i) to select the best growth substrate for the aerobic cometabolic process, and to develop and characterize an effective CAH-degrading microbial consortium, obtained from the site’s indigenous biomass by exposition to the selected substrate; (ii) to select the best carrier for the PBR process, and to evaluate the effect of bacterial adhesion on the developed suspended-cell consortium; and (iii) to identify and test suitable chemical-physical remediation alternatives in the case of presence of CAHs poorly biodegradable through AC. With regard to the growth substrate selection, the comparison of the CAH degradation performances obtained with 5 candidate substrates (methane, propane, butane, pentane and phenol) led to the selection of butane and to the development from the site’s indigenous biomass of a suspended-cell consortium capable to degrade TCE (TCE first-order constant (k1,TCE) = 96 L gprotein-1 d-1 at 30 °C and 4.3 L gprotein-1 d-1 at 15 °C) with a 90% mineralization of the organic Cl. Based on PCR-DGGE analysis of the 16S rRNA genes followed by band excision and sequencing, the microbial consortium enriched was mainly composed of Bacteroidetes and Alpha- and Beta-Proteobacteria that were distantly related to known CAH-cometabolizing bacteria. With regard to the selection of the best-performing biofilm carrier, a preliminary screening based on the previous experience of the research group led to the pre-selection of four candidate biofilm carriers (porous materials specifically designed for biofilm processes): Biomax, Biomech, Biopearl and Cerambios. The choice of the best-performing carrier was made by means of a 2-level procedure. The 1st level consisted of batch tests, operated both at 30 and 15 °C, whereas the 2nd level consisted of continuous-flow tests, operated at 30 °C. The 30 °C continuous-flow tests were conducted in four 1 L packed columns, connected to a feeding system designed so as to attain a pulsed feed of both oxygen and the selected growth substrate (butane). The four columns were operated in continuous mode for about 100 days. The results of the attached-cell tests were compared on the basis of the TCE normalized degradation rate ( and ) and of the attached cell concentration attained, at the two temperatures, at the end of the biofilm development process. On the basis of both the batch and the continuous-flow tests, Biomax resulted the best-performing biofilm carrier. Biomass immobilization on the carrier changed remarkably the structure of the microbial consortium. The effect on k1,TCE of biomass attachment depended on temperature: at 15 °C the attached consortium performed slightly better than the suspended one, whereas at 30 °C an opposite trend was noticed. On the basis of a 1st-order simulation, a 99.9% TCE conversion can be attained, at the site’s temperature, with a 9-hour HRT. Lastly, the low TeCA degradation rate by the developed consortium suggested the introduction of a chemical pre-treatment based on the TeCA to TCE conversion via -elimination, a very fast reaction at alkaline pH. On the basis of the overall results, the procedure for the development of a PBR AC process appears to be correctly designed and generally applicable to CAH contaminated sites.

The constant increase of the plastic production over the world has become a serious problem, since most conventional plastic materials come from fossil resources and are not biodegradable. This causes significant plastics accumulation in the environment, whose end-life must be managed. An effective and eco-friendly approach to solve such problem is the use of biodegradable materials. Biopolymers such as poly(butylene succinate) (PBS), poly(butylene succinate-co-adipate) (PBSA), poly(caprolactone) (PCL), poly(lactic acid) (PLA) and poly(propylene carbonate) (PPC) are among the most promising biodegradable commercial polyesters thanks to their high susceptibility to hydrolytic enzymes and to many microorganisms naturally occurring in the environment. The current study is an investigation of the degradation ability of several hydrolytic enzymes (i.e. lipase, esterase, etc.) against some common aliphatic commercial polyesters. Thedegradation is studied through preliminary film opacity assay, then weight loss measurements were combined to other analyses (i.e., GPC, 1H NMR, FT-IR, DSC and TGA) performed on the residual solids of polyester films after treatment with the different enzymes, in order to understand molecular and chemical modifications induced at the surface and/or in the bulk of polymer materials over time.The results showed that the enzymatic degradation occurred homogenously from the surface through an erosion mechanism and involved both the amorphous and the crystalline regions. Cleaving action mode for each enzyme (endo- and/or exo-type) is also proposed.&nbsp

