In the present work, a remediation bioprocess based on the use of a native isolate of Chlorella vulgaris immobilized in an alginate matrix inside a polylactic acid (PLA) device is proposed. This microalga immobilized in alginate beads was previously shown to be useful for the reduction of several chemical and microbial contaminants present in the highly polluted water from the Matanza–Riachuelo watershed. However, these beads had a relatively short shelf life in the natural environment. To overcome this limitation, a 3D-printed PLA device was designed. PLA is a biocompatible and biodegradable material suitable for biotechnological applications. We used Erlenmeyers and stirred-tank bioreactors fed batch with Murashige Skoog (MS) culture medium or water from the Cildáñez stream (one of the water bodies of the aforementioned watershed) to estimate the growth kinetics parameters and the bioremediation capacity of immobilized-microalgal cells as an unconfined system (UcS) or a confined system (CfS) inside PLA devices on Cildáñez water. Although alga’s growth parameters were maximum in the UcS fed with MS medium as substrate, successful bioremediation of the target water was possible using the CfS: all inorganic nitrogen forms and total phosphorus were reduced at least by 90% after 5 days of bioprocess in an agitated bioreactor, whereas aerobic mesophilic bacteria decreased by about 85%. The number of coliforms also decreased. Standardized cytotoxicity tests using Allium cepa seeds carried out to prove the effectiveness of the bioremediation process, confirmed the high degree of decontamination achieved by the use of immobilized microalga confined in a 3D-printable PLA-device.

In the present work, a remediation bioprocess based on the use of a native isolate of Chlorella vulgaris immobilized in an alginate matrix inside a polylactic acid (PLA) device is proposed. This microalga immobilized in alginate beads was previously shown to be useful for the reduction of several chemical and microbial contaminants present in the highly polluted water from the Matanza-Riachuelo watershed. However, these beads had a relatively short shelf life in the natural environment. To overcome this limitation, a 3D-printed PLA device was designed. PLA is a biocompatible and biodegradable material suitable for biotechnological applications. We used Erlenmeyers and stirred-tank bioreactors fed batch with Murashige Skoog (MS) culture medium or water from the Cildáñez stream (one of the water bodies of the aforementioned watershed) to estimate the growth kinetics parameters and the bioremediation capacity of immobilized-microalgal cells as an unconfined system (UcS) or a confined system (CfS) inside PLA devices on Cildáñez water. Although alga’s growth parameters were maximum in the UcS fed with MS medium as substrate, successful bioremediation of the target water was possible using the CfS: all inorganic nitrogen forms and total phosphorus were reduced at least by 90% after 5 days of bioprocess in an agitated bioreactor, whereas aerobic mesophilic bacteria decreased by about 85%. The number of coliforms also decreased. Standardized cytotoxicity tests using Allium cepa seeds carried out to prove the effectiveness of the bioremediation process, confirmed the high degree of decontamination achieved by the use of immobilized microalga confined in a 3D-printable PLA-device.

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Extended AbstractA native strain of the microalgae Chlorella vugaris was shown to be effective for the reduction of several chemical and microbial contaminants present in polluted streams from the Matanza-Riachuelo Basin (M-R, 1). The microalgae immobilization in calcium alginate beads protects the cells from the toxic effects of numerous substances and from extreme pH and temperatures, allowing better survival and greater efficiency in the production of biomass [2,3]. However, cells trapped in the matrix are easy to be devoured by the fauna existent. Saladita Norte lagoon is part of M-R, this watercourse exhibits a wide range of contaminants levels along the year, mainly depending on the seasonal regime. At certain periods of the year, these levels usually exceed the limits established in international standards of water quality [4][5][6]. Similarly to that observed in other streams of M-R, several nitrogenated compounds, phosphorus, metals, and saprophytic and pathogenic bacteria are the main contaminants of Saladita Norte. In order to protect the alginate beads from the birds and fishes predation, a 3D-printed PLA (polylactic acid) device was designed. PLA is a biocompatible and biodegradable material that has been described as suitable for biotechnological applications. [7]. Bioprocesses carried out with this material must be checked for mechanical stability and optimal pore size to allow an effective bioremediation process. For this purpose, we used a stirredtank bioreactor supplied with a marine propeller and aeration during 6 days. Growth index and biomass production were compared between algae immobilized in alginate beads versus immobilized in alginate beads placed into a complex 3D printed PLA device.As previously observed, our native isolate of C. vulgaris could grow inside the alginate beads based on the consumption of several of the above-mentioned contaminants, such as phosphorus and nitrogenated ions [1]. The results obtained in control treatments with immobilized cells in alginate beads were: A-MS synthetic culture media supplemented with sucrose (3% w/v) and indolacetic acid (1 mg/L) as growth regulator [1] (=0.737, dt= 24 h) and B-as A-but growing in Saladita Norte water (=0.668, dt= 24 h). Also, treatments using 3D-device were: C-like B, but Chlorella cultures immobilized in alginate beads growing into a PLA3D scaffold coloured red (=0.341, dt= 2 d); D-like C, but coloured white (=0.206, dt= 3 d); and E-like C, but uncoloured (=0.342, dt= 2 d) were effective and successful to bioremediation in vitro. All cultures were kept at 24±2ºC, with a photoperiod of 16 h under PAR (above 400 μmol photon m-2 s-1). The data obtained were complemented with standardized cytotoxicity tests using Allium cepa and Lactuca sativa seeds. Germination rate and mitotic index of onion seeds and germination rate and length of lettuce seeds using polluted water or bioremediated water were compared. The results obtained demonstrate the potential of this technology to be used in integrated processes that seek removal ...