Attachment of Acidithiobacillus ferrooxidans to pyrite in fresh and saline water and its ﬁtting to Langmuir and Freundlich isotherms

Attachment of bacteria Acidithiobacillus ferrooxidans to pyrite was investigated in two different environment: fresh water and saline water (water with 35 g/L of NaCl), in both cases at pH 4. Adsorption isotherms were ﬁtted to the Langmuir and Freundlich models. The results showed that the bacteria adhere to pyrite to a greater extent in fresh water than in saline water. The Langmuir and Freundlich models ﬁtted well the data obtained in fresh water, showing a coeﬃcient of determination (R 2 ) approximately equal to 0.8 for both models. On the other hand, in saline water the models did not show a good coeﬃcient of determination with a value approximately equal to 0.4 for both models.


Introduction
Acidithiobacillus ferrooxidans is a chemolithoautotrophic bacterium that obtains the carbon from the CO 2 of the air, to generate their cellular mass, and the energy (electrons) via the oxidation of inorganic compounds (e.g. minerals). Due to its ability to oxidize ferrous and reduced sulfur compunds, A. ferrooxidans has been widely used in leaching when processing 5 copper oxides and copper sulfides (Donati et al., 2007;Rawlings, 2005). Moreover, A. ferrooxidans has been studied as pyrite depressant in the flotation process (Chandraprabha et al., 2004(Chandraprabha et al., , 2005Hosseini et al., 2005;Mehrabani et al., 2011;Misra et al., 1996;Nagaoka et al., 1999;Ohmura et al., 1993;San Martín et al., 2018). In general, bacteria tend to adhere to solid surfaces forming biofilms rather than remain free in solution as planktonic cells because they 10 find protection against hostile environments and a high concentration of nutrient (Flemming and Wingender, 2010;Schaechter, 2009;Chandraprabha and Natarajan, 2006).
In Chile most of the mines are located in the north of the country where arid conditions prevail. Due to the water scarcity in this region, mining companies have started to change the source of water that they currently use, from groundwater to seawater. The use of seawater is 15 becoming increasingly important in mining, especially in the leaching and flotation processes.
Langmuir isotherm is an adsorption model that has been used to determine qualitatively the degree of attachment of A. ferrooxidans to minerals (Xia et al., 2013). The model assumes that: (i) bacteria form a monolayer over the mineral, (ii) all sites on the surface are energetically equivalent, (iii) the attached bacteria do not interact with each other and (iv) the bacteria do not move on the surface. The Langmuir isotherm model is presented in Equation (1): Equation (1) can be easily linearized as follows: where, X e is the attached bacteria per gram of mineral (bacteria/g), C e is the equilibrium concentration of bacteria in solution (bacteria/mL), K L is the Langmuir adsorption equilibrium constant (mL/bacteria) and X m is the maximum adsorption capacity per unit mass of adsorbent (bacteria/g). X m and K L are fitted from experimental data.
Freundlich isotherm is an empirical model given by the Equation (3): Equation (3) can be written in the logarithmic form as follows: where, X e is the attached bacteria per gram of mineral (bacteria/g), C e is the equilibrium concentration of bacteria in solution (bacteria/mL), n indicates the adsorption effectiveness and represents the inherent properties of the adsorbent and K F is a constant related to the adsorption capacity ((bacteria/g)(mL/bacteria) 1/n ). K F and n are fitted from experimental 40 data.
Because the use of bacteria and seawater in mining is gaining attention, it is important to study the adsorption behavior to pyrite in a saline environment. The present work studies the kinetics of attachment of Acidithiobacillus ferrooxidans to pyrite in fresh water and in water with 35 g/L NaCl (saline water), which is the concentration of sodium chloride in seawater. In 45 addition, the attachment behavior will be modeled using Langmuir and Freundlich equations to determine which of these two models represent the real behavior.

Mineral
The pyrite used corresponds to hand picked mineral samples that were manually crushed.

50
The samples were dry screened between 37 µm-212 µm (mesh # 70 and # 400) and cleaned with 6N hydrochloric acid solution to remove the oxidized species from their surfaces. The purity of the pyrite was ascertained by X-ray diffraction and it was determined to be higher than 99%.

55
The bacteria used correspond to Acidithiobacillus ferrooxidans strain ATCC19859. The bacteria were grown at 30℃ in sterile basal medium containing 0.4 g/L of ammonium sulfate

85
The adsorption isotherms were obtained by plotting the data at equilibrium for each initial concentration of bacteria. Subsequently, the Langmuir and Freundlich adsorption isotherms models were fitted to the experimental results.   Figure 3 shows the linear fitting of isotherms obtained in fresh water and saline water to the Langmuir model, while, Figure 4 shows the linear fitting to Freundlich model. Adsorption isotherms were plotted with data of bacteria concentrations at the equilibrium from the attachment kinetics experiments. From the slope and the intersection with the Y axis, the constants of 110 both models were obtained. In Table 1 the values of the Langmuir and Freundlich constants are presented. It is observed that the two models showed a better coefficient of determination for the experimental data obtained in fresh water than in saline water. In fresh water both models presented a R 2 >0.8, while, in saline water, R 2 were approximately 0.4. Therefore, according to the Langmuir model, bacteria could attach as a monolayer to pyrite in fresh water. In saline 115 water is not possible to assume that because the coefficient of determination (R 2 ) was too low.

Adsorption isotherms
The Freundlich model is appropriate to predict the attachment behavior of bacteria to pyrite in fresh water but not in saline water. The value of X m from the Langmuir model, indicates the maximum adsorption capacity per unit mass of adsorbent (bacteria/g). This value was equal to 5 × 10 9 bacteria/g in fresh water and in saline water. However, due to the low determination 120 coefficient in saline water, the result in this case should not be considered. Figure 5 shows the adsorption isotherms and their adjustments to the Langmuir and Freundlich models. Is possible to observe that in fresh water the adsorption of A. ferrooxidans is higher than in saline water, showing higher attachment densities (number of attached bacteria per mass of mineral).

125
The attachment density of bacteria A. ferrooxidans to pyrite was higher in fresh water than in saline water (35 g/L NaCl), showing that not only the characteristics of the adsorbent and the absorbate influence the adherence, but also the characteristics of the environment where it is carried out the adsorption.
Freundlich and Langmuir models show to predict well the adhesion behavior of A. ferroox-130 idans to pyrite in fresh water, however, in saline water the models show a low coefficient of determination indicating that they do not predict the adhesion behavior in this type of solution.
The good fit of the experimental data to the Langmuir model suggests that the bacteria adhere as a monolayer on the surface of the pyrite when the solution correspond to fresh water.