Currently, multilayer packaging is widely used for the preservation and distribution of food, beverages, pharmaceuticals and other consumable products. However, if packaging materials are excellent in preserving, protecting and extending the useful life of the packaged goods, on the other hand their recycling is very demanding because of multimaterial layers, which cannot be simply separated and reused. Actually the majority of such packaging is either incinerated or bound for landfills. The European project H2020 TERMINUS [1] aims at designing a smart multilayer packaging able to self-delaminate at the end of its life, thus enabling the recycling of the different layers. The delamination strategy involves the use of protected enzymes within the multilayer formulation. After a specific trigger, the enzyme is released and degrades the adhesive anchoring the layers, causing debonding, thus layers can be separated and recycled. In detail, in the present study some enzymes with high degrading activity against some polymeric adhesives were protected, in order to increase their thermal stability and allow their use during the processing of the multilayer. With the successful implementation of TERMINUS project, the expected results will lead to a 15% of improved economy efficiency, 80% reduction of the landfilling rates for multilayer packaging as well as 50% decrease of overall landfilling rates of plastic, along with a 65% decrease in carbon dioxide emissions. [1] TERMINUS has received funding from the European Union’s Horizon 2020 research and innovation programme, under grant agreement number 814400.

The production and use of plastics have been growing in the past decades. Most of these plastics are fossil fuel-based and non-degradable, thus they accumulate in the environment causing severe problems. The use of plastic-degrading enzymes in the treatment of plastic waste, thus the use of biodegradable polymers has gained growing attention in different fields, including packaging, agriculture and medicine. Aliphatic polyesters are the most promising biodegradable plastics because of their high susceptibility to the attack of hydrolytic enzymes and of many microorganisms naturally occurring in the environment. Among them, poly(ε-caprolactone) (PCL) is a semicrystalline aliphatic polyester, widely used in biomedical and packaging applications, because of its biocompatibility and biodegradability. This study investigates the degradation mechanism of two commercial PCL with different molecular weights (150*103 and 215*103 g/mol) by a commercial lipase from Candida sp. (CALB). The degradation mechanism is studied through weight loss measurements of polyester films combined to other analyses (i.e., GPC, 1H NMR and DSC) performed on the residual solids after incubation with the enzyme, in order to understand molecular and chemical modifications induced by lipase. Moreover, a cleaving action mode for the enzyme (endo- and/or exo-type) on both PCL was also proposed based on the identification and quantification, via GPC, 1H NMR and HPLC-RID analyses, of residual oligomers and monomers released as degradation products in the liquid fraction. Results showed that lipase CALB degrade the polymers homogeneously from the surface through an erosion mechanism, with an exo-type cleaving action mode, and the degradation rate is slower with a higher molecular weight.

La produzione e il consumo di plastica sono largamente cresciuti negli ultimi decenni. La maggior parte di queste plastiche derivano da fonti fossili e non sono degradabili, per cui tendono ad accumularsi nell���ambiente. L���uso di enzimi e di plastiche biodegradabili per il trattamento di rifiuti plastici si sta sviluppando in diversi settori, quali packaging, agricoltura e medicina. Tra le plastiche biodegradabili, i poliesteri alifatici risultano essere i pi�� suscettibili all���attacco di enzimi idrolitici e microrganismi. Questo studio indaga la degradazione enzimatica di due polimeri commerciali biodegradabili, i.e. poli(butilene succinato-co-adipato) (PBSA) e poli(��-caprolattone) (PCL) e il meccanismo d���azione degli enzimi cutinasi da Fusarium solani e lipasi da Alcaligene sp. (QLM). Il PBSA �� un poliestere alifatico che deriva da fonti rinnovabili. Si tratta di un copolimero semicristallino del poli(butilene succinato) (PBS), con l���acido adipico come co-monomero. L���alta biodegradabilit�� deriva dalla bassa cristallinit�� e dalla maggiore flessibilit�� delle catene polimeriche rispetto al PBS. Il PCL �� un poliestere alifatico semicristallino che deriva da fonti fossili. �� largamente utilizzato nel settore biomedico e nel packaging per via della sua biodegradabilit�� e biocompatibilit��. Lo studio del meccanismo di degradazione enzimatica di questi due polimeri pu�� essere utile nella gestione dei rifiuti plastici, e nella formulazione di nuovi materiali plastici biodegradabili., {"references":["Liu, M., Zhang, T., Long, L., Zhang, R., & Ding, S. (2019). Efficient enzymatic degradation of poly (ɛ-caprolactone) by an engineered bifunctional lipase-cutinase. Polymer Degradation and Stability, 160, 120-125.","Lim, H. A., Raku, T., & Tokiwa, Y. (2005). Hydrolysis of polyesters by serine proteases. Biotechnology letters, 27(7), 459-464."]}

