# Adsorption of Heavy Metals from Low Concentration Solutions onto Dried _Chlamydomonas reinhardtii_
## 4 Discussion  
Heavy metals, such as Pb(II), Cr(III), and Cd(II), can be found at very low concentrations in the wastewaters of different productive sectors, e.g., ceramic manufacturing, ferrous metals processing, and surface treatment of metals and plastics [46; 47; 48]. The aim of this work is to understand the applicability of the microalga _C. reinhardtii_ to treat low concentrated wastewaters (0.5 mg/L Pb(II), 0.2 mg/L Cr(III) and Cd(II)), before reintroducing them into the environment, following European regulation and limitations [49]. Despite the observed cells damage due to the applied freeze-drying process, pristine _C. reinhardtii_ maintains its original metal ion adsorption capability. This result is not surprising when considering the dried biomass as a natural sorbent, consisting mainly of cellulose or proteins. Both biopolymers, are reported to be able to interact and capture metal ions [29; 35; 41]. The sorption capability is strictly connected to the residual functional groups present in the dried biomass as well as to the operating conditions, contact time, and pH adopted during the adsorption process. Two days of contact were found appropriate to reach the 65-80% (\(w/w\)) of metal capture depending on the considered metal ion. Apparently, the algae have a larger affinity for Pb(II) and Cd(II), respectively, 80% (\(w/w\)) and 76% (\(w/w\)) of adsorption, while only 65% (\(w/w\)) of Cr(III) is adsorbed. However, looking at \(\mu\)molion captured by 1 g of sorbent material, it is possible to observe that Cr(III) is the most captured ion, nominally 8.3 \(\mu\)mol\({}_{\rm Cr}\)/gmicrolgae, while Pb(II) and Cd(II) have, respectively, been adsorbed 6.5 \(\mu\)mol\({}_{\rm Pb}\)/gmicrolgae and 4.5 \(\mu\)mol\({}_{\rm Cd}\)/gmicrolgae. A prolonged contact time of 4 days did not influence the adsorption yields of Pb(II) and Cr(III), which remained almost constant, but it had a negative effect on Cd(II) adsorption, which decreased from 76% (\(w/w\)) down to 63% (\(w/w\)), thus from 4.5 \(\mu\)mol\({}_{\rm Cd}\)/gmicrolgae to 3.7 \(\mu\)mol\({}_{\rm Cd}\)/gmicrolgae, suggesting a reversibility in the adsorption-desorption reaction of Cd. Similar results were reported in the literature by [10], on HMs sorption by different algal strains (_Chorella vulgaris_ and _Spirulina maxima_) studied over 10 days. By evaluating the metal removal every 2 days, the authors showed that the adsorption equilibrium was dependent on the metal type, the alga strain, and the amount of the biosorbent used in the experiment. Regarding the observed pH effect, the affinity between the biosorbent surface and the ions species in the solution should be considered to explain the different behaviour. As a matter of fact, according to [PERSON] et al. [56], solution pH impacts two aspects: ion solubility and biosorbent total charge, which both affect the equilibrium of the system [57; 58]. Therefore, the influence of pH on the metal sorption is dependent on the functional groups of the biosorbent and on the presence of different metal species in the solution, thus governing the adsorption behaviour [59; 60]. To reach a deeper insight, chemical speciation in solution was calculated as a function of pH by means of the Hydra-Medusa software [50], the plots for the single-ion system are reported in Figure S7a-c. As a general consideration, at every pH, positive charged or neutral metal species are present; therefore, the interaction with the negative charged surface of the dried alga is favoured. Considering the complexity of the equilibria and species distribution of Figure S7a-c, in Figure 6 the possible main ion species present at pH 6 and 7 are summarized. At pH 6, the Pb ion is present in the stable form of Pb\({}^{2+}\), and in a negligible amount in the form of PbOH\({}^{+}\). The main species in solution of Cr-containing system are Cr\({}_{2}\)O\({}_{3}\) and CrOH\({}^{2+}\), accompanied by a limited amount of Cr(OH)\({}_{2}\)\({}^{+}\) and Cr(OH)\({}_{3}\). Cd metal, instead, is always present as Cd\({}^{2+}\).