Currently, multilayer packaging is widely used for the preservation and distribution of food, beverages, pharmaceuticals and other consumable products. However, if packaging materials are excellent in preserving, protecting and extending the useful life of the packaged goods, on the other hand their recycling is very demanding because of multimaterial layers, which cannot be simply separated and reused. Actually the majority of such packaging is either incinerated or bound for landfills. The European project H2020 TERMINUS [1] aims at designing a smart multilayer packaging able to self-delaminate at the end of its life, thus enabling the recycling of the different layers. The delamination strategy involves the use of protected enzymes within the multilayer formulation. After a specific trigger, the enzyme is released and degrades the adhesive anchoring the layers, causing debonding, thus layers can be separated and recycled. In detail, in the present study some enzymes with high degrading activity against some polymeric adhesives were protected, in order to increase their thermal stability and allow their use during the processing of the multilayer. With the successful implementation of TERMINUS project, the expected results will lead to a 15% of improved economy efficiency, 80% reduction of the landfilling rates for multilayer packaging as well as 50% decrease of overall landfilling rates of plastic, along with a 65% decrease in carbon dioxide emissions. [1] TERMINUS has received funding from the European Union’s Horizon 2020 research and innovation programme, under grant agreement number 814400., poster for Ecomondo The Green Technology Expo

The constant increase of the plastic production over the world has become a serious problem, since most conventional plastic materials come from fossil resources and are not biodegradable. This causes significant plastics accumulation in the environment, whose end-life must be managed. An effective and eco-friendly approach to solve such problem is the use of biodegradable materials. Biopolymers such as poly(butylene succinate) (PBS), poly(butylene succinate-co-adipate) (PBSA), poly(caprolactone) (PCL), poly(lactic acid) (PLA) and poly(propylene carbonate) (PPC) are among the most promising biodegradable commercial polyesters thanks to their high susceptibility to hydrolytic enzymes and to many microorganisms naturally occurring in the environment. The current study is an investigation of the degradation ability of several hydrolytic enzymes (i.e. lipase, esterase, etc.) against some common aliphatic commercial polyesters. The degradation is studied through preliminary film opacity assay, then weight loss measurements were combined to other analyses (i.e., GPC, 1H NMR, FT-IR, DSC and TGA) performed on the residual solids of polyester films after treatment with the different enzymes, in order to understand molecular and chemical modifications induced at the surface and/or in the bulk of polymer materials over time. The results showed that the enzymatic degradation occurred homogenously from the surface through an erosion mechanism and involved both the amorphous and the crystalline regions. Cleaving action mode for each enzyme (endo- and/or exo-type) is also proposed., {"references":["Shah, A.A.; Kato, S.; Shintani, N. et al. Microbial degradation of aliphatic and aliphatic-aromatic co-polyesters. Appl Microbiol Biotechnol 98, 2014, 3437–3447","Azevedo, H. S.; & Reis, R. L. Understanding the enzymatic degradation of biodegradable polymers and strategies to control their degradation rate. Chapter 12 in Biodegradable systems in tissue engineering and regenerative medicine, 2005, 177-201"]}

The production and use of plastics have been growing in the past decades. Most of these plastics are fossil fuel-based and non-degradable, thus they accumulate in the environment causing severe problems. The use of plastic-degrading enzymes in the treatment of plastic waste, thus the use of biodegradable polymers has gained growing attention in different fields, including packaging, agriculture and medicine. Aliphatic polyesters are the most promising biodegradable plastics because of their high susceptibility to the attack of hydrolytic enzymes and of many microorganisms naturally occurring in the environment. Among them, poly(��-caprolactone) (PCL) is a semicrystalline aliphatic polyester, widely used in biomedical and packaging applications, because of its biocompatibility and biodegradability. This study investigates the degradation mechanism of two commercial PCL with different molecular weights (150*103 and 215*103 g/mol) by a commercial lipase from Candida sp. (CALB). The degradation mechanism is studied through weight loss measurements of polyester films combined to other analyses (i.e., GPC, 1H NMR and DSC) performed on the residual solids after incubation with the enzyme, in order to understand molecular and chemical modifications induced by lipase. Moreover, a cleaving action mode for the enzyme (endo- and/or exo-type) on both PCL was also proposed based on the identification and quantification, via GPC, 1H NMR and HPLC-RID analyses, of residual oligomers and monomers released as degradation products in the liquid fraction. Results showed that lipase CALB degrade the polymers homogeneously from the surface through an erosion mechanism, with an exo-type cleaving action mode, and the degradation rate is slower with a higher molecular weight., {"references":["Liu, M., Zhang, T., Long, L., Zhang, R., & Ding, S. (2019). Efficient enzymatic degradation of poly (ɛ-caprolactone) by an engineered bifunctional lipase-cutinase. Polymer Degradation and Stability, 160, 120-125.","Lim, H. A., Raku, T., & Tokiwa, Y. (2005). Hydrolysis of polyesters by serine proteases. Biotechnology letters, 27(7), 459-464."]}

La produzione e il consumo di plastica sono largamente cresciuti negli ultimi decenni. La maggior parte di queste plastiche derivano da fonti fossili e non sono degradabili, per cui tendono ad accumularsi nell’ambiente. L’uso di enzimi e di plastiche biodegradabili per il trattamento di rifiuti plastici si sta sviluppando in diversi settori, quali packaging, agricoltura e medicina. Tra le plastiche biodegradabili, i poliesteri alifatici risultano essere i più suscettibili all’attacco di enzimi idrolitici e microrganismi. Questo studio indaga la degradazione enzimatica di due polimeri commerciali biodegradabili, i.e. poli(butilene succinato-co-adipato) (PBSA) e poli(ε-caprolattone) (PCL) e il meccanismo d’azione degli enzimi cutinasi da Fusarium solani e lipasi da Alcaligene sp. (QLM). Il PBSA è un poliestere alifatico che deriva da fonti rinnovabili. Si tratta di un copolimero semicristallino del poli(butilene succinato) (PBS), con l’acido adipico come co-monomero. L’alta biodegradabilità deriva dalla bassa cristallinità e dalla maggiore flessibilità delle catene polimeriche rispetto al PBS. Il PCL è un poliestere alifatico semicristallino che deriva da fonti fossili. È largamente utilizzato nel settore biomedico e nel packaging per via della sua biodegradabilità e biocompatibilità. Lo studio del meccanismo di degradazione enzimatica di questi due polimeri può essere utile nella gestione dei rifiuti plastici, e nella formulazione di nuovi materiali plastici biodegradabili.

Commercial hydrolytic enzymes belonging to different subclasses (several lipases, proteinase k, cutinase) were investigated for their ability to degrade different aliphatic polyesters, i.e., poly(butylene succinate) (PBS), poly(butylene succinate-co-adipate) (PBSA), two poly(caprolactone), having two different molecular weights, poly(lactic acid) (PLA) and poly(propylene carbonate) (PPC). The enzyme screening was first carried out by investigating the capacity of fully degrading the target polymers in 24 h, then weight loss measurements of selected polyesters and target enzymes were performed. Solid residues after enzyme degradation were characterized by proton nuclear magnetic resonance (1H NMR), gel permeation chromatography (GPC), infrared spectroscopy (FT-IR), differential scanning calorimetry (DSC) and thermogravimetry (TGA). Liquid fractions were studied via GPC, 1H NMR and high-performance liquid chromatography (HPLC). PCL and PBSA were found to be the most biodegradable polyesters, under the conditions used in this study. PBS was fully degraded only by cutinase, whereas none of the tested enzymes were able to completely degrade PLA and PPC, in the conditions assessed here. Cutinase exhibited the highest hydrolytic activity on PBSA, while lipase from Candida sp. (CALB) on low molecular weight PCL. Chemical analyses on residual solids showed that the enzymatic degradation occurred homogeneously from the surface through an erosion mechanism and did not significantly affect the macromolecular structure and thermal stability. Cleaving action mode for each enzyme (endo- and/or exo-type) on the different polyesters were also proposed based on the evaluation of the degradation products in the liquid fraction.

Plastic debris dispersed into the environment provide a substrate for microbial colonization, constituting a new human-made ecosystem called "plastisphere", and altering the microbial species distribution in aquatic, coastal and benthic ecosystems. The study aims at exploring the interaction among microplastics (MPs) made of different polymers, a persistent organic contaminant (polychlorinated biphenyls, PCBs), and the environmental microbial communities, in an anoxic marine sediment. Plastic pellets were incubated in the field in a salt marsh anoxic sediment, to observe the stages of plastisphere formation, by quantitative PCR and 16S rRNA gene sequencing, and PCB dechlorination activity on the MPs surface. Microbes from the sediment rapidly colonized the different microplastics types, with PVC recruiting a peculiar community enriched in sulfate-reducing bacteria. The composition of the plastisphere varied along the 1-year incubation possibly in response either to warmer temperatures in spring-summer or to microhabitat's changes due to the progressive plastic surface weathering. Even if PCB contaminated MPs were able to recruit potentially dehalogenating taxa, actual dechlorination was not detectable after 1 year. This suggests that the concentration of potentially dehalorespiring bacteria in the natural environment could be too low for the onset of the dechlorination process on MP-sorbed contaminants. Our study, which is among very few available longitudinally exploring the plastisphere composition in an anoxic sediment context, is the first exploring the fate and possible biodegradation of persistent organic pollutants sorbed on MPs reaching the seafloor.